The Induction of Acetylcholine Synthesis in Primary Cultures of Dissociated Rat Sympathetic Neurons I. Effects of Conditioned Medium

Abstract

Neurotransmitter metabolism in primary cultures of dissociated sympathetic neurons from the newborn rat has been examined. Previous studies have shown that neurons grown in the virtual absence of nonneuronal cells developed the ability to synthesize and accumulate radioactive norepinephrine (NE) from [^3H]tyrosine, but synthesized little [^3H]acetylcholine (ACh) from labeled choline. However, in the presence of certain types of nonneuronal cells, or in medium conditioned (CM) by them, the neurons produced considerable ACh from choline. Here we show that ACh production from choline depends on the concentration of CM in the growth medium. At 62% CM, the highest level of CM studied, ACh production is >40-fold higher than the control value. Furthermore, homogenates of mixed neuronal/nonneuronal cultures or CM-treated neuronal cultures contain 100- to 1000-fold more choline acetyltransferase activity than neuron-alone sister cultures. On the other hand, the ability to synthesize and accumulate NE is inversely related to the percentage CM in the growth medium; at high concentrations of CM, catecholamine production is depressed about 25-fold from the control. The effects of CM are rather specific, since CM does not affect the number of neurons surviving or their overall growth. These results suggest that CM acts by altering the differentiated fate of individual sympathetic neurons rather than by causing the selective survival or growth of a second population of neurons. CM from primary cultures of several tissues taken from the newborn rat induces neuronal ACh synthesis, as does CM from cultures of several, but not all types of rat cell lines. In contrast, CM from several non-rat cell lines as well as from primary cultures of mouse and chick heart is less effective in causing neuronal ACh production, suggesting some degree of species specificity in this phenomenon.