Purification and characterization of synaptic vesicles from the electric organ of Torpedo ocellata

Highly purified synaptic vesicles have been isolated from the electric organ of Torpedo ocellata by a rapid procedure which enables the concurrent isolation of synaptic vesicles and of intact presynaptic nerve endings (synaptosomes). The purification procedure consists of homogenization of fresh electric tissue in iso-osmotic glycine in the presence of EGTA, differential and density gradient centrifugation, and gel permeation on a glass beads column of 2500 a pore size. The purity of the vesicles was evaluated both biochemically and morphologically. The vesicles contain acetylcholine (ACh) and ATP in a ratio of 3:1 and at specific concentrations of 2,100nmol ACh/mg protein and 1,010nmol ACh/mg phospholipid. They are associated with Ca+2/Mg+2 ATPase activity and are devoid of the ouabain sensitive Na+/K+ ATPase. The relatively high yields as well as the short preparation time (about 9h for the vesicles and 4h for the synaptosomes) enables the employment of large samples of the isolated material on the day of preparation.