Characterization of cellular transport, subcellular distribution, and secretion of the neurotoxicant 1-methyl-4-phenylpyridinium in bovine adrenomedullary cell cultures.
Cultures of bovine adrenomedullary chromaffin cells accumulated 1-[methyl-3H]methyl-4-phenylpyridinium ([3H]MPP+) in a time- and concentration-dependent manner with an apparent Km of 0.7 microM and a Vmax of 3 pmol/min/10(6) cells. The uptake was sodium dependent and sensitive to inhibitors of the cell-surface catecholamine transporter. At low concentrations of MPP+, the subcellular distribution was identical to that of endogenous catecholamines in the catecholamine-containing chromaffin vesicles. However, at a higher concentration of MPP+, a larger proportion of the toxicant was recovered in the cytosolic fraction, with less in the chromaffin vesicle fractions. When cells were prelabeled with [3H]MPP+, at 1 and 300 microM, and then permeabilized with digitonin in the absence of Ca2+, there was a proportionally greater release of MPP+ from the cells labeled at the higher concentration of the toxicant. In the presence of Ca2+, cell permeabilization induced a time-dependent secretion of catecholamines and a parallel secretion of MPP+. Under these conditions, the secretion of endogenous catecholamines was unaffected by the presence of MPP+. When the permeabilization studies were carried out in the presence of tetrabenazine, a massive release of MPP+ was observed in the absence of Ca2+ and was not further increased by Ca2+. In intact cells prelabeled with 300 microM [3H]MPP+, the secretagogues nicotine and veratridine elicited a Ca2+ -dependent secretion of catecholamines and MPP+ from the cells in similar proportions to their cellular contents. Barium-induced release of both species was independent of external Ca2+.(ABSTRACT TRUNCATED AT 250 WORDS)
Reinhard
J F
JF
Division of Medicinal Biochemistry, Wellcome Research Laboratories, Research Triangle Park, North Carolina 27709.
Diliberto
E J
EJ
Daniels
A J
AJ
D000313:Adrenal Medulla / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002395:Catecholamines / Q000037:antagonists & inhibitors / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D002838:Chromaffin System / Q000166:cytology / Q000378:metabolism; D009538:Nicotine / Q000494:pharmacology; D009997:Osmotic Pressure; D011726:Pyridinium Compounds / Q000378:metabolism*; D013268:Stimulation, Chemical; D013347:Subcellular Fractions / Q000378:metabolism; D014018:Tissue Distribution
D002395:Catecholamines; D011726:Pyridinium Compounds; C026560:1-(4-methoxyphenyl)pyridinium; D009538:Nicotine