The activity of ROMK channels in rat kidney tubule cells was assessed as tertiapin-Q (TPNQ)-sensitive current under whole cell clamp conditions. With an external K⁺ concentration of 5 mM and an internal K⁺ concentration of 140 mM and the membrane potential clamped to 0 mV, TPNQ blocked outward currents in principal cells of the cortical collecting duct (CCD) outer medullary collecting duct and connecting tubule (CNT). The apparent K_i was 5.0 nM, consistent with its interaction with ROMK. The TPNQ-sensitive current reversed at voltages close to the equilibrium potential for K⁺. The currents were reduced when the pipette solution contained ATP. In the CCD, the average TPNQ-sensitive outward current (I_SK) was 476 ± 48 pA/cell in control animals on a 1% KCl diet. I_SK increased to 1,255 ± 140 pA when animals were maintained on a high-K (10% KCl) diet for 7 days and decreased to 314 ± 46 pA after 7 days on a low-K (0.1% KCl) diet. In the CNT, I_SK was 360 ± 30 pA on control, 1,160 ± 110 on high-K, and 166 ± 16 pA on low-K diets. The results indicate that ROMK channel activity is highly regulated by dietary K in both the CCD and the CNT.