FIGURE 4.

Effect of cAMP analogs on spontaneous diastolic Ca²⁺ transients. Cardiac myocytes were loaded with indo-1/AM, and Ca²⁺ imaging was carried out as described under “Experimental Procedures.” For activation of Epac, cells were incubated 5 min with 10 μm 8-pCPT before recording of transients. For activation of PKA, cells were incubated for 20 min with 300 μm 6-Bnz-cAMP. Because of its higher membrane permeability, 8-pCPT was used at a lower concentration and shorter incubation periods according to recent publications (49, 50). Iso (200 nm) was added 5 min before recording of transients as positive control. Untreated cells were used as negative control. Cells were electrically stimulated at 0.5 s⁻¹. a–d, characteristic trains of transients obtained at 0.5 s⁻¹ electrical pacing are shown. aU, arbitrary units. Peak number (e), amplitude (f), and reciprocal time constant (g) are presented as mean ± S.E. Significant differences are indicated by ***, p < 0.001, n = 20–26 as indicated in the bars. An ANOVA model followed by LSD post hoc tests was applied to investigate different effects of Iso compared with 8-pCPT and 6-Bnz-cAMP on Ca²⁺ amplitude and reciprocal time constant. A Mann-Whitney rank sum test was applied to investigate different effects of Iso compared with 8-pCPT and 6-Bnz-cAMP on peak number.