Release of [3H]norepinephrine from rat hippocampal slices by N-methyl-D-aspartate: comparison of the inhibitory effects of Mg2+ and MK-801

Abstract

The excitatory amino acid receptor subtype activated by N-methyl-D-aspartic acid (NMDA) was studied using superfused slices from the rat hippocampus preloaded with [3H]norepinephrine. NMDA-induced release was inhibited by the direct receptor antagonist CPP, and was sensitive to physiological concentrations of Mg2+. NMDA-induced transmitter release in the presence of Mg2+ was demonstrable if the slices were first depolarized by exposure to elevated K+ or kainic acid to relieve the voltage-dependent Mg2+ blockade. Transmitter release was also inhibited by the indirectly acting antagonists MK-801 and phencyclidine. This effect of MK-801 showed use dependence, while inhibition of release by Mg2+ remained at a constant level with repeated agonist application. Kinetic analysis indicated the mechanism of MK-801 inhibition was uncompetitive in that agonist was required for the association of the inhibitor with the receptor-channel complex. In contrast, Mg2+ inhibited NMDA-induced transmitter release through a noncompetitive process. The two antagonists also differed in terms of reversibility with inhibition by Mg2+ being evident only in the presence of the cation. The effect of MK-801, however, was still apparent for several stimuli after removal of the drug. These results demonstrate the utility in this in vitro release system for studying the unique characteristics of the NMDA receptor complex.