atractyloside and 4-5-6-7-tetrachloro-2-trifluoromethylbenzimidazole

The effect of long-chain fatty acids (LCFA) on respiration and transmembrane potential (delta psi) in the resting state, and the rate of delta psi dissipation [d delta psi/dt)i) was investigated with oligomycin-inhibited rat liver mitochondria using succinate (plus rotenone) as substrate. The results obtained were compared with those of classical protonophores such as 2,4-dinitrophenol (DNP) and 4,5,6,7-tetrachloro-2-trifluoromethylbenzimidazole (TTFB). The effects of oleate or palmitate and that of DNP or TTFB on respiration and delta psi can be described by a common force-flow relationship. These facts all in all are not compatible with a decoupler-type uncoupling mechanism of LCFA; still, they indicate that the latter are protonophores. Moreover, the oleate-induced increase in the rate of delta psi dissipation closely correlates with that in respiration, suggesting that the uncoupling activity and the protonophoric activity of LCFA are interrelated. Carboxyatractyloside (CAT) exerted only a small inhibitory effect on oleate-induced respiration and delta psi dissipation, indicating that the adenine nucleotide translocase contributes to the uncoupling effect of LCFA to a minor extent only. Proton transport through the lipid region of the membrane as mediated by permeation of the protonated and deprotonated forms of LCFA is interpreted as the main process of the uncoupling of LCFA.

Topics: 2,4-Dinitrophenol; Animals; Atractyloside; Benzimidazoles; Biological Transport; Dinitrophenols; Fatty Acids; Female; Membrane Potentials; Mitochondria, Liver; Mitochondrial ADP, ATP Translocases; Oleic Acid; Oleic Acids; Oligomycins; Oxidative Phosphorylation; Oxygen Consumption; Palmitic Acid; Palmitic Acids; Protons; Rats; Uncoupling Agents

An improved method for the isolation of rat brain mitochondria is described. The preparation exhibits a respiratory control index (RCI) of 6 or 7.3 in the presence of pyruvate and malate or glutamate and malate, respectively. RCI decreases to 2.5 in the presence of Mg++. When the phosphorylation of extramitochondrially added or formed ADP is suppressed by carboxyatractyloside (CAT) inhibition of the adenine nucleotide translocator, the remaining respiration amounts to 6 nmol O2/min X mg mitochondrial protein. These results and the ratio of 16 to 19 from the quotient of phosphorylating active-state respiration to CAT inhibited respiration refer to a high degree of mitochondrial coupling of respiration. Therefore the remaining respiration in the presence of Mg++ is due to a phosphokinase activity located outside the inner membrane of intact mitochondria or at nonphosphorylating mitochondrial fragments. The following activities were observed: Oligomycin sensitive ATPase, 47 mU/mg protein; hexokinase, 272 mU/mg protein; creatinphosphokinase, 116 mU/mg protein; and a surprisingly low activity of adenylatekinase, 57 mU/mg protein.

Topics: Adenosine Monophosphate; Adenosine Triphosphate; Adenylate Kinase; Animals; Atractyloside; Benzimidazoles; Brain; Brain Chemistry; Creatine Kinase; Hexokinase; Magnesium; Male; Mitochondria; Rats; Rats, Inbred Strains; Substrate Specificity; Uncoupling Agents