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In the first two of this series of papers, a sensitive microculture procedure was used to show that rat sympathetic neurons grown singly on small islands of heart cells release norepinephrine (NE) and/or acetylcholine (ACh). We report here the release of a third transmitter in response to stimulation of these neurons. This agent was recognized by its effect on the cocultured cardiac myocytes: an inhibition of beating or a hyperpolarization that, in contrast to cholinergic inhibition, was unaffected by atropine (up to 5 microM). Evidence described here indicates that this agent was primarily adenosine (or a closely related compound): the atropine-resistant myocyte inhibition was antagonized by adenosine-receptor blockers [8-phenyltheophylline, theophylline, 7-(2-chloroethyl) theophylline] and was attenuated by an enzyme (adenosine deaminase) that hydrolyzes adenosine to pharmacologically inactive inosine. Many of the neurons, whether initially dissociated from ganglia of newborn or adult rats, evoked this purinergic response, almost always in combination with adrenergic and cholinergic responses. In a few cases it was the only detectable response. The relative strength of the adrenergic, cholinergic, and purinergic responses varied widely from neuron to neuron, suggesting that the adrenergic and purinergic or the cholinergic and purinergic agents were not stored at constant stoichiometric ratios.

Synaptic functions in rat sympathetic neurons in microcultures. III. A Purinergic effect on cardiac myocytes