Short‐term Regulation of Catecholamine Biosynthesis in a Nerve Growth Factor Responsive Clonal Line of Rat Pheochromocytoma Cells

A clonal cell line (designated PC12) has been previously established from a transplantable rat adrenal medullary pheochromocytoma. Tissue cultures of PC12 cells synthesize, store, release and take up catecholamines. PC12 cells also respond to nerve growth factor (NGF) protein by cessation of mitosis and extension of neurites. The present studies concern the comparison of several aspects of catecholamine metabolism in PC12 cultures with that in normal noradrenergic tissues. One question was why the ratio of dopamine to norepinephrine in PC12 cultures (in contrast to that in normal noradrenergic tissue) is considerably more than one. The presence of exogenous reduced ascorbate (a cofactor for dopamine‐β‐monooxygenase) enhanced by 5–10‐fold the rate at which PC12 cultures converted [ 3 H]tyrosine to [ 3 H]norepinephrine. Under such conditions, the rate of synthesis of [ 3 H]do‐pamine was unchanged. It was also found that the ratio of norepinephrine to dopamine increased by 10‐fold when the cells were grown in vivo as tumors. Since tissue culture medium is essentially free of reduced ascorbate, it is likely that the absence of this cofactor is responsible for the low norepinephrine to dopamine ratio in PC12 cultures. Experiments were also carried out on short‐term regulation of catecholamine synthesis in PC12 cultures. These studies revealed the following: (1) The rate of conversion of [ 3 H]tyrosine to [ 3 H]catechols was increased 2–3‐fold (as compared with controls) in the presence of depolarizing levels of K + (51.5 mM), and by 2‐fold in the presence of 0.5–2 mM‐dibutyryl cyclic adenosine 3′, 5’monophosphoric acid (db‐cAMP). (2) Similar increases occurred in cultures which had been treated with (and had responded to) nerve growth factor. (3) The stimulatory effects of 51.5 mM‐K + rapidly returned toward control levels when the cultures were returned to control medium and (4) required the presence of Ca 2+ in the extracellular medium. (5) Stimulation of catechol synthesis by 51.5 mM‐K + and db‐cAMP also occurred in the presence of an inhibitor of DOPA decar‐boxylase. Thus, the ultimate effects of these agents were probably at the level of conversion of tyrosine to dopa by tyrosine 3‐monooxygenase. (6) Simultaneous exposure of cultures to 51.5 mM‐K + and mM‐db‐cAMP gave additive levels of stimulation. Such findings demonstrate that catecholamine synthesis in cultures of PC12 cells undergoes short‐term regulation which is similar to that previously demonstrated in normal monoaminergic tissues. As a homogeneous tissue culture line, the PC12 bears certain advantages for studying the primary mechanisms of such effects.