Role of P-450 Arachidonic Acid Epoxygenase in the Response of Cerebral Blood Flow to Glutamate in Rats

Abstract

Background and Purpose Glutamate, a major excitatory neurotransmitter in the brain, has been implicated in the hyperemic response to increases in the activity of neurons, but the mechanism of glutamate-induced dilation of cerebral blood vessels is unknown. Glutamate has been shown to enhance the release of arachidonic acid (AA) in brain tissue and cultured astrocytes. We have previously shown that astrocytes metabolize AA to vasodilator products, epoxyeicosatrienoic acids (EETs), and express a P-450 AA epoxygenase, P-450 2C11. We tested the hypothesis that glutamate-induced dilation of cerebral arterioles is mediated in part by changes in the formation and release of EETs by perivascular astrocytes. Methods Primary astrocyte cultures were prepared from 3-day-old rat pups. The cells were labeled with [ 14 C]AA, and the effect of glutamate on the formation of EETs from [ 14 C]AA by cultured astrocytes was studied. The expression of P-450 2C11 protein in the microsomal fractions of cultured astrocytes was assessed by Western blot. In vivo cerebral blood flow measurements were made in adult rats by laser-Doppler flowmetry after administration of glutamate into the subdural space of the rat before and after treatment with miconazole. Results Glutamate treatment (100 μmol/L for 30 minutes) induced a threefold increase in the formation of EETs from [ 14 C]AA by cultured astrocytes, and the increase was inhibited by miconazole (20 μmol/L), an inhibitor of P-450 AA epoxygenase. Treatment with glutamate (100 μmol/L) for 12 hours increased the expression of P-450 2C11 protein in the microsomal fraction of cultured astrocytes. The response of laser-Doppler cerebral blood flow to administration of glutamate (500 μmol/L) into the subdural space of the rat was significantly attenuated after treatment with miconazole (20 μmol/L for 30 minutes). Conclusions These findings suggest a role for a P-450 AA epoxygenase in astrocytes in the coupling between the metabolic activity of neurons and regional blood flow in the brain.