TRANSPORT AND METABOLISM OF THIAMINE IN RAT BRAIN CORTEX IN VITRO

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Abstract
Aeorbic incubation at 37" of rat brain-cortex slices in Krebs-Ringer phosphate medium containing glucose and labeled thiamine results in accumulation in the tissue of labeled thiamine and labeled thiamine phosphates. The concentration of the labeled thiamine in the tissue cell water increases with increase of external labeled thiamine concentration in an approximately linear manner; the concentration ratio for labeled thiamine (tissue: medium) exceeding unity with low external thiamine concentrations (e.g. 0.2 [mu][image]) and diminishing to about unity as the external thiamine concentration is increased to 1 [mu][image]. The concentration of labeled phosphorylated thiamine in the tissue is at least double that of the labeled thiamine present and its amount increases with increase of external thiamine concentration. Labeled phosphorylated thiamine appears in the medium, its amount being about 1/5 that in the tissue. Phosphorylation of thiamine in the tissue proceeds during incubation for 3 hr. and, with an external labeled thiamine concentration of 0.2 [mu][image], about 48% conversion of thiamine takes place. In the presence of ouabain (0.1 m[image]), which does not inhibit thiamine phosphorylation in rat brain extract, there is a fall in the uptake of labeled thiamine by brain-cortex slices and the concentration ratio for the labeled thiamine (tissue: medium) falls to below unity. Anaerobiosis, lack of Na+ or the presence of Amprol (0.01 m[image]) leads to marked inhibition of thiamine phosphorylation, and the concentration ratio for labeled thiamine (tissue: medium) falls to about unity. The facts lead to the conclusion that thiamine is conveyed into the brain cell against a concentration gradient by an energy-assisted process mediated by a membrane carrier. Pyrithiamine is a marked inhibitor of thiamine phosphorylation in brain extract. Thiamine mono-phosphate and thiamine diphosphate inhibit thiamine phosphorylation in brain extract. They diminish "total" thiamine (free and phosphorylated) uptake into brain cortex slices and inhibit the transport of thiamine into the brain cell, possibly by competition for the carrier. Phosphorylation of labeled thiamine in brain extract is not only brought about by adenosine triphosphate (in the presence of Mg2+) but apparently by adenosine diphosphate and uridine triphosphate.

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