PC12 rat pheochromocytoma cells synthesize dynorphin. Its secretion is modulated by nicotine and nerve growth factor

The PC12 is a cloned rat pheochromocytoma cell line that retains a number of chromaffin cell characteristics, such as the presence of nicotinic cholinergic receptors, the synthesis and secretion of catecholamines, and the expression of a number of neuropeptide genes. The PC12 cell line is a useful model for the study of neuronal development, since PC12 cells can be induced to differentiate toward sympathetic neurons after exposure to nerve growth factor (NGF). PC12 cells can also be induced to differentiate toward the opposite direction, i.e. toward mature chromaffin cells. Morphological and biochemical changes mark the differentiation of PC12 cells toward either direction. Among the substances proposed as biochemical markers of PC12 cell differentiation toward chromaffin cells is the endogenous opioid precursor proenkephalin and its posttranslational peptide products. Indeed, the proenkephalin gene is expressed in both adrenal chromaffin and PC12 cells. The secretion of enkephalins from PC12 cells increases by several-fold after differentiation toward chromaffin cells. On the other hand, prodynorphin (another endogenous opioid precursor) is not present in normal adrenal chromaffin cells, but it is synthesized by human pheochromocytomas. It, thus, appears that dedifferentiation of chromaffin cells induces expression of the prodynorphin gene. Consequently, the aim of this study was to determine whether the rat pheochromocytoma-derived PC12 cell line expresses the prodynorphin gene, if it secretes dynorphins, and if the NGF-induced differentiation of PC12 cells toward sympathetic neurons affects the secretion of the latter. We found the following. 1) PC12 cells synthesize prodynorphin and secrete its peptide products. 2) The size of the prodynorphin transcript and the mol wt of its dominant form of dynorphin appear to be similar or identical to those of prodynorphin in rat anterior pituitary. 3) PC12 dynorphin secretion is increased, in a dose-dependent manner, after nicotinic cholinergic stimulation, an effect blocked by the specific nicotine antagonist hexamethonium. Thus, it appears that after cholinergic stimulation, PC12 dynorphin is cosecreted with catecholamines, a phenomenon described for a number of neuropeptides, including the proenkephalin-derived opioids. 4) The NGF-mediated differentiation of PC12 cells into sympathetic neurons exerted a stimulatory effect on basal, nicotine-induced, and depolarization-dependent dynorphin secretion. However, NGF did not shift the nicotine dose-response curve of dynorphin secretion. In conclusion, it appears that while changes in the secretion of proenkephalin-derived peptides may serve as a marker of PC12 cell differentiation toward chromaffin cells, an increase in the secretion of prodynorphin-derived peptides may represent a marker of NGF-induced differentiation of PC12 cells toward sympathetic neurons.