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• Most rat cerebrovascular pericytes are α-SMA positive. • CYP4A and α-SMA coexpress in rat cerebral VSMCs and most pericytes. • 20-HETE is positively correlated with cerebral mural cell contractility. • 20-HETE promotes the myogenic response of rat parenchymal arterioles. • 20-HETE enhances CBF autoregulation in rat surface and deep cortex. We previously reported that deficiency in 20-HETE or CYP4A impaired the myogenic response and autoregulation of cerebral blood flow (CBF) in rats. The present study demonstrated that CYP4A was coexpressed with alpha-smooth muscle actin (α-SMA) in vascular smooth muscle cells (VSMCs) and most pericytes along parenchymal arteries (PAs) isolated from SD rats. Cell contractile capabilities of cerebral VSMCs and pericytes were reduced with a 20-HETE synthesis inhibitor, HET0016, but restored with 20-HETE analog WIT003. Similarly, intact myogenic responses of the middle cerebral artery and PA of SD rats decreased with HET0016 and were rescued by WIT003. The myogenic response of the PA was abolished in SS and was restored in SS.BN5 and SS. Cyp4a1 rats. HET0016 enhanced CBF and impaired its autoregulation in the surface and deep cortex of SD rats. These results demonstrate that 20-HETE has a direct effect on cerebral mural cell contractility that may play an essential role in controlling cerebral vascular function. [ABSTRACT FROM AUTHOR]