Abstract 210: Titin Phosphorylation by Protein Kinase G as a Novel Mechanism of Diastolic Adaptation to Acute Hemodynamic Overload

Abstract

Titin is the main determinant of myocardial passive tension and its distensibility is increased via phosphorylation by protein kinase G (PKG), activated by nitric oxide (NO) and natriuretic peptides (NP) upon acute overload. We hypothesized whether myocardial stretch led to decreased stiffness, optimizing diastolic filling along with the usual increase in contractility. Intact rat Langendorff hearts, strips dissected from the LV or right atrium of cardiac surgery patients and rabbit papillary muscles were acutely stretched for 15min. Passive tension (PT) was measured in skinned cardiomyocytes extracted from the LV of control and stretched rat hearts for sarcomere lengths (SL) 1.8-2.3μm before and after incubation with PKG. Rabbit muscles were incubated with a PKG inhibitor or, simultaneously, a NO synthase inhibitor, a NO scavenger, a NP receptor A antagonist. All-total titin phosphorylation was stained with Pro-Q Diamond and indexed to total-protein signals using SYPRO Ruby. Values are given as mean±SEM and statistical significance was set to p<0.05. After acute stretch there was a progressive decrease in passive tension/diastolic pressure over 15min: 27±8% and 27±6% in human atrium and ventricular muscles, respectively, and 43±2% in rabbit papillary muscles and isolated hearts. This decrease in myocardial stiffness was significantly blunted by PKG inhibition (40%) and NO/NP pathway inhibition (29%). PT of cardiomyocytes was significantly lower (≈60%) in the previously stretched group for all SL. A similar effect, only significant in the control group, was observed after incubation with PKG. Titin phosphorylation increased markedly 15 minutes after acute myocardial stretch in human (atrial: 11±1% vs 41±8%; LV: 27±8% vs 71±21%) and rabbit (13±2% vs 23±3%) myocardium. The progressive decrease in myocardial stiffness after acute hemodynamic overload is preserved at the myofilamental level and seems to depend on PKG activity, representing a potential therapeutic target in patients with pathologically rigid myocardium. Moreover, blocking PKG activation seems to attenuate this adaptive diastolic response. Therefore, titin phosphorylation by this kinase is probably involved in this new myocardial response to stretch in both animals and humans.