Calmodulin kinase is functionally targeted to the action potential plateau for regulation of L-type Ca²⁺ current in rabbit cardiomyocytes

L-type Ca²⁺ current (I_Ca-L) triggers Ca²⁺ release from the sarcoplasmic reticulum (SR) and both SR and I_Ca-L are potential sources of intracellular Ca²⁺ (Ca_i²⁺) for feedback regulation of I_Ca-L-. Ca_i²⁺ bound to calmodulin (Ca²⁺-CaM) can inhibit I_Ca-L, while Ca²⁺-CaM can also activate Ca²⁺-CaM-dependent protein kinase II (CaMK) to increase I_Ca. However, it is not known whether I_Ca-L or the SR is the primary source of Ca²⁺ for I_Ca-L regulation. The L-type Ca²⁺ channel C terminus is implicated as a critical transduction element for I_Ca-L responses to Ca²⁺-CaM and CaMK, and the C terminus undergoes voltage-dependent steric changes, suggesting that Ca_i²⁺ control of I_Ca-L may also be regulated by cell membrane potential. We developed conditions to separately test the relationship of Ca²⁺-CaM and CaMK to I_Ca-L and SR Ca_i²⁺ release during voltage clamp conditions modelled upon time and voltage domains relevant to the cardiac action potential. Here we show that CaMK increases I_Ca-L after brief positive conditioning pulses, whereas Ca²⁺-CaM reduces I_Ca-L over a broad range of positive and negative conditioning potentials. SR Ca²⁺ release was required for both Ca²⁺-CaM and CaMK I_Ca-L responses after strongly positive conditioning pulses (+10 and +40 mV), while Ca_i²⁺ from I_Ca-L was sufficient for Ca²⁺-CaM during weaker depolarizations. These findings show that I_Ca-L responses to CaMK are voltage dependent and suggest a new model of L-type Ca²⁺ channel regulation where voltage-dependent changes control I_Ca-L responses to Ca²⁺-CaM and CaMK signalling.