Diffusible sodium, potassium, magnesium, calcium and phosphorus in frog skeletal muscle

Abstract

A microvolumetric analytical method has been developed to measure the endogenous concentrations of diffusible elements in muscle cells. Single twitch fibres from frog muscle were skinned under oil and 0.2 nl drops of isosmotic sucrose solution, held in the tips of specially constructed pipettes, were placed in contact with the skinned fibres. After 0-10 min, the microdrops were removed and analysed with a wave-length dispersive X-ray spectrometer. The uptake of Na, K, Mg, and P into the microdrops was well fitted by a single exponential function, while the uptake of Ca was better represented by the sum of two exponential functions. All elements analysed except Ca reached diffusional equilibrium within 5 min of placing the microdrop on the fibre, while Ca was still not equilibrated at 10 min. For freshly isolated muscle fibres, diffusible element concentrations in the microdrops at equilibrium were (in mM, mean +/- S.D.): Na, 7.6 +/- 7.2; K, 82 +/- 36; Mg, 5.8 +/- 3.0; P, 51 +/- 19. Diffusible Ca concentration (at 10 min elapsed sampling time) was 0.7 +/- 0.4 mM. Results from experiments in which microdrops were equilibrated with skinned fibres pre-soaked in an artificial (Ca-free) solution support the notion that the exogenous solutes can replace the endogenous diffusible contents of a skinned fibre by soaking the skinned fibre in a relatively large volume of the artificial solution. Total Na, K, Mg, Ca, and P content of whole muscle was measured by electron probe analysis of muscle extracts. In freshly isolated muscle, whole muscle element content was (in mmol/kg wet weight, mean +/- S.D.): Na, 21 +/- 8; K, 120 +/- 26; Mg, 9.7 +/- 2.6; Ca, 2.2 +/- 0.5; P, 76 +/- 18. Extracellular fluid volumes of freshly isolated whole muscles were estimated by compartmental analysis of Na efflux. Extracellular element concentrations were measured by electron probe analysis of frog plasma. Using the extracellular fluid volume and concentration estimates, extracellular contributions were subtracted from measurements of the element content of whole muscle to yield estimates of total intracellular element concentration (in mmol/l myoplasmic water). Based on the values for the intracellular total and diffusible element concentrations, the diffusible/total content fraction in freshly isolated muscle is estimated to be: Na, 0.38; K, 0.48; Mg, 0.42; Ca, 0.22; and P, 0.47.