A nitric oxide synthase activity is involved in the modulation of acetylcholine release in Aplysia ganglion neurons: A histological, voltammetric and electrophysiological study

The role of nitric oxide or related molecules as neuromodulators was investigated in the buccal and the abdominal ganglia of the mollusc Aplysia californica. In a first step we showed that reduced nicotinamide adenine dinucleotide phosphate-diaphorase histochemistry and specific nitric oxide synthase immunohistochemistry labelled the same neurons and fibres in both ganglia, pointing to the presence of a neuronal nitric oxide synthase. In a second step, we performed voltammetric detection of nitric oxide-related molecules using a microcarbon electrode in a reduction mode. A peak identified as N-nitroso- l-arginine was detected at −1.66 V in both ganglia. The identification of this compound as a product of endogenous nitric oxide synthase activity was reinforced by the fact that its peak amplitude was decreased in the presence of N G-monomethyl- l-arginine, an inhibitor of nitric oxide synthase, and increased with its substrate, l-arginine. An additional proof of a nitric oxide synthase activity was the detection of nitrites and nitrates in high concentrations (millimolar range) by capillary electrophoresis. We also showed that these nitric oxide-related molecules modulated acetylcholine release at two identified synapses in these ganglia. l-Arginine decreased acetylcholine release it the inhibitory synapse (buccal ganglion), whereas it increased acetylcholine release at the excitatory synapse (abdominal ganglion). The nitric oxide synthase inhibitors, N ω-nitro- l-arginine and N G-monomethyl- l-arginine, had opposite effects. Moreover, the exogenous nitric oxide donor, 3-morpholinosydnonimine hydrochloride mimicked the effects of l-arginine on both inhibitory and excitatory cholinergic synapses. The identification of two cholinergic synapses where nitric oxide affects acetylcholine release in opposite ways provides a useful tool to study the cellular mechanisms through which nitric oxide-related molecules modulate transmitter release.