Activities and Expression of PKA, PKS and PDE.

<p><b>A</b>. PKA activity in isolated cardiac myocytes without stimulation (Basal) or stimulated with isoproterenol (Iso; 10 µM, 10 min) or NKH477 (NKH; 10 µM, 10 min). AC6mut expression reduced basal PKA activity (p = 0.01) and both Iso (p = 0.001) and NKH (p = 0.001) activities were reduced as well (n = 3, each group). <b>B</b>. The expression of key signaling proteins and their phosphorylation are shown in immunoblots using left ventricular homogenates from AC6mut and control mice. No group differences were observed. Shown are PKA catalytic unit, phospho (P) and Total (T) PKA regulatory subunits II-α and II-β, PKCα, phosphodiesterase type 3A (PDE3A), phospho-troponin I (P22/23-TnI), and total TnI. <b>C</b>. Phosphorylation of RyR2, PLB and TnI before and after isoproterenol stimulation was assessed in cardiac myocytes isolated from each group. Basal phosphorylation of RyR2, PLB and TnI showed no group differences. Isoproterenol stimulation was associated with increased phosporylation of RyR2, PLB, and TnI in both groups, but was more extensive in cardiac mycoytes from AC6mut mice. <b>D</b>. The data from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0072151#pone-0072151-g002" target="_blank"><b>Fig. 2C</b></a> indicating that isoproterenol stimulation was associated with increased phosporylation of RyR2, PLB, and TnI in cardiac mycoytes from AC6mut mice are shown in graphic format, normalized for loading (GAPDH). The increase in TnI phosphorylation was not statistically significant (p = 0.07).