Ontogeny of glutamate-stimulated phosphoinositide turnover in brain and its response to neuronal injury.

Abstract

This dissertation describes the effects of neuronal injury on glutamate receptor-coupled phosphoinositide (PPI) turnover in immature rats. Glutamate is the major excitatory neurotransmitter in the mammalian CNS. Overstimulation of glutamate receptors elicits excitotoxicity that may contribute to the neuronal injury resulting from cerebral ischemia. In the immature rat brain, the glutamate agonist quisqualate (QUIS) potently stimulates PPI hydrolysis. To study the mechanisms of ischemic brain injury and glutamate excitotoxicity, I characterized QUIS-stimulated PPI turnover in the in vivo animal models of perinatal brain injury: (1) ischemic-hypoxic brain injury and (2) N-methyl-D-aspartate (NMDA)-induced brain injury. In the first model, 7-day-old rats were exposed to 8% O$\sb2$ for 2 h after the right carotid artery was ligated. This insult produced neuronal injury in the ipsilateral hippocampus and striatum. In the second model, NMDA (17 nmole) was injected into the right striatum of 7-day-old rats. Iscehmia-hypoxia and NMDA produced brain lesions with similar pathology. QUIS-stimulated $\sp3$H-inositol phosphate release from hippocampal and striatal tissue slices in animals sacrificed 1 day after the ischemic-hypoxic insult or 3 days after NMDA injection was assayed. Both ischemia-hypoxia and NMDA-induced brain lesions doubled QUIS-stimulated PPI hydrolysis in the lesioned hippocampal and striatal slices. Carbachol, a cholinergic agonist which elicits PPI hydrolysis, did not preferentially stimulate lesioned tissue; this observation suggested that the enhanced PPI turnover was specific to the glutamate receptor. The EC$\sb{\rm 50s}$ of QUIS in stimulating $\sp3$H-IP$\sb1$ release were the same for both lesioned and unlesioned tissue. Extracellular Ca$\sp{2+}$ concentration influenced the extent of QUIS-stimulated PPI hydrolysis in lesioned tissue. Low extracellular Ca$\sp{2+}$ concentration (1 $\mu$M) further increased QUIS-stimulated PPI hydrolysis in NMDA-lesioned hippocampal and striatal slices by 50-60%, whereas carbachol-stimulated $\sp3$H-IP$\sb1$ release was unchanged. These results indicate that in two models of neuronal injury in perinatal rats the efficiency of glutamate receptor coupling to PPI hydrolysis increases acutely. However, the molecular mechanisms underlying this enhanced coupling are still unclear.