Acetylcholine release evoked by single or a few nerve impulses in the electric organ of Torpedo.

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Abstract
The acetylcholine (ACh) store in the T. marmorata electric organ was partially labeled with choline and acetate at the same molar concentration but with different isotopes. Under these conditions the 2 precursors were incorporated into ACh in a ratio 1:1. After a single electrical stimulus, or a brief burst of stimuli, the compound electroplaque potential (EPP) was recorded and the radioactive choline and/or acetate counted in the perfusion fluid, providing a sensitive assay for ACh release in the absence of anticholinesterase drugs. The so-called depression of transmission was due to progressive impairment of ACh release in the successive impulses evoked by repeated stimuli. In a pair of impulses separated by 50 ms interval, less ACh was released by the 2nd than by the 1st impulse; this explained why the size of the 2nd EPP was depressed, using a direct measurement of ACh. In repetitive stimulations of longer duration, the maximum rate of release declined as the activity was prolonged. Thus the tissue progressively lost its ability to ensure release at high frequencies. An unexpected finding was that anticholinesterases like eserine or pre-treatment with fluostigmine (DFP) greatly reduced ACh release even by a single impulse. Evoked ACh release and EPP amplitude were both maximum between 10 and 20.degree. C. At higher temperatures, the evoked release decreased as the spontaneous release increased. Changes in external Ca2+ and Mg2+ produced similar changes in the EPP and evoked ACh release. The dose-response curve for Ca dependency of ACh release was very steep with a Hill''s coefficient of 3.2. With a single stimulus in the presence of 4-aminopyridine, there was a dramatic enlargement of the EPP and a still larger potentiation of the evoked ACh release. With this approach it was possible to avoid the inconveniences often encountered in similar studies, i.e., repetitive stimulation, low Ca solutions and cholinesterase inhibition. This permitted a good correlation between electrophysiological and biochemical estimates of transmitter release even by a single nerve impulse.