Molecular and functional characterization of inwardly rectifying K+ currents in murine proximal colon.

Abstract

Interstitial cells of Cajal (ICC) from murine colonic muscles express genes encoding inwardly rectifying K+ channels. Transcripts of Kcnj2 (Kir2.1), Kcnj4 (Kir2.3), Kcnj14 (Kir2.4), Kcnj5 (Kir3.4), Kcnj8 (Kir 6.1) and Kcnj11 (Kir6.2) were found in colonic ICC. A conductance with properties consistent with Kir2 channels was observed in ICC but not in smooth muscle cells (SMC). Despite expression of gene transcripts, G-protein gated K+ channel (Kir3) and KATP (Kir6) currents were not resolved in ICC. KATP is a conductance prominent in SMC. Kir2 antagonist caused depolarization of freshly dispersed ICC and colonic smooth muscles, suggesting that this conductance is active under resting conditions in colonic muscles. The conclusion of the present study is that ICC express the Ba2+ -sensitive, inwardly rectifying K+ conductance in colonic muscles. This conductance is most probably a result of heterotetramers of Kir2 gene products, with this regulating resting potentials and the excitability of colonic muscles. Membrane potentials of gastrointestinal muscles are important because voltage-dependent Ca2+ channels in smooth muscle cells (SMC) provide the Ca2+ that triggers contraction. Regulation of membrane potential is complicated because SMC are electrically coupled to interstitial cells of Cajal (ICC) and PDGFRα+ cells. Activation of conductances in any of these cells affects the excitability of the syncytium. We explored the role of inward rectifier K+ conductances in colonic ICC that might contribute to regulation of membrane potential. ICC expressed Kcnj2 (Kir2.1), Kcnj4 (Kir2.3), Kcnj14 (Kir2.4), Kcnj5 (Kir3.4), Kcnj8 (Kir 6.1) and Kcnj11 (Kir6.2). Voltage clamp experiments showed activation of inward current when extracellular K+ ([K+ ]o ) was increased. The current was inwardly rectifying and inhibited by Ba2+ (10μm) and ML-133 (10μm). A similar current was not available in SMC. The current activated in ICC by elevated [K+ ]o was not affected by Tertiapin-Q. Gβγ, when dialysed into cells, failed to activate a unique, Tertiapin-Q-sensitive conductance. Freshly dispersed ICC showed no evidence of functional KATP . Pinacidil failed to activate current and the inward current activated by elevated [K+ ]o was insensitive to glibenclamide. Under current clamp, ML-133 caused the depolarization of isolated ICC and also that of cells impaled with microelectrodes in intact muscle strips. These findings show that ICC, when isolated freshly from colonic muscles, expressed a Ba2+ -sensitive, inwardly rectifying K+ conductance. This conductance is most probably a result of the expression of multiple Kir2 family paralogues, and the inwardly rectifying conductance contributes to the regulation of resting potentials and excitability of colonic muscles.