On the relationships between membrane potential, calcium transient and tension in single barnacle muscle fibres

Abstract

SUMMARY1. The calcium-sensitive photoprotein aequorin has been used to follow the rapid changes in intracellular calcium concentration that occur during the contraction of single muscle fibres from the barnacle Balanus nubilus, Darwin. 2. The transient change in calcium-mediated light emission (calcium transient) and the changes in membrane potential and tension were recorded simultaneously, thus permitting an examination of the relationships between the chemical, electrical, and mechanical events ofexcitationcontraction coupling.3. With short-duration stimuli (< 200 msec), the calcium transient shows an S-shaped rising phase reaching a maximum soon after the cessation of the stimulus pulse. During membrane repolarization the calcium transient begins an exponential falling phase which has a time constant of 50-80 msec at 11-12°C.4. The shape of the calcium transient resembles the first derivative of the rising phase of the isometric tension response, thus suggesting that calcium controls the rate of tension development.5. There is no detectable increase of the light emission above resting values, during the falling phase of isometric tension.6. A plot of the calcium transient area (lumen x see) versus peak isometric force (g. cm-2) is linear over, at least, a range of forces from ca. 50-400 g. cm-2.7. When the fibre is capable of producing an active membrane response following the intracellular injection of potassium citrate, the onset and cessation of the calcium transient follow closely the onset and cessation * Present address. C. C. ASHLEY AND E. B. BIDG WA Y of the active membrane response. Tension responses under these conditions are much suppressed, suggesting that excitation-contraction coupling may be partially blocked between calcium release and the development of tension.8. Hypertonic salines (1 M sucrose or 1 M glycerol) cause little change in the membrane response, but greatly suppress the calcium transient and completely abolish the tension responses. These effects are readily reversible when normal saline is reintroduced, suggesting that excitationcontraction coupling may be temporarily blocked between the membrane response and calcium release.9. If the stimulus is prolonged (> 250-300 msec), the calcium transient falls slowly from its maximum value despite continued membrane depolarization, suggesting a time-dependent change in the ratio of the rate of release of calcium to the rate of calcium binding. The results from brief tetanic stimulation also support this suggestion.

Ouabain Potentiation of Rapid Cooling Contracture of Caffeinized Cardiac Muscles in Calcium Deprived Medium

“…As shown in Table 1 According to recent views, it is clear that increase in the contractile response relates to the increase of myoplasmic free Ca2+ concentrations, that is, an increase of a Ca release from the Ca-storage site within the cell produces an augmentation of contractions (1)(2)(3)(4)24).…”

“…In view of recent studies on the excitation-contraction (E-C) coupling mechanism in the heart and skeletal muscles (1)(2)(3)(4)(5), there are several possible mechanisms underlying the positive inotropic action of cardiac glycosides on the heart muscle. Lee and Klaus (6) and Nayler (7,8) concluded that the inotropic response to the glycosides reflects the effects on E-C coupling, and not the effect on either energy metabolism or the contractile protein.…”

Fluorescence changes from single striated muscle fibres injected with labelled troponin C (TnCDANZ)

“…Another notion was that the magnesium displacement rate constant could be much altered by the ionic environment within the muscle cell but this possibility has not so far been examined. In addition, it will be of great interest to employ another indicator within the cell whose response to free calcium changes is well documented, such as aequorin [3,4,7] so that the signal from both the aequorin and TnCDANZ signal can be recorded in addition to the force response.…”

“…Single muscle fibres from the barnacle (B. nubilus) were isolated and cannulated [7] and axially microinjected with a stock solution of TnCDANZ (13 mM nominally) in 100 mM KC1 (pH 7.0). The volume injected was 0.4~1, so that after dilution the final intracellular concentration was in the range 50-150/M.…”

“…platinised Pt wire and the voltage sensing part was a lOO/cm capillary filled with 0.5 M KC1 and containing a 25 pm diam. bright Pt wire [7,8]. The voltage clamp circuit was standard and force was recorded with an Akers force transducer (sensitivity 12 mN.…”