The data indicate that 1) phosphorylation of phospholamban at Ser16 by cAMP-dependent protein kinase is the main regulator of beta-adrenergic-induced cardiac relaxation definitely preceding Thr17 phosphorylation and 2) the beta-adrenergic-mediated phosphorylation of Thr17 by Ca2+-calmodulin-dependent protein kinase required influx of Ca2+ through the L-type Ca2+ channel