We cultured smooth muscle cells from rat renal preglomerular arterioles by injecting a suspension of iron oxide into the left ventricle, separating the arterioles magnetically, and growing cells from explants. In passaged cultures we ascertained vascular smooth muscle purity of >98% by morphology; contraction to norepinephrine and angiotensin; positive immunofluorescence staining through the sixth passage with monoclonal antibodies to smooth muscle-specific α- and γ-isoactins, myosin, and desmin; and the absence of von Willebrand factor. Angiotensin II (10−12-10−5M) induced dose-dependent DNA synthesis and proliferation of subcultured (three times) arteriolar smooth muscle cells from a growth-arrested state (p<0.01). Angiotensin II (10−5M) also induced the cells to expressc-fosmRNA. We find no previous report of culture of smooth muscle cells from renal preglomerular arterioles. Our findings also provide evidence that angiotensin II is mitogenic to arteriolar muscle cells and thus may be involved in their hyperplasia accompanying hypertension.