Neuroprotective effects of NMDA receptor glycine recognition site antagonism: dependence on glycine concentration.

High-affinity NMDA receptor glycine recognition site antagonists protect brain tissue from ischemic damage. The neuroprotective effect of 5-nitro-6,7-dichloro-2,3-quinoxalinedione (ACEA 1021), a selective NMDA receptor antagonist with nanomolar affinity for the glycine binding site, was examined in rat cortical mixed neuronal/glial cultures. ACEA 1021 alone did not alter spontaneous lactate dehydrogenase (LDH) release. Treatment with ACEA 1021 (0.1-10 microM) before 500 microM glutamate, 30 microM NMDA, or 300 microM kainate exposure was found to reduce LDH release in a concentration-dependent fashion. These effects were altered by adding glycine to the medium. Glycine (1 mM) partially reversed the effect of ACEA 1021 on kainate cytotoxicity. Glycine (100 microM-1 mM) completely blocked the effects of ACEA 1021 on glutamate and NMDA cytotoxicity. The glycine concentration that produced a half-maximal potentiation of excitotoxin-induced LDH release in the presence of 1.0 microM ACEA 1021 was similar for glutamate and NMDA (18 +/- 3 and 29 +/- 9 microM, respectively). ACEA 1021 also reduced kainate toxicity in cultures treated with MK-801. The effects of glycine and ACEA 1021 on glutamate-induced LDH release were consistent with a model of simple competitive interaction for the strychnine-insensitive NMDA receptor glycine recognition site, although nonspecific effects at the kainate receptor may be of lesser importance.

Effect of halothane in cortical cell cultures exposed to N-methyl-D-aspartate.

In vivo studies have shown potent protection by volatile anesthetic agents against cerebral ischemic insults. Volatile agents have also been shown to antagonize glutamatergic neurotransmission at the N-methyl-D-aspartate (NMDA) receptor. This study examined the potential for halothane to reduce neuronal excitotoxic lesions caused by NMDA. Fetal rat cortical cell cultures were allowed to mature 13-16 d. Culture wells (n = 13-16) were treated with 0 mM - 3.96 mM halothane in the presence/absence of 30 microM NMDA. Additional cultures were exposed to 30 microM NMDA in the presence/absence of 10 microM MK-801 or 10 microM ACEA 1021. Cellular lethality was assessed by measurement of lactate dehydrogenase (LDH) 24 hrs later. A maximal effect of halothane was observed at 0.70 mM (2.1 vol%) wherein a 36% reduction in NMDA-stimulated LDH release occurred relative to untreated controls. Both MK-801 and ACEA 1021 caused complete inhibition of NMDA-stimulated LDH release. These data confirm that halothane has modulatory effects at the NMDA receptor but potency of this drug is less than that of specific antagonists of either glutamate or glycine. These findings suggest that halothane protection in vivo can be partially explained by anti-excitotoxic properties although other mechanisms of action are probably also important.