Action potential prolongation in cardiac myocytes of old rats is an adaptation to sustain youthful intracellular Ca²⁺ regulation

Advanced age in rats is accompanied by reduced expression of the sarcoplasmic reticulum (SR) Ca²⁺ pump (SERCA-2). The amplitudes of intracellular Ca²⁺ (Ca_i²⁺) transients and contractions in ventricular myocytes isolated from old (23-24-months) rats (OR), however, are similar to those of young (4-6-months) rat myocytes (YR). OR myocytes also manifest slowed inactivation of L-type Ca²⁺ current (I_CaL) and marked prolongation of action potential (AP) duration. To determine whether and how age-associated AP prolongation preserves the Ca_i²⁺ transient amplitude in OR myocytes, we employed an AP-clamp technique with simultaneous measurements of I_CaL (with Na⁺ current, K⁺ currents and Ca²⁺ influx via sarcolemmal Na⁺-Ca²⁺ exchanger blocked) and Ca_i²⁺ transients in OR rat ventricular myocytes dialyzed with the fluorescent Ca²⁺ probe, indo-1. Myocytes were stimulated with AP-shaped voltage clamp waveforms approximating the configuration of prolonged, i.e. the native, AP of OR cells (AP-L), or with short AP waveforms (AP-S), typical of YR myocytes. Changes in SR Ca²⁺ load were assessed by rapid, complete SR Ca²⁺ depletions with caffeine. As expected, during stimulation with AP-S vs AP-L, peak I_CaL increased, by 21±4%, while the I_CaL integral decreased, by 19±3% (P<0.01 for each). Compared to AP-L, stimulation of OR myocytes with AP-S reduced the amplitudes of the Ca_i²⁺ transient by 31±6%, its maximal rate of rise (+dCa_i²⁺/dt_max; a sensitive index of SR Ca²⁺ release flux) by 37±4%, and decreased the SR Ca²⁺ load by 29±4% (P<0.01 for each). Intriguingly, AP-S also reduced the maximal rate of the Ca_i²⁺ transient relaxation and prolonged its time to 50% decline, by 35±5% and 33±7%, respectively (P<0.01 for each). During stimulation with AP-S, the gain of Ca²⁺-induced Ca²⁺ release (CICR), indexed by +dCa_i²⁺/dt_max/I_CaL, was reduced by 46±4% vs AP-L (P<0.01). We conclude that the effects of an application of a shorter AP to OR myocytes to reduce +dCa_i²⁺/dt_max and the Ca²⁺ transient amplitude are attributable to a reduction in SR Ca²⁺ load, presumably due to a reduced I_CaL integral and likely also to an increased Ca²⁺ extrusion via sarcolemmal Na⁺-Ca²⁺ exchanger. The decrease in the Ca_i²⁺ transient relaxation rate in OR cells stimulated with shorter APs may reflect a reduction of Ca²⁺/calmodulin-kinase II-regulated modulation of Ca²⁺ uptake via SERCA-2, consequent to a reduced local Ca²⁺ release in the vicinity of SERCA-2, also attributable to reduced SR Ca²⁺ load. Thus, the reduction of CICR gain during stimulation with AP-S is the net result of both a diminished SR Ca²⁺ release and an increased peak I_CaL. These results suggest that ventricular myocytes of old rats utilize AP prolongation to preserve an optimal SR Ca²⁺ loading, CICR gain and relaxation of Ca_i²⁺ transients.