N-methyl-D-aspartate increases cytoplasmic free calcium in mouse hippocampus

The effect of N-methyl-D-aspartate (NMDA) and L-glutamate on the concentration of intracellular free calcium (Cai) and on uptake of the calcium was determined in microsacs and synaptosomes isolated from mouse brain. L-Glutamate and NMDA increased Cai in hippocampal microsacs but had little or no effect on Cai in microsacs, isolated from cortex or cerebellum or in synaptosomes. N-Methyl-D-aspartate also increased uptake of calcium, measured using 45Ca into hippocampal microsacs. The EC50 values for NMDA-stimulated increases in Cai and uptake of calcium in microsacs were about 30 microM. Maximum responses were observed with 100 microM NMDA. Increases in Cai stimulated by NMDA were dependent on extracellular calcium, indicating that NMDA increased Cai in microsacs by increasing conductance through an NMDA receptor-operated cation channel, rather than by releasing calcium from intracellular stores. The NMDA antagonists, 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (MK-801), 2-amino-5-phosphonopentanoic acid (AP-5), magnesium and zinc blocked responses to NMDA. This demonstrates NMDA-mediated effects on ion flux in a cell-free preparation from brain. This preparation may be useful for study of the in vitro effects of drugs or toxins on NMDA receptors in brain.