Both, the decreased L-type Ca current (I) density and increased spontaneous Ca release from the sarcoplasmic reticulum (SR), have been associated with atrial fibrillation (AF). In this study, we tested the hypothesis that remodeling of 3,5-cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) signaling is linked to these compartment-specific changes (up-or down-regulation) in Ca-handling. Perforated patch-clamp experiments were performed in atrial myocytes from 53 patients with AF and 104 patients in sinus rhythm (Ctl). A significantly higher frequency of transient inward currents (I) activated by spontaneous Ca release was confirmed in myocytes from AF patients. Next, inhibition of PKA by H-89 promoted a stronger effect on the I frequency in these myocytes compared to myocytes from Ctl patients (7.6-fold vs. 2.5-fold reduction), while the β-agonist isoproterenol (ISO) caused a greater increase in Ctl patients (5.5-fold vs. 2.1-fold). I density was larger in myocytes from Ctl patients at baseline (p < 0.05). However, the effect of ISO on I density was only slightly stronger in AF than in Ctl myocytes (3.6-fold vs. 2.7-fold). Interestingly, a significant reduction of I and Ca sparks was observed upon Ca /Calmodulin-dependent protein kinase II inhibition by KN-93, but this inhibition had no effect on I. Fluorescence resonance energy transfer (FRET) experiments showed that although AF promoted cytosolic desensitization to β-adrenergic stimulation, ISO increased cAMP to similar levels in both groups of patients in the L-type Ca channel and ryanodine receptor compartments. Basal cAMP signaling also showed compartment-specific regulation by phosphodiesterases in atrial myocytes from 44 Ctl and 43 AF patients. Our results suggest that AF is associated with opposite changes in compartmentalized PKA/cAMP-dependent regulation of I (down-regulation) and I (up-regulation).