Acetylcholine measured at short time intervals during transmission of nerve impulses in the electric organ of Torpedo

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Abstract
The amounts of total acetylcholine (ACh), ATP and vesicle-bound ACh were measured at short time intervals in the electrogenic tissue of T. marmorata. This study is to approach with biochemical analysis the speed of electrophysiological phenomena. A stimulator coupled to a rapid freezer device was used to quench a number of tissue samples simultaneously, at different time intervals during transmission of a brief train of impulses at 100 Hz. The level of total ACh decreased significantly with the first 10 impulses. Then a rapid but transient increase in total ACh occurred, reaching a maximum value by impulse 15-16. Vesicle-bound ACh did not exhibit any changes parallel to those of total ACh and did not decrease beyond the control level during transmission of 20 impulses at 100 Hz. The amount of ATP in the tissue varied in close relation to that of total ACh. No significant phase shift was observed between the transmitter and the nucleotide and the ACh/ATP molar ratio was not significantly different from 1. The shortest time interval investigated in this work was 10 ms. The rate at which the pieces of tissue are quenched for biochemical measurements when plunged into a liquid at low temperature was estimated. To what extent the freezing rate may distort measurements of the biochemical changes occurring in the tissue was also evaluated. Fast freezing appears to be a valuable approach for investigating the rapid biochemical changes underlying cholinergic transmission; a better time resolution might be reached at the price of greatly reducing the size of the samples. Transmission of a brief train of impulses is accompanied by significant changes in the amount of extravesicular ACh.