Local Ca2+ entry through L-type Ca2+ channels activates Ca2+- dependent K+ channels in rabbit coronary myocytes

Abstract

Large-conductance Ca2+-dependent K+ channels (K(Ca), which are abundant on the sarcolemma of vascular myocytes, provide negative feedback via membrane hyperpolarization that limits Ca2+ entry through L-type Ca2+ channels (I(CaL)). We hypothesize that local accumulation of subsarcolemmal Ca2+ during I(CaL) openings amplifies this feedback. Our goal was to demonstrate that Ca2+ entry through voltage-gated I(CaL) channels can stimulate adjacent K(Ca) channels by a localized interaction in enzymatically isolated rabbit coronary arterial myocytes voltage clamped in whole-cell or in cell-attached patch clamp mode. During slow-voltage-ramp protocols, we identified an outward K(Ca) current that is activated by a subsarcolemmal Ca2+ pool dissociated from bulk cytosolic Ca2+ pool (measured with indo 1) and is dependent on L-type Ca2+ channel activity. Transient activation of unitary K(Ca) channels in cell-attached patches could be detected during long step depolarizations to +40 mV (holding potential, -40 mV; 219 pS in near-symmetrical K+). This local interaction between the channels required the presence of Ca2+ in the pipette solution, was enhanced by the I(CaL) agonist Bay K 8644, and persisted after impairment of the sarcoplasmic reticulum by incubation with 10 μmol/L ryanodine and 30 μmol/L cyclopiazonic acid for at least 60 minutes. Furthermore, we provide the first direct evidence of simultaneous openings of single K(Ca) (67 pS) and I(CaL) (3.9 pS) channels in near-physiological conditions, near resting membrane potential. Our data imply a novel sensitive mechanism for regulating resting membrane potential and tone in vascular smooth muscle.