Effects of isoflurane and hypothermia on glutamate receptor-mediated calcium influx in brain slices

To understand how volatile anesthetics protect neurons during cerebral ischemia, we studied the effects of isoflurane on cerebral glutamate receptor-mediated calcium influx. Calcium influx via these key excitatory receptors may mediate pain transmission, memory, and the pathophysiologic sequelae of cerebral anoxia or ischemia. Because cerebral protection by hypothermia may involve a decrease in glutamate receptor activity, we also examined the interaction of temperature and isoflurane on glutamate receptor inhibition. We measured glutamate receptor-mediated changes in cytosolic calcium in 300-microns-thick rat cortical brain slices. Temperature was varied to 28, 34, 37, or 39 degrees C and isoflurane partial pressure to 0.016-0.019 atm (equivalent to 1.16 minimum alveolar concentration [MAC], adjusted for temperature and age). Brain slices were loaded with fura-2 to permit measurement of cytosolic free calcium. Calcium changes due to the glutamate receptor agonist N-methyl-D-aspartate (NMDA) (50 microM), to ischemia levels of L-glutamate (1.0 mM) or to simulated ischemia (1.0 mM glutamate, 100 microM NaCN, and 3.5 mM iodoacetate) was then measured. Slice lactate dehydrogenase leakage and adenosine triphosphate were measured as indices of cellular integrity. Isoflurane reduced both L-glutamate and NMDA-mediated calcium fluxes by approximately 60%. Neither the activity of the NMDA receptor nor its inhibition by isoflurane was altered by temperature. The rate of calcium influx during ischemia was significantly reduced both by temperature and by isoflurane (P < 0.05). Adenosine triphosphate loss and lactate dehydrogenase leakage were reduced by isoflurane during simulated ischemia by 37% and 73% (P < 0.05), respectively. (1) At 1.16 MAC, isoflurane potently inhibits glutamate receptors and delays cellular injury induced by simulated ischemia, and (2) hypothermia does not reduce the intrinsic activity of cortical glutamate receptors but delays calcium accumulation during simulated ischemia. Isoflurane reduces the severity of key pathophysiologic events in an in vitro model of simulated cerebral ischemia.