Abstract 17264: β-Adrenergic Signaling Promotes Calcium Oscillations in Human Cardiac Progenitor Cells
Catecholamines enhance cardiac function by activating effector molecules targeting Ca2+ handling proteins in myocytes, but the impact of these neurohumoral factors on the pool of resident primitive cells is poorly defined. The aim of this study was to establish whether catecholaminergic stimulation of human cardiac progenitor cells (hCPCs) promotes intracellular Ca2+ oscillations, critical events for cell cycle progression. For this purpose, c-kit-positive hCPCs were loaded with the Ca2+ indicator Fluo-4 and monitored by two-photon microscopy. After documenting that intracellular Ca2+ elevations in hCPCs are mediated by Ca2+ translocation from the endoplasmic reticulum to the cytoplasm via IP3 receptor channels, the effects of adrenergic receptor (AR) stimulation were tested. In Tyrode solution, Ca2+ spikes were observed in 12% of the cell population over a period of 33 minutes, and exposure to norepinephrine, agonist of α- and β-AR, increased the fraction of active cells to 26%. To dissect the contribution of the two families of adrenergic receptors to the observed response, phenylephrine (Phe) and isoproterenol (ISO) were employed. Phe stimulates α-AR, which is coupled to Gq protein promoting phospholipase C activity and IP3 production; in contrast, ISO activates β-AR, which is coupled to Gs/Gi proteins, modulating adenylyl cyclase and cAMP levels. Phe did not alter the occurrence of Ca2+ events, whereas ISO led to a 2.1-fold increase in the number of active hCPCs. To define the β-AR subtypes involved in this positive response, selective stimulators were used. The β1-AR agonist denopamine enhanced the incidence of Ca2+ spikes, whereas activation of β2-AR with fenoterol or β3-AR with CL 316243 did not alter Ca2+ levels. The cell-permeable cAMP analog dibutyryl-cAMP had effects on Ca2+ oscillations comparable to those mediated by β-AR stimulation. Moreover, inhibition of the cAMP-dependent protein kinase A (PKA) with H-89 abrogated the response of hCPCs to ISO. By immunocytochemistry, IP3 receptor phosphorylation at a PKA serine site was found to be increased following β-AR stimulation, consistent with enhanced IP3R channel activity. Thus, β-AR signaling in hCPCs promotes the kinase activity of PKA and enhances Ca2+ oscillatory events.
Author Disclosures: N. Alesi: None. C. Mangiaracina: None. P. Goichberg: None. G. Borghetti: None. S. Signore: None. S. Bardelli: None. M. Moccetti: None. A. Sorrentino: None. E. Wybieralska: None. L. Graciotti: None. A. Cannata: None. K. Waight: None. S. Shin: None. J. Kim: None. A. Leri: None. P. Anversa: None. T. Moccetti: None. M. Rota: None.
- © 2014 by American Heart Association, Inc.