Microglial Transient Receptor Potential Melastatin 2 Deficiency Accelerates Seizure Development via Increasing AMPAR-Mediated Neuronal Excitability

Abstract

Epilepsy is one of the most common neurological disorders, characterized by the enhancement of neural excitability from a neurocentric perspective. Emerging evidence indicates that microglia play a pivotal role in the pathogenesis of epilepsy through complex and various mechanisms that is still not fully understood. In this study, we demonstrate that the deficiency of transient receptor potential melastatin 2 (TRPM2) channel, a calcium-permeable nonselective cation channel, significantly accelerates seizure development in multiple mouse seizure models, including MES- and pentylenetetrazole(PTZ)-induced seizure model, intrahippocampal KA model, hippocampal kindling model, without affecting seizure susceptibility in initial acute seizure. Notably, it is the deficiency of TRPM2 specifically in microglia, rather than in CaMKIIα⁺ excitatory neurons or PV⁺ interneurons, that primarily responsible for seizure development. Moreover, microglial TRPM2 deficiency increases the excitability of hippocampal pyramidal neurons by enhancing the AMPAR-mediated excitatory synaptic transmission independent of changes in the expression of inflammatory cytokines. These findings reveal a previously unrecognized, inflammation-independent mechanism by which microglial instead of neuronal TRPM2 channel contributes to seizure development, highlighting microglial TRPM2 as a novel potential therapeutic target for epilepsy by specifically targeting microglial TRPM2 channel.

Keywords

TRPM2 channel / seizure / microglia / AMPAR / neuron

References

[1] O. Devinsky, A. Vezzani, T. J. O'Brien, et al., “Epilepsy,” Nature Reviews Disease primers 4 (2018): 18024.

[2] Y. Wang and Z. Chen, “An Update for Epilepsy Research and Antiepileptic Drug Development: Toward Precise Circuit Therapy,” Pharmacology & Therapeutics 201 (2019): 77-93.

[3] J. T. Paz and J. R. Huguenard, “Microcircuits and Their Interactions in Epilepsy: Is the Focus out of Focus?,” Nature Neuroscience 18, no. 3 (2015): 351-359.

[4] B. P. Bean, “The Action Potential in Mammalian central Neurons,” Nature Reviews Neuroscience 8, no. 6 (2007): 451-465.

[5] A. V. Tzingounis and J. I. Wadiche, “Glutamate Transporters: Confining Runaway Excitation by Shaping Synaptic Transmission,” Nature Reviews Neuroscience 8, no. 12 (2007): 935-947.

[6] G. Buzsaki, K. Kaila, and M. Raichle, “Inhibition and Brain Work,” Neuron 56, no. 5 (2007): 771-783.

[7] J. Oyrer, S. Maljevic, I. E. Scheffer, S. F. Berkovic, S. Petrou, and C. A. Reid, “Ion Channels in Genetic Epilepsy: From Genes and Mechanisms to Disease-Targeted Therapies,” Pharmacological Reviews 70, no. 1 (2018): 142-173.

[8] R. H. Thomas and S. F. Berkovic, “The Hidden Genetics of Epilepsy-a Clinically Important New Paradigm,” Nature Reviews Neurology 10, no. 5 (2014): 283-292.

[9] R. D. Thijs, R. Surges, T. J. O'Brien, and J. W. Sander, “Epilepsy in Adults,” Lancet 393, no. 10172 (2019): 689-701.

[10] F. A. C. Azevedo, L. R. B. Carvalho, L. T. Grinberg, et al., “Equal Numbers of Neuronal and Nonneuronal Cells Make the human Brain an Isometrically Scaled-up Primate Brain,” Journal of Comparative Neurology 513, no. 5 (2009): 532-541.

[11] S. Herculano-Houzel, “The Glia/Neuron Ratio: How It Varies Uniformly Across Brain Structures and Species and What That Means for Brain Physiology and Evolution,” Glia 62, no. 9 (2014): 1377-1391.

[12] P. G. Haydon, “GLIA: Listening and Talking to the Synapse,” Nature Reviews Neuroscience 2, no. 3 (2001): 185-193.

[13] X. Yang, S. Xu, Y. Qian, and Q. Xiao, “Resveratrol Regulates Microglia M1/M2 Polarization via PGC-1α in Conditions of Neuroinflammatory Injury,” Brain, Behavior, and Immunity 64 (2017): 162-172.

[14] R. Mancuso, J. Van Den Daele, N. Fattorelli, et al., “Stem-cell-derived human Microglia Transplanted in Mouse Brain to Study human Disease,” Nature Neuroscience 22, no. 12 (2019): 2111-2116.

[15] B. Bassett, S. Subramaniyam, Y. Fan, et al., “Minocycline Alleviates Depression-Like Symptoms by Rescuing Decrease in Neurogenesis in Dorsal Hippocampus via Blocking Microglia Activation/Phagocytosis,” Brain, Behavior, and Immunity 91 (2021): 519-530.

[16] D. C. Patel, B. P. Tewari, L. Chaunsali, and H. Sontheimer, “Neuron-glia Interactions in the Pathophysiology of Epilepsy,” Nature Reviews Neuroscience 20, no. 5 (2019): 282-297.

[17] P. B. Andersson, V. H. Perry, and S. Gordon, “The Kinetics and Morphological Characteristics of the Macrophage-microglial Response to Kainic Acid-induced Neuronal Degeneration,” Neuroscience 42, no. 1 (1991): 201-214.

[18] E. Avignone, L. Ulmann, F. Levavasseur, F. Rassendren, and E. Audinat, “Status Epilepticus Induces a Particular Microglial Activation state Characterized by Enhanced Purinergic Signaling,” Journal of Neuroscience 28, no. 37 (2008): 9133-9144.

[19] E. Avignone, M. Lepleux, J. Angibaud, and U. V. Nägerl, “Altered Morphological Dynamics of Activated Microglia After Induction of Status Epilepticus,” Journal of Neuroinflammation 12 (2015): 202.

[20] T. Hiragi, Y. Ikegaya, and R. Koyama, “Microglia After Seizures and in Epilepsy,” Cells 7, no. 4 (2018): 26.

[21] M. Andoh, Y. Ikegaya, and R. Koyama, “Synaptic Pruning by Microglia in Epilepsy,” Journal of Clinical Medicine 8, no. 12 (2019): 2170.

[22] U. B. Eyo, N. Gu, S. De, H. Dong, J. R. Richardson, and L. J. Wu, “Modulation of Microglial Process Convergence Toward Neuronal Dendrites by Extracellular Calcium,” Journal of Neuroscience 35, no. 6 (2015): 2417-2422.

[23] P. Malko and L. H. Jiang, “TRPM2 channel-mediated Cell Death: An Important Mechanism Linking Oxidative Stress-inducing Pathological Factors to Associated Pathological Conditions,” Redox Biology 37 (2020): 101755.

[24] C. R. Lee, R. P. Machold, P. Witkovsky, and M. E. Rice, “TRPM2 channels Are Required for NMDA-induced Burst Firing and Contribute to H(2)O(2)-dependent Modulation in substantia nigra Pars Reticulata GABAergic Neurons,” Journal of Neuroscience 33, no. 3 (2013): 1157-1168.

[25] J. Zou, J. F. Ainscough, W. Yang, et al., “A Differential Role of Macrophage TRPM2 Channels in Ca(2)(+) Signaling and Cell Death in Early Responses to H(2)O(2),” American Journal of Physiology. Cell Physiology 305, no. 1 (2013): C61-C69.

[26] J. Miyanohara, M. Kakae, K. Nagayasu, et al., “TRPM2 Channel Aggravates CNS Inflammation and Cognitive Impairment via Activation of Microglia in Chronic Cerebral Hypoperfusion,” Journal of Neuroscience 38, no. 14 (2018): 3520-3533.

[27] P. Zong, J. Feng, Z. Yue, et al., “Functional Coupling of TRPM2 and Extrasynaptic NMDARs Exacerbates Excitotoxicity in Ischemic Brain Injury,” Neuron 110, no. 12 (2022): 1944-1958.

[28] S. Y. Ko, S. E. Wang, H. K. Lee, et al., “Transient Receptor Potential Melastatin 2 Governs Stress-induced Depressive-Like Behaviors,” Proceedings of the National Academy of Sciences of the United States of America 116, no. 5 (2019): 1770-1775.

[29] J. N. Pearson-Smith and M. Patel, “Metabolic Dysfunction and Oxidative Stress in Epilepsy,” International Journal of Molecular Sciences 18, no. 11 (2017): 2365.

[30] P. B. McElroy, L. P. Liang, B. J. Day, and M. Patel, “Scavenging Reactive Oxygen Species Inhibits Status Epilepticus-induced Neuroinflammation,” Experimental Neurology 298, no. Pt A (2017): 13-22.

[31] E. J. Shin, J. H. Jeong, Y. H. Chung, et al., “Role of Oxidative Stress in Epileptic Seizures,” Neurochemistry International 59, no. 2 (2011): 122-137.

[32] E. W. Lothman and J. M. Williamson, “Rapid Kindling With Recurrent Hippocampal Seizures,” Epilepsy Research 14, no. 3 (1993): 209-220.

[33] H. Zhang, P. Yu, H. Lin, et al., “The Discovery of Novel ACA Derivatives as Specific TRPM2 Inhibitors That Reduce Ischemic Injury both in Vitro and in Vivo,” Journal of Medicinal Chemistry 64, no. 7 (2021): 3976-3996.

[34] B. Chen, C. Xu, Y. Wang, et al., “A Disinhibitory nigra-parafascicular Pathway Amplifies Seizure in Temporal Lobe Epilepsy,” Nature Communications 11, no. 1 (2020): 923.

[35] K. Yildizhan and M. Naziroglu, “Glutathione Depletion and Parkinsonian Neurotoxin MPP(+)-Induced TRPM2 Channel Activation Play Central Roles in Oxidative Cytotoxicity and Inflammation in Microglia,” Molecular Neurobiology 57, no. 8 (2020): 3508-3525.

[36] S. Smajic, C. A. Prada-Medina, Z. Landoulsi, et al., “Single-cell Sequencing of human Midbrain Reveals Glial Activation and a Parkinson-specific Neuronal state,” Brain 145, no. 3 (2022): 964-978.

[37] A. Chatzikonstantinou, “Epilepsy and the Hippocampus,” Frontiers of Neurology and Neuroscience 34 (2014): 121-142.

[38] K. Borst, A. A. Dumas, and M. Prinz, “Microglia: Immune and Non-immune Functions,” Immunity 54, no. 10 (2021): 2194-2208.

[39] Q. Wang, X. Xue, Z. Huang, and Y. Wang, “Microglia Share the Burden,” Neuroscience Bulletin 38, no. 6 (2022): 695-698.

[40] Y. Ying, L. Gong, X. Tao, et al., “Genetic Knockout of TRPM2 Increases Neuronal Excitability of Hippocampal Neurons by Inhibiting Kv7 Channel in Epilepsy,” Molecular Neurobiology 59, no. 11 (2022): 6918-6933.

[41] N. Spruston, “Pyramidal Neurons: Dendritic Structure and Synaptic Integration,” Nature Reviews Neuroscience 9, no. 3 (2008): 206-221.

[42] Y. F. Xie, J. C. Belrose, G. Lei, et al., “Dependence of NMDA/GSK-3beta Mediated Metaplasticity on TRPM2 Channels at Hippocampal CA3-CA1 Synapses,” Molecular Brain 4 (2011): 44.

[43] I. Alim, L. Teves, R. Li, Y. Mori, and M. Tymianski, “Modulation of NMDAR Subunit Expression by TRPM2 Channels Regulates Neuronal Vulnerability to Ischemic Cell Death,” Journal of Neuroscience 33, no. 44 (2013): 17264-17277.

[44] Q. Zheng, T. Zhu, H. Hu, et al., “TRPM2 ion Channel Is Involved in the Aggravation of Cognitive Impairment and Down Regulation of Epilepsy Threshold in Pentylenetetrazole-induced Kindling Mice,” Brain Research Bulletin 155 (2020): 48-60.

[45] A. Rana and A. E. Musto, “The Role of Inflammation in the Development of Epilepsy,” Journal of Neuroinflammation 15, no. 1 (2018): 144.

[46] A. Vezzani, T. Ravizza, P. Bedner, E. Aronica, C. Steinhäuser, and D. Boison, “Astrocytes in the Initiation and Progression of Epilepsy,” Nature Reviews Neurology 18, no. 12 (2022): 707-722.

[47] Z. Wang, L. Zhou, D. An, et al., “TRPV4-induced Inflammatory Response Is Involved in Neuronal Death in Pilocarpine Model of Temporal Lobe Epilepsy in Mice,” Cell Death & Disease 10, no. 6 (2019): 386.

[48] X. Zhao, Y. Liao, S. Morgan, et al., “Noninflammatory Changes of Microglia Are Sufficient to Cause Epilepsy,” Cell Reports 22, no. 8 (2018): 2080-2093.

[49] S. N. Mandhane, K. Aavula, and T. Rajamannar, “Timed Pentylenetetrazol Infusion Test: A Comparative Analysis With s.c.PTZ and MES Models of Anticonvulsant Screening in Mice,” Seizure: The Journal of the British Epilepsy Association 16, no. 7 (2007): 636-644.

[50] T. Chachua, K. L. Poon, M. S. Yum, et al., “Rapamycin Has Age-, Treatment Paradigm-, and Model-specific Anticonvulsant Effects and Modulates Neuropeptide Y Expression in Rats,” Epilepsia 53, no. 11 (2012): 2015-2025.

[51] P. S. Buckmaster, E. A. Ingram, and X. Wen, “Inhibition of the Mammalian Target of Rapamycin Signaling Pathway Suppresses Dentate Granule Cell Axon Sprouting in a Rodent Model of Temporal Lobe Epilepsy,” Journal of Neuroscience 29, no. 25 (2009): 8259-8269.

[52] P. S. Buckmaster and F. H. Lew, “Rapamycin Suppresses Mossy fiber Sprouting but Not Seizure Frequency in a Mouse Model of Temporal Lobe Epilepsy,” Journal of Neuroscience 31, no. 6 (2011): 2337-2347.

[53] R. Han, X. Lan, Z. Han, et al., “Improving Outcomes in Intracerebral Hemorrhage Through Microglia/Macrophage-targeted IL-10 Delivery With Phosphatidylserine Liposomes,” Biomaterials 301 (2023): 122277.

[54] X. Zheng, K. Sun, Y. Liu, et al., “Resveratrol-loaded Macrophage Exosomes Alleviate Multiple Sclerosis Through Targeting Microglia,” Journal of Controlled Release 353 (2023): 675-684.

[55] J. Gao, Q. Song, X. Gu, et al., “Intracerebral Fate of Organic and Inorganic Nanoparticles Is Dependent on Microglial Extracellular Vesicle Function,” Nature Nanotechnology 19, no. 3 (2024): 376-386.

[56] D. E. Tylawsky, H. Kiguchi, J. Vaynshteyn, et al., “P-selectin-targeted Nanocarriers Induce Active Crossing of the Blood-brain Barrier via Caveolin-1-dependent Transcytosis,” Nature Materials 22, no. 3 (2023): 391-399.

[57] A. M. Vargason, A. C. Anselmo, and S. Mitragotri, “The Evolution of Commercial Drug Delivery Technologies,” Nature Biomedical Engineering 5, no. 9 (2021): 951-967.

[58] L. H. Jiang, X. Li, S. A. Syed Mortadza, M. Lovatt, and W. Yang, “The TRPM2 Channel Nexus From Oxidative Damage to Alzheimer's Pathologies: An Emerging Novel Intervention Target for Age-related Dementia,” Ageing Research Reviews 47 (2018): 67-79.

[59] H. W. Liu, L. N. Gong, K. Lai, et al., “Bilirubin Gates the TRPM2 Channel as a Direct Agonist to Exacerbate Ischemic Brain Damage,” Neuron 111, no. 10 (2023): 1609-1625. e6.

[60] S. Werneburg, P. A. Feinberg, K. M. Johnson, and D. P. Schafer, “A Microglia-cytokine Axis to Modulate Synaptic Connectivity and Function,” Current Opinion in Neurobiology 47 (2017): 138-145.

[61] X. Zhou, R. Zhao, M. Lv, et al., “ACSL4 promotes Microglia-mediated Neuroinflammation by Regulating Lipid Metabolism and VGLL4 Expression,” Brain, Behavior, and Immunity 109 (2023): 331-343.

[62] C. Chen, T. Zhu, L. Gong, et al., “Trpm2 deficiency in Microglia Attenuates Neuroinflammation During Epileptogenesis by Upregulating Autophagy via the AMPK/mTOR Pathway,” Neurobiology of Disease 186 (2023): 106273.

[63] C. Lauro, M. Catalano, F. Trettel, and C. Limatola, “Fractalkine in the Nervous System: Neuroprotective or Neurotoxic Molecule?,” Annals of the New York Academy of Sciences 1351 (2015): 141-148.

[64] Y. Zhan, R. C. Paolicelli, F. Sforazzini, et al., “Deficient Neuron-microglia Signaling Results in Impaired Functional Brain Connectivity and Social Behavior,” Nature Neuroscience 17, no. 3 (2014): 400-406.

[65] R. L. Doser, G. C. Amberg, and F. J. Hoerndli, “Reactive Oxygen Species Modulate Activity-Dependent AMPA Receptor Transport in C. elegans,” The Journal of Neuroscience: the Official Journal of the Society For Neuroscience 40, no. 39 (2020): 7405-7420.

[66] J. Zhang, A. Malik, H. B. Choi, R. W. Y. Ko, L. Dissing-Olesen, and B. A. MacVicar, “Microglial CR3 Activation Triggers Long-term Synaptic Depression in the Hippocampus via NADPH Oxidase,” Neuron 82, no. 1 (2014): 195-207.

[67] S. N. Rakhade, C. Zhou, P. K. Aujla, R. Fishman, N. J. Sucher, and F. E. Jensen, “Early Alterations of AMPA Receptors Mediate Synaptic Potentiation Induced by Neonatal Seizures,” Journal of Neuroscience 28, no. 32 (2008): 7979-7990.

[68] U. Pfisterer, V. Petukhov, S. Demharter, et al., “Identification of Epilepsy-associated Neuronal Subtypes and Gene Expression Underlying Epileptogenesis,” Nature Communications 11, no. 1 (2020): 5038.

[69] T. Eiro, T. Miyazaki, M. Hatano, et al., “Dynamics of AMPA Receptors Regulate Epileptogenesis in Patients With Epilepsy,” Cell Reports Medicine 4, no. 5 (2023): 101020.

[70] F. Fei, X. Wang, C. Xu, et al., “Discrete Subicular Circuits Control Generalization of Hippocampal Seizures,” Nature Communications 13, no. 1 (2022): 5010.

[71] K. B. J. Franklin and G. Paxions, The Mouse Brain in Stereotaxic Coordinates. 2nd ed.. (San Diego: Academic Press, 2011): 1-350.

[72] J. Zhao, Y. Wang, C. Xu, et al., “Therapeutic Potential of an Anti-high Mobility Group Box-1 Monoclonal Antibody in Epilepsy,” Brain, Behavior, and Immunity 64 (2017): 308-319.

[73] S.-J. Dai, Y.-Y. Shao, Y. Zheng, et al., “Inflachromene Attenuates Seizure Severity in Mouse Epilepsy Models via Inhibiting HMGB1 Translocation,” Acta Pharmacologica Sinica 44, no. 9 (2023): 1737-1747.

[74] J. Engel, L. Wolfson, and L. Brown, “Anatomical Correlates of Electrical and Behavioral Events Related to Amygdaloid Kindling,” Annals of Neurology 3, no. 6 (1978): 538-544.

[75] Y. Wang, Y. Wang, C. Xu, et al., “Direct Septum-Hippocampus Cholinergic Circuit Attenuates Seizure through Driving Somatostatin Inhibition,” Biological Psychiatry 87, no. 9 (2020): 843-856.

[76] M. Sato, R. J. Racine, and D. C. McIntyre, “Kindling: Basic Mechanisms and Clinical Validity,” Electroencephalography and Clinical Neurophysiology 76, no. 5 (1990): 459-472.

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