Choline Metabolism in Discrete Areas of the Brain: the Effect of Hemicholinium-3

Abstract

When [Me-14C]choline is injected into the brain of the rat it is rapidly phosphorylated to give phosphorylcholine; subsequently the radioactivity appears in the phosphatidylcholine (Ansell & Spanner, 1968). The incorporation into discrete areas of the brain has now been investigated, together with the effect of injecting the labelled choline at different depths from the surface. The object of this was to study the effect of hemicholinium-3, which interferes with the transport and utilization of choline for acetylcholine synthesis. Adult female rat brains were dissected as described by Glowinski & Iversen (1966), and in Table 1 the specific activities of acetylcholinesterase (EC 3.1.1.7) and choline kinase (EC 2.7.1.32) and the concentrations of total phospholipid, phosphatidylcholine and phosphorylcholine are given. Acetylcholinesterase activity was unevenly distributed (cf. cerebellum and striatum) and paralleled acetylcholine distribution (Schmidt et al., 1972), but choline kinase activity, largely present in areas rich in neurons (McCaman, 1962), was, like the phosphorylcholine, evenly distributed (Table 1). When [Me-14C]choline (1 pCi; 0.016pmol) was injected into the region of the lateral ventricle there was an uneven distribution of radioactivity between the six regions of the brain. A distribution of the same order was found for [3H]noradrenaline by Carr & Moore (1969). Of the radioactivity recovered in the brain, 67 % was distributed between the hypothalamus and the midbrain, but there was less than 0.3 % in the striatum. It was therefore decided to vary the depth of injection to test whether this uneven distribution was merely a reflection of the site of injection or was, in fact, a preferential uptake. The choline was injected at the same point of entry as before but the distance from the surface varied from 1.5 to 4mm. After an exchange time of 0.5h the total radioactivity in the brain was slightly higher after the shallower injections than after those at 4mm. The distribution of radioactivity among the regions was very different from that found after the injections at 4mm. For example, after the injection at 1.5mm the percentage of the recovered activity in the hypothalamus was only 8 % compared with 29 % after that at 4mm. However, the cortex contained 35 % compared with 14% after the injection at 4mm. It is noteworthy that the percentage distribution in the midbrain was almost unaffected by the depth of injection. It had been found previously (Ansell & Spanner, 1968) that there was a very rapid phosphorylation of choline after intraventricular injection. This was borne out when the individual regions were examined, each region showing a high percentage of labelling in the phosphorylcholine after 0.5-4.0h of exchange. However, when the labelled choline was injected into the brain tissue above the ventricle there was a very considerable depression of the phosphorylation, i.e. to an average of 15 % after the injection at 1 Smm and of 19.5 % after the injection at 3.0mm compared with over 75 % after that at 4mm. The distribution of choline kinase activity when measured in vitro showed little variation from region to region (Table l), so that the reason for this discrepency has yet to be explained. The ratio of the specific radioactivity of phosphatidylcholine to that of phosphorylcholine in vivo was unaffected. The transfer of the labelled choline and its metabolites from the injected to the uninjected side of the brain was also examined after different depths of injection. The steep gradient found by Carr & Moore (1969) with [3H]noradrenaline and by Fibiger et al. (1972) with ~-[U-'~C]leucine was not so marked with [Me-14C]choline. The side of injection contained 70 % of the isotopic label compared with 30 % in the uninjected side after 1-4h of exchange. This ratio was unaffected by the depth of injection. The cortex, striatum and midbrain showed the highest gradient. The medulla and hypothalamus contained the same percentage of radioactivity on both sides, whereas the cerebellum