Discontinued stimulation of cardiomyocytes provides protection against hypothermia–rewarming-induced disruption of excitation–contraction coupling

New Findings: What is the central question of this study? Will discontinued stimulation of isolated cardiomyocytes (asystole) during hypothermia mitigate hypothermia–rewarming-induced cytosolic Ca²⁺ overload? What is the main finding and its importance? Mimicking asystole or hypothermic cardiac arrest by discontinued stimulation of cardiomyocytes during hypothermia resulted in normal contractile function after rewarming. This result suggests that asystole during severe hypothermia provides protection from hypothermia–rewarming-induced contractile dysfunction in cardiomyocytes. Abstract: After exposure of spontaneously beating hearts or electrically stimulated isolated cardiomyocytes to hypothermia–rewarming (H/R), cardiac dysfunction or alteration in excitation–contraction coupling, respectively, is a consequence. In contrast, hypothermic cardiac arrest, as routinely applied during cardiac surgery, will not impose any hazard to cardiac function after rewarming. We hypothesize that by maintaining asystole during H/R, cardiomyocytes will avoid Ca²⁺ overload attributable to the transient stimulation-evoked elevation of [Ca²⁺]_i and thus, H/R-induced elevation of phosphorylated cardiac troponin I and reduced Ca²⁺ sensitivity after rewarming. To test this hypothesis, the aim of the study was to determine whether discontinued electrical stimulation (to imitate hypothermic cardiac arrest) versus stimulation during 3 h of H/R prevents disruption of excitation–contraction coupling in our established cardiomyocyte H/R model. Cytosolic Ca²⁺ and the contractile response (sarcomere length shortening) were measured using an IonOptix system, and the dynamic assessment of Ca²⁺ sensitivity of contraction was conducted using a phase-loop plot. Cardiomyocytes were divided into three groups. Group 1 (time-matched control) was continuously stimulated at 0.5 Hz for 3 h at 35°C. Group 2 was continuously stimulated during H/R at 0.5 Hz, whereas in group 3 stimulation was discontinued during H/R and thus the cells remained quiescent until the resumption of stimulation after rewarming. The results demonstrate that discontinued stimulation of cardiomyocytes during H/R, imitating hypothermic cardiac arrest during cardiac surgery, provides protection against H/R-induced disruption of excitation–contraction coupling. We suggest that protective effects are caused by preventing the protein kinase A-induced elevation of phosphorylated cardiac troponin I, which is a key mechanism to reduce myofilament Ca²⁺ sensitivity of contraction.