| 870085 |
Lefresne P, Hamon M, Beaujouan JC, Glowinski J: Origin of the acetyl moiety of acetylcholine synthesized in rat striatal synaptosomes. Biochimie. 1977;59(2):197-215.
The subcellular localization of the AcCoA compartment supplying the cytoplasmic choline acetyltransferase (ChAc, EC 2.3.1.6) was investigated using a purified preparation of rat striatal synaptosomes (B fraction). It was first demonstrated that the SRA of the [14C] ACh synthesized during a 10 min incubation period was equal to the SRA of the [2-14C] and the [3-14C] pyruvate added to the isolated nerve terminal suspension. The experimental results can be summarised as follows: (i) No modification in the amount of [14C] ACh synthesized from [2-14C] pyruvatetion in the amount of [14C] ACh synthesized from [2-13C] pyruvate could be detected after the addition of high concentrations of either carnitine, acetylcarnitine or acetyl phosphate to the synaptosomal suspension. (ii) Under experimental conditions in which the amount of [1,5-14C] citrate taken up by passive diffusion into the cholinergic nerve endings would allow detection of the possible formation of the labelled ester, no [14C] ACh could be recovered. (iii) The SRA's of the individual carbon atoms of the Krebs cycle intermediary compounds when the cycle is fed with [2-14C] and [3-14C] pyruvate were calculated as a function of the STA's of each of these two precursors (a and a' respectively), of the number of 14CO2 dpm produced in the Krebs cycle from each of these two labelled compounds (D2 and D3 respectively), and as the function of the rate y of exchanges of molecules between the tricarboxylic acid cycle and other metabolic compartments. The experimental value obtained from a 10 min incubation, after the nerve endings had reached a steady metabolic activity, indicate that if the acetyl moiety of ACh was derived from some Krebs cycle intermediary compounds, its SRA could never exceed 55 per cent that of the [2-14C] pyruvate from which it is produced, (iv) No correlation could be found between the rate of [14C] ACh formation and changes in the Krebs cycle activity induced by sodium cyanide, 2-4 dinitrophenol and Ca2+ free medium. (v) The lack of significant [14C] ACh synthesis from [1-14C] acetate in striatal synaptosomes is consistent with the failure of fluoroacetate to modify the amounts of 14CO2 as well as of [14C] ACh formed from [2-14C] pyruvate. These results were interpreted as a confirmation of the presence of a low AcCoA synthetase activity in the nerve terminals. To reconcile all these data, it is proposed that pyruvate is transformed into AcCoA outside the mitochondria by the action of some cytoplasmic pyruvate dehydrogenase-like enzyme. |
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