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

Abstract

The relative isometric tension-pCa relationship was determined for isolated bundles of barnacle [Balanus nubilus] myofibrils under a variety of ionic conditions using [Ca2+]-buffered solutions which also contained an ATP regenerating system (creatine phosphate and creatine kinase). The results agree better with the consecutive scheme of reaction rather than with the independent alternative (Ashley et Moisescu, 1972) for the co-operative action of two Ca2+ in the process of tension activation in crustacean skeletal muscle. Variations in the pH of the activating solutions had 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 [units] in the range 6.6-7.6 shifted the Ca2+-activation curve by 0.5 log u towards lower free Ca2+ concentrations. 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 .apprx. 0.7 log u to higher free Ca2+ concentrations, without significantly modifying its steepness. 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. Varying the K+ concentration in the activating solutions had a marked effect upon the tension-pCa relationship in barnacle. An increase in the K+ concentration from 90 to 170 mM shifted the curve by some 0.6 log u towards higher free Ca2+ concentrations. Cooling the standard activating solutions from room temperature to +4.degree. C made no apparent difference to the relative tension-pCa relationship, but decreased significantly the absolute tension responses. Tonicity by itself had a marked effect upon the absolute steady-state tension levels in isolated bundles of myofibrils. Maximum isometric tension in this preparation was not simply related to ionic strength, or to the monovalent cation concentration, but it depended, as well, upon the anionic composition of the activating solution. A change in ionic strength of 25 mM over the range 245-270 mM did not appear to modify the relative tension-pCa relationship. The effect of the physiologically occurring cations H+, K+ and Mg2+ upon the relative isometric tension-pCa relationship can be accounted for on the basis of a model of competitive inhibition between these cations and Ca2+ for the functional unit for tension. This inhibitory effect appears to involve at least one H+, one Mg2+ and two K+ per Ca2+ participating in the activation process of the functional unit for tension.