This study explores the role of K⁺ and aldosterone in the regulation of mRNA of the ATP-sensitive, inwardly rectifying K⁺ channel, ROMK, in the rat kidney. K⁺ deficiency downregulated ROMK mRNA in cortex to 47.1 ± 5.1% of control (P < 0.001) and in medulla to 56.1 ± 3.4% (P < 0.001). High-K⁺ diet slightly increased ROMK mRNA in medulla to 122 ± 9% (P < 0.05 vs. control). Adrenalectomy (Adx) downregulated cortical ROMK mRNA to 30.7 ± 6.8% (P < 0.001 vs. control), and increased it in medulla to 138 ± 12.9% (P < 0.02 vs. control). In Adx rats, K⁺ deficiency decreased ROMK mRNA in cortex and medulla similar to intact rats. The α₁- and β₁-Na-K-ATPase subunits were regulated in parallel to that of ROMK. In medulla, ROMK mRNA correlated with serum K⁺ concentration atR = 0.9406 (n = 6,P < 0.001) and α₁-Na-K-ATPase mRNA atR = 0.9756 (n = 6,P < 0.001). ROMK2 also correlated with serum K⁺ concentration (R = 0.895;n = 6,P < 0.01). These results show that cortical ROMK expression is regulated by aldosterone and K⁺, whereas the medullary ROMK mRNA is regulated by serum K⁺.