The action of ATP/ADP-antiporter inhibitors upon the uncoupling effect of pahnitate, detergents and 'classical' uncouplers has been studied. The uncoupling effect was estimated by stimulation of succinate oxidation and of Hf permeability of rat liver mitochondria in the presence of oligomytin. It is shown that carboxyatractylate (CAtr) and pyridoxal 5phosphate (PLP) suppress the uncoupling induced by pahnitate and the anionic detergents SDS and cholate, but do not affect that induced by the cationic detergents CTAB, by the non-ionic detergent Triton X-100, as well as by the 'classical' uncouplers FCCP and DNP. The results are discussed in terms of a concept assuming that the ATP/ADP-antiporter facilitates the electrophoretic export of hydrophobic anions from mitochondria.Fatty acid; Uncoupling; ATP/ADP-antiporter

The effect of melittin, an activator of phospholipase A 2 , on proliferation and death of rat thymocytes in a broad concentration range was studied. Cell proliferation was estimated by the accumulation of colchicin metaphases, necrotic death was determined from lysis and staining of cells with trypan blue, and apoptosis was assessed from the type of DNA fragmentation, the amount of fragmented DNA, and the percentage of cells with subdiploid DNA. It was shown that low melittin concentrations (below 5 ug/ml) stimulate thymocyte proliferation. At high melittin concentrations, thymocytes die by the primary necrosis type. Throughout the concentration range studied, melittin does not produce apoptosis in thymocytes. Conversely, high melittin concentrations even inhibit thymocyte apoptosis in the control and after irradiation. An inhibitor of RNA synthesis actinomycin D does not affect thymocyte death in the presence of melittin. It is concluded that the activation of phospholipase A 2 can induce necrosis but not apoptosis and thus is not a necessary step in the signaling cascade that initiates apoptosis in thymocytes.

Thermoregulatory uncoupling of oxidative phosphorylation has been studied in heart and skeletal muscle mitochondria of ground squirrels. The respiratory rate ofmitochondria in the presence of oligornycin was found to be much higher in winter (in hibernating, arousing, or aroused animals) than in summer. This additional respiration is strongly (arousing animals) or completely (hibernating and aroused anhnals) inhibited by carboxyntractylate (CAtr) and bovine serum albumin (USA). The CAtr-and BSA-induced decreases in the rate of respiration arc accompanied by membrane potential increases, The rate of the CAtr-and BSA-sensitive respiration is proportional to the content of free fatty acids which, in the heart, dccrcascs in the order: arousing Z+ aroused = hibernating > summer animals. Maximal respiratory rates observed in the prcxncc of dinitrophenol (arousing > aroused > summer > hibernating animals) do not parallel the fatty acid Ievcl. It is assumed that some heat production in the winter animals is due to fatty acid-induced, ATP/ADP-antiportcr-mediated uncoupling in heart and skeletal muscle mitochondria. The peak of heat production during arousal after hibernation also includes some other stimulatory effect on mitochondrial respiration.

Energy coupling parameters were studied in liver mitochondria of ground squirrel during arousal from hibernation. It was found that such mitochondria become uncoupled during incubation with phosphate in a salt medium. The uncoupling was revealed by respiration rate increase and membrane potential decrease in the presence of oligomycin. Both effects were reversed by addition of cyclosporin A. Under the same in vitro conditions, mitochondria from aroused (active) animals showed no uncoupling but could be uncoupled by addition of palmitate in the cyclosporin A-sensitive fashion. It is proposed that formation ofeyclosporin A-sensitive pores can be involved in urgent heat production in arousing hibernators.

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