Downregulation of calcium-dependent NMDA receptor desensitization by sodium-calcium exchangers: a role of membrane cholesterol

Dmitry A Sibarov,Sergei M Antonov,Ekaterina E Poguzhelskaya

doi:10.1186/s12868-018-0475-3

Dmitry A Sibarov, Sergei M Antonov + Show 1 more

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https://doi.org/10.1186/s12868-018-0475-3

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Abstract

BackgroundThe plasma membrane Na+/Ca2+-exchanger (NCX) has recently been shown to regulate Ca2+-dependent N-methyl-d-aspartate receptor (NMDAR) desensitization, suggesting a tight interaction of NCXs and NMDARs in lipid nanoclasters or “rafts”. To evaluate possible role of this interaction we studied effects of Li+ on NMDA-elicited whole-cell currents and Ca2+ responses of rat cortical neurons in vitro before and after cholesterol extraction by methyl-β-cyclodextrin (MβCD).ResultsSubstitution Li+ for Na+ in the external solution caused a concentration-dependent decrease of steady-state NMDAR currents from 440 ± 71 pA to 111 ± 29 pA in 140 mM Na+ and 140 mM Li+, respectively. The Li+ inhibition of NMDAR currents disappeared in the absence of Ca2+ in the external solution (Ca2+-free), suggesting that Li+ enhanced Ca2+-dependent NMDAR desensitization. Whereas the cholesterol extraction with MβCD induced a decrease of NMDAR currents to 136 ± 32 pA in 140 mM Na+ and 46 ± 15 pA in 140 mM Li+, the IC50 values for the Li+ inhibition were similar (about 44 mM Li+) before and after this procedure. In the Ca2+-free Na+ solution the steady-state NMDAR currents after the cholesterol extraction were 47 ± 6% of control values. Apparently this amplitude decrease was not Ca2+-dependent. In the Na+ solution containing 1 mM Ca2+ the Ca2+-dependent NMDAR desensitization was greater when cholesterol was extracted. Obviously, this procedure promoted its development. In agreement, Li+ and KB-R7943, an inhibitor of NCX, both considerably reduced NMDA-activated Ca2+ responses. The cholesterol extraction itself caused a decrease of NMDA-activated Ca2+ responses and, in addition, abolished the effects of Li+ and KB-R7943. The cholesterol loading into the plasma membrane caused a recovery of the KB-R7943 effects.ConclusionsTaken together our data suggest that NCXs downregulate the Ca2+-dependent NMDAR desensitization. Most likely, this is determined by a tight functional interaction of NCX and NMDAR molecules because of their co-localization in membrane lipid rafts. The destruction of these rafts is accompanied by an enhancement of NMDAR desensitization and a loss of NCX-selective agent effects on NMDARs.

Highlights

The plasma membrane Na+/Ca2+-exchanger (NCX) has recently been shown to regulate Ca2+-dependent N-methyl-d-aspartate receptor (NMDAR) desensitization, suggesting a tight interaction of N a+/Ca2+ exchanger (NCX) and N-methyl-d-aspartate activated glutamate receptors (NMDARs) in lipid nanoclasters or “rafts”
The stepwise proportional substitution of Li+ for N a+ in the bathing solution was used to obtain the dose-inhibition curve of NMDA-evoked currents for L i+. With this particular aim the NMDA-activated currents were measured at 0, 21, 42, 70, 112 and 140 mM Li+ in the bathing solution in the same experiment, where 140 mM Li+ corresponded to 100% substitution of L i+ for N a+
The control NMDA-evoked currents, measured at the steady state in the bathing solutions (140 mM Na+) had the amplitude of 440.4 ± 71.9 pA (n = 10), that was significantly (p < 0.001, Student’s two-tailed t test) larger compared to the corresponding value of 111.4 ± 29.1 pA (n = 10) measured at 140 mM Li+ in the external solution

Summary

Introduction

The plasma membrane Na+/Ca2+-exchanger (NCX) has recently been shown to regulate Ca2+-dependent N-methyl-d-aspartate receptor (NMDAR) desensitization, suggesting a tight interaction of NCXs and NMDARs in lipid nanoclasters or “rafts”. N-methyl-d-aspartate activated glutamate receptors (NMDARs) are ligand gated ion channels which naturally transfer currents determined by Na+, K+ and Ca2+ permeation. Ca2+-dependent desensitization of NMDARs represents a feedback regulation of the NMDAR open probability by the C a2+ entry into neurons [4,5,6,7,8]. Calmodulin binds free Ca2+ and interacts with C-terminal domains of NMDAR GluN1 subunits causing the decrease of the channel open probability in the Ca2+ concentration-dependent manner because of Ca2+-dependent NMDAR desensitization [9, for review see 10]. As Li+ is a substrate inhibitor of Na+-dependent neurotransmitter transporters [12, 13]

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