Synthesis, subcellular distribution and turnover of dopamine beta-hydroxylase in organ cultures of sympathetic ganglia and adrenal medullae.
The synthesis, subcellular distribution and turnover of dopamine beta-hydroxylase was studied in organ cultures of rat adrenal medullae and superior cervical ganglia. After exposure to [3H]leucine for 1 or 3 h, the tissues were homogenized at various time intervals and the amount of labelled dopamine beta-hydroxylase in different subcellular fractions (cytosol, soluble and membrane-bound fraction of catecholamine storage vesicles) was determined by immunoprecipitation and subsequent electrophoresis. In cultured adrenal medullae, induction of dopamine beta-hydroxylase initiated in vivo by administration of reserpine affected both soluble and membrane-bound pools of dopamine beta-hydroxylase to a similar extent after pulse-labelling for 1 or 3 h. The half-lives of dopamine beta-hydroxylase, which amounted to 6 h for the cytosol, 7.5 h for the soluble vesicular and 32 h for the membrane-bound vesicular pools were not altered by pretreatment with reserpine. In superior cervical ganglia the half-lives of the soluble pools were 2-3 times longer than in the adrenal medulla, whereas the half-life of the membrane-bound fraction was the same as in the adrenal medulla. In both organs the most heavily labelled fraction (both after a pulse of 1 or 3 h) was always that of the vesicular membrane, suggesting that newly-synthesized dopamine beta-hydroxylase is immediately incorporated into the storage vesicles and not via release into the cytosol from the site of synthesis. The fact that the half-life of membrane-bound dopamine beta-hydroxylase is markedly longer than that of the two soluble pools suggests that the single pools are not only independently supplied by newly-synthesized DBH but there is also no appreciable subsequent exchange between soluble and membrane-bound pools.
Gagnon
C
C
Department of Pharmacology, Biocenter of the University, Basel, Switzerland.
Schatz
R
R
Otten
U
U
Thoenen
H
H
D000313:Adrenal Medulla / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology*; D018759:Adrenergic Uptake Inhibitors / Q000494:pharmacology; D000818:Animals; D002458:Cell Fractionation; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D046508:Culture Techniques; D004299:Dopamine beta-Hydroxylase / Q000032:analysis / Q000378:metabolism*; D004586:Electrophoresis; D004790:Enzyme Induction / Q000187:drug effects; D007930:Leucine / Q000378:metabolism; D008297:Male; D011233:Precipitin Tests; D051381:Rats; D017207:Rats, Sprague-Dawley; D012110:Reserpine / Q000494:pharmacology; D013347:Subcellular Fractions / Q000737:chemistry / Q000201:enzymology; D017783:Superior Cervical Ganglion / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology*; D014316:Tritium
D018759:Adrenergic Uptake Inhibitors; D017077:Culture Media, Conditioned; D014316:Tritium; D012110:Reserpine; D004299:Dopamine beta-Hydroxylase; D007930:Leucine