Determination of the Intravesicular pH of Fragmented Sarcoplasmic Reticulum with 5, 5-Dimethyl-2, 4-oxazolidinedione

抄録

The intravesicular pH (pH_i) of fragmented sarcoplasmic reticulum (SR) of the skeletal muscle was determined from the distribution of 5, 5-dimethyl-2, 4-oxazolidinedione (DMO), a weak organic acid, between the intra- and extravesicular spaces. The pH_i's thus obtained were found to be slightly lower (0.02-0.17 pH unit) than the pH's of the external medium (pH_e, ) at pH 6.5-8.5 in the presence of 105 mM KCl and 40 mM Tris-maleate buffer. The higher the pH_e, the greater the pH gradient. When pH_e was changed, pH_i attained equilibrium within about 20 min, the time necessary for the separation of the SR by centrifugation. When 0.25 M sucrose and 5 mM Tris-maleate buffer were used instead of 105 mM KCl and 40 mM buffer, the pH gradient increased to 0.56.
It was also demonstrated by direct measurements of pH_e, with a glass-electrode pH meter that K⁺ ions added to the external medium exchanged the intravesicular H⁺ ions.
From these results it appears that the pH gradient across the SR membrane was at the Donnan equilibrium. In this state, the Donnan potentials corresponding to pH gradients of 0.17 and 0.56 were -9.3 and -30.6 mV, respectively.