The separation of synaptic vesicles from nerve-ending particles (‘synaptosomes’)

Abstract

A density-gradient procedure has been devised for fractionating pinched-off nerve endings ("synaptosomes") after disruption by suspension in water. Seven fractions are obtained. These consist of soluble cytoplasmic components, synaptic vesicles, microsomes, larger membrane fragments, partially disrupted synaptosomes and mitochondria. The various fractions have been characterized morphologically and biochemically. Lac-tate dehydrogenase (known to be a soluble-cytoplasmic marker), potassium and choline acetyltransferase were recovered mainly in the soluble-cytoplasmic fraction; succinate dehydrogenase in the mitochon-drial fraction; and cholinesterase in the microsome and large-mem- brane fractions. Bound acetylcholine was distributed bimodally, part being recovered in the synaptic-vesicle fraction and part in the fraction containing incompletely disrupted synaptosomes. The isolated synaptic vesicles had a mean diameter of 469 [plus or minus] 110 A, close to values reported in the literature for synaptic vesicles in whole-tissue preparations. The acetylcholine in the vesicle fraction was in the bound form, but was readily released on incubation or acid treatment. The specific activity of this fraction was higher than that of preparations of intact synaptosomes. The "compartmentalization" of acetylcholine, choline acetyl-transferase and cholinesterase in the cholinergic synapse is discussed in the light of these findings.