Cholinergic synaptic vesicle heterogeneity: evidence for regulation of acetylcholine transport.
Crude cholinergic synaptic vesicles from a homogenate of the electric organ of Torpedo californica were centrifuged to equilibrium in an isosmotic sucrose density gradient. The classical VP1 synaptic vesicles banding at 1.055 g/mL actively transported [3H]acetylcholine (AcCh). An organelle banding at about 1.071 g/mL transported even more [3H]AcCh. Transport by both organelles was inhibited by the known AcCh storage blockers trans-2-(4-phenylpiperidino)cyclohexanol (vesamicol, formerly AH5183) and nigericin. Relative to VP1 vesicles the denser organelle was slightly smaller as shown by size-exclusion chromatography. It is concluded that the denser organelle corresponds to the recycling VP2 synaptic vesicle originally described in intact Torpedo marmorata electric organ [Zimmermann, H., & Denston, C.R. (1977) Neuroscience (Oxford) 2, 695-714; Zimmermann, H., & Denston, C.R. (1977) Neuroscience (Oxford) 2, 715-730]. The properties of the receptor for vesamicol were studied by measuring binding of [3H]vesamicol, and the amount of SV2 antigen characteristic of secretory vesicles was assayed with a monoclonal antibody directed against it. Relative to VP1 vesicles the VP2 vesicles had a ratio of [3H]AcCh transport activity to vesamicol receptor concentration that typically was 4-7-fold higher, whereas the ratio of SV2 antigen concentration to vesamicol receptor concentration was about 2-fold higher. Based on an antibody standardization, in a typical preparation the VP1 vesicles contained 237 +/- 15 pmol of receptor/mg of protein whereas VP2 vesicles contained 102 +/- 3 pmol of receptor/mg of protein, and VP2 vesicles transported AcCh 2-3-fold more actively than VP1 vesicles.(ABSTRACT TRUNCATED AT 250 WORDS)
Gracz
L M
LM
Department of Chemistry, University of California, Santa Barbara 93106.
Wang
W C
WC
Parsons
S M
SM
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D001693:Biological Transport, Active; D002799:Cholinergic Fibers / Q000648:ultrastructure*; D004557:Electric Organ / Q000648:ultrastructure; D009550:Nigericin / Q000378:metabolism; D010622:Phencyclidine / Q000031:analogs & derivatives / Q000378:metabolism; D010880:Piperidines; D013572:Synaptic Vesicles / Q000378:metabolism*; D014101:Torpedo
D010880:Piperidines; C006189:vesamicol; D010622:Phencyclidine; D000109:Acetylcholine; D009550:Nigericin