Effects of pH and inorganic phosphate on force production in alpha-toxin-permeabilized isolated rat uterine smooth muscle.

1. Strips of longitudinal smooth muscle isolated from rat uterus were permeabilized using crude alpha-toxin from the bacterium Staphylococcus aureus. This treatment rendered the surface membrane permeable to small molecular weight substances. Simultaneous measurements of tension and calcium concentration ([Ca2+]) (using indo-1 fluorescence) were used to investigate the effects of pH and inorganic phosphate concentration ([Pi]) on Ca(2+)-activated force generated by the contractile proteins. 2. Raising the [Pi] from 1 to 11 mM at a pH of 7.2 depressed both maximal and submaximal Ca(2+)-activated force. This effect of Pi was concentration dependent having the majority of its effect by 6 mM. 3. Further experiments at a submaximal [Ca2+] showed that Ca(2+)-activated force was enhanced by raising [Pi] from 6 to 11 mM suggesting that Pi increased the Ca2+ sensitivity of tension production. Based on these results, calculations indicate that the apparent affinity constant of Ca2+ for the contractile proteins increased from 4 x 10(6) M-1 to 6 x 10(6) M-1 on raising [Pi] from 1 to 11 mM. 4. Lowering pH from 7.2 to 6.7 at a [Pi] of 1 mM potentiated Ca(2+)-activated force with a small depression in the apparent Ca2+ sensitivity of tension production. This effect of pH on maximum (100 microM Ca2+) and submaximum (0.3 microM Ca2+) Ca(2+)-activated force was observed over a range of acidic pHs (7.0-6.7). 5. Increasing pH from 7.2 to 7.7 at a [Pi] of 1 mM depressed Ca(2+)-activated force with no effect on Ca2+ sensitivity of tension production. 6. Spontaneous contractions in intact rat myometrium are abolished under hypoxic conditions. Under these same conditions intracellular [Pi] rises and pH falls. The results of this study suggest that taken individually neither the effect of a rise in [Pi] nor a fall in pH on Ca(2+)-activated force generated by the contractile proteins can account for the effect of hypoxia on spontaneous contractions.