Sarcomere length dependence of power output is increased after PKA treatment in rat cardiac myocytes

Abstract

The Frank-Starling relationship of the heart yields increased stroke volume with greater end-diastolic volume, and this relationship is steeper after beta-adrenergic stimulation. The underlying basis for the Frank-Starling mechanism involves length-dependent changes in both Ca(2+) sensitivity of myofibrillar force and power output. In this study, we tested the hypothesis that PKA-induced phosphorylation of myofibrillar proteins would increase the length dependence of myofibrillar power output, which would provide a myofibrillar basis to, in part, explain the steeper Frank-Starling relations after beta-adrenergic stimulation. For these experiments, adult rat left ventricles were mechanically disrupted, permeabilized cardiac myocyte preparations were attached between a force transducer and position motor, and the length dependence of loaded shortening and power output were measured before and after treatment with PKA. PKA increased the phosphorylation of myosin binding protein C and cardiac troponin I, as assessed by autoradiography. In terms of myocyte mechanics, PKA decreased the Ca(2+) sensitivity of force and increased loaded shortening and power output at all relative loads when the myocyte preparations were at long sarcomere length ( approximately 2.30 mum). PKA had less of an effect on loaded shortening and power output at short sarcomere length ( approximately 2.0 mum). These changes resulted in a greater length dependence of myocyte power output after PKA treatment; peak normalized power output increased approximately 20% with length before PKA and approximately 40% after PKA. These results suggest that PKA-induced phosphorylation of myofibrillar proteins explains, in part, the steeper ventricular function curves (i.e., Frank-Starling relationship) after beta-adrenergic stimulation of the left ventricle.

Cardiac Myosin-binding Protein C and Troponin-I Phosphorylation Independently Modulate Myofilament Length-dependent Activation

“…In particular, cMyBP-C AAA phospho-null myocytes in which endogenous troponin was exchanged for DD phospho-mimetic cTnI (n ϭ 4) displayed a 10.5 Ϯ 1.0% reduction in passive force (14.0 Ϯ 0.5 versus 12.5 Ϯ 0.3 millinewtons/mm 2 ) upon PKA treatment that was associated with a significant reduction in ⌬EC 50 (0.67 Ϯ 0.08 versus 0.49 Ϯ 0.03 M), supporting the notion that a PKA-mediated reduction in titin strain by itself reduces LDA. In addition, maximum Ca 2ϩ -activated myofilament force was reduced upon treatment with PKA in all muscle groups (NTG, AAA, and DDD), consistent with previous reports by others and us (1,8,10,11). In contrast, exchange of either AAA or DDD myocytes with the cTnI phospho-mimetic containing troponin resulted in an increase in maximum force (Table 1).…”

“…2 summarizes average ⌬EC 50 , a convenient parameter to index LDA, and the impact of PKA treatment on this parameter. LDA increased upon PKA treatment in NTG myocytes, consistent with previous reports (1,8,10,11,13) but was without impact in either the AAA or DDD myocytes. In addition, after PKA treatment, LDA in NTG myocytes approached LDA recorded in DDD myocytes regardless of PKA treatment, whereas it was ϳ50% lower in AAA myocytes.…”

“…This is caused, in part, by the expression of the unique cardiac-specific isoform of troponin-I (cTnI) (7), the inhibitory subunit of the striated muscle regulatory troponin complex (8,9). Cardiac LDA has been shown to be modulated by contractile protein phosphorylation by most (1,8,10,11) but not all (12) reports. Moreover, LDA is affected by cardiac disease-associated mutations within various contractile proteins, including the troponins and cardiac myosin binding protein C (cMyBP-C) (13).…”

Elevated Ca2+transients and increased myofibrillar power generation cause cardiac hypercontractility in a model of Noonan syndrome with multiple lentigines

“…Posttranslational modification of sarcomeric proteins is a major mechanism for fine tuning myofibrillar function. Importantly, PKA-mediated phosphorylation of myofibrillar proteins increases power generation (12). Furthermore, interaction of SHP2 and PKA in a signalosome complex induced by shear stress has been reported (7), suggesting that similar interactions could also play a role in cardiomyocytes expressing Q510E-SHP2.…”

Restrictive Cardiomyopathy Troponin I R145W Mutation Does Not Perturb Myofilament Length-dependent Activation in Human Cardiac Sarcomeres

“…In many cases, but notably not all, depressed LDA was reversed by treatment of the skinned myocytes with protein kinase A (PKA). Cardiac LDA has been shown to be modulated by contractile protein phosphorylation by most (7,(15)(16)(17)(18) although not all (19,20) reports. Moreover, selective introduction of a PKA phosphomimetic isoform of cTnI into the cardiac sarcomere increases LDA (15,17,(21)(22)(23)(24), a finding we confirm in the present study on isolated multicellular skinned human myocardium.…”