The synthesis, storage, and release of acetylcholine (ACh) were studied in perfused and intact superior cervical ganglia of cats. ACh was determined by bio-assay in ganglion extracts and in the venous effluent from ganglia perfused with fluid containing an anticholinesterase drug. Of the extractable ACh in a normal ganglion, about 85% is "depot ACh" available for release by nerve impulses. This must be located in the nerve endings; most of the remainder is in the intraganglionic portions of the preganglionic axons. The depot ACh exists as two fractions, of which one, the smaller, is the more readily available for release by nerve impulses. ACh synthesis and release go on at a measurable low rate in the absence of nerve impulses; both are greatly accelerated by activity. Under physiological conditions of excitation and perfusion, ACh release does not outrun ACh synthesis; but synthesis is slowed, with consequent depletion of depot ACh and reduction in ACh release, if choline is absent from the extracellular fluid, or if the hemicholinium base HC-3 is present. The latter compound specifically inhibits ACh synthesis by competing with choline; as a result it produces delayed block in repetitively activated cholinergic pathways. For efficient synthesis of ACh during experiments lasting an hour or so, a ganglion need be supplied with no substances other than choline and the constituents of Locke's solution; for the efficient release of ACh, the perfusion fluid must also contain CO2and an unidentified factor present in plasma. ACh accumulates above the resting level in a ganglion whose cholinesterase has been inactivated, provided that the perfusing fluid is one that supports ACh synthesis; the additional ACh is not immediately available for release by nerve impulses. Under physiological conditions of perfusion the amount of ACh set free by each maximal preganglionic volley is highest in a ganglion that has been at rest, and is then independent of stimulation frequency; after repetitive activation for several minutes the volley output is lower and is only frequency-independent at rates of excitation below 20/second. Consideration is given to the probable intracellular locations of the several fractions of the ganglionic ACh, and to their interrelationship.

Methods have been developed for depositing thin stable semipermeable polymer membranes around aqueous microdroplets (mean diameter down to 5 μ or less) by either interfacial polymerization or interfacial coacervation. The enclosed aqueous phase may contain enzymes or other proteins, cell fragments, or intact cells. Examples of methods for preparing such microcapsules are given in detail, and some of their properties are described.

COLLIER, B., AND MACINTOSH, F. C. 1969. The source of choline for acetylcholine synthesis in a sympathetic ganglion. Can. J. Physiol. Pharmacol. 47, Choline (Ch) and acetylcholine (ACh) uptake and release were measured by a combination of tracer and bioassay techniques in perfused superior cervical ganglia of the cat during rest and repetitive preganglionic stimulation. The uptake of labelled ACh as such was small; but when Ch ( m e t h~l -~H labelled) was present at physiological concentration (1.5 vg/ml) in the perfusion fluid, its incorporaticbn into the ACh and free Ch pools of the ganglion proceeded linearly in the absence of stimulation, was accelerated by stimulation, and was inhibited by hemicholinium but not by hexamcthonium. Up to 85 % of ganglionic ACh could be replaced by labelled ACh during a 1-h stin~ulation period. On subsequent perfusion with fluid containing unlabclled Ch, labelled Ch was lost but labelled ACh was retained and could be rcleased by stimulation, either as ACh in the presence or as Ch in the absence of eserine. The output of label during stimulation was approximately doubled by eserine but was unaffected by hemicholiniurn, which, however, prevented the formation and release of newly synthesized ACh. It appcars that about half the Ch formed from released ACh is immediately recaptured and resynthesized into ACh. Newly synthesized ACh rapidly gains access to the releasable transmitter pool and may be preferentially released.

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