Adult male rats were injected intraventricularly with N-[3H]acetylmannosamine. After different time intervals the rats were killed and free sialic acid, CMP-sialic acid, lipid- and protein-bound sialic acid were isolated from brain and the specific radioactivities determined. Maximal specific radioactivity was reached after approximately 4 h for CMP-sialic acid, after 10-12 h for free sialic acid and after approximately 42 h for lipid- and protein-bound sialic acid. After some days the specific radioactivities of all four pools were the same and decreased equally, with a calculated turnover rate of approximately 3.5 weeks. The conclusion was that this phenomenon was the result of reutilisation of sialic acid and/or precursors. Therefore, the calculated turnover is not the turnover of bound sialic acid, but merely the rate of leakage of sialic acid and/or precursors out of the brain, so that no real turnover can be measured by this method. The first few hours after injection the specific radioactivity of CMP-sialic acid rose above that of free sialic acid. It is supposed that a compartmentalization exists of free sialic acid. The newly synthesised sialic acid molecules are not secreted into the cytoplasmic pool but are preferentially used for the synthesis of CMP-sialic acid. The results and conclusions are discussed in view of the general problems concerning turnover measurements of glycoconjugates.
Adult male rats, under starving and normal conditions, were injected intravenously with N-acetyl[3H]mannosamine and after various time intervals the specific radioactivities of free N-acetylneuraminic acid (NeuAc) and CMP-N-acetylneuraminic acid were determined in the liver. The specific radioactivity of free NeuAc was high even within 20s after injection; the maximum was reached between 7 and 10 min. The specific radioactivity of CMP-NeuAc showed a lag phase of approx. 1 min. Thereafter it increased quickly and rose above the specific radioactivity of free NeuAc, reaching a maximum about 20 min after injection. These results point to a channelling of the newly synthesized NeuAc molecules into a special compartment, from which they are preferentially used by the enzyme CMP-sialic acid synthetase. It is suggested that the cytosolic enzyme N-acetylneuraminic acid 9-phosphate phosphatase is working in concert with the nuclear localized enzyme CMP-N-acetylneuraminic acid synthetase. Incorporation of radioactive sialic acid into sialoglycoproteins in liver occurred 2 min after injection, and after 10 min bound radioactivity began to appear in the circulation, indicating a transport time of 8 min of sialoglycoproteins from the point of attachment of sialic acid to the point of excretion.
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