Inotropic Response of Cardiac Ventricular Myocytes to β-Adrenergic Stimulation With Isoproterenol Exhibits Diurnal Variation Involvement of Nitric Oxide

Rationale: Although > 10% of cardiac gene expression displays diurnal variations, little is known of their impact on excitation-contraction coupling. Objective: To determine whether the time of day affects excitation-contraction coupling in rat ventricles. Methods and Results: Left ventricular myocytes were isolated from rat hearts at 2 opposing time points, corresponding to the animals resting or active periods. Basal contraction and [Ca2+](i) was significantly greater in myocytes isolated during the resting versus active periods (cell shortening 12.4+/-0.3 versus 11.0+/-0.2%; P<0.05 and systolic [Ca2+](i) 422+/-12 versus 341+/-9 nmol/L; P<0.01. This corresponded to a greater sarcoplasmic reticulum (SR) Ca2+ load (672+/-20 versus 551+/-13 nmol/L P<0.001). The increase in systolic [Ca2+](i) in response to isoproterenol (> 3 nmol/L) was also significantly greater in resting versus active period myocytes, reflecting a greater SR Ca2+ load at this time. This diurnal variation in response of Ca2+-homeostasis to isoproterenol translated to a greater incidence of arrhythmic activity in resting period myocytes. Inhibition of neuronal NO synthase during stimulation with isoproterenol, further increased systolic [Ca2+](i) and the percentage of arrhythmic myocytes, but this effect was significantly greater in active period versus resting period myocytes. Quantitative RT-PCR analysis revealed a 2.65-fold increase in neuronal NO synthase mRNA levels in active over resting period myocytes (P<0.05). Conclusions: The threshold for the development of arrhythmic activity in response to isoproterenol is higher during the active period of the rat. We suggest this reflects a reduction in SR Ca2+ loading and a diurnal variation in neuronal NO synthase signaling. (Circ Res. 2010;106:1244-1252.)