Dependence of calcium release, tension generation and restoring forces on sarcomere length in skinned cardiac cells

Abstract

The ascending limb of the length-tension diagram was studied in single skinned (sarcolemma-free) cardiac cells of rat ventricle. The free (Ca2+) in the perfusing solutions was buffered with ethyleneglycol-bis(beta-aminoethyl)N,N'-tetraacetic acid (EGTA). Tension was recorded with a photo-diode force transducer (detection limit: 1 mug, compliance 2-3 mum/mg) and was expressed as a function of the sarcomere length (SL) measured during contraction with a high-speed movie camera. When the SL was decreased by a brief (1-2 sec) exposure to high free (Ca2+), a restoring force qas observed upon return of the cell to a relaxing solution. The restoring force was comprised of two components: (1) a rapid elongation to 1.57 identical to 0.06 mum developing a negative tension (relaxing force) as large as 4% of the maximum positive tension, which was observed in both skinned cells and in single myofibrils, and (2) a slow elongation from 1.57 identical to 0.06 to 1.93 identical to 0.14 mum, which was observed in skinned cells but not in single myofibrils. When the SL was kept extremely short by continuous Ca2+ activation for more than 30 sec, a shortening of the A band much below 1.0 mum persisted for several minutes after imposed re-elongation of the sarcomers (delta state). The amplitude of the tonic tension developed by direct activation of the myofilaments in the presence of high total (EGTA) was maximum for 2.20 mum SL and decreased very little when the SL was decreased. However, the decrease of tension became more pronounced at SL shorter than 1.55 mum when polyvinylpyrrolidone was added to the solutions at a concentration reducing the swelling of the myofilament lattice. This finding imposes some caution in applying the results obtained in skinned cardiac cells to intact tissue. The SL-tension diagram of the phasic contractions induced by Ca2+-triggered of Ca2+ in the presence of a slight EGTA buffering was similar to the SL-tension diagram of the intact rat ventricular muscle when the latter was expressed as a function of the active SL. In contrast, the SL-tension diagram of caffeine-induced phasic contractions was similar to that of the tonic tension produced by direct activation of the myofilaments. Decreasing SL results therefore in a partial inhibition of the Ca2+ triggered release process. It was concluded that Starling's Law may correspond to a length-dependence of several mechanisms including Ca2+-triggered release from the SR, interaction between thick and thin filaments and restoring forces.