Abstract
The possibility that the large H+ electrochemical potential of chromaffin granules, the secretory granules of adrenal medullary chromaffin cells, plays an important role in exocytosis was investigated in cultures of chromaffin cells from bovine adrenal medulla. Methylamine uptake into the cells, [gamma-31P]phosphate nmr of ATP within intracellular chromaffin granules, O2 consumption of intracellular mitochondria, and MgATP-stimulated catecholamine uptake into chromaffin granules isolated from cultured chromaffin cells were assessed to determine whether various manipulations altered the H+ electrochemical gradients of intracellular chromaffin granules or mitochondria. Catecholamine secretion was not significantly altered by ammonium, methylamine, nigericin, carbonyl cyanide p-trifluoromethoxyphenylhydrazone, or dicyclohexylcarbodiimide under conditions when the pH of intracellular chromaffin granules was reduced or when granular or mitochondrial processes were uncoupled from H+ electrochemical gradients. The data indicate that the H+ electrochemical gradient across the chromaffin granule membrane does not play a role in exocytosis.
Highlights
RESULTSMethylamine was concentrateadpproximately 20-fold within chromaffin cells after a 100-min incubation in 0.1 mM methylamine (Fig. 1A)
Tential of chromaffin granules, the secretory granules of adrenal medullary chromaffin cells, playsan important role in exocytosis was investigated in cultures of chromaffin cells from bovine adrenal medulla
The data indicate that the H+ electrochemical gradient across the chromaffin granule membrane does not play a role in exocytosis
Summary
Methylamine was concentrateadpproximately 20-fold within chromaffin cells after a 100-min incubation in 0.1 mM methylamine (Fig. 1A). Methv lamine uptake into chromaffin cells was determined as described under "Materials and Methods'; af'er incubation for various times in medium containing 0.1 mM [3H]methylamine (0,0)or 10 mM [3H]methylamine (A).Some cells were incubated in solution containing 0.1 mM [3H]methylamine, 100 mM KC1, and 51 mM NaCl (0).The time course in the presence of 10 mM [3H]methylamine was performed on a different preparation of cells than the time courses in 0.1 mM [3H]methylamine. B, the effect of medium pH on methylamine uptake into chromaffin cells. Methylamine uptake into chromaffin cells was determined after a 45-min (0)or 60-min (0)incubation in 0.1 mM [3H]metbylamine in media of different pHs. Logarithm (base 10) of the methylamine concentration ratio (cell/ medium) is plotted uersus pH in the inset. The standard error of the mean bars were smaller than the symbols and were omitted
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