Effect of a deficiency of thiamine on brain pyruvate dehydrogenase: enzyme assay by three different methods.
A simple and rapid method based on the NADH-linked reduction of a tetrazolium dye was described for the determination of pyruvate dehydrogenase activity in rat brain homogenates. The method (method 3) gave a value of 36.06 +/- 1.24 nmol of pyruvate utilised/min/mg of whole brain protein. This value was higher than that obtained by measurement of the rate of decarboxylation of [1-14C]pyruvate (15.10 +/- 0.88 nmol/min/mg of protein; method 1) and was comparable with the rate of transfer of acetyl groups to an arylamine (39.04 +/- 1.32 nmol/min/mg of protein; method 2). A critique of the values reported by others by different methods was given. The pyruvate dehydrogenase activity in the mitochondria isolated from rat brain was in the "active" (nonphosphorylated) form. A deficiency of thiamine in rats was produced by treatment with pyrithiamine, an antagonist of thiamine. This treatment resulted in abnormal neurological signs, such as ataxia and convulsions. The measurement of the total activity of pyruvate dehydrogenase in the brain by all three methods showed no significant change in the enzymic activity in thiamine-deficient rats after treatment with pyrithiamine. The activities of the enzyme in the brains of pair-fed animals were similar to those in the controls.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002245:Carbon Dioxide / Q000378:metabolism; D004396:Coloring Agents; D004789:Enzyme Activation; D007700:Kinetics; D008297:Male; D008928:Mitochondria / Q000201:enzymology; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D011745:Pyrithiamine; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism*; D011773:Pyruvates / Q000378:metabolism; D019289:Pyruvic Acid; D051381:Rats; D011919:Rats, Inbred Strains; D013778:Tetrazolium Salts; D013832:Thiamine Deficiency / Q000139:chemically induced / Q000201:enzymology*; D013835:Thiamine Pyrophosphate / Q000494:pharmacology
D004396:Coloring Agents; D011768:Pyruvate Dehydrogenase Complex; D011773:Pyruvates; D013778:Tetrazolium Salts; D009243:NAD; D002245:Carbon Dioxide; C013574:iodonitrotetrazolium; D011745:Pyrithiamine; D019289:Pyruvic Acid; D013835:Thiamine Pyrophosphate