Extracellular Application of the N-Methyl-D-Aspartate Receptor Allosteric Modulator Rapastinel Acts Remotely to Regulate Ca2+ Inactivation at an Intracellular Locus

Authors

Xiao-Lei Zhang, Department of Cell Biology & Anatomy, New York Medical College, Valhalla, New York, USA.Follow
Nils A. Berglund, Department of Chemistry, Aarhus University, Aarhus, Denmark.
Jeffrey S. Burgdorf, Gate Neurosciences Inc., Carmel, Indiana.
John E. Donello, Gate Neurosciences Inc., Carmel, Indiana.
Joseph R. Moskal, Falk Center for Molecular Therapeutics, Department of Biomedical Engineering, McCormick School of Engineering and Applied Sciences, Northwestern University, Evanston, Illinois, USA.
Patric K. Stanton, Department of Cell Biology & Anatomy, New York Medical College, Valhalla, New York, USA.Follow

Author Type(s)

Faculty

DOI

10.1097/WNR.0000000000001787

Journal Title

Neuroreport

First Page

312

Last Page

319

Document Type

Article

Publication Date

5-4-2022

Department

Cell Biology and Anatomy

Abstract

BACKGROUND: A novel N-methyl-D-aspartate receptor (NMDAR) allosteric modulator, rapastinel (RAP, formerly GLYX-13), elicits long-lasting antidepressant-like effects by enhancing long-term potentiation (LTP) of synaptic transmission. RAP elicits these effects by binding to a unique site in the extracellular region of the NMDAR complex, transiently enhancing NMDAR-gated current in pyramidal neurons of both hippocampus and medial prefrontal cortex. METHODS: We compared efficacy of RAP in modulating Schaffer collateral-evoked NMDAR-currents as a function of kinetics of the Ca2+ chelator in the intracellular solution, using whole-cell patch-clamp recordings. The intracellular solution contained either the slow Ca2+ chelator EGTA [3,12-bis(carboxymethyl)-6,9-dioxa-3,12-diazatetradecane-1,14-dioic acid, 0.5 mmol/l] or the 40-500-fold kinetically faster, more selective Ca2+ chelator BAPTA {2,2',2″,2‴-[ethane-1,2-diylbis(oxy-2,1-phenylenenitrilo)] tetraacetic acid, 5 mmol/l}. NMDAR-gated currents were pharmacologically isolated by bath application of the 2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid receptor antagonist 6-nitro-2,3-dioxo-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide (10 μmol/l) plus the GABA receptor blocker bicuculline (20 μmol/l). RESULTS: When the slow Ca2+ chelator EGTA was in the intracellular solution, RAP elicited significant enhancement of NMDAR-gated current at a 1 μmol/l concentration, and significantly reduced current at 10 μmol/l. In contrast, when recording with the 40-500-fold kinetically faster, more selective Ca2+ chelator BAPTA, NMDAR current increased in magnitude by 84% as BAPTA washed into the cell, and the enhancement of NMDAR current by 1 μmol/l RAP was completely blocked. Interestingly, the reduction in NMDAR current from 10 μmol/l RAP was not affected by the presence of BAPTA in the recording pipette, indicating that this effect is mediated by a different mechanism. CONCLUSION: Extracellular binding of RAP to the NMDAR produces a novel, long-range reduction in affinity of the Ca2+ inactivation site on the NMDAR C-terminus accessible to the intracellular space. This action underlies enhancement in NMDAR-gated conductance elicited by RAP.

Recommended Citation

Zhang, X., Berglund, N. A., Burgdorf, J. S., Donello, J. E., Moskal, J. R., & Stanton, P. K. (2022). Extracellular Application of the N-Methyl-D-Aspartate Receptor Allosteric Modulator Rapastinel Acts Remotely to Regulate Ca2+ Inactivation at an Intracellular Locus. Neuroreport, 33 (7), 312-319. https://doi.org/10.1097/WNR.0000000000001787