Central infusions of benzamil prevent/reverse salt-induced hypertension in genetic models of salt-sensitive hypertension. Benzamil acts by blockade of ion—presumably sodium—channels. In the present study, we assessed in Dahl salt-sensitive (S) rats on high salt intake whether these channels mediate increases in brain “ouabain” and, thereby, hypertension. Intracerebroventricular (icv) infusions of a low (1.2 &mgr;g/kg per hour) or high (4.0 &mgr;g/kg per hour) dose of benzamil were given to Dahl S rats on high salt diet (1370 &mgr;mol Na+/g food) for 2 or 4 weeks. “Ouabain” content was measured using a specific enzyme-linked immunosorbent assay (ELISA). Systolic blood pressure (BP) in Dahl S rats on high salt for 4 weeks increased markedly (188±10 versus 128±4 mm Hg, n=8,P<0.05). Benzamil fully blocked this increase (131±7 mm Hg after the high dose of benzamil). Hypothalamic and pituitary “ouabain” increased significantly (22±7 versus 12±3 and 151±38 versus 69±6 ng/g tissue, respectively,P<0.05) in Dahl S rats on high salt versus regular salt diet for 2 weeks. Benzamil blocked these increases of brain “ouabain” to high salt intake. Similarly, high salt intake for 4 weeks increased hypothalamic (18±2 versus 13±1 ng/g tissue,P<0.05) and pituitary (183±30 versus 78±8 ng/g tissue,P<0.05) “ouabain.” Benzamil also inhibited these increases of brain “ouabain.” Both hypothalamic and pituitary “ouabain” showed significant positive correlations with BP. In contrast, high salt intake did not affect “ouabain” levels in the adrenal gland or plasma in Dahl S rats on high salt for either 2 or 4 weeks. These findings indicate that in Dahl S rats high salt intake only increases brain and not peripheral “ouabain” and that benzamil-blockable brain sodium channels mediate the increases in brain “ouabain” and the subsequent hypertension.