Preparation of synaptosomes and vesicles with sodium diatrizoate

Abstract— Synaptosomes were prepared from rat brain on a continuous gradient of sodium diatrizoate and compared with synaptosomes obtained on a discontinuous sucrose gradient. The yield of synaptosomal protein using diatrizoate was double that obtained with sucrose. No significant differences in quality between the two preparations were found based on measurement of: β-glucuronidase, RNA polymerase, 2′,3′-cyclic nucleotide 3′-phosphohydrolase, total and Na⁺, K⁺ -ATPase, acetylcholinesterase, lactic acid dehydrogenase, glucose utilization, and serotonin uptake. Electron microscopy showed the vesicles in the diatrizoate synaptosomes to be better preserved. Vesicles prepared on diatrizoate segregated into two distinct bands which differed in electron microscopic appearance and enzyme activity. The less dense vesicles were smaller, had much higher Mg²⁺ -ATPase activity, and a lower content of acetylcholinesterase than the more dense vesicles. The less dense vesicle preparation was very homogeneous morphologically, free of myelin and mitochondria, and contained occasional organelle fragments and double membrane structures.