Choline into brain subcellular fractions

In some preliminary studies, the uptake of [Me-14C]choline into the subcellular fractions of brain has been measured. Rats were injected intracerebrally with 1 Ci (0.018 moles) of [Afe-I4C]choline, and after 5 h the animals were killed and the brains were homogenized to obtain the subcellular fractions. Fig. 3 demonstrates that the largest percentage incorporated was into the P2 fraction, and, of this fraction, nearly 50 % was found in the plasma membranes. The uptake into synaptosomes was about 12% of the uptake into the P2 fraction. There was no significant difference between the two synaptosomal peaks in this experiment. It seems clear that there is an active transport of free choline into isolated synaptosomes in vitro (Diamond and Kennedy, 1969 Marchbanks, 1968) but only a small amount of the incorporated choline is converted to ACh. In vivo we found that 40 % of the labelled choline in the synaptosomes was in a lipid-bound form 5 h after injection, and work is in progress... 

Fig. 3. The uptake of [A/e-14C]choline into the subcellular fractions of whole rat brain 5 h after intracerebral injection. A, The distribution of radioactivity as a percentage of that in the homogenate in the nuclei + debris (Pi), crude mitochondrial fraction (P2), microsomal fraction (Ps) and soluble cell sap (Sol.) and B, The distribution of radioactivity in the components of the Pa fraction. Peaks are the same as those described in Fig. 2.

Our "in vitro" experiments were done using brain slices. The incorporation of precursor (either radioactive choline or Na2S04) into brain subcellular fractions was studied up to 120 min. Chase experiments were done washing the slices 15 min. after starting the incubation and re-incubating in the presence of unlabelled precursor. With choline, there is a short initial lag period followed by linear incorporation during the time studied. In chase experiments, the incorporation rapidly reaches a plateau, without decreasing thereafter (Fig. 6). 

Incorporation of radioactive choline into PC of neuronal and glial enriched fractions and turnover of the labelled phospholipids was studied at different times up to 20 days after the intracranial injection of the precursors. The pattern of incorporation and decay was very similar to that found in brain subcellular fractions (Fig. 

Fig. 5. "In vivo" incorporation of choline (%-methyl) in subcellular fractions of rat brain. (A) Incorporation into lipids of the different subcellular fractions was measured at various times after the intracranial injection of 2 pCi of the radioactive precursor. (B)...

Fig. 1. Incorporation of radioactive choline into subcellular fractions of rat brain. Adult rats were injected intracranially with 2 )xCi of choline (%-methyl). Labelling of PC was studied in subcellular fractions at times between 0-48 h. Results are expressed as dpm/ iig of P of phosphatidyl choline. Each point represents the mean value of five different experiments. S.E.M. for each point was 10% or lower.

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