Effects of osmolality and ionic strength on the mechanism of Ca2+ release in skinned skeletal muscle fibres of the toad

Abstract

1. The effects of increased osmolality and ionic strength on the mechanism of Ca2+ release were examined in mechanically skinned skeletal muscle fibres of the toad at 23 degrees C. Ca2+ release was induced by depolarizing the transverse tubular (T-) system by ionic substitution. 2. Increasing the osmolality of the 'myoplasmic' solution about four times (to 955 mosmol/kg), by addition of 700 mM sucrose to the standard potassium (K-)HDTA solution (HDTA: hexamethylenediamine-tetraacetate), only depressed the depolarization-induced response by about 46%. Much of this decrease could be attributed to a reduction in the Ca(2+)-sensitivity of the contractile proteins at this high osmolality. 3. Addition of > 400 mM sucrose itself often induced substantial Ca2+ release and a transient tension response. This 'spontaneous' release was (a) greatly enhanced when the sarcoplasmic reticulum (SR) had been heavily loaded with Ca2+, (b) little affected by inactivation of the voltage sensors by prolonged or permanent depolarization of the T-system and (c) blocked by Ruthenium Red (10 microM). 4. When both the osmolality and ionic strength were increased, by increasing the K-HDTA concentration, the depolarization-induced force was greatly reduced (to 35% at 818 mosmol/kg and 5% at 1095 mosmol/kg). Most of this reduction could be directly attributed to the substantially reduced maximum force and Ca2+ sensitivity of the contractile apparatus. 5. The small amount of releasable Ca2+ remaining in the SR after a single depolarization in a high-HDTA solution with 1 mM EGTA (to chelate the released Ca2+), indicated that depolarization could still elicit massive Ca2+ release at high ionic strength and osmolality (at 1 mM free Mg2+). 6. In contrast, when the total Mg2+ and ATP concentrations were raised about threefold (free [Mg2+] increased 2.7-fold) along with the osmolality and ionic strength, the ability of depolarization to elicit Ca2+ release was greatly hindered. 7. Osmotic compression of the skinned fibres to their in situ diameter by addition of 4% polyvinylpyrrolidone (PVP-40), substantially potentiated the depolarization-induced force responses, due partly to an increase in the Ca(2+)-sensitivity of the contractile apparatus. 8. These results indicate how increased intracellular osmolality, ionic strength and [Mg2+] produce the transient contraction and subsequent inhibition of tetanic tension in intact muscle fibres exposed to hypertonic solutions.