Preferential release of newly synthesized 3 H-acetylcholine from rat cerebral cortex slices in vitro

1. Slices of rat cerebral cortex after treatment with the irreversible cholinesterase inhibitor soman, were incubated for 5 min in a Krebs-Henseleit solution containing 25 mM KCl and (3)H-choline. Subsequently incubation was continued in a medium containing non-radioactive choline and this medium was replaced at 5 min intervals. The amounts of labelled and total acetylcholine (ACh) released into the medium and extracted from the slices were determined at intervals.2. After the initial 5 min contact with (3)H-choline, 44% of the newly synthesized ACh contained a choline moiety originating from the choline in the medium. During the initial 5 min and the subsequent incubation part of the labelled ACh was released. While the rate of total ACh release remained constant, that of the release of labelled ACh was highest in the 5 min period following the initial incubation with (3)H-choline and then declined exponentially.3. The ratio of labelled ACh/total ACh in the ACh released during the initial 5 min incubation with (3)H-choline and during the subsequent 5 min was about three times as high as that in the ACh extracted from the slices at the end of these incubation periods.4. The ratio of labelled ACh/total ACh in superficial layers of the slices was not higher than that in the total slices.5. The rates of release of labelled and unlabelled ACh decreased when calcium was omitted from the incubation medium and were restored when the calcium was added. This suggests that both labelled and unlabelled ACh were released from nerve endings. The efflux of (3)H-choline was not calcium dependent.6. It is concluded that labelled ACh newly synthesized from externally applied (3)H-choline does not exchange immediately with all other ACh in the tissue and has a greater chance of being released than unlabelled ACh.