Subcellular localization of chromogranins, calcium channels, amine carriers, and proteins of the exocytotic machinery in bovine splenic nerve.
Subcellular fractionation of bovine splenic nerves, which consist mainly of sympathetic nerve fibers, has been useful for characterizing cellular organelles en route to the terminal. In the present study we have characterized the subcellular distribution of both secretory and membrane proteins. A newly discovered chromogranin-like protein, NESP55, was found in large dense-core vesicles. The endogenous processing of NESP55 was comparable to that of chromogranins but more limited than that of secretogranin II and chromogranin B. For membrane proteins three major types of distribution were found. The amine carrier VMAT2 was confined to large dense-core vesicles. VAMP or synaptobrevin was present both in large dense-core vesicles and in lighter vesicles, whereas SNAP-25, syntaxin, and two types (N and L) of Ca2+ channels were found in a special population of lighter vesicles but were not present in large dense-core vesicles or at the most in very low concentrations. The plasma membrane norepinephrine transporter was apparently present in a separate type of vesicle, but this requires further study. These results further characterize vesicles en route to the terminal and establish for the first time that peptides involved in exocytosis (syntaxin, SNAP-25, and N- and L-type Ca2+ channels) are apparently transported to the terminal in a special type of vesicle. The exclusive presence of the amine carrier in large dense-core vesicles indicates that the formation of small dense-core vesicles in the terminals requires a reuse of membrane components of large dense-core vesicles.
Department of Pharmacology, University of Innsbruck, Austria.
B
B
Leitner
P
P
Lovisetti-Scamihorn
J
J
Heilmann
J
J
Striessnig
R D
RD
Blakely
L E
LE
Eiden
H
H
Winkler
D000588:Amines / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D015220:Calcium Channels / Q000378:metabolism*; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002851:Chromatography, High Pressure Liquid; D002864:Chromogranins / Q000378:metabolism*; D005089:Exocytosis / Q000502:physiology; D019205:GTP-Binding Protein alpha Subunits, Gs; D015151:Immunoblotting; D066298:In Vitro Techniques; D008562:Membrane Glycoproteins / Q000378:metabolism*; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009479:Neuropeptides; D011863:Radioimmunoassay; D013154:Spleen / Q000294:innervation* / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism*; D050492:Vesicular Biogenic Amine Transport Proteins
D000588:Amines; D015220:Calcium Channels; D002864:Chromogranins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D009479:Neuropeptides; D050492:Vesicular Biogenic Amine Transport Proteins; C498644:GNAS1 protein, Bos taurus; D019205:GTP-Binding Protein alpha Subunits, Gs