Electron microscopic localization of choline acetyl transferase activity in the electric organ of Torpedo marmorata.

The light microscopic method for demonstration of choline acetyltransferase (CAT) activity based on the formation of a lead mercaptide of free SH-acetyl Coenzyme A was adapted for electron microscopy. In samples of electric organ of Torpedo marmorata CAT activity was found to be restricted to synaptic vesicles and cysternae. The precipitate formed was mostly fine grained and distributed more or less evenly throughout the vesicles. Generally, the reaction product seemed not to adhere to the inner side of the vesicle membrane. CAT activity was found only in the presynaptic region of the synapse, neither the synaptic cleft nor the postsynaptic region reacted positively. CAT activity was found also within synaptic vesicles in nerve endings prepared from electric organ. Samples of Torpedo brain reacted positively too. Complete suppression of CAT activity with inhibitors, judged on the basis of lead mercaptide deposited, was rather difficult to achieve. From a group of 10 presumed enzyme inhibitors, only 2 compounds reacted satisfactorily, namely trans-1,2-dihydro-2-imino-4-(1-naphthylvinyl)-1-pyridine-ethanol hydrobromide and 5,5-dithio-bis-(2-nitrobenzoic acid) (3,3'-6). On the whole, the results obtained show the viability of the method used and furthermore it offers also some new insight into the turnover of acetylcholine, since it may be deduced from the results that under certain circumstances acetylcholine may be synthesized in synaptic vesicles.