Insulin and IGF-1 activate Kir4.1/5.1 channels in cortical collecting duct principal cells to control basolateral membrane voltage

Potassium K(ir)4.1/5.1 channels are abundantly expressed at the basolateral membrane of principal cells in the cortical collecting duct (CCD), where they are thought to modulate transport rates by controlling transepithelial voltage. Insulin and insulin-like growth factor-1 (IGF-1) stimulate apically localized epithelial sodium channels (ENaC) to augment sodium reabsorption in the CCD. However, little is known about their actions on potassium channels localized at the basolateral membrane. In this study, we implemented patch-clamp analysis in freshly isolated murine CCD to assess the effect of these hormones on K(ir)4.1/5.1 at both single channel and cellular levels. We demonstrated that K+-selective conductance via K(ir)4.1/5.1 is the major contributor to the macroscopic current recorded from the basolateral side in principal cells. Acute treatment with 10 mu M amiloride (ENaC blocker), 100 nM tertiapin-Q (TPNQ; ROMK inhibitor), and 100 mu M ouabain (Na+-K+-ATPase blocker) failed to produce a measurable effect on the macroscopic current. In contrast, K(ir)4.1 inhibitor nortriptyline (100 mu M), but not fluoxetine (100 mu M), virtually abolished whole cell K+-selective conductance. Insulin (100 nM) markedly increased the open probability of K(ir)4.1/5.1 and nortriptyline-sensitive whole cell current, leading to significant hyperpolarization of the basolateral membrane. Inhibition of the phosphatidylinositol 3-kinase cascade with LY294002 (20 mu M) abolished action of insulin on K(ir)4.1/5.1. IGF-1 had similar stimulatory actions on K(ir)4.1/5.1-mediated conductance only when applied at a higher (500 nM) concentration and was ineffective at 100 nM. We concluded that both insulin and, to a lesser extent, IGF-1 activate K(ir)4.1/5.1 channel activity and open probability to hyperpolarize the basolateral membrane, thereby facilitating Na+ reabsorption in the CCD.