scholarly journals The tritium isotope effect of sn-glycerol 3-phosphate oxidase and the effects of clofenapate and N-(2-benzoyloxyethyl)norfenfluramine on the esterification of glycerol phosphate and dihydroxyacetone phosphate by rat liver mitochondria

1973 ◽  
Vol 136 (2) ◽  
pp. 421-427 ◽  
Author(s):  
Mariana Bowley ◽  
Ray Manning ◽  
David N. Brindley

1. Owing to a 3H isotope effect, the mitochondrial sn-glycerol 3-phosphate oxidase (EC 1.1.99.5) had a mean activity which was 8.4 times less with sn-[2-3H]-rather than with sn-[1-14C]glycerol 3-phosphate as a substrate. 2. A method for measuring the simultaneous synthesis of lipid from glycerol phosphate and dihydroxyacetone phosphate in rat liver mitochondria is described. 3. The lipid synthesized by rat liver mitochondria from sn-[1-14C]glycerol 3-phosphate was mainly phosphatidate and lysophosphatidate, whereas that synthesized from dihydroxy[1-14C]acetone phosphate was mainly acyldihydroxyacetone phosphate. 4. Additions of NADPH facilitated the conversion of acyldihydroxyacetone phosphate into lysophosphatidate and phosphatidate. 5. Hydrazine (1.4mm) or KCN (1.4mm) inhibited the synthesis of lipids from dihydroxyacetone phosphate but not from glycerol phosphate. 6. Clofenapate (1–2.5mm) inhibited the synthesis of lipids from dihydroxyacetone phosphate but slightly stimulated synthesis from glycerol phosphate. 7. The methanesulphonate of N-(2-benzoyloxyethyl)norfenfluramine, at 0.25–0.75mm, inhibited lipid synthesis from both glycerol phosphate and dihydroxyacetone phosphate.

1972 ◽  
Vol 130 (4) ◽  
pp. 1003-1012 ◽  
Author(s):  
Ray Manning ◽  
David N. Brindley

1. Rat liver slices were employed to study the relative rates of incorporation of a mixture of [2-3H]- or [1,3-3H]-glycerol and [1-14C]glycerol into lipids. 2. With 0.1mm-glycerol approx. 82% of the newly synthesized lipid, calculated from 14C incorporation, was present as neutral lipid, 13% as phosphatidylcholine and 5% as phosphatidylethanolamine. Increasing the glycerol concentration to 40mm caused a decrease in the percentage of neutral lipid to 59% and a corresponding increase in the percentage of phosphatidylcholine to 36% of the newly synthesized lipid. 3. The (d.p.m. of 2-3H)/(d.p.m. of 1-14C) ratio in glycerolipid was considerably higher than that in precursor glycerol throughout the range of experimental conditions. In contrast the incorporation of a mixture of [1,3-3H]glycerol and [1-14C]glycerol into lipid occurred with little or no change in the 3H/14C ratio. 4. Respiring rat liver mitochondria were found to oxidize a mixture of sn-[2-3H]- and sn-[1-14C]-glycerol 3-phosphate with a resultant increase in the 3H/14C ratio of the remaining sn-glycerol 3-phosphate. This increase is due to a 3H isotope effect of the mitochondrial sn-glycerol 3-phosphate dehydrogenase (EC 1.1.99.5), which discriminates against sn-[2-3H]glycerol 3-phosphate during oxidation. 5. A method is described for the simultaneous determination of the relative contributions of the glycerol phosphate and dihydroxyacetone phosphate pathways of glycerolipid biosynthesis in rat liver slices. The method involves measurement of the (d.p.m. of 2-3H)/(d.p.m. of 1-14C) ratio in both sn-glycerol 3-phosphate and glycerolipid after incubation of rat liver slices with a mixture of [2-3H]glycerol and [1-14C]glycerol for various times. 6. By using this method it was shown that 40–50% of the glycerol incorporated into lipid by rat liver slices proceeded via the sn-glycerol 3-phosphate pathway and 50–60% was incorporated via dihydroxyacetone phosphate.


Author(s):  
E. A. Elfont ◽  
R. B. Tobin ◽  
D. G. Colton ◽  
M. A. Mehlman

Summary5,-5'-diphenyl-2-thiohydantoin (DPTH) is an effective inhibitor of thyroxine (T4) stimulation of α-glycerophosphate dehydrogenase in rat liver mitochondria. Because this finding indicated a possible tool for future study of the mode of action of thyroxine, the ultrastructural and biochemical effects of DPTH and/or thyroxine on rat liver mere investigated.Rats were fed either standard or DPTH (0.06%) diet for 30 days before T4 (250 ug/kg/day) was injected. Injection of T4 occurred daily for 10 days prior to sacrifice. After removal of the liver and kidneys, part of the tissue was frozen at -50°C for later biocheailcal analyses, while the rest was prefixed in buffered 3.5X glutaraldehyde (390 mOs) and post-fixed in buffered 1Z OsO4 (376 mOs). Tissues were embedded in Araldlte 502 and the sections examined in a Zeiss EM 9S.Hepatocytes from hyperthyroid rats (Fig. 2) demonstrated enlarged and more numerous mitochondria than those of controls (Fig. 1). Glycogen was almost totally absent from the cytoplasm of the T4-treated rats.


Author(s):  
Olga A. Gonchar ◽  
Valentina I. Nosar ◽  
Larisa. V. Bratus ◽  
I. N. Tymchenko ◽  
N. N. Steshenko ◽  
...  

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