BETA-ADRENERGIC-RECEPTOR STIMULATION INCREASES UNLOADED SHORTENING VELOCITY OF SKINNED SINGLE VENTRICULAR MYOCYTES FROM RATS

In vitro biochemical experiments have suggested that stimulation of be ta-adrenergic receptors may increase the rate of crossbridge cycling i n mammalian myocardium, but recent attempts to demonstrate a mechanica l correlate have yielded conflicting results. To investigate this issu e, we measured the effect of isoproterenol (ISO) and cAMP-dependent pr otein kinase (PKA) on unloaded shortening velocity (V-o). V-o is thoug ht to be determined by the rate-limiting step of the crossbridge cycle , ie, the rate of crossbridge detachment from actin, and is therefore an index of the cycling rate. Single rat ventricular myocytes were enz ymatically isolated, incubated in Ringer's solution without (control) or with 0.1 mu mol/L ISO, and then rapidly skinned. Some control cells were subsequently treated with 3 mu g/mL PKA for 40 minutes. V-o was then measured during maximal activation (pCa 4.5) in control, ISO-trea ted, and PKA-treated cells using the slack-test method. To test the ef ficacy of the agonist treatments, Ca2+ sensitivity of isometric tensio n was also assessed for each treatment by determining the [Ca2+] requi red for half-maximal tension tie, pCa(50)). Both ISO and PKA treatment reduced the Ca2+ sensitivity of isometric tension compared with same- day control cells, in agreement with previous studies in intact and in skinned preparations. V-o was increased 38% by ISO treatment and 41% by PKA treatment compared with same-day control cells. P-32 autoradiog raphy showed that troponin I and C protein were the principal proteins phosphorylated by PKA treatment. We conclude that P-adrenergic stimul ation increases the rate of crossbridge release from actin, by a mecha nism that most likely involves the phosphorylation of troponin I and/o r C protein by PKA. (Circ Res. 1994;74:542-549.)