α2-Adrenergic agonists reduce glutamate release and glutamate receptor-mediated calcium changes in hippocampal slices during hypoxia

Abstract

The mechanisms by which α2-adrenergic agonists reduce ischemic brain damage are not clear. In ischemia-vulnerable hippocampal neurons we tested whether α2-agonists reduce glutamate efflux and glutamate receptor-mediated increase of cytosolic free calcium. Brain slices (300 μm thick) from rat hippocampus were loaded with fura-2 for measurements of cytosolic free calcium with a microscope fluorometer. Change of cytosolic calcium in CA1 neurons during application of N-methyl-D-aspartate (NMDA) was measured, as were calcium changes during simulated ischemia (hypoxia, NaCN, iodoacetate) or hypoxia plus high glutamate concentration (pO2 = 25 mmHg, 3 mM glutamate). In other slices, glutamate efflux evoked by anoxia (pO2 = 25 mmHg, 100 μM NaCN) was measured. The selective α2-agonists mivazerol (1 μM) decreased NMDA receptor-mediated calcium changes in hippocampal CA1 neurons by 28% (p = 0.0079). With hypoxia and 3 mM glutamate, 1 μM mivazerol reduced early peak calcium changes in CA1 neurons by 57% (p = 0.0007). An α2-antagonist (rauwolscine, 1 μM) blocked this. Mivazerol did not reduce the rate of calcium change during simulated ischemia. Clonidine (0.1 μM), a partial α2-agonists, decreased glutamate/hypoxia-mediated calcium changes in CA1 (p = 0.01), but 1 μM clonidine, which stimulates α1-receptors, did not. Mivazerol decreased hypoxia and KCl-evoked glutamate release by 50% and 75% (p < 0.01), respectively. In addition, 1 μM mivazerol reduced lactate dehydrogenase leakage rate from brain slices during anoxia by 61% (p = 0.018). Thus, α2-receptors influence glutamate release, calcium changes, and cell damage in ischemia-vulnerable hippocampal neurons. These effects may contribute to the cerebroprotective actions of α2-agonists.