Newly synthesized L-[14C]dopamine failed to accumulate immediately in synaptic vesicles within isolated rat brain synaptosomal cytoplasm.
It was examined whether or not enzymically synthesized L-dopamine could be immediately incorporated and stored in synaptic vesicles within a synaptosomal cytoplasm fraction isolated from the rat brain. The synaptosomal cytoplasm was isotonically prepared by disrupting the synaptosomes in 0.32 M sucrose using an N2-gas pressure homogenizer. Synaptic vesicles present in the isolated synaptosomal cytoplasm appeared to be osmotically sensitive (i.e., not leaky vesicles) under iso-osmotic conditions. DOPA decarboxylase [EC 4.1.1.28] present in the soluble cytosol region synthesized L-[14C]dopamine from added L-[14C]DOPA. However, the newly synthesized L-[14C]dopamine was not immediately incorporated in the synaptic vesicles within the synaptosomal cytoplasm during the reaction period of 20 min at 30 degrees C. Plain synaptic vesicles prepared by the method of Kadota and Kadota (10) also failed to take up the newly synthesized dopamine under the same conditions. It was found that soluble protein(s) in the synaptosomal cytoplasm was able to bind as much added L-[3H]dopamine as 35 pmol per mg of protein, and the binding ability seemed to inhibit the amine uptake by synaptic vesicles within the synaptosomal cytoplasm. The [3H]dopamine bound to the protein(s) became unbound in the presence of Ca2+ (5 mM). This effect of Ca2+ may have an important role in the releasing process of neurotransmitters.
D000818:Animals; D001692:Biological Transport; D001923:Brain Chemistry; D002118:Calcium / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism*; D004296:Dopa Decarboxylase / Q000378:metabolism; D004298:Dopamine / Q000378:metabolism*; D005260:Female; D008297:Male; D011485:Protein Binding; D051381:Rats; D011919:Rats, Inbred Strains; D013572:Synaptic Vesicles / Q000378:metabolism*; D013574:Synaptosomes / Q000378:metabolism
D004296:Dopa Decarboxylase; D002118:Calcium; D004298:Dopamine