Phosphorylation of Excitation-Contraction Coupling Components in a Guinea-Pig Model of Heart Failure

Abstract

Phosphorylation status appears to be a key determinant of excitation-contraction coupling ion channel and pump function. Dysfunction of the ryanodine receptor (RyR) secondary to catecholaminergic drive and phosphorylation has been proposed as a factor in contractile dysfunction and arrhythmia patho-physiology in the failing heart. The phosphorylation states of RyR, along with those of phospholamban and troponin I have been investigated by immunoblotting, and quantitated by comparing levels in failing hearts with basal levels, minimum levels after beta-blocker treatment and maximal levels achieved by ex vivo treatment with isoprenaline. We found that RyR residue Ser2809 was phosphorylated to 124 ± 11 % (n = 5, P > 0.05) of control (sham-operated, basal) in heart failure under basal conditions and 143 ± 12 % (n = 6, P 0.05) for heart failure and 199 ± 9 % (n=6, P < 0.05) for isoprenaline treatment. Phosphorylation levels at Ser16 of phospholamban were higher: 159 ± 17 % (heart failure, n = 7, P < 0.05) and 366 ± 95 % (isoprenaline treatment, n = 5, P < 0.05). At Ser23/24 of troponin I there is no significant change in heart failure (n = 5, P < 0.05) but a 230 ± 92 % increase with isoprenaline treatment (n = 6, P < 0.05). Basal levels of phosphorylation are thus relatively low at RyR Ser2030, phospholamban Ser16, and troponin I Ser23/24, and are not significantly increased in heart failure, but are substantially increased by isoprenaline treatment. In contrast, the phosphorylation level at Ser 2809 is already high and can be increased only moderately by isoprenaline.