DETERMINANTS OF LOADED SHORTENING VELOCITY IN SINGLE CARDIAC MYOCYTESPERMEABILIZED WITH ALPHA-HEMOLYSIN

Force-velocity relations were obtained from single cardiac myocytes is olated by enzymatic digestion of rat myocardium and permeabilized with the pore-forming staphylococcal toxin alpha-hemolysin. Single cardiac myocytes were attached to a force transducer and piezoelectric transl ator and viewed with an inverted microscope to allow periodic monitori ng of sarcomere length during experiments. Permeabilized cells were ac tivated by immersion in a bath of known [Ca2+]. We report that the Ca2 + sensitivity of cells obtained by enzymatic digestion and permeabiliz ed using alpha-hemolysin is similar to that reported previously for me chanically disrupted ventricular myocardium; however, the tension-pCa relation is less steep in the new preparation. During isotonic measure ments, force was clamped to various loads using a rapid-response servo system. All recordings of shortening under load were distinctly curvi linear, and analysis of data involved fitting each shortening recordin g with a single exponential curve and calculating the value of the slo pe at the initial time of the load clamp. In addition, the presence of significant resting force at initial sarcomere lengths in these cells required that the possibility of alteration of velocity due to the pr esence of resting force be addressed. The maximum shortening velocity in fully Ca2+-activated single ventricular myocytes studied by this me thod was 2.83 muscle lengths per second on average. The basis for curv ilinear shortening is postulated to be multifactorial in cardiac muscl e, involving a combination of shortening inactivation and one or more passive elasticities that resist stretch or compression depending on s arcomere length. Shortening velocity shows a dependence on myosin isof orm content when cells from a single heart are compared; however, this relation does not hold when cells from different hearts are compared. The behavior of single alpha-hemolysin-permeabilized myocyte shorteni ng under loaded conditions at lower levels of Ca2+ is also described. During submaximal Ca2+ activation, initial shortening velocities are f aster than those observed in maximally activated cells. This may be du e to contributions of high passive force to increase shortening veloci ty under conditions of low active force generation, when passive force in the cell is a greater proportion of the total force and there are fewer bound crossbridges.