Regulatory Mechanism of Endothelin Receptor B in the Cerebral Arteries after Focal Cerebral Ischemia

BACKGROUND AND PURPOSE: Increased expression of endothelin receptor type B (ET(B)R), a vasoactive receptor, has recently been implied in the reduced cerebral blood flow and exacerbated neuronal damage after ischemia-reperfusion (I/R). The study explores the regulatory mechanisms of ET(B)R to identify drug targets to restore normal cerebral artery contractile function as part of successful neuroprotective therapy. METHODS: We have employed in vitro methods on human and rat cerebral arteries to study the regulatory mechanisms and the efficacy of target selective inhibitor, Mithramycin A (MitA), to block the ET(B)R mediated contractile properties. Later, middle cerebral artery occluded (MCAO) rats were used to substantiate the observations. Quantative PCR, immunohistochemistry, western blot and wire myograph methods were employed to study the expression and contractile properties of cerebral arteries. RESULTS: Increased expression of specificity protein (Sp1) was observed in human and rat cerebral arteries after organ culture, strongly correlating with the ET(B)R upregulation. Similar observations were made in MCAO rats. Treatment with MitA, a Sp1 specific inhibitor, significantly downregulated the ET(B)R mRNA and protein levels. It also significantly reduced the ET(B)R mediated cerebrovascular contractility. Detailed analysis indicated that ERK1/2 mediated phosphorylation of Sp1 might be essential for ET(B)R transcription. CONCLUSION: Transcription factor Sp1 regulates the ET(B)R mediated vasoconstriction in focal cerebral ischemia via MEK-ERK signaling, which is also conserved in humans. The results show that MitA can effectively be used to block ET(B)R mediated vasoconstriction as a supplement to an existing ischemic stroke therapy.