Treatment with tetrahydrobiopterin (BH4) reduces blood pressure in SHR. In the present study we tested the hypothesis that chronic BH4 reduces blood pressure in male SHR by reducing testosterone biosynthesis mediated by increasing NO. Male SHR, aged 17-18 weeks, intact or castrated, were treated for 1 week with BH4 (20 mg/kg/d, i.p.). After 1 week, mean arterial pressure (MAP), serum testosterone, and nitrate/nitrite excretion (NOx) were measured. MAP was significantly higher in intact males than castrated males (179±2 vs 155±4 mm Hg, p<0.001). In intact males, BH4 caused a 17% reduction in MAP (148±2 mm Hg), had no effect on NOx, and reduced serum testosterone by 85% (24.09±2.37 vs. 3.72±0.73 ng/dL; p<0.001). In castrated males, BH4 had no effect on MAP (152±5 mm Hg), but increased NOx by 38%. When castrated males were supplemented with testosterone, MAP increased to the same level as in intact males (180±7) and BH4 had no effect on MAP (182±7 mm Hg) or NOx. Nitric oxide (NO) has been shown to decrease testosterone biosynthesis. Chronic sodium nitrite (70 mg/kg/d x 1 wk) decreased MAP in intact males (150±4 mm Hg), but had no effect on serum testosterone (21.46±3.08 ng/dL). The data suggest that BH4 reduces testosterone synthesis and thereby reduces MAP in male SHR, an androgen-dependent model of hypertension. The mechanism(s) by which BH4 reduces serum testosterone levels are not clear but the data do not support a role for NO as a mediator.