Ethanol inhibition of NMDA receptors in calcium-dependent and -independent modes

N-methyl-d-aspartate receptor (NMDAR) is an essential target for ethanol action in the central nervous system (CNS). Whereas an alcohol addiction treatment represents a severe medical problem, many aspects of ethanol action at physiologically relevant concentrations on NMDARs are still unclear. Here using the whole-cell patch-clamp recording on cortical neurons in the primary culture, we studied inhibition of NMDAR currents by different ethanol concentrations ([Et]s) and its dependence on extracellular Ca²⁺. The ethanol action on NMDA-activated currents exhibited a biphasic concentration-inhibition relationship in the presence of extracellular Ca²⁺. The high-affinity region of the curve was found within the range of [Et]s from 9 mM to 30 mM and was characterized by IC_50,H of about 20 mM. The low-affinity region was observed within the range of [Et]s from 85 mM to 200 mM with IC_50,L of about 150 mM. In the absence of extracellular Ca²⁺, the ethanol concentration-inhibition relationship became monophasic, with IC_50,L of about 200 mM, since the high-affinity component disappeared. A substitution of Li⁺ for Na⁺ in the bathing solution and an extraction of cholesterol from the plasma membrane with methyl-β-cyclodextrin, which are the treatments that both promote the Ca²⁺-dependent desensitization (CDD) of NMDARs, abolished the high-affinity Ca²⁺-dependent component of the NMDAR ethanol inhibition. Besides, this component was not observed when neurons were loaded with BAPTA. These data suggest that most likely, ethanol at low concentrations enhances the NMDAR CDD. In agreement when the dependence of the NMDAR CDD on extracellular Ca²⁺ was directly measured 22 mM ethanol enhanced the NMDAR CDD since an extracellular Ca²⁺ concentration that caused 50% of the NMDAR CDD decreased almost 3-folds from 0.81 mM to 0.28 mM, and an extent of the CDD was also more pronounced. The low-affinity component of the NMDAR ethanol inhibition was resistant to the above treatments suggesting CDD-independent direct action on NMDARs. Thus, at a physiologically relevant concentration of extracellular Ca²⁺ and ethanol that could be reached in the blood during light-mild human alcohol intoxication, ethanol causes an enhancement of the NMDAR CDD, which could be in general accompanied by some disruptions of the CNS excitatory system.