Autonomic regulation of calcium cycling in developing embryonic mouse hearts

In the present study, we combined optical Ca(2+) imaging with immunocytochemistry studies to characterize autonomic regulation of Ca(2+) cycling during early development in isolated embryonic mouse hearts. At embryonic days 9.5-11.5 (E9.5-E11.5), the Ca(2+) transient originated in the superior portion of the right atrium, propagated rapidly through both atria, slowly through the atrio-ventricular (AV) ring, and rapidly through both ventricles. Isoproterenol (ISO) significantly increased heart rate, increased Ca(2+) transient amplitude, rate of rise (RR) and a rate of decay, and shortened AV conduction time, indicating the presence of functional beta-adrenergic receptors. The muscarinic agonist carbachol (CCh) had no effects until 1 day later at E10.5. Both beta1-adrenergic and M2 muscarinic receptors were detected in ventricular muscle sections by immunochemistry at E10.5. Growing nerves, labeled using growth-associated protein 43 antibodies, were detected at the E14.5 stage, but not at E10.5, whereas mature sympathetic nerves, detected by tyrosine hydroxylase (TH) labeling, were not yet present at E14.5. These results demonstrate that functional regulation of Ca(2+) cycling by beta-adrenergic receptors occurs earliest in developing embryonic mouse hearts, followed a day later by muscarinic receptor responsiveness, with autonomic innervation developing later. These results define the functional and structural sequence of autonomic regulation of Ca(2+) transient in the embryonic mouse heart.