Effect of magnesium on NMDA mediated toxicity and increases in [Ca2+]i and cGMP in cultured neocortical neurons: evidence for distinct regulation of different responses.

The effect of varying external concentrations of Mg2+ has been studied on NMDA induced toxicity, increases in the intracellular calcium concentration, [Ca2+]i, and cGMP production in cultured neocortical neurons. The neurotoxic potency of NMDA during a 5 h exposure period in 7-day-old cultures was examined in three different exposure media: phosphate buffered saline, PBS (137 mM NaCl, 2.7 mM KCl, 7.3 mM Na2HPO4, 1.5 mM KH2PO4, 0.9 mM CaCl2, 0.6 mM MgCl2; pH 7.4); PBS without addition of Mg2+; and Neuronal Dulbecco's Minimal Essential Medium (NDMEM = DMEM with a final concentration of KCl of 25.5 mM KCl; cf. Experimental Procedures). In the presence of Mg2+, no toxicity of NMDA was observed in PBS (ED50 > 1000 microM) whereas omission of Mg2+ in PBS resulted in an ED50 value for NMDA of 9 +/- 3 microM. Using NDMEM as the exposure medium, an intermediate neurotoxic potency of NMDA (ED50 = 40 +/- 5 microM) was observed. This intermediate value is probably due to partial attenuation of the Mg2+ block of the NMDA associated cation channel by the depolarizing conditions in NDMEM (with 25.5 mM KCl). Diminishing the external Mg2+ concentration also potentiated the increase in [Ca2+]i after stimulation with NMDA. This potentiation was maximally 3-fold (obtained at 300 microM NMDA), and the IC50 was 15 +/- 2 microM. Surprisingly, the NMDA induced production of cGMP was not sensitive to variations in the external Mg2+ concentration. These findings of distinct regulatory mechanisms of different NMDA receptor coupled responses may indicate the existence of several types of NMDA receptors.