Effect of changing the composition of the bathing solution upon the isometric tension—pCa relationship in bundles of crustacean myofibrils

Abstract

SUMMARY1. The relative isometric tension-pCa relationship has been determined for isolated bundles of barnacle myofibrils under a variety of ionic conditions using [Ca2+]-buffered solutions which also contained an ATP regenerating system (creatine phosphate and creatine kinase).2. The results are in better agreement with the 'consecutive' scheme of reaction rather than with the 'independent' alternative (Ashley & Moisescu, 1972) for the co-operative action of two Ca2+ ions in the process of tension activation in crustacean skeletal muscle.3. Variations in the pH of the activating solutions did have a marked effect on the relative tension-pCa curve, although no effect was observed on the absolute maximum value for isometric tension. A shift in pH by 0-5 u. in the range 6-6-7-6 shifted the Ca2+-activation curve by 0.5 log u. towards lower free Ca2+ concentrations.4. Changes in the free Mg2+ concentration of the activating solutions in the millimolar range produced a pronounced shift of the relative tension-pCa curve along the pCa axis. Increasing [Mg2+] from 1 to 5 mM shifted the curve by about 0-7 log u. to higher free Ca2+ concentrations, without significantly modifying its steepness.5. Changes in the MgATP concentration of the activating solutions in the range of 1-13 mm had no significant effect on the relative tension-pCa relationship.6. Varying the K+ concentration in the activating solutions was also

Effect of cross-bridge kinetics on apparent Ca2+ sensitivity.

“…An increase in maximum tension does accompany the left shifts of the curve induced by ionic strength reduction and reduction in phosphate concentration. Similar increases in tension with reduction in ionic strength have been reported by others for skinned fibers (Ashley and Moisescu, 1977 ;Julian and Moss, 1981 ;Gordon et al, 1973 ;Thames et al, 1974) and for intact fibers (April et al, 1968 ;Edman and Hwang, 1977 ;Gordon and Godt, 1970) . We observe a large increase in tension with reduction of phosphate from 7.5 to 0 mM in 5 mM substrate but no such increase in 0.25 mM substrate.…”

“…Some of the curves used to obtain data for Table II (those for experiments with 0.25 mM substrate) were conducted at 10 and 20°C in addition to 15°C. We found, as had others (Ashley and Moisescu, 1977;Stevenson and Williams, 1980), that, with the exception of maximum tension, there is no effect of temperature change in this range on the normalized steady state parameters of fibers bathed in 0 phosphate saline and only a small effect on fibers bathed in 7.5 mM phosphate saline. Consequently, in Table II we pooled the results from these experiments with the data from experiments at 15°C.…”

Calcium and strontium concentration changes within skinned muscle preparations following a change in the external bathing solution.

“…The mechanical arrangement of the tension transducer is similar to that previously described (Ashley & Moisescu, 1977). The output from the bridge circuit went either to a Tektronix storage oscilloscope or to a pen recorder (Schwarzer,Varioscript V 242).…”

“…This set of movements could be carried out in 1-2 sec. The preparation does not appear to be damaged by being moved through an air-solution interface (Ford & Podolsky, 1972a;Ashley & Moisescu, 1977).…”

“…Subsequently this end of the preparation was securely tied under oil to a stainless-steel rod attached directly to the anode peg of the RCA 5734 mechanoelectric transducer. The other end was clamped in a pair of fine stainless steel Barcroft forceps (J. Weiss & Sons, London) which when closed do not hold liquid between the points (see Ashley & Moisescu, 1977 for the diagram of the experimental set-up). The myofibrillar bundle was then cut behind the forceps, the dissecting chamber was removed, and the preparation was lowered into a standard relaxing solution B (Table 2).…”