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

BACKGROUND: The plasma membrane Na(+)/Ca(2+)-exchanger (NCX) has recently been shown to regulate Ca(2+)-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 Ca(2+) responses of rat cortical neurons in vitro before and after cholesterol extraction by methyl-β-cyclodextrin (MβCD). RESULTS: Substitution 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 Ca(2+) in the external solution (Ca(2+)-free), suggesting that Li(+) enhanced Ca(2+)-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 IC(50) values for the Li(+) inhibition were similar (about 44 mM Li(+)) before and after this procedure. In the Ca(2+)-free Na(+) solution the steady-state NMDAR currents after the cholesterol extraction were 47 ± 6% of control values. Apparently this amplitude decrease was not Ca(2+)-dependent. In the Na(+) solution containing 1 mM Ca(2+) the Ca(2+)-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 Ca(2+) responses. The cholesterol extraction itself caused a decrease of NMDA-activated Ca(2+) 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. CONCLUSIONS: Taken together our data suggest that NCXs downregulate the Ca(2+)-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.