Choline uptake, acetylcholine synthesis and release by Limulus abdominal ganglia

Isolated ganglia from the ventral nerve cord of the horseshoe crab, Limulus polyphemus, were incubated in ³H]choline and subsequently analyzed for choline uptake, conversion of choline to acetylcholine and the release of the newly synthesized acetylcholine. The ganglia readily accumulated radioactivity when incubated in Chao's solution containing 2 μM ³H]choline. The rate of uptake was 0.08 pmols/min/mg tissue and 26% of the ³H]choline taken up during a 1 h period was converted to ³H]acetylcholine. A 15 min exposure of ganglia to 90 mM K⁺ prior to incubation in ³H]choline caused an 89% increase in choline uptake and a 150% increase in its conversion to ³H]acetylcholine. The presence of unlabeled acetylcholine in the uptake medium inhibited both uptake and conversion of ³H]choline.There was a 5-fold increase in the efflux of radioactivity when ganglia incubated in 2 μM ³H]choline were superperfused with 90 mM K⁺. The increased efflux of radioactivity was Ca²⁺-dependent and was inhibited by Mg²⁺ (44%) and by Co²⁺ (72%). Similarly, addition of veratridine caused a Ca²⁺ and Na⁺-dependent release of radioactivity from prelabeled ganglia. Analysis of the superperfusate revealed that virtually all of the released radioactivity was ³H]acetylcholine.The abdominal ganglia of Limulus take up choline at micromolar concentrations, convert substantial amounts of it to acetylcholine and possess a depolarization-triggered, Ca²⁺-requiring mechanism for the specific release of acetylcholine. These results give further support to the view that the abdominal ganglia of Limulus contain a population of cholinergic terminals.