beta-adrenergic receptor (beta-AR) stimulation increases cardiac L-type Ca2+ channel (CaCh) currents via cAMP-dependent phosphorylation. We report here that the affinity and maximum response of CaCh to isoproterenol (Iso), in mouse ventricular myocytes were significantly higher when Ba2+ was used as the charge carrier (IBa) instead of Ca2+ (ICa). The EC50 and maximum increase of peak currents were 43.7 +/- 7.9 nM and 1.8 +/- 0.1-fold for ICa and 23.3 +/- 4.7 nM and 2.4 +/- 0.1-fold for IBa. When cells were dialyzed with the faster Ca2+ chelator, BAPTA, both sensitivity and maximum response of ICa to Iso were significantly augmented compared to cells with EGTA (EC50 of 23.1 +/- 5.2 nM and maximal increase of 2.2 +/- 0.1-fold). Response of ICa to forskolin was also significantly increased when cells were dialyzed with BAPTA or when currents were measured in Ba2+. In contrast, depletion of the sarcoplasmic reticulum (SR) Ca2+ stores by ryanodine did not alter sensitivity of ICa to Iso or forskolin. These results suggest that the Ca2+ entering through CaCh regulates cAMP-dependent phosphorylation, and such negative feedback may play a significant role in cellular Ca2+ homeostasis and contraction in cardiac cells during beta-AR stimulation.