We studied gender differences in NCC activity and expression in wild-type (WT) and AT1a receptor knockout (KO) mice. In renal clearance experiments, urine volume (UV), GFR, absolute Na and K (ENa; EK) and fractional Na and K (FENa; FEK) excretion were measured and compared at peak changes after bolus iv injection of hydrochlorothiazide (HCTZ; 30mg/kg). In WT, females responded more strongly than males to HCTZ, with larger fractional increases of UV (7.8- vs. 3.4-fold), ENa (11.7- vs. 5.7-fold), FENa (7.9-vs. 4.9-fold) and EK (2.8- vs. 1.4-fold). In contrast, there were no gender differences in the responses to the diuretic in KO mice; HCTZ produced greater effects on male KO than on WT, but similar effects on females. In WT, total (tNCC) and phosphorylated (pNCC) NCC protein expression were 1.8- fold and 4.6- fold higher in females compared to the males (P<0.05), consistent with the larger response to HCTZ. In KO mice, tNCC and pNCC increased significantly in males, to levels not different from those in females. There were no gender differences in the expression of the Na/H-Exchanger (NHE3) expression in WT; NHE3 protein decreased to similar extents in male and female KO animals, suggesting AT1a-mediated NHE3 expression in proximal tubules. The resulting increase in delivery of NaCl to the distal nephron may underlie increased NCC expression and activity in mice lacking AT1aR.