Occurrence of two types of secretory vesicles in the human neuroblastoma SH-SY5Y.
Western blot analysis showed that the human neuroblastoma SH-SY5Y expresses the proteins synaptotagmin l, synaptobrevin, synapsin-l, rab3a, syntaxin, SNAP-25, NSF, alpha-SNAP, and munc-18, which have been implicated in the movement, docking, and fusion of vesicles during exocytosis from other neuroendocrine cells. The subcellular localization of secretogranins I and II, synaptotagmin l, neuropeptide Y, rab3a, synaptobrevin, synaptophysin, and syntaxin was investigated by immunofluorescence microscopy and revealed punctate staining patterns characteristic of secretory vesicles. The comigration of noradrenaline, secretogranin II, and dopamine-beta-hydroxylase on sucrose-D2O gradient fractions indicates the presence of a population of noradrenaline-containing large dense-cored vesicles (LDCVs). In addition, a lighter vesicle population is also present that does not appear to be noradrenergic and contains a 48-kDa synaptophysin antigen absent from the large dense-cored vesicles. Immunocytochemical experiments show that not all of the vesicles that express synaptotagmin l contain secretogranin II. Thus, our studies suggest that two types of vesicle are present in SH-SY5Y cells, one of which, the LDCVs, contains noradrenaline. These findings confirm our previous studies suggesting that depolarization-evoked release of noradrenaline from SH-SY5Y occurs by LDCV exocytosis. This enhances the value of SH-SY5Y as a cell line in which to study the mechanism by which noradrenaline release is regulated.
Goodall
A R
AR
Institute for Cardiovascular Research, University of Leeds, England.
Danks
K
K
Walker
J H
JH
Ball
S G
SG
Vaughan
P F
PF
D000918:Antibody Specificity; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000032:analysis / Q000276:immunology; D002864:Chromogranins; D003594:Cytoplasmic Granules / Q000737:chemistry* / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D015151:Immunoblotting; D008562:Membrane Glycoproteins / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis / Q000276:immunology; D052066:N-Ethylmaleimide-Sensitive Proteins; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000276:immunology; D009447:Neuroblastoma; D009479:Neuropeptides / Q000032:analysis; D009638:Norepinephrine / Q000032:analysis; D011506:Proteins / Q000032:analysis; D050765:Qa-SNARE Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D013347:Subcellular Fractions / Q000737:chemistry; D050825:Synaptosomal-Associated Protein 25; D050857:Synaptotagmins; D014316:Tritium; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D033921:Vesicular Transport Proteins
D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D002864:Chromogranins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D009479:Neuropeptides; D011506:Proteins; D050765:Qa-SNARE Proteins; C494208:SNAP25 protein, human; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D050825:Synaptosomal-Associated Protein 25; D033921:Vesicular Transport Proteins; D014316:Tritium; D050857:Synaptotagmins; D052066:N-Ethylmaleimide-Sensitive Proteins; D009638:Norepinephrine