Newly synthesized dopamine as the precursor for norepinephrine synthesis in bovine adrenomedullary chromaffin cells.
The precursor pool of dopamine for norepinephrine synthesis was investigated in cultured bovine adrenomedullary chromaffin cells incubated with [14C]tyrosine. Under conditions where the intracellular [14C]tyrosine specific activity was constant and [14C]dopamine synthesis was maximal, [14C]dopamine and [14C]norepinephrine accumulated over time, and the total intracellular dopamine content more than doubled within 120 min. When [14C]norepinephrine synthesis was calculated at different times based on the specific activity of [14C]dopamine, this rate was approximately equal to the rate of [14C]dopamine synthesis and was, thus, inconsistent with the observed dopamine accumulation. However, the rate of [14C]norepinephrine synthesis based on the [14C]tyrosine specific activity accounted for the dopamine accumulation, an observation suggesting that newly synthesized dopamine, i.e., dopamine with a specific activity equivalent to that of its precursor, [14C]tyrosine, is preferentially utilized for norepinephrine synthesis. Further studies showed that the subcellular distribution of [14C]dopamine was identical to that of norepinephrine and epinephrine and that the accumulated [14C]dopamine could be converted to norepinephrine within the chromaffin vesicle if dopamine uptake was blocked. Taken together, these results suggest that a small intravesicular dopamine pool, rapidly replenished by newly synthesized dopamine, serves as the substrate for dopamine beta-hydroxylase. Several mechanisms to account for this observation are discussed.