Membrane and calcium clock mechanisms contribute variably as a function of temperature to setting cardiac pacemaker...

Here, we show that heart rate in zebrafish Danio rerio is dependent upon two pacemaking mechanisms and it possesses a limited ability to reset the cardiac pacemaker with temperature acclimation. Electrocardiogram recordings, taken from individual, anaesthetised zebrafish that had been acclimated to 18, 23 or 28°C were used to follow the response of maximum heart rate (f_Hmax ) to acute warming from 18°C until signs of cardiac failure appeared (up to c. 40°C). Because f_Hmax was similar across the acclimation groups at almost all equivalent test temperatures, warm acclimation was limited to one significant effect, the 23°C acclimated zebrafish had a significantly higher (21%) peak f_Hmax and reached a higher (3°C) test temperature than the 18°C acclimated zebrafish. Using zatebradine to block the membrane hyperpolarisation-activated cyclic nucleotide-gated channels (HCN) and examine the contribution of the membrane clock mechanisms to cardiac pacemaking, f _Hmax was significantly reduced (by at least 40%) at all acute test temperatures and significantly more so at most test temperatures for zebrafish acclimated to 28°C vs. 23°C. Thus, HCN channels and the membrane clock were not only important, but could be modified by thermal acclimation. Using a combination of ryanodine (to block sarcoplasmic calcium release) and thapsigargin (to block sarcoplasmic calcium reuptake) to examine the contribution of sarcoplasmic reticular handling of calcium and the calcium clock, f _Hmax was again consistently reduced independent of the test temperature and acclimation temperature, but to a significantly lesser degree than zatebradine for zebrafish acclimated to both 28 and 18°C. Thus, the calcium clock mechanism plays an additional role in setting pacemaker activity that was independent of temperature. In conclusion, the zebrafish cardiac pacemaker has a limited temperature acclimation ability compared with known effects for other fishes and involves two pacemaking mechanisms, one of which was independent of temperature.