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Wood JN, Yan N, Huang J, Zhao J, Akopian A, Cox JJ, Woods CG, Nassar MA. Sensory neuron sodium channels as pain targets; from cocaine to Journavx (VX-548, suzetrigine). J Gen Physiol 2025; 157:e202513778. [PMID: 40294084 PMCID: PMC12036950 DOI: 10.1085/jgp.202513778] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2025] [Revised: 03/20/2025] [Accepted: 03/31/2025] [Indexed: 04/30/2025] Open
Abstract
Voltage-gated sodium channels underpin electrical signaling in sensory neurons. Their activity is an essential element in the vast majority of pain conditions, making them significant drug targets. Sensory neuron sodium channels play roles not only in afferent signaling but also in a range of efferent regulatory mechanisms. Side effects through actions on other cell types and efferent signaling are thus important issues to address during analgesic drug development. As an example, the human genetic evidence for NaV1.7 as an ideal pain target contrasts with the side effects of NaV1.7 antagonists. In this review, we describe the history and progress toward the development of useful analgesic drugs and the renewed focus on NaV1.8 as a key target in pain treatment. NaV1.8 antagonists alone or in combination with other analgesics are likely to provide new opportunities for pain relief for the vast number of people (about 33% of the population) impacted by chronic pain, particularly present in aging populations.
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Affiliation(s)
- John N. Wood
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, London, UK
| | - Nieng Yan
- Beijing Frontier Research Center for Biological Structures, State Key Laboratory of Membrane Biology, Tsinghua-Peking Joint Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China
- Institute of Bio-Architecture and Bio-Interactions (IBABI), Shenzhen Medical Academy of Research and Translation, Shenzhen, China
| | - Jian Huang
- Institute of Bio-Architecture and Bio-Interactions (IBABI), Shenzhen Medical Academy of Research and Translation, Shenzhen, China
| | - Jing Zhao
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, London, UK
| | - Armen Akopian
- Department of Endodontics, The School of the Dentistry, UTHSCSA, San Antonio, TX, USA
| | - James J. Cox
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, London, UK
| | | | - Mohammed A. Nassar
- School of Biosciences, Firth Court, University of Sheffield, Sheffield, UK
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2
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Allen HN, Hestehave S, Duran P, Nelson TS, Khanna R. Uncoupling the CRMP2-Ca V2.2 Interaction Reduces Pain-Like Behavior in a Preclinical Joint-Pain Model. THE JOURNAL OF PAIN 2024; 25:104664. [PMID: 39233208 PMCID: PMC11560641 DOI: 10.1016/j.jpain.2024.104664] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/06/2024] [Revised: 08/29/2024] [Accepted: 08/29/2024] [Indexed: 09/06/2024]
Abstract
Osteoarthritis (OA) represents a significant pain challenge globally, as current treatments are limited and come with substantial and adverse side effects. Voltage-gated calcium channels have proved to be pharmacologically effective targets, with multiple Food and Drug Administration-approved CaV2.2 modulators available for the treatment of pain. Although effective, drugs targeting CaV2.2 are complicated by the same obstacles facing other pain therapeutics-invasive routes of administration, narrow therapeutic windows, side effects, and addiction potential. We have identified a key regulator of CaV2.2 channels, collapsin response mediator protein 2, that allows us to indirectly regulate CaV2.2 expression and function. We previously developed a peptidomimetic modulator of collapsin response mediator protein 2, CBD3063, that effectively reverses neuropathic and inflammatory pain without negative side effects by reducing membrane expression of CaV2.2. The potent analgesic properties of CBD3063, combined with the lack of negative side effects, prompted us to assess the efficacy of CBD3063 in a rodent model of OA pain. Here, we demonstrate the intraperitoneal administration of CBD3063 alleviates both evoked and nonevoked behavioral hallmarks of OA pain. Further, we reveal that CBD3063 reduces OA-induced increased neural activity in the parabrachial nucleus, a key supraspinal site modulating the pain experience. Together, these studies suggest that CBD3063 is an effective analgesic for OA pain. PERSPECTIVE: Despite the high prevalence of OA pain worldwide, current treatment options remain limited. We demonstrate that CBD3063-mediated disruption of the CaV2.2-collapsin response mediator protein 2 interaction alleviates pain in a preclinical joint pain model, providing a promising basis for the development of new OA pain treatments.
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Affiliation(s)
- Heather N Allen
- Department of Pharmacology & Therapeutics, University of Florida College of Medicine, Gainesville, Florida
| | - Sara Hestehave
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York; Department of Experimental Medicine, University of Copenhagen, 2200 Copenhagen N, Denmark
| | - Paz Duran
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York
| | - Tyler S Nelson
- Department of Pharmacology & Therapeutics, University of Florida College of Medicine, Gainesville, Florida
| | - Rajesh Khanna
- Department of Pharmacology & Therapeutics, University of Florida College of Medicine, Gainesville, Florida.
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3
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Thio BJ, Titus ND, Pelot NA, Grill WM. Reverse-engineered models reveal differential membrane properties of autonomic and cutaneous unmyelinated fibers. PLoS Comput Biol 2024; 20:e1012475. [PMID: 39374306 PMCID: PMC11486378 DOI: 10.1371/journal.pcbi.1012475] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2024] [Revised: 10/17/2024] [Accepted: 09/11/2024] [Indexed: 10/09/2024] Open
Abstract
Unmyelinated C-fibers constitute the vast majority of axons in peripheral nerves and play key roles in homeostasis and signaling pain. However, little is known about their ion channel expression, which controls their firing properties. Also, because of their small diameters (~ 1 μm), it has not been possible to characterize their membrane properties using voltage clamp. We developed a novel library of isoform-specific ion channel models to serve as the basis functions of our C-fiber models. We then developed a particle swarm optimization (PSO) framework that used the isoform-specific ion channel models to reverse engineer C-fiber membrane properties from measured autonomic and cutaneous C-fiber conduction responses. Our C-fiber models reproduced experimental conduction velocity, chronaxie, action potential duration, intracellular threshold, and paired pulse recovery cycle. The models also matched experimental activity-dependent slowing, a property not included in model optimization. We found that simple conduction responses, characterizing the action potential, were controlled by similar membrane properties in both the autonomic and cutaneous C-fiber models, but complicated conduction response, characterizing the afterpotenials, were controlled by differential membrane properties. The unmyelinated C-fiber models constitute important tools to study autonomic signaling, assess the mechanisms of pain, and design bioelectronic devices. Additionally, the novel reverse engineering approach can be applied to generate models of other neurons where voltage clamp data are not available.
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Affiliation(s)
- Brandon J. Thio
- Department of Biomedical Engineering Duke University Durham, North Carolina, United States of America
| | - Nathan D. Titus
- Department of Biomedical Engineering Duke University Durham, North Carolina, United States of America
| | - Nicole A. Pelot
- Department of Biomedical Engineering Duke University Durham, North Carolina, United States of America
| | - Warren M. Grill
- Department of Biomedical Engineering Duke University Durham, North Carolina, United States of America
- Duke University, Department of Electrical and Computer Engineering, Durham, North Carolina, United States of America
- Duke University School of Medicine, Department of Neurobiology, Durham, North Carolina, United States of America
- Duke University School of Medicine, Department of Neurosurgery, Durham, North Carolina, United States of America
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4
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Perez-Miller S, Gomez K, Khanna R. Peptide and Peptidomimetic Inhibitors Targeting the Interaction of Collapsin Response Mediator Protein 2 with the N-Type Calcium Channel for Pain Relief. ACS Pharmacol Transl Sci 2024; 7:1916-1936. [PMID: 39022365 PMCID: PMC11249630 DOI: 10.1021/acsptsci.4c00181] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2024] [Revised: 05/13/2024] [Accepted: 05/23/2024] [Indexed: 07/20/2024]
Abstract
Ion channels serve pleiotropic functions. Often found in complexes, their activities and functions are sculpted by auxiliary proteins. We discovered that collapsin response mediator protein 2 (CRMP2) is a binding partner and regulator of the N-type voltage-gated calcium channel (CaV2.2), a genetically validated contributor to chronic pain. Herein, we trace the discovery of a new peptidomimetic modulator of this interaction, starting from the identification and development of CBD3, a CRMP2-derived CaV binding domain peptide. CBD3 uncouples CRMP2-CaV2.2 binding to decrease CaV2.2 surface localization and calcium currents. These changes occur at presynaptic sites of nociceptive neurons and indeed, CBD3 ameliorates chronic pain in preclinical models. In pursuit of a CBD3 peptidomimetic, we exploited a unique approach to identify a dipeptide with low conformational flexibility and high solvent accessibility that anchors binding to CaV2.2. From a pharmacophore screen, we obtained CBD3063, a small-molecule that recapitulated CBD3's activity, reversing nociceptive behaviors in rodents of both sexes without sensory, affective, or cognitive effects. By disrupting the CRMP2-CaV2.2 interaction, CBD3063 exerts these effects indirectly through modulating CaV2.2 trafficking, supporting CRMP2 as an auxiliary subunit of CaV2.2. The parent peptide CBD3 was also found by us and others to have neuroprotective properties at postsynaptic sites, through N-methyl-d-aspartate receptor and plasmalemmal Na+/Ca2+ exchanger 3, potentially acting as an auxiliary subunit for these pathways as well. Our new compound is poised to address several open questions regarding CRMP2's role in regulating the CaV2.2 pathways to treat pain with the potential added benefit of neuroprotection.
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Affiliation(s)
- Samantha Perez-Miller
- Department
of Pharmacology & Therapeutics, College of Medicine, University of Florida, 1200 Newell Drive, ARB R5-234, Gainesville, Florida 32610-0267, United States
| | - Kimberly Gomez
- Department
of Pharmacology & Therapeutics, College of Medicine, University of Florida, 1200 Newell Drive, ARB R5-234, Gainesville, Florida 32610-0267, United States
| | - Rajesh Khanna
- Department
of Pharmacology & Therapeutics, College of Medicine, University of Florida, 1200 Newell Drive, ARB R5-234, Gainesville, Florida 32610-0267, United States
- Pain
and Addiction Therapeutics (PATH) Collaboratory, College of Medicine, University of Florida, Gainesville, Florida 32610, United States
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Iseppon F, Kanellopoulos AH, Tian N, Zhou J, Caan G, Chiozzi R, Thalassinos K, Çubuk C, Lewis MJ, Cox JJ, Zhao J, Woods CG, Wood JN. Sodium channels Na v1.7, Na v1.8 and pain; two distinct mechanisms for Na v1.7 null analgesia. NEUROBIOLOGY OF PAIN (CAMBRIDGE, MASS.) 2024; 16:100168. [PMID: 39559752 PMCID: PMC11570969 DOI: 10.1016/j.ynpai.2024.100168] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 06/27/2024] [Revised: 10/03/2024] [Accepted: 10/03/2024] [Indexed: 11/20/2024]
Abstract
Genetic deletion and pharmacological inhibition are distinct approaches to unravelling pain mechanisms, identifying targets and developing new analgesics. Both approaches have been applied to the voltage-gated sodium channels Nav1.7 and Nav1.8. Genetic deletion of Nav1.8 in mice leads to a loss of pain and antagonists are effective analgesics. The situation with Nav1.7 is more complex. Complete embryonic loss of Nav1.7 in humans or in mouse sensory neurons leads to anosmia as well as profound analgesia as a result of diminished neurotransmitter release. This is mediated by enhanced endogenous opioid signaling in humans and mice. In contrast, anosmia is opioid-independent. Sensory neuron excitability and autonomic function appear to be normal. Adult deletion of Nav1.7 in sensory neurons also leads to analgesia, but through diminished sensory and autonomic neuron excitability. There is no opioid component of analgesia or anosmia as shown by a lack of effect of naloxone. Pharmacological inhibition of Nav1.7 in mice and humans leads both to analgesia and dramatic side-effects on the autonomic nervous system with no therapeutic window. These data demonstrate that specific Nav1.7 channel blockers will fail as analgesic drugs. The viability of embryonic null mutants suggests that there are compensatory changes to replace the lost Nav1.7 channel. Here we show that sensory neuron sodium channels Nav1.1, Nav1.2 and β4 subunits detected by Mass Spectrometry are upregulated in Nav1.7 embryonic null neurons and, together with other proteome changes, potentially compensate for the loss of Nav1.7. Interestingly, many of the upregulated proteins are known to interact with Nav1.7.
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Affiliation(s)
- Federico Iseppon
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
| | - Alexandros H. Kanellopoulos
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
| | - Naxi Tian
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
| | - Jun Zhou
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
| | - Gozde Caan
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
| | - Riccardo Chiozzi
- Institute of Structural and Molecular Biology, Division of Biosciences, University College London, London WC1E 6BT, UK
- University College London Mass Spectrometry Science Technology Platform, Division of Biosciences, University College London, London, UK
| | - Konstantinos Thalassinos
- Institute of Structural and Molecular Biology, Division of Biosciences, University College London, London WC1E 6BT, UK
- University College London Mass Spectrometry Science Technology Platform, Division of Biosciences, University College London, London, UK
| | - Cankut Çubuk
- Centre for Experimental Medicine and Rheumatology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London EC1M 6BQ, UK
| | - Myles J. Lewis
- Centre for Experimental Medicine and Rheumatology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London EC1M 6BQ, UK
| | - James J. Cox
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
| | - Jing Zhao
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
| | - Christopher G. Woods
- Cambridge Institute for Medical Research, Keith Peters Building, Biomedical Campus, Hills Rd, Cambridge CB2 0XY, UK
| | - John N. Wood
- Molecular Nociception Group, Wolfson Institute for Biomedical Research, UCL, Gower Street, London WC1E 6BT, UK
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6
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Allen HN, Hestehave S, Duran P, Nelson TS, Khanna R. Uncoupling the CRMP2-Ca V2.2 interaction reduces pain-like behavior in a preclinical osteoarthritis model. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.06.05.596514. [PMID: 38895294 PMCID: PMC11185632 DOI: 10.1101/2024.06.05.596514] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/21/2024]
Abstract
Osteoarthritis (OA) represents a significant pain challenge globally, as current treatments are limited and come with substantial and adverse side effects. Voltage-gated calcium channels have proved to be pharmacologically effective targets, with multiple FDA-approved CaV2.2 modulators available for the treatment of pain. Although effective, drugs targeting CaV2.2 are complicated by the same obstacles facing other pain therapeutics-invasive routes of administration, narrow therapeutic windows, side effects, and addiction potential. We have identified a key regulator of CaV2.2 channels, collapsing response mediator protein 2 (CRMP2), that allows us to indirectly regulate CaV2.2 expression and function. We developed a peptidomimetic modulator of CRMP2, CBD3063, that effectively reverses neuropathic and inflammatory pain without negative side effects by reducing membrane expression of CaV2.2. Using a rodent model of OA, we demonstrate the intraperitoneal administration of CBD3063 alleviates both evoked and non-evoked behavioral hallmarks of OA pain. Further, we reveal that CBD3063 reduces OA-induced increased neural activity in the parabrachial nucleus, a key supraspinal site modulating the pain experience. Together, these studies suggest CBD3063 is an effective analgesic for OA pain.
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Affiliation(s)
- Heather N. Allen
- Department of Pharmacology & Therapeutics, University of Florida College of Medicine, Gainesville, Florida 32610, USA
| | - Sara Hestehave
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York 10010, USA
| | - Paz Duran
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York 10010, USA
| | - Tyler S. Nelson
- Department of Pharmacology & Therapeutics, University of Florida College of Medicine, Gainesville, Florida 32610, USA
| | - Rajesh Khanna
- Department of Pharmacology & Therapeutics, University of Florida College of Medicine, Gainesville, Florida 32610, USA
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7
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Salib AMN, Crane MJ, Lee SH, Wainger BJ, Jamieson AM, Lipscombe D. Interleukin-1α links peripheral Ca V2.2 channel activation to rapid adaptive increases in heat sensitivity in skin. Sci Rep 2024; 14:9051. [PMID: 38643253 PMCID: PMC11032389 DOI: 10.1038/s41598-024-59424-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2023] [Accepted: 04/10/2024] [Indexed: 04/22/2024] Open
Abstract
Neurons have the unique capacity to adapt output in response to changes in their environment. Within seconds, sensory nerve endings can become hypersensitive to stimuli in response to potentially damaging events. The underlying behavioral response is well studied, but several of the key signaling molecules that mediate sensory hypersensitivity remain unknown. We previously discovered that peripheral voltage-gated CaV2.2 channels in nerve endings in skin are essential for the rapid, transient increase in sensitivity to heat, but not to mechanical stimuli, that accompanies intradermal capsaicin. Here we report that the cytokine interleukin-1α (IL-1α), an alarmin, is necessary and sufficient to trigger rapid heat and mechanical hypersensitivity in skin. Of 20 cytokines screened, only IL-1α was consistently detected in hind paw interstitial fluid in response to intradermal capsaicin and, similar to behavioral sensitivity to heat, IL-1α levels were also dependent on peripheral CaV2.2 channel activity. Neutralizing IL-1α in skin significantly reduced capsaicin-induced changes in hind paw sensitivity to radiant heat and mechanical stimulation. Intradermal IL-1α enhances behavioral responses to stimuli and, in culture, IL-1α enhances the responsiveness of Trpv1-expressing sensory neurons. Together, our data suggest that IL-1α is the key cytokine that underlies rapid and reversible neuroinflammatory responses in skin.
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Affiliation(s)
- Anne-Mary N Salib
- Department of Neuroscience, Carney Institute for Brain Science, Brown University, Providence, RI, 02912, USA
| | - Meredith J Crane
- Department of Molecular Microbiology and Immunology, Brown University, Providence, RI, 02912, USA
| | - Sang Hun Lee
- Department of Neurology, Massachusetts General Hospital, Boston, MA, 02114, USA
| | - Brian J Wainger
- Department of Neurology, Massachusetts General Hospital, Boston, MA, 02114, USA
| | - Amanda M Jamieson
- Department of Molecular Microbiology and Immunology, Brown University, Providence, RI, 02912, USA
| | - Diane Lipscombe
- Department of Neuroscience, Carney Institute for Brain Science, Brown University, Providence, RI, 02912, USA.
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8
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Salib AMN, Crane MJ, Lee SH, Wainger BJ, Jamieson AM, Lipscombe D. Interleukin-1α links peripheral Ca V2.2 channel activation to rapid adaptive increases in heat sensitivity in skin. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2023.12.17.572072. [PMID: 38585803 PMCID: PMC10996502 DOI: 10.1101/2023.12.17.572072] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 04/09/2024]
Abstract
Neurons have the unique capacity to adapt output in response to changes in their environment. Within seconds, sensory nerve endings can become hypersensitive to stimuli in response to potentially damaging events. The underlying behavioral response is well studied, but several of the key signaling molecules that mediate sensory hypersensitivity remain unknown. We previously discovered that peripheral voltage-gated CaV2.2 channels in nerve endings in skin are essential for the rapid, transient increase in sensitivity to heat, but not to mechanical stimuli, that accompanies intradermal capsaicin. Here we report that the cytokine interleukin-1α (IL-1α), an alarmin, is necessary and sufficient to trigger rapid heat and mechanical hypersensitivity in skin. Of 20 cytokines screened, only IL-1α was consistently detected in hind paw interstitial fluid in response to intradermal capsaicin and, similar to behavioral sensitivity to heat, IL-1α levels were also dependent on peripheral CaV2.2 channel activity. Neutralizing IL-1α in skin significantly reduced capsaicin-induced changes in hind paw sensitivity to radiant heat and mechanical stimulation. Intradermal IL-1α enhances behavioral responses to stimuli and, in culture, IL-1α enhances the responsiveness of Trpv1-expressing sensory neurons. Together, our data suggest that IL-1α is the key cytokine that underlies rapid and reversible neuroinflammatory responses in skin.
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Affiliation(s)
- Anne-Mary N Salib
- Department of Neuroscience, Carney Institute for Brain Science, Brown University, Providence, RI 02912, USA
| | - Meredith J Crane
- Department of Molecular Microbiology and Immunology, Brown University, Providence, RI 02912, USA
| | - Sang Hun Lee
- Department of Neurology, Massachusetts General Hospital, Boston, MA 02114, USA
| | - Brian J Wainger
- Department of Neurology, Massachusetts General Hospital, Boston, MA 02114, USA
| | - Amanda M Jamieson
- Department of Molecular Microbiology and Immunology, Brown University, Providence, RI 02912, USA
| | - Diane Lipscombe
- Department of Neuroscience, Carney Institute for Brain Science, Brown University, Providence, RI 02912, USA
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Loya-Lopez SI, Allen HN, Duran P, Calderon-Rivera A, Gomez K, Kumar U, Shields R, Zeng R, Dwivedi A, Saurabh S, Korczeniewska OA, Khanna R. Intranasal CRMP2-Ubc9 inhibitor regulates Na V 1.7 to alleviate trigeminal neuropathic pain. Pain 2024; 165:573-588. [PMID: 37751532 PMCID: PMC10922202 DOI: 10.1097/j.pain.0000000000003053] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2023] [Accepted: 07/25/2023] [Indexed: 09/28/2023]
Abstract
ABSTRACT Dysregulation of voltage-gated sodium Na V 1.7 channels in sensory neurons contributes to chronic pain conditions, including trigeminal neuropathic pain. We previously reported that chronic pain results in part from increased SUMOylation of collapsin response mediator protein 2 (CRMP2), leading to an increased CRMP2/Na V 1.7 interaction and increased functional activity of Na V 1.7. Targeting this feed-forward regulation, we developed compound 194 , which inhibits CRMP2 SUMOylation mediated by the SUMO-conjugating enzyme Ubc9. We further demonstrated that 194 effectively reduces the functional activity of Na V 1.7 channels in dorsal root ganglia neurons and alleviated inflammatory and neuropathic pain. Here, we used a comprehensive array of approaches, encompassing biochemical, pharmacological, genetic, electrophysiological, and behavioral analyses, to assess the functional implications of Na V 1.7 regulation by CRMP2 in trigeminal ganglia (TG) neurons. We confirmed the expression of Scn9a , Dpysl2 , and UBE2I within TG neurons. Furthermore, we found an interaction between CRMP2 and Na V 1.7, with CRMP2 being SUMOylated in these sensory ganglia. Disrupting CRMP2 SUMOylation with compound 194 uncoupled the CRMP2/Na V 1.7 interaction, impeded Na V 1.7 diffusion on the plasma membrane, and subsequently diminished Na V 1.7 activity. Compound 194 also led to a reduction in TG neuron excitability. Finally, when intranasally administered to rats with chronic constriction injury of the infraorbital nerve, 194 significantly decreased nociceptive behaviors. Collectively, our findings underscore the critical role of CRMP2 in regulating Na V 1.7 within TG neurons, emphasizing the importance of this indirect modulation in trigeminal neuropathic pain.
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Affiliation(s)
- Santiago I. Loya-Lopez
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York, United States of America
- NYU Pain Research Center, 433 First Avenue, New York, NY 10010, United States of America
| | - Heather N. Allen
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York, United States of America
- NYU Pain Research Center, 433 First Avenue, New York, NY 10010, United States of America
| | - Paz Duran
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York, United States of America
- NYU Pain Research Center, 433 First Avenue, New York, NY 10010, United States of America
| | - Aida Calderon-Rivera
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York, United States of America
- NYU Pain Research Center, 433 First Avenue, New York, NY 10010, United States of America
| | - Kimberly Gomez
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York, United States of America
- NYU Pain Research Center, 433 First Avenue, New York, NY 10010, United States of America
| | - Upasana Kumar
- Center for Orofacial Pain and Temporomandibular Disorders, Department of Diagnostic Sciences, Rutgers School of Dental Medicine, Newark, NJ 07101, United States of America
| | - Rory Shields
- Rutgers School of Graduate Studies, Newark Health Science Campus, Newark, NJ 07101, United States of America
| | - Rui Zeng
- Department of Chemistry, College of Arts and Sciences, New York University, 100 Washington Square East, New York, NY 10003, United States of America
| | - Akshat Dwivedi
- Department of Chemistry, College of Arts and Sciences, New York University, 100 Washington Square East, New York, NY 10003, United States of America
| | - Saumya Saurabh
- Department of Chemistry, College of Arts and Sciences, New York University, 100 Washington Square East, New York, NY 10003, United States of America
| | - Olga A. Korczeniewska
- Center for Orofacial Pain and Temporomandibular Disorders, Department of Diagnostic Sciences, Rutgers School of Dental Medicine, Newark, NJ 07101, United States of America
- Rutgers School of Graduate Studies, Newark Health Science Campus, Newark, NJ 07101, United States of America
| | - Rajesh Khanna
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, New York, United States of America
- NYU Pain Research Center, 433 First Avenue, New York, NY 10010, United States of America
- Department of Neuroscience and Physiology and Neuroscience Institute, School of Medicine, New York University, New York, NY, 10010, USA
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10
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Gomez K, Santiago U, Nelson TS, Allen HN, Calderon-Rivera A, Hestehave S, Rodríguez Palma EJ, Zhou Y, Duran P, Loya-Lopez S, Zhu E, Kumar U, Shields R, Koseli E, McKiver B, Giuvelis D, Zuo W, Inyang KE, Dorame A, Chefdeville A, Ran D, Perez-Miller S, Lu Y, Liu X, Handoko, Arora PS, Patek M, Moutal A, Khanna M, Hu H, Laumet G, King T, Wang J, Damaj MI, Korczeniewska OA, Camacho CJ, Khanna R. A peptidomimetic modulator of the Ca V2.2 N-type calcium channel for chronic pain. Proc Natl Acad Sci U S A 2023; 120:e2305215120. [PMID: 37972067 PMCID: PMC10666126 DOI: 10.1073/pnas.2305215120] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2023] [Accepted: 10/09/2023] [Indexed: 11/19/2023] Open
Abstract
Transmembrane Cav2.2 (N-type) voltage-gated calcium channels are genetically and pharmacologically validated, clinically relevant pain targets. Clinical block of Cav2.2 (e.g., with Prialt/Ziconotide) or indirect modulation [e.g., with gabapentinoids such as Gabapentin (GBP)] mitigates chronic pain but is encumbered by side effects and abuse liability. The cytosolic auxiliary subunit collapsin response mediator protein 2 (CRMP2) targets Cav2.2 to the sensory neuron membrane and regulates their function via an intrinsically disordered motif. A CRMP2-derived peptide (CBD3) uncouples the Cav2.2-CRMP2 interaction to inhibit calcium influx, transmitter release, and pain. We developed and applied a molecular dynamics approach to identify the A1R2 dipeptide in CBD3 as the anchoring Cav2.2 motif and designed pharmacophore models to screen 27 million compounds on the open-access server ZincPharmer. Of 200 curated hits, 77 compounds were assessed using depolarization-evoked calcium influx in rat dorsal root ganglion neurons. Nine small molecules were tested electrophysiologically, while one (CBD3063) was also evaluated biochemically and behaviorally. CBD3063 uncoupled Cav2.2 from CRMP2, reduced membrane Cav2.2 expression and Ca2+ currents, decreased neurotransmission, reduced fiber photometry-based calcium responses in response to mechanical stimulation, and reversed neuropathic and inflammatory pain across sexes in two different species without changes in sensory, sedative, depressive, and cognitive behaviors. CBD3063 is a selective, first-in-class, CRMP2-based peptidomimetic small molecule, which allosterically regulates Cav2.2 to achieve analgesia and pain relief without negative side effect profiles. In summary, CBD3063 could potentially be a more effective alternative to GBP for pain relief.
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Affiliation(s)
- Kimberly Gomez
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Ulises Santiago
- Department of Computational and Systems Biology, University of Pittsburgh, Pittsburgh, PA15261
| | - Tyler S. Nelson
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Heather N. Allen
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Aida Calderon-Rivera
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Sara Hestehave
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Erick J. Rodríguez Palma
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Yuan Zhou
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ85724
| | - Paz Duran
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Santiago Loya-Lopez
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Elaine Zhu
- Department of Anesthesiology, Perioperative Care and Pain Medicine, New York University Grossman School of Medicine, New York, NY10016
- Interdisciplinary Pain Research Program, New York University Langone Health, New York, NY10016
| | - Upasana Kumar
- Department of Diagnostic Sciences, Center for Orofacial Pain and Temporomandibular Disorders, Rutgers School of Dental Medicine, Newark, NJ07101
| | - Rory Shields
- Rutgers School of Graduate Studies, Newark Health Science Campus, Newark, NJ07101
| | - Eda Koseli
- Department of Pharmacology and Toxicology and Translational Research Initiative for Pain and Neuropathy, Virginia Commonwealth University, Richmond, VA23298
| | - Bryan McKiver
- Department of Pharmacology and Toxicology and Translational Research Initiative for Pain and Neuropathy, Virginia Commonwealth University, Richmond, VA23298
| | - Denise Giuvelis
- Department of Biomedical Sciences, College of Osteopathic Medicine, Center for Excellence in the Neurosciences, University of New England, Biddeford, ME04005
| | - Wanhong Zuo
- Department of Anesthesiology, Rutgers New Jersey Medical School, Newark, NJ07103
| | | | - Angie Dorame
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ85724
| | - Aude Chefdeville
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ85724
| | - Dongzhi Ran
- Department of Pharmacology, School of Pharmacy, Chongqing Medical University, Chongqing400016, China
| | - Samantha Perez-Miller
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Yi Lu
- Department of Pharmacology, School of Pharmacy, Chongqing Medical University, Chongqing400016, China
| | - Xia Liu
- Department of Pharmacology, School of Pharmacy, Chongqing Medical University, Chongqing400016, China
| | - Handoko
- Department of Chemistry, New York University, New York, NY10003
| | | | - Marcel Patek
- Bright Rock Path Limited Liability Company, Tucson, AZ85724
| | - Aubin Moutal
- Department of Pharmacology and Physiology, School of Medicine, St. Louis University, St. Louis, MO63104
| | - May Khanna
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
| | - Huijuan Hu
- Department of Anesthesiology, Rutgers New Jersey Medical School, Newark, NJ07103
| | - Geoffroy Laumet
- Department of Physiology, Michigan State University, East Lansing, MI48824
| | - Tamara King
- Department of Biomedical Sciences, College of Osteopathic Medicine, Center for Excellence in the Neurosciences, University of New England, Biddeford, ME04005
| | - Jing Wang
- Department of Anesthesiology, Perioperative Care and Pain Medicine, New York University Grossman School of Medicine, New York, NY10016
- Interdisciplinary Pain Research Program, New York University Langone Health, New York, NY10016
- Department of Neuroscience and Physiology and Neuroscience Institute, School of Medicine, New York University, New York, NY10010
| | - M. Imad Damaj
- Department of Pharmacology and Toxicology and Translational Research Initiative for Pain and Neuropathy, Virginia Commonwealth University, Richmond, VA23298
| | - Olga A. Korczeniewska
- Department of Diagnostic Sciences, Center for Orofacial Pain and Temporomandibular Disorders, Rutgers School of Dental Medicine, Newark, NJ07101
- Rutgers School of Graduate Studies, Newark Health Science Campus, Newark, NJ07101
| | - Carlos J. Camacho
- Department of Computational and Systems Biology, University of Pittsburgh, Pittsburgh, PA15261
| | - Rajesh Khanna
- Department of Molecular Pathobiology, College of Dentistry, New York University, New York, NY10010
- New York University Pain Research Center, New York, NY10010
- Department of Neuroscience and Physiology and Neuroscience Institute, School of Medicine, New York University, New York, NY10010
- Chemical, and Biomolecular Engineering Department, Tandon School of Engineering, New York University, New York City, NY11201
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11
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Feuer KL, Peng X, Yovo CK, Avramopoulos D. DPYSL2/CRMP2 isoform B knockout in human iPSC-derived glutamatergic neurons confirms its role in mTOR signaling and neurodevelopmental disorders. Mol Psychiatry 2023; 28:4353-4362. [PMID: 37479784 PMCID: PMC11138811 DOI: 10.1038/s41380-023-02186-w] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/05/2023] [Revised: 07/05/2023] [Accepted: 07/07/2023] [Indexed: 07/23/2023]
Abstract
The DPYSL2/CRMP2 gene encodes a microtubule-stabilizing protein crucial for neurogenesis and is associated with numerous psychiatric and neurodegenerative disorders including schizophrenia, bipolar disorder, and Alzheimer's disease. DPYSL2 generates multiple RNA and protein isoforms, but few studies have differentiated between them. We previously reported an association of a functional variant in the DPYSL2-B isoform with schizophrenia (SCZ) and demonstrated in HEK293 cells that this variant reduced the length of cellular projections and created transcriptomic changes that captured schizophrenia etiology by disrupting mTOR signaling-mediated regulation. In the present study, we follow up on these results by creating, to our knowledge, the first models of endogenous DPYSL2-B knockout in human induced pluripotent stem cells (iPSCs) and neurons. CRISPR/Cas9-faciliated knockout of DPYSL2-B in iPSCs followed by Ngn2-induced differentiation to glutamatergic neurons showed a reduction in DPYSL2-B/CRMP2-B RNA and protein with no observable impact on DPYSL2-A/CRMP2-A. The average length of dendrites in knockout neurons was reduced up to 58% compared to controls. Transcriptome analysis revealed disruptions in pathways highly relevant to psychiatric disease including mTOR signaling, cytoskeletal dynamics, immune function, calcium signaling, and cholesterol biosynthesis. We also observed a significant enrichment of the differentially expressed genes in SCZ-associated loci from genome-wide association studies (GWAS). Our findings expand our previous results to neuronal cells, clarify the functions of the human DPYSL2-B isoform and confirm its involvement in molecular pathologies shared between many psychiatric diseases.
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Affiliation(s)
- Kyra L Feuer
- Predoctoral Training Program in Human Genetics, McKusick-Nathans Department of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, MD, USA
- McKusick-Nathans Department of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, MD, USA
| | - Xi Peng
- McKusick-Nathans Department of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, MD, USA
| | - Christian K Yovo
- McKusick-Nathans Department of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, MD, USA
| | - Dimitrios Avramopoulos
- McKusick-Nathans Department of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, MD, USA.
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12
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Kioutchoukova IP, Foster DT, Thakkar RN, Foreman MA, Burgess BJ, Toms RM, Molina Valero EE, Lucke-Wold B. Neurologic orphan diseases: Emerging innovations and role for genetic treatments. World J Exp Med 2023; 13:59-74. [PMID: 37767543 PMCID: PMC10520757 DOI: 10.5493/wjem.v13.i4.59] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/17/2023] [Revised: 07/16/2023] [Accepted: 08/11/2023] [Indexed: 09/15/2023] Open
Abstract
Orphan diseases are rare diseases that affect less than 200000 individuals within the United States. Most orphan diseases are of neurologic and genetic origin. With the current advances in technology, more funding has been devoted to developing therapeutic agents for patients with these conditions. In our review, we highlight emerging options for patients with neurologic orphan diseases, specifically including diseases resulting in muscular deterioration, epilepsy, seizures, neurodegenerative movement disorders, inhibited cognitive development, neuron deterioration, and tumors. After extensive literature review, gene therapy offers a promising route for the treatment of neurologic orphan diseases. The use of clustered regularly interspaced palindromic repeats/Cas9 has demonstrated positive results in experiments investigating its role in several diseases. Additionally, the use of adeno-associated viral vectors has shown improvement in survival, motor function, and developmental milestones, while also demonstrating reversal of sensory ataxia and cardiomyopathy in Friedreich ataxia patients. Antisense oligonucleotides have also been used in some neurologic orphan diseases with positive outcomes. Mammalian target of rapamycin inhibitors are currently being investigated and have reduced abnormal cell growth, proliferation, and angiogenesis. Emerging innovations and the role of genetic treatments open a new window of opportunity for the treatment of neurologic orphan diseases.
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Affiliation(s)
| | - Devon T Foster
- Florida International University Herbert Wertheim College of Medicine, Florida International University Herbert Wertheim College of Medicine, Miami, FL 33199, United States
| | - Rajvi N Thakkar
- College of Medicine, University of Florida, Gainesville, FL 32611, United States
| | - Marco A Foreman
- College of Medicine, University of Florida, Gainesville, FL 32611, United States
| | - Brandon J Burgess
- College of Medicine, University of Florida, Gainesville, FL 32611, United States
| | - Rebecca M Toms
- College of Medicine, University of Florida, Gainesville, FL 32611, United States
| | | | - Brandon Lucke-Wold
- Department of Neurosurgery, University of Florida, Gainesville, FL 32611, United States
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13
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Loya-Lopez SI, Allen HN, Duran P, Calderon-Rivera A, Gomez K, Kumar U, Shields R, Zeng R, Dwivedi A, Saurabh S, Korczeniewska OA, Khanna R. Intranasal CRMP2-Ubc9 Inhibitor Regulates Na V 1.7 to Alleviate Trigeminal Neuropathic Pain. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.07.16.549195. [PMID: 37502910 PMCID: PMC10370107 DOI: 10.1101/2023.07.16.549195] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/29/2023]
Abstract
Dysregulation of voltage-gated sodium Na V 1.7 channels in sensory neurons contributes to chronic pain conditions, including trigeminal neuropathic pain. We previously reported that chronic pain results in part from increased SUMOylation of collapsin response mediator protein 2 (CRMP2), leading to an increased CRMP2/Na V 1.7 interaction and increased functional activity of Na V 1.7. Targeting this feed-forward regulation, we developed compound 194 , which inhibits CRMP2 SUMOylation mediated by the SUMO-conjugating enzyme Ubc9. We further demonstrated that 194 effectively reduces the functional activity of Na V 1.7 channels in dorsal root ganglia neurons and alleviated inflammatory and neuropathic pain. Here, we employed a comprehensive array of investigative approaches, encompassing biochemical, pharmacological, genetic, electrophysiological, and behavioral analyses, to assess the functional implications of Na V 1.7 regulation by CRMP2 in trigeminal ganglia (TG) neurons. We confirmed the expression of Scn9a , Dpysl2 , and UBE2I within TG neurons. Furthermore, we found an interaction between CRMP2 and Na V 1.7, with CRMP2 being SUMOylated in these sensory ganglia. Disrupting CRMP2 SUMOylation with compound 194 uncoupled the CRMP2/Na V 1.7 interaction, impeded Na V 1.7 diffusion on the plasma membrane, and subsequently diminished Na V 1.7 activity. Compound 194 also led to a reduction in TG neuron excitability. Finally, when intranasally administered to rats with chronic constriction injury of the infraorbital nerve (CCI-ION), 194 significantly decreased nociceptive behaviors. Collectively, our findings underscore the critical role of CRMP2 in regulating Na V 1.7 within TG neurons, emphasizing the importance of this indirect modulation in trigeminal neuropathic pain.
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14
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Brustovetsky T, Khanna R, Brustovetsky N. CRMP2 Participates in Regulating Mitochondrial Morphology and Motility in Alzheimer's Disease. Cells 2023; 12:cells12091287. [PMID: 37174687 PMCID: PMC10177167 DOI: 10.3390/cells12091287] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2023] [Revised: 04/05/2023] [Accepted: 04/27/2023] [Indexed: 05/15/2023] Open
Abstract
Mitochondrial bioenergetics and dynamics (alterations in morphology and motility of mitochondria) play critical roles in neuronal reactions to varying energy requirements in health and disease. In Alzheimer's disease (AD), mitochondria undergo excessive fission and become less motile. The mechanisms leading to these alterations are not completely clear. Here, we show that collapsin response mediator protein 2 (CRMP2) is hyperphosphorylated in AD and that is accompanied by a decreased interaction of CRMP2 with Drp1, Miro 2, and Mitofusin 2, which are proteins involved in regulating mitochondrial morphology and motility. CRMP2 was hyperphosphorylated in postmortem brain tissues of AD patients, in brain lysates, and in cultured cortical neurons from the double transgenic APP/PS1 mice, an AD mouse model. CRMP2 hyperphosphorylation and dissociation from its binding partners correlated with increased Drp1 recruitment to mitochondria, augmented mitochondrial fragmentation, and reduced mitochondrial motility. (S)-lacosamide ((S)-LCM), a small molecule that binds to CRMP2, decreased its phosphorylation at Ser 522 and Thr 509/514, and restored CRMP2's interaction with Miro 2, Drp1, and Mitofusin 2. This was paralleled by decreased Drp1 recruitment to mitochondria, diminished mitochondrial fragmentation, and improved motility of the organelles. Additionally, (S)-LCM-protected cultured cortical AD neurons from cell death. Thus, our data suggest that CRMP2, in a phosphorylation-dependent manner, participates in the regulation of mitochondrial morphology and motility, and modulates neuronal survival in AD.
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Affiliation(s)
- Tatiana Brustovetsky
- Department of Pharmacology and Toxicology, Indiana University School of Medicine, 635 Barnhill Drive, Medical Science Building, Room 362, Indianapolis, IN 46202, USA
| | - Rajesh Khanna
- Department of Molecular Pathobiology, New York University, New York, NY 10010, USA
- College of Dentistry, NYU Pain Research Center, New York University, New York, NY 10010, USA
- Department of Neuroscience and Physiology and Neuroscience Institute, School of Medicine, New York University, New York, NY 10010, USA
| | - Nickolay Brustovetsky
- Department of Pharmacology and Toxicology, Indiana University School of Medicine, 635 Barnhill Drive, Medical Science Building, Room 362, Indianapolis, IN 46202, USA
- Stark Neurosciences Research Institute, Indiana University School of Medicine, Indianapolis, IN 46202, USA
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15
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Ovsepian SV, Waxman SG. Gene therapy for chronic pain: emerging opportunities in target-rich peripheral nociceptors. Nat Rev Neurosci 2023; 24:252-265. [PMID: 36658346 DOI: 10.1038/s41583-022-00673-7] [Citation(s) in RCA: 10] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/22/2022] [Indexed: 01/20/2023]
Abstract
With sweeping advances in precision delivery systems and manipulation of the genomes and transcriptomes of various cell types, medical biotechnology offers unprecedented selectivity for and control of a wide variety of biological processes, forging new opportunities for therapeutic interventions. This perspective summarizes state-of-the-art gene therapies enabled by recent innovations, with an emphasis on the expanding universe of molecular targets that govern the activity and function of primary sensory neurons and which might be exploited to effectively treat chronic pain.
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Affiliation(s)
- Saak V Ovsepian
- School of Science, Faculty of Engineering and Science, University of Greenwich London, Chatham Maritime, UK.
| | - Stephen G Waxman
- Department of Neurology, Yale University School of Medicine, New Haven, CT, USA.
- Department of Neuroscience, Yale University School of Medicine, New Haven, CT, USA.
- Department of Pharmacology, Yale University School of Medicine, New Haven, CT, USA.
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16
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Zong P, Yue L. Regulation of Presynaptic Calcium Channels. ADVANCES IN NEUROBIOLOGY 2023; 33:171-202. [PMID: 37615867 DOI: 10.1007/978-3-031-34229-5_7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/25/2023]
Abstract
Voltage-gated calcium channels (VGCCs), especially Cav2.1 and Cav2.2, are the major mediators of Ca2+ influx at the presynaptic membrane in response to neuron excitation, thereby exerting a predominant control on synaptic transmission. To guarantee the timely and precise release of neurotransmitters at synapses, the activity of presynaptic VGCCs is tightly regulated by a variety of factors, including auxiliary subunits, membrane potential, G protein-coupled receptors (GPCRs), calmodulin (CaM), Ca2+-binding proteins (CaBP), protein kinases, various interacting proteins, alternative splicing events, and genetic variations.
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Affiliation(s)
- Pengyu Zong
- Department of Cell Biology, Calhoun Cardiology Center, University of Connecticut School of Medicine, Farmington, CT, USA
| | - Lixia Yue
- Department of Cell Biology, Calhoun Cardiology Center, University of Connecticut School of Medicine, Farmington, CT, USA.
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17
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Dupree JL, Paez PM, Tiwari-Woodruff SK, Denton TT, Hensley K, Angeliu CG, Boullerne AI, Kalinin S, Egge S, Cheli VT, Denaroso G, Atkinson KC, Feri M, Feinstein DL. Lanthionine Ketimine Ethyl Ester Accelerates Remyelination in a Mouse Model of Multiple Sclerosis. ASN Neuro 2022; 14:17590914221112352. [PMID: 35791633 PMCID: PMC9272172 DOI: 10.1177/17590914221112352] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
Although over 20 disease modifying therapies are approved to treat Multiple Sclerosis (MS), these do not increase remyelination of demyelinated axons or mitigate axon damage. Previous studies showed that lanthionine ketenamine ethyl ester (LKE) reduces clinical signs in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS and increased maturation of oligodendrocyte (OL) progenitor cells (OPCs) in vitro. In the current study, we used the cuprizone (CPZ) demyelination model of MS to test if LKE could increase remyelination. The corpus callosum (CC) and somatosensory cortex was examined by immunohistochemistry (IHC), electron microscopy and for mRNA expression changes in mice provided 5 weeks of CPZ diet followed by 2 weeks of normal diet in the presence of LKE or vehicle. A significant increase in the number of myelinated axons, and increased myelin thickness was observed in the CC of LKE-treated groups compared to vehicle-treated groups. LKE also increased myelin basic protein and proteolipid protein expression in the CC and cortex, and increased the number of mature OLs in the cortex. In contrast, LKE did not increase the percentage of proliferating OPCs suggesting effects on OPC survival and differentiation but not proliferation. The effects of LKE on OL maturation and remyelination were supported by similar changes in their relative mRNA levels. Interestingly, LKE did not have significant effects on GFAP or Iba1 immunostaining or mRNA levels. These findings suggest that remyelinating actions of LKE can potentially be formulated to induce remyelination in neurological diseases associated with demyelination including MS.
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Affiliation(s)
- Jeffrey L. Dupree
- Department of Anatomy and Neurobiology, Virginia Commonwealth University, Richmond, VA, USA,Research Service, HH McGuire VA Medical Center, Richmond, VA, USA
| | - Pablo M. Paez
- Institute for Myelin and Glia Exploration, Department of Pharmacology and
Toxicology, University at Buffalo, NY, USA
| | - Seema K. Tiwari-Woodruff
- Division of Biomedical Sciences, School of Medicine at the University of California
Riverside, Riverside, CA, USA
| | - Travis T. Denton
- Department of Pharmaceutical Sciences, College of Pharmacy &
Pharmaceutical Sciences, Washington State University Health Sciences Spokane, Spokane, WA,
USA,Department of Translational Medicine and Physiology, Elson S. Floyd College
of Medicine, Washington State University Health Sciences Spokane, Spokane, WA, USA,Steve Gleason Institute for Neuroscience, Washington State University Health Sciences
Spokane, Spokane, WA, USA
| | - Kenneth Hensley
- Arkansas College of Osteopathic
Medicine, Fort Smith, AR, USA
| | - Christina G. Angeliu
- Institute for Myelin and Glia Exploration, Department of Pharmacology and
Toxicology, University at Buffalo, NY, USA
| | | | - Sergey Kalinin
- Department Anesthesiology, University of Illinois, Chicago, IL, USA
| | - Sophia Egge
- Department of Anatomy and Neurobiology, Virginia Commonwealth University, Richmond, VA, USA
| | - Veronica T. Cheli
- Institute for Myelin and Glia Exploration, Department of Pharmacology and
Toxicology, University at Buffalo, NY, USA
| | - Giancarlo Denaroso
- Institute for Myelin and Glia Exploration, Department of Pharmacology and
Toxicology, University at Buffalo, NY, USA
| | - Kelley C. Atkinson
- Division of Biomedical Sciences, School of Medicine at the University of California
Riverside, Riverside, CA, USA
| | - Micah Feri
- Division of Biomedical Sciences, School of Medicine at the University of California
Riverside, Riverside, CA, USA
| | - Douglas L. Feinstein
- Department Anesthesiology, University of Illinois, Chicago, IL, USA,Jesse Brown VA Medical Center, Chicago, IL, USA,Douglas L. Feinstein, Department of Anesthesiology,
University of Illinois, 835 South Wolcott Avenue, MC 513, Chicago IL, 60612, USA.
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18
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IQM-PC332, a Novel DREAM Ligand with Antinociceptive Effect on Peripheral Nerve Injury-Induced Pain. Int J Mol Sci 2022; 23:ijms23042142. [PMID: 35216258 PMCID: PMC8876042 DOI: 10.3390/ijms23042142] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2021] [Revised: 02/07/2022] [Accepted: 02/12/2022] [Indexed: 02/04/2023] Open
Abstract
Neuropathic pain is a form of chronic pain arising from damage of the neural cells that sense, transmit or process sensory information. Given its growing prevalence and common refractoriness to conventional analgesics, the development of new drugs with pain relief effects constitutes a prominent clinical need. In this respect, drugs that reduce activity of sensory neurons by modulating ion channels hold the promise to become effective analgesics. Here, we evaluated the mechanical antinociceptive effect of IQM-PC332, a novel ligand of the multifunctional protein downstream regulatory element antagonist modulator (DREAM) in rats subjected to chronic constriction injury of the sciatic nerve as a model of neuropathic pain. IQM-PC332 administered by intraplantar (0.01–10 µg) or intraperitoneal (0.02–1 µg/kg) injection reduced mechanical sensitivity by ≈100% of the maximum possible effect, with ED50 of 0.27 ± 0.05 µg and 0.09 ± 0.01 µg/kg, respectively. Perforated-patch whole-cell recordings in isolated dorsal root ganglion (DRG) neurons showed that IQM-PC332 (1 and 10 µM) reduced ionic currents through voltage-gated K+ channels responsible for A-type potassium currents, low, T-type, and high voltage-activated Ca2+ channels, and transient receptor potential vanilloid-1 (TRPV1) channels. Furthermore, IQM-PC332 (1 µM) reduced electrically evoked action potentials in DRG neurons from neuropathic animals. It is suggested that by modulating multiple DREAM–ion channel signaling complexes, IQM-PC332 may serve a lead compound of novel multimodal analgesics.
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19
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Boinon L, Yu J, Madura CL, Chefdeville A, Feinstein DL, Moutal A, Khanna R. Conditional knockout of CRMP2 in neurons, but not astrocytes, disrupts spinal nociceptive neurotransmission to control the initiation and maintenance of chronic neuropathic pain. Pain 2022; 163:e368-e381. [PMID: 35029600 PMCID: PMC8760468 DOI: 10.1097/j.pain.0000000000002344] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2021] [Accepted: 05/13/2021] [Indexed: 02/03/2023]
Abstract
ABSTRACT Mechanistic studies principally focusing on primary afferent nociceptive neurons uncovered the upregulation of collapsin response mediator protein 2 (CRMP2)-a dual trafficking regulator of N-type voltage-gated calcium (Cav2.2) as well as Nav1.7 voltage-gated sodium channels-as a potential determinant of neuropathic pain. Whether CRMP2 contributes to aberrant excitatory synaptic transmission underlying neuropathic pain processing after peripheral nerve injury is unknown. Here, we interrogated CRMP2's role in synaptic transmission and in the initiation or maintenance of chronic pain. In rats, short-interfering RNA-mediated knockdown of CRMP2 in the spinal cord reduced the frequency and amplitude of spontaneous excitatory postsynaptic currents, but not spontaneous inhibitory postsynaptic currents, recorded from superficial dorsal horn neurons in acute spinal cord slices. No effect was observed on miniature excitatory postsynaptic currents and inhibitory postsynaptic currents. In a complementary targeted approach, conditional knockout of CRMP2 from mouse neurons using a calcium/calmodulin-dependent protein kinase II alpha promoter to drive Cre recombinase expression reduced the frequency and amplitude of spontaneous excitatory postsynaptic currents, but not miniature excitatory SCss. Conditional knockout of CRMP2 from mouse astrocytes using a glial fibrillary acidic protein promoter had no effect on synaptic transmission. Conditional knockout of CRMP2 in neurons reversed established mechanical allodynia induced by a spared nerve injury in both male and female mice. In addition, the development of spared nerve injury-induced allodynia was also prevented in these mice. Our data strongly suggest that CRMP2 is a key regulator of glutamatergic neurotransmission driving pain signaling and that it contributes to the transition of physiological pain into pathological pain.
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Affiliation(s)
- Lisa Boinon
- Department of Pharmacology, College of Medicine, the University of Arizona, Tucson, Arizona 85724 United States of America
| | - Jie Yu
- Department of Pharmacology, College of Medicine, the University of Arizona, Tucson, Arizona 85724 United States of America
| | - Cynthia L. Madura
- Department of Pharmacology, College of Medicine, the University of Arizona, Tucson, Arizona 85724 United States of America
| | - Aude Chefdeville
- Department of Pharmacology, College of Medicine, the University of Arizona, Tucson, Arizona 85724 United States of America
| | - Douglas L. Feinstein
- Department of Anesthesiology, University of Illinois, Chicago, Chicago, Illinois 60612, United States of America
- Jesse Brown VA Medical Center, Chicago, Illinois, 60612, United States of America
| | - Aubin Moutal
- Department of Pharmacology, College of Medicine, the University of Arizona, Tucson, Arizona 85724 United States of America
| | - Rajesh Khanna
- Department of Pharmacology, College of Medicine, the University of Arizona, Tucson, Arizona 85724 United States of America
- Comprehensive Pain and Addiction Center, The University of Arizona, Tucson, Arizona, 85724, United States of America
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20
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Jiang YP, Wang S, Lai WD, Wu XQ, Jin Y, Xu ZH, Moutal A, Khanna R, Park KD, Shan ZM, Wen CP, Yu J. Neuronal CRMP2 phosphorylation inhibition by the flavonoid, naringenin, contributes to the reversal of spinal sensitization and arthritic pain improvement. Arthritis Res Ther 2022; 24:277. [PMID: 36564853 PMCID: PMC9783725 DOI: 10.1186/s13075-022-02975-8] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2022] [Accepted: 12/10/2022] [Indexed: 12/25/2022] Open
Abstract
BACKGROUND Rheumatoid arthritis patients usually suffer from arthritic chronic pain. However, due to an incomplete understanding of the mechanisms underlying autoimmune disorders, the management of arthritic pain is unsatisfactory. Here, we investigated the analgesic effect and underlying mechanism of the natural flavonoid naringenin (NAR) in collagen-induced arthritis (CIA) pain. METHODS NAR was injected (i.p.) once per day for 42 days after initial immunization, and rats were sacrificed on the 28th (the 21st day after final immunization, PID 21) and 42nd days (PID 35). The inflammatory factors, central sensitization indicators, and CRMP2 phosphorylation, as well as the anti-rheumatoid activity and analgesic effect of NAR, were further investigated. RESULTS We found that NAR decreased the arthritis score and paw swelling, as well as the mechanical and thermal pain. The immunofluorescence results also showed a dose dependent effect of NAR on reducing the expressions of spinal cFos, IBA-1, and GFAP on the 28th (PID 21) and 42nd day (PID 35). NAR decreased the phosphorylation of CRMP2 S522 and the expression of the kinase CDK5 in the spinal dorsal horn, but pCRMP2 Y479 was unchanged. In addition, CRMP2 was co-localized with NEUN, but not IBA-1 or GFAP, indicating the involvement of neural CRMP2 phosphorylation in CIA-related pain. Finally, CRMP2 S522 phosphorylation selective inhibitor (S)-lacosamide also alleviated arthritic pain. CONCLUSIONS Taken together, our results demonstrate that NAR alleviates inflammation and chronic pain in CIA model, which might be related to its inhibition of neuronal CRMP2 S522 phosphorylation, potentially mitigating the central sensitization. Our study provide evidence for the potential use of NAR as non-opioid-dependent analgesia in arthritic pain.
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Affiliation(s)
- Yue-Peng Jiang
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
| | - Song Wang
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
| | - Wei-Dong Lai
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
| | - Xue-Qing Wu
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
| | - Yan Jin
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
| | - Zheng-Hao Xu
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
| | - Aubin Moutal
- grid.262962.b0000 0004 1936 9342Department of Pharmacology and Physiology, Saint Louis University - School of Medicine, Saint Louis, MO 63104 USA
| | - Rajesh Khanna
- grid.137628.90000 0004 1936 8753Department of Molecular Pathobiology, College of Dentistry, and NYU Pain Research Center, New York University, New York, 10010 USA
| | - Ki Duk Park
- grid.35541.360000000121053345Korea Institute of Science and Technology, Seoul, South Korea
| | - Zhi-Ming Shan
- grid.440218.b0000 0004 1759 7210Department of Anesthesiology, Shenzhen People’s Hospital (The First Affiliated Hospital, Southern University of Science and Technology, The Second Clinical Medical College, Jinan University), Shenzhen, 518020 China
| | - Cheng-Ping Wen
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
| | - Jie Yu
- grid.268505.c0000 0000 8744 8924College of Basic Medical Science, College of Pharmaceutical Science, Key Laboratory of Neuropharmacology and Translational Medicine of Zhejiang Province, Zhejiang Chinese Medical University, Hangzhou, 310058 China
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21
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Wei Y, Wang G, Chen J, Xiao L, Wu Z, He J, Zhang N. Maternal deprivation induces cytoskeletal alterations and depressive-like behavior in adult male rats by regulating the AKT/GSK3β/CRMP2 signaling pathway. Physiol Behav 2021; 242:113625. [PMID: 34666114 DOI: 10.1016/j.physbeh.2021.113625] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2020] [Revised: 08/21/2021] [Accepted: 10/13/2021] [Indexed: 12/25/2022]
Abstract
Early-life adverse events exert persistent effects on brain functions and may increase the risk of psychopathology in adulthood. However, the underlying mechanism remains unclear. The purpose of our study was to study the long-lasting effects of maternal deprivation (MD) on depression-related behaviors and microtubule dynamics, and to illuminate the underlying molecular mechanism. Rat pups were separated from the dams for 360 min (MD) or 15 min (brief maternal separation) each day from postnatal day 4 through 10. Rats with MD experience showed significant depressive-like behaviors in adulthood, while brief maternal separation did not alter the behaviors. Behavioral alterations in the MD group were accompanied by alterations in the AKT/GSK3β/CRMP2 signaling pathway and hyperphosphorylation of CRMP2. CRMP2 interacted and colocalized with the cytoskeleton in the hippocampus, and the overlap of CRMP2 and tubulin staining in the hippocampus of MD rats was decreased. In MD rats, the expression of the α-tubulin isoforms Acet-tubulin and Tyr-tubulin changed, and the ratio of Tyr/Acet-tubulin, which is an important marker of microtubule dynamics, was decreased, indicating decreased microtubule dynamics. Furthermore, regulation of the AKT/GSK3β/CRMP2 signaling pathway by an LY294002 microinjection in the hippocampus resulted in cytoskeletal alterations and depressive-like behaviors in rats. These findings suggest that early-life MD induces depressive-like behaviors and cytoskeletal alterations in adult male rats and that the effects may be partly mediated by the AKT/GSK3β/CRMP2 signaling pathway. An understanding of the mechanism underlying the effect of MD on behaviors is crucial for developing pharmacological and psychological interventions for childhood neglect.
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Affiliation(s)
- Yanyan Wei
- Beijing Hui-Long-Guan Hospital, Peking University, Beijing, 100096, China
| | - Gaohua Wang
- Department of Psychiatry, Renmin Hospital of Wuhan University, Jiefang Road 238#, Wuhan 430060, Hubei, PR China.
| | - Jingxu Chen
- Beijing Hui-Long-Guan Hospital, Peking University, Beijing, 100096, China
| | - Ling Xiao
- Department of Psychiatry, Renmin Hospital of Wuhan University, Jiefang Road 238#, Wuhan 430060, Hubei, PR China
| | - Zuotian Wu
- Department of Psychiatry, Renmin Hospital of Wuhan University, Jiefang Road 238#, Wuhan 430060, Hubei, PR China
| | - Jing He
- Department of Psychiatry, Renmin Hospital of Wuhan University, Jiefang Road 238#, Wuhan 430060, Hubei, PR China
| | - Nan Zhang
- Department of Psychiatry, Renmin Hospital of Wuhan University, Jiefang Road 238#, Wuhan 430060, Hubei, PR China
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22
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Zhang Y, Yao L, Li X, Meng M, Shang Z, Wang Q, Xiao J, Gu X, Xu Z, Zhang X. Schizophrenia risk-gene Crmp2 deficiency causes precocious critical period plasticity and deteriorated binocular vision. Sci Bull (Beijing) 2021; 66:2225-2237. [PMID: 36654114 DOI: 10.1016/j.scib.2021.02.011] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2020] [Revised: 12/15/2020] [Accepted: 01/29/2021] [Indexed: 02/03/2023]
Abstract
Brain-specific loss of a microtubule-binding protein collapsin response mediator protein-2 (CRMP2) in the mouse recapitulates many schizophrenia-like behaviors of human patients, possibly resulting from associated developmental deficits in neuronal differentiation, path-finding, and synapse formation. However, it is still unclear how the Crmp2 loss affects neuronal circuit function and plasticity. By conducting in vivo and ex vivo electrophysiological recording in the mouse primary visual cortex (V1), we reveal that CRMP2 exerts a key regulation on the timing of postnatal critical period (CP) for experience-dependent circuit plasticity of sensory cortex. In the developing V1, the Crmp2 deficiency induces not only a delayed maturation of visual tuning functions but also a precocious CP for visual input-induced ocular dominance plasticity and its induction activity - coincident binocular inputs right after eye-opening. Mechanistically, the Crmp2 deficiency accelerates the maturation process of cortical inhibitory transmission and subsequently promotes an early emergence of balanced excitatory-inhibitory cortical circuits during the postnatal development. Moreover, the precocious CP plasticity results in deteriorated binocular depth perception in adulthood. Thus, these findings suggest that the Crmp2 deficiency dysregulates the timing of CP for experience-dependent refinement of circuit connections and further leads to impaired sensory perception in later life.
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Affiliation(s)
- Yuan Zhang
- State Key Laboratory of Cognitive Neuroscience & Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing 100875, China
| | - Li Yao
- State Key Laboratory of Cognitive Neuroscience & Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing 100875, China
| | - Xiang Li
- State Key Laboratory of Cognitive Neuroscience & Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing 100875, China
| | - Meizhen Meng
- State Key Laboratory of Cognitive Neuroscience & Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing 100875, China
| | - Ziwei Shang
- State Key Laboratory of Cognitive Neuroscience & Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing 100875, China
| | - Qin Wang
- State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China
| | - Jiaying Xiao
- State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China
| | - Xiang Gu
- State Key Laboratory of Cognitive Neuroscience & Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing 100875, China
| | - Zhiheng Xu
- State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China
| | - Xiaohui Zhang
- State Key Laboratory of Cognitive Neuroscience & Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing 100875, China.
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23
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Cheng L, Chen K, Li J, Wu J, Zhang J, Chen L, Guo G, Zhang J. Phosphorylation of CRMP2 by Cdk5 Negatively Regulates the Surface Delivery and Synaptic Function of AMPA Receptors. Mol Neurobiol 2021; 59:762-777. [PMID: 34773219 DOI: 10.1007/s12035-021-02581-w] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2020] [Accepted: 09/24/2021] [Indexed: 11/28/2022]
Abstract
AMPA receptor mediate most fast excitatory synaptic transmission and play a key role in synaptic plasticity in the central nervous system (CNS) by trafficking and targeting of its subunits to individual postsynaptic membrane. Collapsing response mediator protein 2 (CRMP2), an intracellular phospho-protein, has been reported to promote the maturation of the dendritic spine and transfer AMPA receptors to the membrane. However, our knowledge about the molecular mechanisms of CRMP2 regulating AMPA receptors trafficking is limited. Here, we reported that CRMP2 promoted the surface expression of AMPA receptor GluA1 subunit in cultured hippocampal neurons and in HEK293T cells expressing GluA1 subunits. Furthermore, we found that CRMP2 interacted with GluA1, and their interaction was inhibited by CRMP2 phosphorylation at ser522. Moreover, our results showed that phosphorylation of CRMP2 at ser522 by cyclin-dependent kinase 5 (Cdk5) decreased the fluorescence intensity of surface GluA1 and the amplitude and frequency of miniature excitatory synaptic currents (mEPSCs) in cultured hippocampal neurons, indicating a reduction levels and synaptic function of AMPA receptors. Taken together, our data demonstrated that phosphorylation of CRMP2 by Cdk5 is important for AMPA receptor surface delivery in hippocampal neurons.
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Affiliation(s)
- Longfei Cheng
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China.,Department of Neurosurgery, The First Affiliated Hospital of Jinan University Guangzhou, Guangzhou, 510630, China
| | - Keen Chen
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China.,Department of Neurosurgery, The First Affiliated Hospital of Jinan University Guangzhou, Guangzhou, 510630, China
| | - Jiong Li
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China
| | - Jiaming Wu
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China.,Department of Neurosurgery, The First Affiliated Hospital of Jinan University Guangzhou, Guangzhou, 510630, China
| | - Jiaqi Zhang
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China
| | - Li Chen
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China
| | - Guoqing Guo
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China.
| | - Jifeng Zhang
- Department of Anatomy, Neuroscience Laboratory for Cognitive and Developmental Disorders, Medical College of Jinan University, Guangzhou, 510630, China.
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24
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CRMP2 Is Involved in Regulation of Mitochondrial Morphology and Motility in Neurons. Cells 2021; 10:cells10102781. [PMID: 34685760 PMCID: PMC8535169 DOI: 10.3390/cells10102781] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2021] [Revised: 10/13/2021] [Accepted: 10/14/2021] [Indexed: 11/16/2022] Open
Abstract
Regulation of mitochondrial morphology and motility is critical for neurons, but the exact mechanisms are unclear. Here, we demonstrate that these mechanisms may involve collapsin response mediator protein 2 (CRMP2). CRMP2 is attached to neuronal mitochondria and binds to dynamin-related protein 1 (Drp1), Miro 2, and Kinesin 1 light chain (KLC1). Treating neurons with okadaic acid (OA), an inhibitor of phosphatases PP1 and PP2A, resulted in increased CRMP2 phosphorylation at Thr509/514, Ser522, and Thr555, and augmented Drp1 phosphorylation at Ser616. The CRMP2-binding small molecule (S)-lacosamide ((S)-LCM) prevented an OA-induced increase in CRMP2 phosphorylation at Thr509/514 and Ser522 but not at Thr555, and also failed to alleviate Drp1 phosphorylation. The increased CRMP2 phosphorylation correlated with decreased CRMP2 binding to Drp1, Miro 2, and KLC1. (S)-LCM rescued CRMP2 binding to Drp1 and Miro 2 but not to KLC1. In parallel with CRMP2 hyperphosphorylation, OA increased mitochondrial fission and suppressed mitochondrial traffic. (S)-LCM prevented OA-induced alterations in mitochondrial morphology and motility. Deletion of CRMP2 with a small interfering RNA (siRNA) resulted in increased mitochondrial fission and diminished mitochondrial traffic. Overall, our data suggest that the CRMP2 expression level and phosphorylation state are involved in regulating mitochondrial morphology and motility in neurons.
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25
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Cuveillier C, Boulan B, Ravanello C, Denarier E, Deloulme JC, Gory-Fauré S, Delphin C, Bosc C, Arnal I, Andrieux A. Beyond Neuronal Microtubule Stabilization: MAP6 and CRMPS, Two Converging Stories. Front Mol Neurosci 2021; 14:665693. [PMID: 34025352 PMCID: PMC8131560 DOI: 10.3389/fnmol.2021.665693] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2021] [Accepted: 04/09/2021] [Indexed: 12/21/2022] Open
Abstract
The development and function of the central nervous system rely on the microtubule (MT) and actin cytoskeletons and their respective effectors. Although the structural role of the cytoskeleton has long been acknowledged in neuronal morphology and activity, it was recently recognized to play the role of a signaling platform. Following this recognition, research into Microtubule Associated Proteins (MAPs) diversified. Indeed, historically, structural MAPs—including MAP1B, MAP2, Tau, and MAP6 (also known as STOP);—were identified and described as MT-binding and -stabilizing proteins. Extensive data obtained over the last 20 years indicated that these structural MAPs could also contribute to a variety of other molecular roles. Among multi-role MAPs, MAP6 provides a striking example illustrating the diverse molecular and cellular properties of MAPs and showing how their functional versatility contributes to the central nervous system. In this review, in addition to MAP6’s effect on microtubules, we describe its impact on the actin cytoskeleton, on neuroreceptor homeostasis, and its involvement in signaling pathways governing neuron development and maturation. We also discuss its roles in synaptic plasticity, brain connectivity, and cognitive abilities, as well as the potential relationships between the integrated brain functions of MAP6 and its molecular activities. In parallel, the Collapsin Response Mediator Proteins (CRMPs) are presented as examples of how other proteins, not initially identified as MAPs, fall into the broader MAP family. These proteins bind MTs as well as exhibiting molecular and cellular properties very similar to MAP6. Finally, we briefly summarize the multiple similarities between other classical structural MAPs and MAP6 or CRMPs.In summary, this review revisits the molecular properties and the cellular and neuronal roles of the classical MAPs, broadening our definition of what constitutes a MAP.
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26
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Ferron L, Koshti S, Zamponi GW. The life cycle of voltage-gated Ca 2+ channels in neurons: an update on the trafficking of neuronal calcium channels. Neuronal Signal 2021; 5:NS20200095. [PMID: 33664982 PMCID: PMC7905535 DOI: 10.1042/ns20200095] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2021] [Revised: 02/10/2021] [Accepted: 02/15/2021] [Indexed: 01/26/2023] Open
Abstract
Neuronal voltage-gated Ca2+ (CaV) channels play a critical role in cellular excitability, synaptic transmission, excitation-transcription coupling and activation of intracellular signaling pathways. CaV channels are multiprotein complexes and their functional expression in the plasma membrane involves finely tuned mechanisms, including forward trafficking from the endoplasmic reticulum (ER) to the plasma membrane, endocytosis and recycling. Whether genetic or acquired, alterations and defects in the trafficking of neuronal CaV channels can have severe physiological consequences. In this review, we address the current evidence concerning the regulatory mechanisms which underlie precise control of neuronal CaV channel trafficking and we discuss their potential as therapeutic targets.
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Affiliation(s)
- Laurent Ferron
- Department of Physiology and Pharmacology, Alberta Children’s Hospital Research Institute, Hotchkiss Brain Institute, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada
| | - Saloni Koshti
- Department of Physiology and Pharmacology, Alberta Children’s Hospital Research Institute, Hotchkiss Brain Institute, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada
| | - Gerald W. Zamponi
- Department of Physiology and Pharmacology, Alberta Children’s Hospital Research Institute, Hotchkiss Brain Institute, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada
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27
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Ji ZS, Li JP, Fu CH, Luo JX, Yang H, Zhang GW, Wu W, Lin HS. Spastin interacts with collapsin response mediator protein 3 to regulate neurite growth and branching. Neural Regen Res 2021; 16:2549-2556. [PMID: 33907047 PMCID: PMC8374569 DOI: 10.4103/1673-5374.313052] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022] Open
Abstract
Cytoskeletal microtubule rearrangement and movement are crucial in the repair of spinal cord injury. Spastin plays an important role in the regulation of microtubule severing. Both spastin and collapsin response mediator proteins can regulate neurite growth and branching; however, whether spastin interacts with collapsin response mediator protein 3 (CRMP3) during this process remains unclear, as is the mechanism by which CRMP3 participates in the repair of spinal cord injury. In this study, we used a proteomics approach to identify key proteins associated with spinal cord injury repair. We then employed liquid chromatography-mass spectrometry to identify proteins that were able to interact with glutathione S-transferase-spastin. Then, co-immunoprecipitation and staining approaches were used to evaluate potential interactions between spastin and CRMP3. Finally, we co-transfected primary hippocampal neurons with CRMP3 and spastin to evaluate their role in neurite outgrowth. Mass spectrometry identified the role of CRMP3 in the spinal cord injury repair process. Liquid chromatography-mass spectrometry pulldown assays identified three CRMP3 peptides that were able to interact with spastin. CRMP3 and spastin were co-expressed in the spinal cord and were able to interact with one another in vitro and in vivo. Lastly, CRMP3 overexpression was able to enhance the ability of spastin to promote neurite growth and branching. Therefore, our results confirm that spastin and CRMP3 play roles in spinal cord injury repair by regulating neurite growth and branching. These proteins may therefore be novel targets for spinal cord injury repair. The Institutional Animal Care and Use Committee of Jinan University, China approved this study (approval No. IACUS-20181008-03) on October 8, 2018.
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Affiliation(s)
- Zhi-Sheng Ji
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong Province, China
| | - Jian-Ping Li
- Guangdong-Hong Kong-Macau Institute of CNS Regeneration, Jinan University, Guangzhou, Guangdong Province, China
| | - Chao-Hua Fu
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou; Department of Orthopedics, Jiangmen Hospital of Sun Yat-sen University, Jiangmen, Guangdong Province, China
| | - Jian-Xian Luo
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong Province, China
| | - Hua Yang
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong Province, China
| | - Guo-Wei Zhang
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong Province, China
| | - Wutian Wu
- Guangdong-Hong Kong-Macau Institute of CNS Regeneration, Jinan University, Guangzhou, Guangdong Province; Re-Stem Biotechnology Co., Ltd., Suzhou, Jiangsu Province; Spine Surgery, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, China
| | - Hong-Sheng Lin
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong Province, China
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Assessment of nociception and related quality-of-life measures in a porcine model of neurofibromatosis type 1. Pain 2020; 160:2473-2486. [PMID: 31246731 DOI: 10.1097/j.pain.0000000000001648] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023]
Abstract
Neurofibromatosis type 1 (NF1) is an autosomal dominant genetic disorder resulting from germline mutations in the NF1 gene, which encodes neurofibromin. Patients experience a variety of symptoms, but pain in the context of NF1 remains largely underrecognized. Here, we characterize nociceptive signaling and pain behaviors in a miniswine harboring a disruptive NF1 mutation (exon 42 deletion). We present the first characterization of pain-related behaviors in a pig model of NF1, identifying unchanged agitation scores, lower tactile thresholds (allodynia), and decreased response latencies to thermal laser stimulation (hyperalgesia) in NF1 (females only) pigs. Male NF1 pigs with tumors showed reduced sleep quality and increased resting, 2 health-related quality-of-life symptoms found to be comorbid in people with NF1 pain. We explore these phenotypes in relationship to suppression of the increased activity of the N-type voltage-gated calcium (CaV2.2) channel by pharmacological antagonism of phosphorylation of a regulatory protein-the collapsin response mediator protein 2 (CRMP2), a known interactor of neurofibromin, and by targeting the interface between the α subunit of CaV2.2 and the accessory β-subunits with small molecules. Our data support the use of NF1 pigs as a large animal model for studying NF1-associated pain and for understanding the pathophysiology of NF1. Our findings demonstrate the translational potential of 2 small molecules in reversing ion channel remodeling seen in NF1. Interfering with CaV2.2, a clinically validated target for pain management, might also be a promising therapeutic strategy for NF1-related pain management.
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Khanna R, Moutal A, Perez-Miller S, Chefdeville A, Boinon L, Patek M. Druggability of CRMP2 for Neurodegenerative Diseases. ACS Chem Neurosci 2020; 11:2492-2505. [PMID: 32693579 DOI: 10.1021/acschemneuro.0c00307] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Collapsin response mediator proteins (CRMPs) are ubiquitously expressed phosphoproteins that coordinate cytoskeletal formation and regulate cellular division, migration, polarity, and synaptic connection. CRMP2, the most studied of the five family members, is best known for its affinity for tubulin heterodimers and function in regulating the microtubule network. Accumulating evidence has also demonstrated a key role for CRMP2 in trafficking of voltage- and ligand-gated ion channels. These functions are tightly regulated by post-translational modifications including phosphorylation and SUMOylation (addition of a small ubiquitin like modifier). Over the past decade, it has become increasingly clear that dysregulated post-translational modifications of CRMP2 contribute to the pathomechanisms of diverse diseases, including cancer, neurodegenerative diseases, chronic pain, and bipolar disorder. Here, we review the discovery, functions, and current putative preclinical and clinical therapeutics targeting CRMP2. These potential therapeutics include CRMP2-based peptides that inhibit protein-protein interactions and small-molecule compounds. Capitalizing on the availability of structural information, we identify druggable pockets on CRMP2 and predict binding modes for five known CRMP2-targeting compounds, setting the stage for optimization and de novo drug discovery targeting this multifunctional protein.
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Affiliation(s)
- Rajesh Khanna
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona 85724, United States
- Graduate Interdisciplinary Program in Neuroscience, College of Medicine, University of Arizona, Tucson, Arizona 85724, United States
- The Center for Innovation in Brain Sciences, The University of Arizona Health Sciences, Tucson, Arizona 85724, United States
- Regulonix LLC, Tucson, Arizona 85718, United States
| | - Aubin Moutal
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona 85724, United States
| | - Samantha Perez-Miller
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona 85724, United States
| | - Aude Chefdeville
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona 85724, United States
| | - Lisa Boinon
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, Arizona 85724, United States
| | - Marcel Patek
- BrightRock Path, LLC, Tucson, Arizona 85704, United States
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Mitchell R, Mikolajczak M, Kersten C, Fleetwood-Walker S. ErbB1-dependent signalling and vesicular trafficking in primary afferent nociceptors associated with hypersensitivity in neuropathic pain. Neurobiol Dis 2020; 142:104961. [DOI: 10.1016/j.nbd.2020.104961] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2020] [Revised: 05/26/2020] [Accepted: 06/08/2020] [Indexed: 02/06/2023] Open
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Cai Z, Zhu X, Zhang G, Wu F, Lin H, Tan M. Ammonia induces calpain-dependent cleavage of CRMP-2 during neurite degeneration in primary cultured neurons. Aging (Albany NY) 2020; 11:4354-4366. [PMID: 31278888 PMCID: PMC6660054 DOI: 10.18632/aging.102053] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2018] [Accepted: 06/19/2019] [Indexed: 01/07/2023]
Abstract
Hyperammonemia in the CNS induces irreversible damages to neurons due to ultimate cell loss. Neurite degeneration, a primary event that leads to neuronal cell death, remains less elucidated especially in hyperammonemia circumstances. Here, we found that the administration of ammonia induced neurite degeneration in cultured cerebellar granule neurons. The resulting altered neuronal morphology, rupture of neurites, and disassembly of the cytoskeleton led to cell death. Calcein and Fluo-4 staining revealed that ammonia induced intracellular calcium dysregulation. Subsequently activated calpain cleaved CRMP-2, a microtubule assembly protein. Pharmacologically inhibition of calpain, but not caspases or GSK-3, suppressed the cleavage of CRMP-2 and reversed neurite degeneration under ammonia treatment. Exposure to ammonia decreased whereas inhibition of calpain restored the amplitude and frequency of miniature excitatory postsynaptic currents. These data suggest a mechanism by which elevated ammonia level may induce neuronal dysfunction via abnormal calcium influx and calpain-dependent CRMP-2 cleavage, leading to abnormal synaptic transmission, cytoskeletal collapse, and neurite degeneration.
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Affiliation(s)
- Zhenbin Cai
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, China
| | - Xiaonan Zhu
- Department of Anatomy, Medical College of Jinan University, Guangzhou, China
| | - Guowei Zhang
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, China
| | - Fengming Wu
- Department of Anatomy, Medical College of Jinan University, Guangzhou, China
| | - Hongsheng Lin
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, China
| | - Minghui Tan
- Department of Orthopedics, The First Affiliated Hospital of Jinan University, Guangzhou, China
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Vicario N, Turnaturi R, Spitale FM, Torrisi F, Zappalà A, Gulino R, Pasquinucci L, Chiechio S, Parenti C, Parenti R. Intercellular communication and ion channels in neuropathic pain chronicization. Inflamm Res 2020; 69:841-850. [PMID: 32533221 DOI: 10.1007/s00011-020-01363-9] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2020] [Revised: 05/13/2020] [Accepted: 05/17/2020] [Indexed: 12/22/2022] Open
Abstract
BACKGROUND Neuropathic pain is caused by primary lesion or dysfunction of either peripheral or central nervous system. Due to its complex pathogenesis, often related to a number of comorbidities, such as cancer, neurodegenerative and neurovascular diseases, neuropathic pain still represents an unmet clinical need, lacking long-term effective treatment and complex case-by-case approach. AIM AND METHODS We analyzed the recent literature on the role of selective voltage-sensitive sodium, calcium and potassium permeable channels and non-selective gap junctions (GJs) and hemichannels (HCs) in establishing and maintaining chronic neuropathic conditions. We finally focussed our review on the role of extracellular microenvironment modifications induced by resident glial cells and on the recent advances in cell-to-cell and cell-to-extracellular environment communication in chronic neuropathies. CONCLUSION In this review, we provide an update on the current knowledge of neuropathy chronicization processes with a focus on both neuronal and glial ion channels, as well as on channel-mediated intercellular communication.
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Affiliation(s)
- Nunzio Vicario
- Section of Physiology, Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy
| | - Rita Turnaturi
- Section of Medicinal Chemistry, Department of Drug Sciences, University of Catania, Catania, Italy
| | - Federica Maria Spitale
- Section of Physiology, Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy
| | - Filippo Torrisi
- Section of Physiology, Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy
| | - Agata Zappalà
- Section of Physiology, Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy
| | - Rosario Gulino
- Section of Physiology, Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy
| | - Lorella Pasquinucci
- Section of Medicinal Chemistry, Department of Drug Sciences, University of Catania, Catania, Italy
| | - Santina Chiechio
- Section of Pharmacology, Department of Drug Sciences, University of Catania, Catania, Italy
- Oasi Research Institute IRCCS, Troina, Italy
| | - Carmela Parenti
- Section of Pharmacology, Department of Drug Sciences, University of Catania, Catania, Italy.
| | - Rosalba Parenti
- Section of Physiology, Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy.
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Zhou Y, Cai S, Gomez K, Wijeratne EMK, Ji Y, Bellampalli SS, Luo S, Moutal A, Gunatilaka AAL, Khanna R. 1-O-Acetylgeopyxin A, a derivative of a fungal metabolite, blocks tetrodotoxin-sensitive voltage-gated sodium, calcium channels and neuronal excitability which correlates with inhibition of neuropathic pain. Mol Brain 2020; 13:73. [PMID: 32393368 PMCID: PMC7216607 DOI: 10.1186/s13041-020-00616-2] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/03/2020] [Accepted: 05/04/2020] [Indexed: 01/03/2023] Open
Abstract
Chronic pain can be the result of an underlying disease or condition, medical treatment, inflammation, or injury. The number of persons experiencing this type of pain is substantial, affecting upwards of 50 million adults in the United States. Pharmacotherapy of most of the severe chronic pain patients includes drugs such as gabapentinoids, re-uptake blockers and opioids. Unfortunately, gabapentinoids are not effective in up to two-thirds of this population and although opioids can be initially effective, their long-term use is associated with multiple side effects. Therefore, there is a great need to develop novel non-opioid alternative therapies to relieve chronic pain. For this purpose, we screened a small library of natural products and their derivatives in the search for pharmacological inhibitors of voltage-gated calcium and sodium channels, which are outstanding molecular targets due to their important roles in nociceptive pathways. We discovered that the acetylated derivative of the ent-kaurane diterpenoid, geopyxin A, 1-O-acetylgeopyxin A, blocks voltage-gated calcium and tetrodotoxin-sensitive voltage-gated sodium channels but not tetrodotoxin-resistant sodium channels in dorsal root ganglion (DRG) neurons. Consistent with inhibition of voltage-gated sodium and calcium channels, 1-O-acetylgeopyxin A reduced reduce action potential firing frequency and increased firing threshold (rheobase) in DRG neurons. Finally, we identified the potential of 1-O-acetylgeopyxin A to reverse mechanical allodynia in a preclinical rat model of HIV-induced sensory neuropathy. Dual targeting of both sodium and calcium channels may permit block of nociceptor excitability and of release of pro-nociceptive transmitters. Future studies will harness the core structure of geopyxins for the generation of antinociceptive drugs.
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Affiliation(s)
- Yuan Zhou
- Department of Clinical Laboratory, the First Hospital of Jilin University, Changchun, 130021, China
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - Song Cai
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - Kimberly Gomez
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - E M Kithsiri Wijeratne
- Southwest Center for Natural Products Research, School of Natural Resources & the Environment, College of Agriculture & Life Sciences, The University of Arizona, Tucson, AZ, 85724, USA
| | - Yingshi Ji
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - Shreya S Bellampalli
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - Shizhen Luo
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - Aubin Moutal
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - A A Leslie Gunatilaka
- Southwest Center for Natural Products Research, School of Natural Resources & the Environment, College of Agriculture & Life Sciences, The University of Arizona, Tucson, AZ, 85724, USA
| | - Rajesh Khanna
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA.
- Neuroscience Graduate Interdisciplinary Program, College of Medicine, Tucson, AZ, 85724, USA.
- The Center for Innovation in Brain Sciences, The University of Arizona Health Sciences, Tucson, AZ, 85724, USA.
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Stratton H, Boinon L, Moutal A, Khanna R. Coordinating Synaptic Signaling with CRMP2. Int J Biochem Cell Biol 2020; 124:105759. [PMID: 32437854 DOI: 10.1016/j.biocel.2020.105759] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2020] [Revised: 03/27/2020] [Accepted: 04/21/2020] [Indexed: 10/24/2022]
Abstract
Synaptic transmission is a complex process, dysregulation of which underlies several neurological conditions. Collapsin response mediator protein 2 (CRMP2) is a microtubule associated protein expressed ubiquitously in the central nervous system. Identified initially in the context of Semaphorin 3A (Collapsin) induced growth cone collapse, more recent findings revealed the involvement of CRMP2 in ion channel trafficking, kinesin-dependent axonal transport and maintenance of intracellular calcium homeostasis. CRMP2 is a synaptic protein, expressed at pre- and post-synaptic sites. Interactions with proteins such as N-methyl-D-aspartate receptors, syntaxin1A as well as voltage-gated calcium and sodium channels, suggest that CRMP2 may control both the electrical and chemical components of synaptic transmission. This short review will outline the known synaptic interactions of CRMP2 and illustrate its role in synaptic transmission, thereby introducing CRMP2 as a prospective target for the pathophysiological modulation of aberrant synaptic activity.
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Affiliation(s)
- Harrison Stratton
- Department of Pharmacology, College of Medicine, University of Arizona, United States
| | - Lisa Boinon
- Department of Pharmacology, College of Medicine, University of Arizona, United States
| | - Aubin Moutal
- Department of Pharmacology, College of Medicine, University of Arizona, United States
| | - Rajesh Khanna
- Department of Pharmacology, College of Medicine, University of Arizona, United States; BIO5 Institute, University of Arizona, United States; The Center for Innovation in Brain Sciences, The University of Arizona Health Sciences, Tucson, Arizona, United States.
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Cai S, Shan Z, Zhang Z, Moutal A, Khanna R. Activity of T-type calcium channels is independent of CRMP2 in sensory neurons. Channels (Austin) 2020; 13:147-152. [PMID: 31025580 PMCID: PMC6527066 DOI: 10.1080/19336950.2019.1608129] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Amongst the regulators of voltage-gated ion channels is the collapsin response mediator protein 2 (CRMP2). CRMP2 regulation of the activity and trafficking of NaV1.7 voltage-gated sodium channels as well as the N-type (CaV2.2) voltage-gated calcium channel (VGCC) has been reported. On the other hand, CRMP2 does not appear to regulate L- (CaV1.x), P/Q- (CaV2.1), and R- (CaV2.3) type high VGCCs. Whether CRMP2 regulates low VGCCs remains an open question. Here, we asked if CRMP2 could regulate the low voltage-gated (T-type/CaV3.x) channels in sensory neurons. Reducing CRMP2 protein levels with short interfering RNAs yielded no change in macroscopic currents carried by T-type channels. No change in biophysical properties of the T-type currents was noted. Future studies pursuing CRMP2 druggability in neuropathic pain will benefit from the findings that CRMP2 regulates only the N-type (CaV2.2) calcium channels.
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Affiliation(s)
- Song Cai
- a Department of Pharmacology, College of Medicine , The University of Arizona Health Sciences , Tucson , AZ , USA
| | - Zhiming Shan
- a Department of Pharmacology, College of Medicine , The University of Arizona Health Sciences , Tucson , AZ , USA.,b Department of Anesthesiology , Shenzhen People's Hospital & Second Clinical Medical College of Jinan University , Shenzhen , P.R. China
| | - Zhongjun Zhang
- b Department of Anesthesiology , Shenzhen People's Hospital & Second Clinical Medical College of Jinan University , Shenzhen , P.R. China
| | - Aubin Moutal
- a Department of Pharmacology, College of Medicine , The University of Arizona Health Sciences , Tucson , AZ , USA
| | - Rajesh Khanna
- a Department of Pharmacology, College of Medicine , The University of Arizona Health Sciences , Tucson , AZ , USA.,c The Center for Innovation in Brain Sciences , The University of Arizona Health Sciences , Tucson , AZ , USA
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Qi B, Yang Y, Cheng Y, Sun D, Wang X, Khanna R, Ju W. Nasal delivery of a CRMP2-derived CBD3 adenovirus improves cognitive function and pathology in APP/PS1 transgenic mice. Mol Brain 2020; 13:58. [PMID: 32272942 PMCID: PMC7144060 DOI: 10.1186/s13041-020-00596-3] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2020] [Accepted: 03/23/2020] [Indexed: 12/12/2022] Open
Abstract
Calcium dysregulation is a key pathological event in Alzheimer's disease (AD). In studying approaches to mitigate this calcium overload, we identified the collapsin response mediator protein 2 (CRMP2), an axonal guidance protein that participates in synapse dynamics by interacting with and regulating activity of N-methyl-D-aspartate receptors (NMDARs). We further identified a 15 amino acid peptide from CRMP2 (designated CBD3, for calcium-binding domain 3), that reduced NMDAR-mediated Ca2+ influx in cultured neurons and post-synaptic NMDAR-mediated currents in cortical slices. Whether targeting CRMP2 could be therapeutically beneficial in AD is unknown. Here, using CBD3, we tested the utility of this approach. Employing the APP/PS1 mouse model of AD which demonstrates robust pathophysiology including Aβ1-42 deposition, altered tau levels, and diminished cognitive functions, we asked if overexpression of CBD3 could rescue these events. CBD3 was engineered into an adeno-associated vector and nasally delivered into APP/PS1 mice and then biochemical (immunohistochemistry, immunoblotting), cellular (TUNEL apoptosis assays), and behavioral (Morris water maze test) assessments were performed. APP/PS1 mice administered adeno-associated virus (AAV, serotype 2) harboring CBD3 demonstrated: (i) reduced levels of Aβ1-42 and phosphorylated-tau (a marker of AD progression), (ii) reduced apoptosis in the hippocampus, and (iii) reduced cognitive decline compared with APP/PS1 mice or APP/PS1 administered a control virus. These results provide an instructive example of utilizing a peptide-based approach to unravel protein-protein interactions that are necessary for AD pathology and demonstrate the therapeutic potential of CRMP2 as a novel protein player in AD.
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Affiliation(s)
- Baochang Qi
- Department of Orthopedic Traumatology, The First Hospital of Jilin University, Changchun, 130021, Jilin Province, China
| | - Yu Yang
- Department of Neurology and neuroscience center, The First Hospital of Jilin University, No.1 Xinmin Street, Chaoyang District, Changchun, 130021, Jilin Province, China
| | - Yingying Cheng
- Department of Neurology and neuroscience center, The First Hospital of Jilin University, No.1 Xinmin Street, Chaoyang District, Changchun, 130021, Jilin Province, China
| | - Di Sun
- Department of Colorectal and Anal Surgery, The First Hospital of Jilin University, Changchun, 130021, Jilin Province, China
| | - Xu Wang
- Department of Neurology and neuroscience center, The First Hospital of Jilin University, No.1 Xinmin Street, Chaoyang District, Changchun, 130021, Jilin Province, China
| | - Rajesh Khanna
- Department of Pharmacology, College of Medicine, The University of Arizona, Tucson, AZ, 85718, USA.
- Center for Innovation in Brain Sciences, University of Arizona, Tucson, AZ, 85721, USA.
| | - Weina Ju
- Department of Neurology and neuroscience center, The First Hospital of Jilin University, No.1 Xinmin Street, Chaoyang District, Changchun, 130021, Jilin Province, China.
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Sun YY, Zhu L, Sun ZL, Feng DF. CRMP2 improves memory deficits by enhancing the maturation of neuronal dendritic spines after traumatic brain injury. Exp Neurol 2020; 328:113253. [PMID: 32084454 DOI: 10.1016/j.expneurol.2020.113253] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/18/2019] [Revised: 02/12/2020] [Accepted: 02/17/2020] [Indexed: 11/28/2022]
Abstract
Our recent study investigated the role of collapsin response mediator protein-2 (CRMP2) on dendritic spine morphology and memory function after traumatic brain injury (TBI). First, we examined the density and morphology of dendritic spines in Thy1-GFP mice on the 1 st day (P1D) and 7th day (P7D) after controlled cortical impact injury (CCI). The dendritic spine density in the hippocampus was decreased on P1D, in which mainly mushroom-type and thin-type spines were lost. The density of dendritic spines was increased on P7D, most of which were of the thin type. Next, we explored the expression of CRMP2 on P1D and P7D. CRMP2 expression was decreased on P1D, but the levels of the CRMP2 breakdown product were increased. On P7D, the expression pattern was the opposite. Then, we constructed CRMP2 overexpression and knockdown plasmids and transfected them into cultured neurons in vitro. CRMP2 increased the dendritic spine density of cultured neurons and the proportion of mushroom-type spines, while CRMP2-shRNA reduced the dendritic spine density and the proportion of mushroom-type spines. To determine the role of CRMP2 in dendritic spines after TBI, we stereotactically injected the CRMP2 overexpression and knockdown viruses into the hippocampus and found that CRMP2 increased the dendritic spine density and the proportion of mushroom-type spines after TBI. Meanwhile, as suggested by the morphological changes, fear conditioning behavioral experiments confirmed that CRMP2 improved memory deficits after TBI.
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Affiliation(s)
- Yi-Yu Sun
- Department of Neurosurgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 201999, China
| | - Liang Zhu
- Department of Neurosurgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 201999, China
| | - Zhao-Liang Sun
- Department of Neurosurgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 201999, China
| | - Dong-Fu Feng
- Department of Neurosurgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 201999, China; Institute of Traumatic Medicine, Shanghai Jiao Tong University School of Medicine, Shanghai 201999, China.
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Towards a neurobiological understanding of pain in neurofibromatosis type 1: mechanisms and implications for treatment. Pain 2020; 160:1007-1018. [PMID: 31009417 DOI: 10.1097/j.pain.0000000000001486] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/10/2023]
Abstract
Neurofibromatosis type 1 (NF1) is the most common of a group of rare diseases known by the term, "Neurofibromatosis," affecting 1 in 3000 to 4000 people. NF1 patients present with, among other disease complications, café au lait patches, skin fold freckling, Lisch nodules, orthopedic complications, cutaneous neurofibromas, malignant peripheral nerve sheath tumors, cognitive impairment, and chronic pain. Although NF1 patients inevitably express pain as a debilitating symptom of the disease, not much is known about its manifestation in the NF1 disease, with most current information coming from sporadic case reports. Although these reports indicate the existence of pain, the molecular signaling underlying this symptom remains underexplored, and thus, we include a synopsis of the literature surrounding NF1 pain studies in 3 animal models: mouse, rat, and miniswine. We also highlight unexplored areas of NF1 pain research. As therapy for NF1 pain remains in various clinical and preclinical stages, we present current treatments available for patients and highlight the importance of future therapeutic development. Equally important, NF1 pain is accompanied by psychological complications in comorbidities with sleep, gastrointestinal complications, and overall quality of life, lending to the importance of investigation into this understudied phenomenon of NF1. In this review, we dissect the presence of pain in NF1 in terms of psychological implication, anatomical presence, and discuss mechanisms underlying the onset and potentiation of NF1 pain to evaluate current therapies and propose implications for treatment of this severely understudied, but prevalent symptom of this rare disease.
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Buchta WC, Moutal A, Hines B, Garcia-Keller C, Smith ACW, Kalivas P, Khanna R, Riegel AC. Dynamic CRMP2 Regulation of CaV2.2 in the Prefrontal Cortex Contributes to the Reinstatement of Cocaine Seeking. Mol Neurobiol 2020; 57:346-357. [PMID: 31359322 PMCID: PMC6980501 DOI: 10.1007/s12035-019-01711-9] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2019] [Accepted: 07/15/2019] [Indexed: 02/06/2023]
Abstract
Cocaine addiction remains a major health concern with limited effective treatment options. A better understanding of mechanisms underlying relapse may help inform the development of new pharmacotherapies. Emerging evidence suggests that collapsin response mediator protein 2 (CRMP2) regulates presynaptic excitatory neurotransmission and contributes to pathological changes during diseases, such as neuropathic pain and substance use disorders. We examined the role of CRMP2 and its interactions with a known binding partner, CaV2.2, in cocaine-seeking behavior. We employed the rodent self-administration model of relapse to drug seeking and focused on the prefrontal cortex (PFC) for its well-established role in reinstatement behaviors. Our results indicated that repeated cocaine self-administration resulted in a dynamic and persistent alteration in the PFC expression of CRMP2 and its binding partner, the CaV2.2 (N-type) voltage-gated calcium channel. Following cocaine self-administration and extinction training, the expression of both CRMP2 and CaV2.2 was reduced relative to yoked saline controls. By contrast, cued reinstatement potentiated CRMP2 expression and increased CaV2.2 expression above extinction levels. Lastly, we utilized the recently developed peptide myr-TAT-CBD3 to disrupt the interaction between CRMP2 and CaV2.2 in vivo. We assessed the reinstatement behavior after infusing this peptide directly into the medial PFC and found that it decreased cue-induced reinstatement of cocaine seeking. Taken together, these data suggest that neuroadaptations in the CRMP2/CaV2.2 signaling cascade in the PFC can facilitate drug-seeking behavior. Targeting such interactions has implications for the treatment of cocaine relapse behavior.
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Affiliation(s)
- William C Buchta
- Department of Neuroscience, Medical University of South Carolina (MUSC), 410C Basic Sciences Building, 173 Ashley Avenue, Charleston, SC, 29425, USA
- Neurobiology of Addiction Research Center, Medical University of South Carolina, Charleston, SC, 29425, USA
| | - Aubin Moutal
- Department of Pharmacology, University of Arizona, Tucson, AZ, 85724, USA
| | - Bethany Hines
- Department of Neuroscience, Medical University of South Carolina (MUSC), 410C Basic Sciences Building, 173 Ashley Avenue, Charleston, SC, 29425, USA
- Neurobiology of Addiction Research Center, Medical University of South Carolina, Charleston, SC, 29425, USA
| | - Constanza Garcia-Keller
- Department of Neuroscience, Medical University of South Carolina (MUSC), 410C Basic Sciences Building, 173 Ashley Avenue, Charleston, SC, 29425, USA
- Neurobiology of Addiction Research Center, Medical University of South Carolina, Charleston, SC, 29425, USA
| | - Alexander C W Smith
- Department of Neuroscience, Medical University of South Carolina (MUSC), 410C Basic Sciences Building, 173 Ashley Avenue, Charleston, SC, 29425, USA
- Neurobiology of Addiction Research Center, Medical University of South Carolina, Charleston, SC, 29425, USA
| | - Peter Kalivas
- Department of Neuroscience, Medical University of South Carolina (MUSC), 410C Basic Sciences Building, 173 Ashley Avenue, Charleston, SC, 29425, USA
- Neurobiology of Addiction Research Center, Medical University of South Carolina, Charleston, SC, 29425, USA
| | - Rajesh Khanna
- Department of Pharmacology, University of Arizona, Tucson, AZ, 85724, USA
- Department of Anesthesiology, University of Arizona, Tucson, AZ, 85724, USA
- The Center for Innovation in Brain Sciences, The University of Arizona Health Sciences, Tucson, AZ, USA
| | - Arthur C Riegel
- Department of Neuroscience, Medical University of South Carolina (MUSC), 410C Basic Sciences Building, 173 Ashley Avenue, Charleston, SC, 29425, USA.
- Neurobiology of Addiction Research Center, Medical University of South Carolina, Charleston, SC, 29425, USA.
- Department of Pharmacology, University of Arizona, Tucson, AZ, 85724, USA.
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Xiang D, Xiao J, Sun S, Fu L, Yao L, Wang G, Liu Z. Differential Regulation of DNA Methylation at the CRMP2 Promoter Region Between the Hippocampus and Prefrontal Cortex in a CUMS Depression Model. Front Psychiatry 2020; 11:141. [PMID: 32256396 PMCID: PMC7093734 DOI: 10.3389/fpsyt.2020.00141] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/08/2019] [Accepted: 02/14/2020] [Indexed: 12/21/2022] Open
Abstract
Current evidence supports the idea that neural plasticity is a potential cause of depression. Abundant studies indicate that CRMP2 has important roles in neural plasticity. Moreover, CRMP2 may contribute to the etiology of depression. However, the regulatory mechanisms underlying the role of CRMP2 remain unclear. DNA methylation alteration is generally acknowledged to be involved in the development of depression. The aim of this study was to explore the relationship between the expression and DNA methylation of CRMP2 in the hippocampus and prefrontal cortex of a rat depression model. Chronic unpredictable mild stress (CUMS) was used to establish a rat depression model, and body weight and behavioral tests were used to evaluate the effects of stress. Real-time PCR and Western blotting were used to test CRMP2 mRNA and protein expression, respectively, in the hippocampus and prefrontal cortex of rats. DNA methylation levels of the CRMP2 promoter were analyzed by bisulfite sequencing PCR (BSP). CUMS caused depressive-like behavior in rats, as evidenced by: decreased body weight and sucrose preference rate; decreases in the total distance traveled, rearing frequency, velocity, and duration in the center in the open field test (OFT); and prolonged immobility in the forced swimming test (FST). CRMP2 mRNA and protein expression in the hippocampus and prefrontal cortex were significantly decreased in the CUMS group compared with the control group. The levels of CRMP2 promoter DNA methylation in the hippocampus of the CUMS group were significantly higher than those of the control group, while these changes were not observed in the prefrontal cortex of CUMS rats. Our data provide evidence that altered expression of CRMP2 in the hippocampus and prefrontal cortex is associated with the pathogenesis of depression. Moreover, the results also suggest regional differences in the regulation of DNA methylation in the CRMP2 promoter between the hippocampus and prefrontal cortex during the development of depression.
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Affiliation(s)
- Dan Xiang
- Department of Psychiatry, Renmin Hospital of Wuhan University, Wuhan, China
| | - Jiawei Xiao
- Department of Psychiatry, Renmin Hospital of Wuhan University, Wuhan, China
| | - Siqi Sun
- Department of Psychiatry, Renmin Hospital of Wuhan University, Wuhan, China
| | - Linyan Fu
- Department of Psychiatry, Renmin Hospital of Wuhan University, Wuhan, China
| | - Lihua Yao
- Department of Psychiatry, Renmin Hospital of Wuhan University, Wuhan, China
| | - Gaohua Wang
- Department of Psychiatry, Renmin Hospital of Wuhan University, Wuhan, China.,Institute of Neuropsychiatry, Renmin Hospital, Wuhan University, Wuhan, China
| | - Zhongchun Liu
- Department of Psychiatry, Renmin Hospital of Wuhan University, Wuhan, China.,Institute of Neuropsychiatry, Renmin Hospital, Wuhan University, Wuhan, China
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41
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Lee S, Jo S, Talbot S, Zhang HXB, Kotoda M, Andrews NA, Puopolo M, Liu PW, Jacquemont T, Pascal M, Heckman LM, Jain A, Lee J, Woolf CJ, Bean BP. Novel charged sodium and calcium channel inhibitor active against neurogenic inflammation. eLife 2019; 8:48118. [PMID: 31765298 PMCID: PMC6877086 DOI: 10.7554/elife.48118] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2019] [Accepted: 11/13/2019] [Indexed: 12/12/2022] Open
Abstract
Voltage-dependent sodium and calcium channels in pain-initiating nociceptor neurons are attractive targets for new analgesics. We made a permanently charged cationic derivative of an N-type calcium channel-inhibitor. Unlike cationic derivatives of local anesthetic sodium channel blockers like QX-314, this cationic compound inhibited N-type calcium channels more effectively with extracellular than intracellular application. Surprisingly, the compound is also a highly effective sodium channel inhibitor when applied extracellularly, producing more potent inhibition than lidocaine or bupivacaine. The charged inhibitor produced potent and long-lasting analgesia in mouse models of incisional wound and inflammatory pain, inhibited release of the neuropeptide calcitonin gene-related peptide (CGRP) from dorsal root ganglion neurons, and reduced inflammation in a mouse model of allergic asthma, which has a strong neurogenic component. The results show that some cationic molecules applied extracellularly can powerfully inhibit both sodium channels and calcium channels, thereby blocking both nociceptor excitability and pro-inflammatory peptide release.
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Affiliation(s)
- Seungkyu Lee
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States
| | - Sooyeon Jo
- Department of Neurobiology, Harvard Medical School, Boston, United States
| | - Sébastien Talbot
- Département de Pharmacologie et Physiologie, Université de Montréal, Montréal, Canada
| | | | - Masakazu Kotoda
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States
| | - Nick A Andrews
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States
| | - Michelino Puopolo
- Department of Anesthesiology, Renaissance School of Medicine at Stony Brook University, Stony Brook, United States
| | - Pin W Liu
- Department of Neurobiology, Harvard Medical School, Boston, United States
| | - Thomas Jacquemont
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States
| | - Maud Pascal
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States
| | - Laurel M Heckman
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States
| | - Aakanksha Jain
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States
| | - Jinbo Lee
- Sage Partner International, Andover, United States
| | - Clifford J Woolf
- FM Kirby Neurobiology Research Center, Boston Children's Hospital, Boston, United States.,Department of Neurobiology, Harvard Medical School, Boston, United States
| | - Bruce P Bean
- Department of Neurobiology, Harvard Medical School, Boston, United States
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42
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In vitro cellular uptake and neuroprotective efficacy of poly-arginine-18 (R18) and poly-ornithine-18 (O18) peptides: critical role of arginine guanidinium head groups for neuroprotection. Mol Cell Biochem 2019; 464:27-38. [DOI: 10.1007/s11010-019-03646-0] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2019] [Accepted: 10/23/2019] [Indexed: 12/13/2022]
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43
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Moutal A, White KA, Chefdeville A, Laufmann RN, Vitiello PF, Feinstein D, Weimer JM, Khanna R. Dysregulation of CRMP2 Post-Translational Modifications Drive Its Pathological Functions. Mol Neurobiol 2019; 56:6736-6755. [PMID: 30915713 PMCID: PMC6728212 DOI: 10.1007/s12035-019-1568-4] [Citation(s) in RCA: 54] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2018] [Accepted: 03/15/2019] [Indexed: 12/13/2022]
Abstract
Collapsin response mediator proteins (CRMPs) are a family of ubiquitously expressed, homologous phosphoproteins best known for coordinating cytoskeletal formation and regulating cellular division, migration, polarity, and synaptic connection. CRMP2, the most studied of the five family members, is best known for its affinity for tubulin heterodimers and function in regulating the microtubule network. These functions are tightly regulated by post-translational modifications including phosphorylation, SUMOylation, oxidation, and O-GlcNAcylation. While CRMP2's physiological functions rely mostly on its non-phosphorylated state, dysregulation of CRMP2 phosphorylation and SUMOylation has been reported to be involved in the pathophysiology of multiple diseases including cancer, chronic pain, spinal cord injury, neurofibromatosis type 1, and others. Here, we provide a consolidated update on what is known about CRMP2 signaling and function, first focusing on axonal growth and neuronal polarity, then illustrating the link between dysregulated CRMP2 post-translational modifications and diseases. We additionally discuss the roles of CRMP2 in non-neuronal cells, both in the CNS and regions of the periphery. Finally, we offer thoughts on the therapeutic implications of modulating CRMP2 function in a variety of diseases.
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Affiliation(s)
- Aubin Moutal
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - Katherine A White
- Pediatrics and Rare Diseases Group, Sanford Research, 2301 E 60th St N, Sioux Falls, SD, 57104, USA
| | - Aude Chefdeville
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
| | - Rachel N Laufmann
- Pediatrics and Rare Diseases Group, Sanford Research, 2301 E 60th St N, Sioux Falls, SD, 57104, USA
| | - Peter F Vitiello
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA
- Department of Pediatrics, Sanford School of Medicine, University of South Dakota, Sioux Falls, SD, USA
| | - Douglas Feinstein
- Department of Veterans Affairs, Jesse Brown VA Medical Center, University of Illinois at Chicago, Chicago, IL, USA
| | - Jill M Weimer
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA.
- Department of Pediatrics, Sanford School of Medicine, University of South Dakota, Sioux Falls, SD, USA.
| | - Rajesh Khanna
- Department of Pharmacology, College of Medicine, University of Arizona, 1501 North Campbell Drive, P.O. Box 245050, Tucson, AZ, 85724, USA.
- Pediatrics and Rare Diseases Group, Sanford Research, 2301 E 60th St N, Sioux Falls, SD, 57104, USA.
- Department of Anesthesiology, University of Arizona, Tucson, AZ, USA.
- The Center for Innovation in Brain Sciences, The University of Arizona Health Sciences, Tucson, AZ, USA.
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Wang L, Ji S. Inhibition of Ubc9-Induced CRMP2 SUMOylation Disrupts Glioblastoma Cell Proliferation. J Mol Neurosci 2019; 69:391-398. [PMID: 31267313 DOI: 10.1007/s12031-019-01368-y] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2019] [Accepted: 06/25/2019] [Indexed: 12/30/2022]
Abstract
Glioblastoma (GBM) is the most aggressive astrocytoma. Despite maximum treatment, the GBM usually recurs and the patient survival is poor. Thus, understanding the molecular mechanism of GBM progression will be meaningful to ameliorate this situation. In this study, collapsin response mediator protein 2 (CRMP2) and Ubc9 protein levels were evaluated in three GBM cell lines. Sumoylated CRMP2 were enriched and immunoprecipitated using SUMO1 and IgG antibodies. CRMP2-K374A mutant was generated by site-direct mutagenesis. All indicated constructs were transfected into GL15 cells, and the corresponding proliferation-promoting effect was assessed through cell proliferation ratio. The t-CSM peptide was used to disturb Ubc9-CRMP2 interaction. CRMP2 is expressed in all tested GBM cell lines. The Ubc9 protein levels are positively correlated with CRMP2 level, and both can promote GBM cell proliferation. Blocking CRMP2 SUMOylation through SUMOylation-incompetent mutant or small peptide suppresses CRMP2-induced GBM cell proliferation. This study demonstrates that the CRMP2 SUMOylation exists widely in GBM cells and drives glioblastoma proliferation. CRMP2 SUMOylation inhibition can significantly suppress GBM proliferation in vitro.
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Affiliation(s)
- Leilei Wang
- Department of Neurosurgery, Cangzhou Central Hospital, Xinhua West Road, Cangzhou, 061000, Hebei, China
| | - Suzhen Ji
- Department of Emergency, Cangzhou Central Hospital, Xinhua West Road, Cangzhou, 061000, Hebei, China.
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Shan Z, Cai S, Yu J, Zhang Z, Vallecillo TGM, Serafini MJ, Thomas AM, Pham NYN, Bellampalli SS, Moutal A, Zhou Y, Xu GB, Xu YM, Luo S, Patek M, Streicher JM, Gunatilaka AAL, Khanna R. Reversal of Peripheral Neuropathic Pain by the Small-Molecule Natural Product Physalin F via Block of CaV2.3 (R-Type) and CaV2.2 (N-Type) Voltage-Gated Calcium Channels. ACS Chem Neurosci 2019; 10:2939-2955. [PMID: 30946560 DOI: 10.1021/acschemneuro.9b00166] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
No universally efficacious therapy exists for chronic pain, a disease affecting one-fifth of the global population. An overreliance on the prescription of opioids for chronic pain despite their poor ability to improve function has led to a national opioid crisis. In 2018, the NIH launched a Helping to End Addiction Long-term plan to spur discovery and validation of novel targets and mechanisms to develop alternative nonaddictive treatment options. Phytochemicals with medicinal properties have long been used for various treatments worldwide. The natural product physalin F, isolated from the Physalis acutifolia (family: Solanaceae) herb, demonstrated antinociceptive effects in models of inflammatory pain, consistent with earlier reports of its anti-inflammatory and immunomodulatory activities. However, the target of action of physalin F remained unknown. Here, using whole-cell and slice electrophysiology, competition binding assays, and experimental models of neuropathic pain, we uncovered a molecular target for physalin F's antinociceptive actions. We found that physalin F (i) blocks CaV2.3 (R-type) and CaV2.2 (N-type) voltage-gated calcium channels in dorsal root ganglion (DRG) neurons, (ii) does not affect CaV3 (T-type) voltage-gated calcium channels or voltage-gated sodium or potassium channels, (iii) does not bind G-protein coupled opioid receptors, (iv) inhibits the frequency of spontaneous excitatory postsynaptic currents (EPSCs) in spinal cord slices, and (v) reverses tactile hypersensitivity in models of paclitaxel-induced peripheral neuropathy and spinal nerve ligation. Identifying CaV2.2 as a molecular target of physalin F may spur its use as a tool for mechanistic studies and position it as a structural template for future synthetic compounds.
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Affiliation(s)
- Zhiming Shan
- Department of Anesthesiology, Shenzhen People’s Hospital & Second Clinical Medical College of Jinan University, Shenzhen 518020, P.R. China
- Department of Anesthesiology, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, P.R. China
| | | | - Jie Yu
- College of Basic Medical Science, Zhejiang Chinese Medical University, Hangzhou 310058, P.R. China
| | - Zhongjun Zhang
- Department of Anesthesiology, Shenzhen People’s Hospital & Second Clinical Medical College of Jinan University, Shenzhen 518020, P.R. China
| | | | | | | | | | | | | | - Yuan Zhou
- The First Hospital of Jilin University, 71 Xinmin Street, Changchun 130021, P. R. China
- BrightRock Path Consulting, LLC, Tucson 85721, Arizona, United States
| | | | | | | | - Marcel Patek
- BrightRock Path Consulting, LLC, Tucson 85721, Arizona, United States
| | | | | | - Rajesh Khanna
- The Center for Innovation in Brain Sciences, The University of Arizona Health Sciences, Tucson, Arizona 85724, United States
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46
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Yu H, Shin SM, Xiang H, Chao D, Cai Y, Xu H, Khanna R, Pan B, Hogan QH. AAV-encoded Ca V2.2 peptide aptamer CBD3A6K for primary sensory neuron-targeted treatment of established neuropathic pain. Gene Ther 2019; 26:308-323. [PMID: 31118475 DOI: 10.1038/s41434-019-0082-7] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/05/2018] [Revised: 02/25/2019] [Accepted: 03/20/2019] [Indexed: 12/17/2022]
Abstract
Transmission of pain signals from primary sensory neurons to secondary neurons of the central nervous system is critically dependent on presynaptic voltage-gated calcium channels. Calcium channel-binding domain 3 (CBD3), derived from the collapsin response mediator protein 2 (CRMP2), is a peptide aptamer that is effective in blocking N-type voltage-gated calcium channel (CaV2.2) activity. We previously reported that recombinant adeno-associated virus (AAV)-mediated restricted expression of CBD3 affixed to enhanced green fluorescent protein (EGFP) in primary sensory neurons prevents the development of cutaneous mechanical hypersensitivity in a rat neuropathic pain model. In this study, we tested whether this strategy is effective in treating established pain. We constructed AAV6-EGFP-CBD3A6K (AAV6-CBD3A6K) expressing a fluorescent CBD3A6K (replacing A to K at position 6 of CBD3 peptide), which is an optimized variant of the parental CBD3 peptide that is a more potent blocker of CaV2.2. Delivery of AAV6-CBD3A6K into lumbar (L) 4 and 5 dorsal root ganglia (DRG) of rats 2 weeks following tibial nerve injury (TNI) induced transgene expression in neurons of these DRG and their axonal projections, accompanied by attenuation of pain behavior. We additionally observed that the increased CaV2.2α1b immunoreactivity in the ipsilateral spinal cord dorsal horn and DRG following TNI was significantly normalized by AAV6-CBD3A6K treatment. Finally, the increased neuronal activity in the ipsilateral dorsal horn that developed after TNI was reduced by AAV6-CBD3A6K treatment. Collectively, these results indicate that DRG-restricted AAV6 delivery of CBD3A6K is an effective analgesic molecular strategy for the treatment of established neuropathic pain.
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Affiliation(s)
- Hongwei Yu
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA. .,Zablocki Veterans Affairs Medical Center, Milwaukee, WI, 53295, USA.
| | - Seung Min Shin
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA.,Zablocki Veterans Affairs Medical Center, Milwaukee, WI, 53295, USA
| | - Hongfei Xiang
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA.,Department of Orthopedic Surgery, Affiliated Hospital of Qingdao University, 266000, Qingdao, PR China
| | - Dongman Chao
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA
| | - Yongsong Cai
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA.,Xi'an Jiaotong University Health Science Center, 710061, Xi'an, Shaanxi, PR China
| | - Hao Xu
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA.,Department of Orthopedic Surgery, Affiliated Hospital of Qingdao University, 266000, Qingdao, PR China
| | - Rajesh Khanna
- Departments of Pharmacology, Neuroscience and Anesthesiology, College of Medicine, University of Arizona, Tucson, AZ, 85724, USA
| | - Bin Pan
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA
| | - Quinn H Hogan
- Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, 53226, USA.,Zablocki Veterans Affairs Medical Center, Milwaukee, WI, 53295, USA
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Castillo C, Martinez JC, Longart M, García L, Hernández M, Carballo J, Rojas H, Matteo L, Casique L, Escalona JL, Rodríguez Y, Rodriguez J, Hernández D, Balbi D, Villegas R. Extracellular Application of CRMP2 Increases Cytoplasmic Calcium through NMDA Receptors. Neuroscience 2019; 376:204-223. [PMID: 29555037 DOI: 10.1016/j.neuroscience.2018.02.002] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2017] [Revised: 01/30/2018] [Accepted: 02/02/2018] [Indexed: 12/27/2022]
Abstract
Collapsin Response Mediator Protein 2 (CRMP2) is an intracellular protein involved in axon and dendrite growth and specification. In this study, CRMP2 was identified in a conditioned media derived from degenerated sciatic nerves (CM). On cultured rat hippocampal neurons, acute extracellular application of CM or partially purified recombinant CRMP2 produced an increase in cytoplasmic calcium. The increase in cytoplasmic calcium was mostly mediated through NMDA receptors, with a minor contribution of N-type VDCC, and it was maintained as long as CM was present. By using live-labeling of CRMP2, Ca2+ channel binding domain 3 (CBD3) peptide derived from CRMP2, and recombinant CRMP2, we demonstrated that that this effect was mediated by an action on the extracellular side of the NMDA receptor. This is the first report of an extracellular action of CRMP2. Prolonged exposure to extracellular CRMP2, may contribute to neuronal calcium dysregulation and neuronal damage.
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Affiliation(s)
- Cecilia Castillo
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela.
| | - Juan Carlos Martinez
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Marines Longart
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Lisbeth García
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Marianela Hernández
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Jeismar Carballo
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Héctor Rojas
- Instituto de Inmunología, Facultad de Medicina, Universidad Central de Venezuela, Caracas 1051, Venezuela
| | - Lorena Matteo
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Liliana Casique
- Depto. de Biología Celular, Universidad Simón Bolívar, Caracas 1080, Venezuela
| | | | - Yuryanni Rodríguez
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Jessica Rodriguez
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Deyanell Hernández
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Domingo Balbi
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
| | - Raimundo Villegas
- Unidad de Neurociencias, Instituto de Estudios Avanzados IDEA, Caracas 1080, Venezuela
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48
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Kim W, Won SY, Yoon BJ. CRMP2 mediates GSK3β actions in the striatum on regulating neuronal structure and mania-like behavior. J Affect Disord 2019; 245:1079-1088. [PMID: 30699850 DOI: 10.1016/j.jad.2018.10.371] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/22/2018] [Revised: 09/29/2018] [Accepted: 10/05/2018] [Indexed: 11/20/2022]
Abstract
BACKGROUND Genetic and physiological studies have implicated the striatum in bipolar disorder (BD). Although Glycogen synthase kinase 3 beta (GSK3β) has been suggested to play a role in the pathophysiology of BD since it is inhibited by lithium, it remains unknown how GSK3β activity might be involved. Therefore we examined the functional roles of GSK3β and one of its substrates, CRMP2, within the striatum. METHODS Using CRISPR-Cas9 system, we specifically ablated GSK3β in the striatal neurons in vivo and in vitro. Sholl analysis was performed for the structural studies of medium spiny neurons (MSNs) and amphetamine-induced hyperlocomotion was measured to investigate the effects of gene ablations on the mania-like symptom of BD. RESULTS GSK3β deficiency in cultured neurons and in neurons of adult mouse brain caused opposite patterns of neurite changes. Furthermore, specific knockout of GSK3β in the MSNs of the indirect pathway significantly suppressed amphetamine-induced hyperlocomotion. We demonstrated that these phenotypes of GSK3β ablation were mediated by CRMP2, a major substrate of GSK3β. LIMITATIONS Amphetamine-induced hyperlocomotion only partially recapitulate the symptoms of BD. It requires further study to examine whether abnormality in GSK3β or CRMP2 is also involved in depression phase of BD. Additionally, we could not confirm whether the behavioral changes observed in GSK3β-ablated mice were indeed caused by the cellular structural changes observed in the striatal neurons. CONCLUSION Our results demonstrate that GSK3β and its substrate CRMP2 critically regulate the neurite structure of MSNs and their functions specifically within the indirect pathway of the basal ganglia network play a critical role in manifesting mania-like behavior of BD. Moreover, our data also suggest lithium may exert its effect on BD through a GSK3β-independent mechanism, in addition to the GSK3β inhibition-mediated mechanism.
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Affiliation(s)
- Wonju Kim
- Division of Life Sciences, School of Life Sciences and Biotechnology, Korea University, Anam-ro 145, Seongbuk-gu, Seoul 02841, Republic of Korea
| | - Seong-Yeon Won
- Division of Life Sciences, School of Life Sciences and Biotechnology, Korea University, Anam-ro 145, Seongbuk-gu, Seoul 02841, Republic of Korea
| | - Bong-June Yoon
- Division of Life Sciences, School of Life Sciences and Biotechnology, Korea University, Anam-ro 145, Seongbuk-gu, Seoul 02841, Republic of Korea.
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Chew LA, Bellampalli SS, Dustrude ET, Khanna R. Mining the Na v1.7 interactome: Opportunities for chronic pain therapeutics. Biochem Pharmacol 2019; 163:9-20. [PMID: 30699328 DOI: 10.1016/j.bcp.2019.01.018] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/02/2019] [Accepted: 01/24/2019] [Indexed: 12/14/2022]
Abstract
The peripherally expressed voltage-gated sodium NaV1.7 (gene SCN9A) channel boosts small stimuli to initiate firing of pain-signaling dorsal root ganglia (DRG) neurons and facilitates neurotransmitter release at the first synapse within the spinal cord. Mutations in SCN9A produce distinct human pain syndromes. Widely acknowledged as a "gatekeeper" of pain, NaV1.7 has been the focus of intense investigation but, to date, no NaV1.7-selective drugs have reached the clinic. Elegant crystallographic studies have demonstrated the potential of designing highly potent and selective NaV1.7 compounds but their therapeutic value remains untested. Transcriptional silencing of NaV1.7 by a naturally expressed antisense transcript has been reported in rodents and humans but whether this represents a viable opportunity for designing NaV1.7 therapeutics is currently unknown. The demonstration that loss of NaV1.7 function is associated with upregulation of endogenous opioids and potentiation of mu- and delta-opioid receptor activities, suggests that targeting only NaV1.7 may be insufficient for analgesia. However, the link between opioid-dependent analgesic mechanisms and function of sodium channels and intracellular sodium-dependent signaling remains controversial. Thus, additional new targets - regulators, modulators - are needed. In this context, we mine the literature for the known interactome of NaV1.7 with a focus on protein interactors that affect the channel's trafficking or link it to opioid signaling. As a case study, we present antinociceptive evidence of allosteric regulation of NaV1.7 by the cytosolic collapsin response mediator protein 2 (CRMP2). Throughout discussions of these possible new targets, we offer thoughts on the therapeutic implications of modulating NaV1.7 function in chronic pain.
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Affiliation(s)
- Lindsey A Chew
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ 85724, USA
| | - Shreya S Bellampalli
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ 85724, USA
| | - Erik T Dustrude
- Stark Neurosciences Research Institute, Indiana University School of Medicine, Indianapolis, IN, USA; Department of Psychiatry, Indiana University School of Medicine, Indianapolis, IN, USA
| | - Rajesh Khanna
- Department of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ 85724, USA; Graduate Interdisciplinary Program in Neuroscience, College of Medicine, University of Arizona, Tucson, AZ, USA; The Center for Innovation in Brain Sciences, The University of Arizona Health Sciences, Tucson, AZ 85724, USA.
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50
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Moutal A, Kalinin S, Kowal K, Marangoni N, Dupree J, Lin SX, Lis K, Lisi L, Hensley K, Khanna R, Feinstein DL. Neuronal Conditional Knockout of Collapsin Response Mediator Protein 2 Ameliorates Disease Severity in a Mouse Model of Multiple Sclerosis. ASN Neuro 2019; 11:1759091419892090. [PMID: 31795726 PMCID: PMC6893573 DOI: 10.1177/1759091419892090] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/20/2019] [Revised: 10/23/2019] [Accepted: 11/02/2019] [Indexed: 01/17/2023] Open
Abstract
We previously showed that treatment with lanthionine ketimine ethyl ester (LKE) reduced disease severity and axonal damage in an experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis and increased neuronal maturation and survival in vitro . A major target of LKE is collapsin response mediator protein 2 (CRMP2), suggesting this protein may mediate LKE actions. We now show that conditional knockout of CRMP2 from neurons using a CamK2a promoter to drive Cre recombinase expression reduces disease severity in the myelin oligodendrocyte glycoprotein (MOG)35–55 EAE model, associated with decreased spinal cord axonal damage, and less glial activation in the cerebellum, but not the spinal cord. Immunohistochemical staining and quantitative polymerase chain reaction show CRMP2 depletion from descending motor neurons in the motor cortex, but not from spinal cord neurons, suggesting that the benefits of CRMP2 depletion on EAE may stem from effects on upper motor neurons. In addition, mice in which CRMP2 S522 phosphorylation was prevented by substitution for an alanine residue also showed reduced EAE severity. These results show that modification of CRMP2 expression and phosphorylation can influence the course of EAE and suggests that treatment with CRMP2 modulators such as LKE act in part by reducing CRMP2 S522 phosphorylation.
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Affiliation(s)
| | | | | | | | | | | | - Kinga Lis
- University of Illinois, Chicago, IL, USA
| | - Lucia Lisi
- Universita Cattolica del Sacro Cuore, Rome,
Italy
| | - Kenneth Hensley
- Arkansas College of Osteopathic Medicine, Fort Smith,
AR, USA
| | | | - Douglas L. Feinstein
- University of Illinois, Chicago, IL, USA
- Jesse Brown VA Medical Center, Chicago, IL, USA
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