Involvement of the N-methyl-D-aspartate receptor in neuronal cell death induced by cytotoxic T cell-derived secretory granules

The mechanisms underlying neurotoxicity mediated by cytotoxic T lymphocytes (CTL) and their secretory granule proteins perforin and granzymes remain u nclear. We evaluated the possible role of the neurotransmitter glutamate in cell death observed in differentiated neurons exposed to CTL-derived granu les. Excitotoxicity induced by excessive concentrations of extracellular gl utamate is associated with a rise in intracellular calcium and can lead to generation of NO through the activation of glutamatergic N-methyl-D-asparta te (NMDA) receptors. Consistent with an involvement of glutamate, we found that cell death in mature cerebral granule cells was inhibited by 65-80% by two NMDA receptor blockers (MK-801 and D-2-amino-5-phosphonovaleric acid) or a NO synthase blocker (N-G-nitro-L-arginine methyl ester). Furthermore, neurons treated with secretory granules responded with a biphasic rise in t he intracellular calcium concentration ([Ca2+](i)). Whereas MK-801 did not interfere with the immediate rise of [Ca2+](i), the second wave of calcium accummulation starting at 40 min was delayed by 20 min and reduced in ampli tude in the presence of MK-801. In immature, NMDA receptor-negative neurons , MK-801 prevented neither the cytotoxicity nor the calcium influx observed 5 min after addition of cytotoxic granules. The demonstration that NMDA re ceptors and NO are involved in granule-mediated killing of mature neurons o pens new avenues in the treatment of neuronal cell death in CTL-mediated di seases such as viral encephalitis.