Characterization of constitutively active GIRK ion channels in rat pulmonary vein

Abstract

In human, pulmonary vein (PV) cardiomyocytes (CM) generate abnormal electrical activity, which is responsible for the triggering of atrial fibrillation. In rat PV, activation of α1-adrenergic receptors results in the depolarization of PV CM that lead to the inhibition of electrical conduction from PV toward left atria (LA). The mechanism underlying the depolarization remain unknown but might involve G-protein activated K+ channels (GIRK) which is responsible for the IkAch current. The aim of this study was to characterize and compare IkAch in PV and LA CM. All experiments were carried out in rat. Patch clamp technique in whole cell configuration was used to measure background and IkAch currents in CM. Membrane potential of PV and LA CM was recorded in tissues using sharp microelectrodes. The expression of Kir3.1 and Kir3.4 subunits of GIRK channels was quantified by western blot experiments. In patch clamp experiments, carbachol (Cch) (1 μM) induced the activation of an inward rectifying current which was fully blocked by tertiapin-Q (TQ, 300 nM), a selective blocker of IkAch in both PV and LA CM. In the absence of Cch, TQ blocked an inward rectifier current, which inverted at equilibrium potential of K+ in the PV but not in the LA CM. This suggests the existence of a constitutively active IKAch only in the PV CM. In accordance with these results, the western blot experiments indicate that the expression level of Kir3.4 subunits which contribute to the constitutive activity of GIRK channels was about 1.73 times more abundant in PV CM than in LA ones ( P < 0.001). Finally, sharp microelectrodes experiments showed that TQ (150 nM) lead to membrane depolarization of PV CM (16.7 ± 1 mV, P < 0.001) but not in LA CM (2.1 ± 0.6 mV, P = 0.173). However, this depolarization was less than that induced by the alpha adrenergic agonist cirazoline (steady state diastolic membrane potential in presence of cirazoline = − 44.1 ± 1.7 mV vs −57.2 ± 0.9 mV in presence of TQ ( P < 0.001)) suggesting that blockade of IkAch is not the only mechanism involved in the PV depolarization. These results suggest the presence of constitutively active GIRK channels only in PV CM. Their blockade could be a promising pharmacological target to block the electrical conduction between PV and LA, and thus containing abnormal electrical activity in PV.