The effect of thiols and disulfides on water uptake and extrusion by rat liver mitochondria.

Reduced glutathione is among the substances of physiological occurrence which stimulate water uptake by isolated rat liver mitochondria (l-3). The water uptake produced by glutathione is of special interest since it differs in some respects from that induced by other agents such as phosphate, thyroxine, and phlorizin (l-6). Perhaps the most significant difference is that glutathione-swollen mitochondria in dilute suspension do not contract1 again, with extrusion of water, on addition of adenosine triphosphate, Mg++, and bovine serum albumin, whereas these agents cause prompt contraction of mitochondria swollen by phosphate, thyroxine, Ca”, and other agents (7, 8). It was later shown, however, that glutathione-swollen mitochondria can be contracted again by adenosine triphosphate if a heat-labile mitochondrial factor, “C-factor,” is added to the swelling medium (9). C-factor is detached or “leaks” from the mitochondria during swelling induced by glutathione, and its subsequent dilution in the medium below an effective level apparently accounts for the failure to contract in the absence of added C-factor, as shown in the preceding papers (9, 10). In this report a more detailed study of the action of various thiols and disulfides on both water uptake and extrusion in rat liver mitochondria is described. This information was necessary in order to develop a more reproducible and specific assay system for the purification of C-factor and to assist in interpreting the chemical nature of the action of glutathione and C-factor on mitochondrial structure. In brief, the effects of thiols and disulfides on water uptake and extrusion are complex and depend strikingly on their structure, concentration, and ratio. These effects, which are presumably mediated through thiol-disulfide interchange reactions with the membranes, permit more rigorous definition of the conditions under which C-factor activity may be assayed. times with 0.25 M sucrose. They were then suspended in cold