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Calcium-Dependent Regulation of the Neuronal Glycine Transporter GlyT2 by M2 Muscarinic Acetylcholine Receptors

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Abstract

The neuronal glycine transporter GlyT2 modulates inhibitory glycinergic neurotransmission and plays a key role in regulating nociceptive signal progression. The cholinergic system acting through muscarinic acetylcholine receptors (mAChRs) also mediates important regulations of nociceptive transmission being the M2 subtype the most abundantly expressed in the spinal cord. Here we studied the effect of M2 mAChRs stimulation on GlyT2 function co-expressed in a heterologous system with negligible levels of muscarinic receptor activity. We found GlyT2 is down-regulated by carbachol in a calcium-dependent manner. Different components involved in cell calcium homeostasis were analysed to establish a role in the mechanism of GlyT2 inhibition. GlyT2 down-regulation by carbachol was increased by thapsigargin and reduced by internal store depletion, although calcium release from endoplasmic reticulum or mitochondria had a minor role on GlyT2 inhibition. Our results are consistent with a GlyT2 sensitivity to intracellular calcium mobilized by M2 mAChRs in the subcortical area of the plasma membrane. A crucial role of the plasma membrane sodium calcium exchanger NCX is proposed.

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Acknowledgments

The authors are grateful to FJ Díez-Guerra and JM Cuezva for generous support. The confocal microscopy facility at the CBMSO (Madrid, Spain) is also acknowledged for valuable and expert help with the confocal microscopy work. Laura Contreras (CBMSO, Madrid) is acknowledged for expert assistance with the calcium imaging in COS7 cells. Authors want to acknowledge the important contribution of Professor Baruch I. Kanner to the field of neurotransmitter transporters and are honored for his friendship.

Funding

This work was supported by grants of the Spanish ‘Ministerio de Economía y Competitividad’, grant number SAF2017-84235-R (AEI/FEDER, EU) to B.L.-C. and by institutional grants from the Fundación Ramón Areces and Banco de Santander to the CBMSO.

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  1. Amparo Fornés

    Present address: Novartis Farmacéutica S.A., Basel, Switzerland

  2. Esperanza Jiménez and Amparo Fornés have Contributed equally to the work.

Authors and Affiliations

  1. Departamento de Biología Molecular and Centro de Biología Molecular “Severo Ochoa”, Consejo Superior de Investigaciones Científicas-Universidad Autónoma de Madrid, 28049, Madrid, Spain

    Esperanza Jiménez, Amparo Fornés, Raquel Felipe, Enrique Núñez, Carmen Aragón & Beatriz López-Corcuera

  2. IdiPAZ-Hospital Universitario La Paz, Madrid, Spain

    Enrique Núñez, Carmen Aragón & Beatriz López-Corcuera

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  1. Esperanza Jiménez
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Conceived the work: BL-C performed the experiments: EJ, AF, RF and EN analyzed the data: BL-C and CA. Wrote the paper: BL-C.

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Correspondence to Beatriz López-Corcuera.

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Jiménez, E., Fornés, A., Felipe, R. et al. Calcium-Dependent Regulation of the Neuronal Glycine Transporter GlyT2 by M2 Muscarinic Acetylcholine Receptors. Neurochem Res 47, 190–203 (2022). https://doi.org/10.1007/s11064-021-03298-x

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