Cerebral vascular endothelial heme oxygenase: expression, localization, and activation by glutamate

Endogenous carbon monoxide (CO)contributes to vasodilator responses of cerebral microvessels innewborn pigs. We investigated the expression, intracellularlocalization, and activity of heme oxygenase (HO), the key enzyme in COproduction, in quiescent cerebral microvascular endothelial cells(CMVEC) from newborn pigs. HO-1 and HO-2 isoforms were detected byRT-PCR, immunoblotting, and immunofluorescence. HO-1 and HO-2 aremembrane-bound proteins that have a strong preference for the nuclearenvelope and perinuclear area of the cytoplasm. Betamethasone(10⁶ to 10⁴ M for 48 h) was associatedwith upregulation of HO-2 protein by ~50% and inhibition of Cox-2but did not alter HO-1 or endothelial nitric oxide synthase expressionin CMVEC. In vivo betamethasone treatment of newborn pigs (0.2 and 5.0 mg/kg im for 48 h) upregulated HO-2 in cerebral microvessels by30-60%. HO activity as ¹⁴CO production from[¹⁴C]glycine-labeled endogenous heme was inhibited bychromium mesoporphyrin (10⁶ to 10⁴ M).L-Glutamate (0.3-1.0 mM) stimulated HO activity1.5-fold. High-affinity specific binding sites forL-[³H]glutamate suggestive of the glutamatereceptors were detected in CMVEC. Altogether, these data suggest that,in cerebral circulation of newborn pigs, endothelium-derived CO maycontribute to basal vascular tone and to responses that involveglutamate receptor activation.