Gating mechanisms of TRPC1 channel in hippocampal neurons.

Event Abstract Back to Event Gating mechanisms of TRPC1 channel in hippocampal neurons. Roberta Gualdani1, Sophie Lepannetier1, Anna Kreis1, Olivier Schakman1 and Philippe Gailly1* 1 Université Catholique de Louvain, Belgium The canonical family of Transient Receptor Potential (TRPC) proteins owns seven members (TRPC1 to TRPC7) that form homo- and/or heterotetrameric Ca2+-permeable, nonselective cation membrane channels (Owsianik et al., 2006). TRPC channels are widely expressed in the central nervous system but their functions and their gating mechanisms remain unclear. They are able to modulate neuronal excitability and could promote excitotoxicity (Phelan et al., 2012; Phelan et al., 2013; Narayanan et al., 2014). In particular, heterotetrameric complexes made of TRPC1, TRPC4 and or TRPC5 seem involved presynaptically in the efficiency of synaptic transmission in the hippocampus (Broker-Lai et al., 2017). Recently, we reported that mice made deficient in TRPC1 expression (Trpc1-/- mice) showed alterations in spatial working memory and fear conditioning (Lepannetier et al., 2017). On brain slices, we also observed that LTP triggered by a theta burst could not maintain over time in Trpc1-/- mice. Interestingly, Li et al. recently found that TRPC1 deletion aggravated learning and memory deficits induced by amyloid-β, the major component of senile plaques observed in the brains of Alzheimer disease patients (Li et al., 2018). The present study was therefore focused on the gating mechanisms of TRPC1. We observed that stimulation of neurons with the selective agonist of group I mGluR dihydroxyphenylglycine (DHPG, 50 µM) induced an intracellular Ca2+ transient that was slightly reduced by pre-treatment with 10µM LY367385, a specific inhibitor of mGluR1 but completely abolished by 50 µM MPEP, a specific inhibitor of mGluR5. The response to DHPG was significantly reduced in neurons from Trpc1-/- mice. Emptying the stores of Ca2+ with the SERCA inhibitor thapsigargin triggered an influx of Ca2+ that was slightly reduced in neurons from Trpc1-/- mice, suggesting that TRPC1 acts at least partially as a store-operated channel, as shown in other cell types (Ong et al., 2017; Tajeddine et al., 2012). However, after inhibition of the mGluR-induced phospholipase C pathway by 10 µM U73122, DHPG still induced a Ca2+ response that depended on the presence of external Ca2+ and that was not observed in neurons from Trpc1-/- mice suggesting that DHPG also induced an entry of Ca2+ through TRPC1 that was independent of ER store depletion. As expected, DHPG was still able to induce an entry of Ca2+ after emptying the stores with thapsigargin in neurons from WT animals. This store-independent entry of Ca2+ induced by DHPG was not observed in neurons from Trpc1-/- mice. Electrophysiological experiments were then performed to characterize DHPG-induced response. Isolated hippocampal neurons from WT and Trpc1-/- mice were voltage-clamped at -60mV. To prevent neuronal activity, experiments were performed in the presence of inhibitors of Na+ voltage-dependent channels, AMPA, NMDA and GABA receptors. Moreover, to prevent the activation of store-dependent entry of Ca2+, the PLC inhibitor U-73122 (5 µM) was also added in the extracellular medium. In these conditions, stimulation of the wild type cells with 50 µM DHPG induced an inward current that was significantly diminished in neurons isolated from Trpc1-/- mice (1.13 ± 0.22 pA/pF, n=5 in WT vs 0.23 ± 0.06 pA/pF, n= 5 in Trpc1-/- neurons). In current clamp mode, stimulation of WT neurons with 50 µM DHPG depolarized the cells by about 11 mV. This depolarization was significantly decreased although not abolished in Trpc1-/- neurons. To examine the consequences on neuronal excitability, we measured spontaneous excitatory postsynaptic current of neurons kept in culture and treated with inhibitors of GABA receptors. We observed that stimulation of the cells with 50 µM DHPG increased by a factor of 3 the firing frequency. In conclusion, our data suggest that, in hippocampal neurons, stimulation of mGluR5 receptor activates both a store-operated and a store-independent entry of cations through TRPC1 channel, resulting in a depolarization of the membrane potential and in an increased synaptic excitability. Acknowledgements This work was supported by the Belgian Fund for Scientific Research (FNRS, grants CDR J0080.17 and EQP U.N011.17), the Interuniversity Poles of Attraction Belgian Science Policy (P7/13), the “Fondation pour la Recherche sur la Maladie d’Alzheimer” (SAO/FRA) and the Concerted Research Action from the General Direction of Scientific Research of the French Community of Belgium (ARC17/22-083). References Broker-Lai J, Kollewe A, Schindeldecker B, Pohle J, Nguyen Chi V, Mathar I, Guzman R, Schwarz Y, Lai A, Weissgerber P, Schwegler H, Dietrich A, Both M, Sprengel R, Draguhn A, Kohr G, Fakler B, Flockerzi V, Bruns D, Freichel M (2017) Heteromeric channels formed by TRPC1, TRPC4 and TRPC5 define hippocampal synaptic transmission and working memory. The EMBO journal 36:2770-2789. Lepannetier S, Schakman O, Seghers F and Gailly P (2017). Role of TRPC1 ion channel in hippocampal neurons. Front. Neurosci. Conference Abstract: 12th National Congress of the Belgian Society for Neuroscience. doi: 10.3389/conf.fnins.2017.94.00031 Li M, Liu E, Zhou Q, Li S, Wang X, Liu Y, Wang L, Sun D, Ye J, Gao Y, Yang X, Liu J, Yang Y, Wang JZ. (2018) TRPC1 Null Exacerbates Memory Deficit and Apoptosis Induced by Amyloid-β. Journal of Alzheimers Disease 63:761-772. Narayanan KL, Subramaniam S, Bengston CP, Irmady K, Unsicker K, von Bohlen und Halbach O (2014) Role of transient receptor potential channel 1 (TRPC1) in glutamate-induced cell death in the hippocampal cell line HT22. Journal of molecular neuroscience 52:425-433. Ong HL, Ambudkar IS (2017) STIM-TRP Pathways and Microdomain Organization: Contribution of TRPC1 in Store-Operated Ca(2+) Entry: Impact on Ca(2+) Signaling and Cell Function. Advances in experimental medicine and biology 993:159-188. Owsianik G, D'Hoedt D, Voets T, Nilius B (2006) Structure-function relationship of the TRP channel superfamily. Reviews of physiology, biochemistry and pharmacology 156:61-90. Phelan KD, Mock MM, Kretz O, Shwe UT, Kozhemyakin M, Greenfield LJ, Dietrich A, Birnbaumer L, Freichel M, Flockerzi V, Zheng F (2012) Heteromeric canonical transient receptor potential 1 and 4 channels play a critical role in epileptiform burst firing and seizure-induced neurodegeneration. Molecular pharmacology 81:384-392. Phelan KD, Shwe UT, Abramowitz J, Wu H, Rhee SW, Howell MD, Gottschall PE, Freichel M, Flockerzi V, Birnbaumer L, Zheng F (2013) Canonical transient receptor channel 5 (TRPC5) and TRPC1/4 contribute to seizure and excitotoxicity by distinct cellular mechanisms. Molecular pharmacology 83:429-438. Tajeddine N, Gailly P (2012) TRPC1 protein channel is major regulator of epidermal growth factor receptor signaling. The Journal of biological chemistry 287:16146-16157. Keywords: TRP (A, C, M, ML, N, P, V), transient receptor potential (ankyrin, canonical, melastatin, mucolipin, no mechanoreceptor potential C, polycystic, vanilloid), working memory, spatial memory, synaptic plasiticty, Store-operated Ca entry (SOCE) 2+ Conference: Belgian Brain Congress 2018 — Belgian Brain Council, LIEGE, Belgium, 19 Oct - 19 Oct, 2018. Presentation Type: e-posters Topic: NOVEL STRATEGIES FOR NEUROLOGICAL AND MENTAL DISORDERS: SCIENTIFIC BASIS AND VALUE FOR PATIENT-CENTERED CARE Citation: Gualdani R, Lepannetier S, Kreis A, Schakman O and Gailly P (2019). Gating mechanisms of TRPC1 channel in hippocampal neurons.. Front. Neurosci. Conference Abstract: Belgian Brain Congress 2018 — Belgian Brain Council. doi: 10.3389/conf.fnins.2018.95.00011 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 30 Jul 2018; Published Online: 17 Jan 2019. * Correspondence: Prof. Philippe Gailly, Université Catholique de Louvain, Louvain-la-Neuve, Walloon Brabant, 1348, Belgium, philippe.gailly@uclouvain.be Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Roberta Gualdani Sophie Lepannetier Anna Kreis Olivier Schakman Philippe Gailly Google Roberta Gualdani Sophie Lepannetier Anna Kreis Olivier Schakman Philippe Gailly Google Scholar Roberta Gualdani Sophie Lepannetier Anna Kreis Olivier Schakman Philippe Gailly PubMed Roberta Gualdani Sophie Lepannetier Anna Kreis Olivier Schakman Philippe Gailly Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.