Downregulation of the renal outer medullary K+ channel ROMK by the AMP-activated protein kinase

The 5′-adenosine monophosphate-activated serine/threonine protein kinase ( AMPK) is stimulated by energy depletion, increase in cytosolic Ca2+ activity, oxidative stress, and nitric oxide. AMPKparticipates in the regulation of the epithelial Na+ channel ENaC and the voltage-gated K+ channel KCNE1/KCNQ1. It is partially effective by decreasing PIP2 formation through the PI3K pathway. The present study explored whether AMPKregulates the renal outer medullary K+ channel ROMK. To this end, cRNA encoding ROMKwas injected into Xenopusoocytes with and without additional injection of constitutively active AMPKγR70Q (AMPKα1-HA+AMPKβ1-Flag+AMPKγ1R70Q), or of inactive AMPKαK45R (AMPKα1K45R+AMPKβ1-Flag+AMPKγ1-HA), and the current determined utilizing two-electrode voltage-clamp and single channel patch clamp. ROMKprotein abundance was measured utilizing chemiluminescence in Xenopusoocytes and western blot in whole kidney tissue. Moreover, renal Na+ and K+ excretion were determined in AMPKα1-deficient mice ( ampk−/− ) and wild-type mice ( ampk+/+ ) prior to and following an acute K+ load (111 mM KCl, 30 mM NaHCO3, 4.7 mM NaCl, and 2.25 g/dl BSA) at a rate of 500 μl/h. As a result, coexpression of AMPKγR70Q but not of AMPKαK45R significantly decreased the current in ROMK1-expressing Xenopusoocytes. Injection of phosphatidylinositol PI(4,5)P2 significantly increased the current in ROMK1-expressing Xenopusoocytes, an effect reversed in the presence of AMPKγR70Q. Under control conditions, no significant differences between ampk−/− and ampk+/+ mice were observed in glomerular filtration rate (GFR), urinary flow rate, serum aldosterone, plasma Na+, and K+ concentrations as well as absolute and fractional Na+ and K+ excretion. Following an acute K+ load, GFR, urinary flow rate, serum aldosterone, plasma Na+, and K+ concentration were again similar in both genotypes, but renal absolute and fractional Na+ and K+ excretion were higher in ampk−/− than in ampk+/+ mice. According to micropuncture following a K+ load, delivery of Na+ to the early distal tubule but not delivery of K+ to late proximal and early distal tubules was increased in ampk−/− mice. The upregulation of renal ROMK1 protein expression by acute K+ load was more pronounced in ampk−/− than in ampk+/+ mice. In conclusion, AMPKdownregulates ROMK, an effect compromising the ability of the kidney to excrete K+ following an acute K+ load.