Abstract 16089: Functional Coupling of SK Channels and L-type Calcium Channels in Cardiomyocytes

Abstract

Background. Small conductance Ca 2+ -activated K + (SK) channels play significant roles in regulating the excitability of cardiomyocytes (CMs). SK channels are unique in that they are gated solely by intracellular Ca 2+ and hence, function to integrate intracellular Ca 2+ and membrane potentials on a beat-to-beat basis in the heart. Our previous studies revealed that cardiac SK2 channels coupled with L-type Ca 2+ channels (LTCCs) through a physical bridge, α-actinin2, suggesting that LTCCs may be functionally coupled with SK2 channels by providing local Ca 2+ domain to activate the SK channels. However, a recent study suggested that sarcoplasmic reticulum (SR) Ca 2+ release is necessary and sufficient for the activation of cardiac SK channels. The objective of the study is to examine the mechanisms of SK channel activation in native CMs. Methods and Results. By using a voltage-clamp protocol in rabbit CMs to activate LTCCs followed immediately by a test voltage to monitor the SK currents, we recorded apamin-sensitive SK currents in response to the Ca 2+ influx. By altering the duration of activation of LTCCs, the activation of SK currents as a function of the increasing Ca 2+ influx was quantified. EGTA or BAPTA were then used to test the intracellular Ca 2+ domain necessary for SK channel activation. BAPTA but not EGTA completely disengaged the coupling between LTCCs and SK channels. SK currents coupled with Ca 2+ influx via LTCCs continued to be elicited after application of caffeine, ryanodine or thapsigargin to deplete SR Ca 2+ store, suggesting that LTCCs provide the immediate Ca 2+ microdomain for the activation of SK channels in CMs. Super-resolution imaging of SK2, Ca v 1.2, and ryanodine receptor 2 (RyR2) using stimulated emission depletion (STED) microscopy was performed to directly quantify the distance among SK, LTCC and RyR2 in CMs. Conclusion. Ca 2+ influx through LTCCs provides the immediate and efficient Ca 2+ microdomain for the activation of SK channels. Furthermore, changes in SK channel activation kinetics following SR Ca 2+ depletion suggest both LTCC-mediated Ca 2+ influx and SR Ca 2+ release are necessary for the activation of cardiac SK channels. Funded by NIH (R01 HL085727 and R01 HL085844 to NC), VA (I01 BX000576 to NC) and AHA (14BGIA18870087 to XZ).