Quantitative assessment of neuroprotection against NMDA-induced brain injury

Abstract

In immature rodent brain, unilateral intrastriatal injections of selected excitatory amino acid (EAA) receptor agonists, such as N-methyl--aspartate (NMDA), produce prominent ipsilateral forebrain lesions. In Postnatal Day (PND) 7 rats that receive a right intrastriatal injection of NMDA (25 nmol) and are sacrificed 5 days later, there is a considerable and consistent reduction in the weight of the injected cerebral hemisphere relative to that of the contralateral side (-28.5 +/- 1.9%, n = 6). In animals treated with specific NMDA receptor antagonists, the severity of NMDA-induced damage is markedly reduced. We have previously reported that the efficacy of potential neuroprotective drugs in limiting NMDA-induced lesions can be assessed quantitatively by comparison of hemisphere weights after a unilateral NMDA injection. In this study, we compared three quantitative methods to evaluate the severity of NMDA-induced brain injury and the degree of neuroprotection provided by NMDA receptor antagonists. We characterized the severity of brain injury resulting from intrastriatal injections of 1-50 nmol NMDA in PND 7 rats sacrificed on PND 12 by (i) comparison of cerebral hemisphere weights; (ii) assay of the activity of the cholinergic neuronal marker, choline acetyltransferase (ChAT) activity; and (iii) measurement of regional brain cross-sectional areas. The severity of the resulting brain injury as assessed by comparison of hemisphere weights increased linearly with the amount of NMDA injected into the striatum up to 25 nmol NMDA. The magnitude of injury was highly correlated with the degree of reduction in ChAT activity (r2 = 0.97). Quantification of neuroprotection against NMDA toxicity by measurement of cerebral hemisphere weight disparities was highly correlated with comparisons of hemisphere and striatal cross-sectional areas (r2 = 0.98). The potency and efficacy of the dissociative anesthetic ketamine was easily distinguished from those of two other NMDA receptor antagonists (MK-801, (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-immine maleate and CPP, 3-((+/-)-2-carboxypiperazine-4-yl)-propyl-1-phosphonic acid) using this model. These data demonstrate that this model can be used to accurately quantitate NMDA-induced brain injury and evaluate neuro-protective properties of glutamate antagonists in vivo.