Nitric Oxide as an Efferent Modulator of Circadian Pacemaker Neurones in the Eye of the Marine Mollusc Bulla gouldiana

An antiserum against the nitric oxide (NO) synthetase labelled varicosities surrounding circadian pacemaker cells (“basal retinal neurons”, BRNs) in the eye of the marine mollusc Bulla gouldiana. These profiles appear to arise from extrinsic neurons with axons entering the eye via the optic nerve and are distinct from FMRFamide containing profiles, which represent the only efferent optic fibres in Bulla. In intact, dark-adapted eyes, the NO-donor SNAP (S-nitroso- N acetylpenicillamine) had no obvious effect on BRN activity recorded from the optic nerve. However, the light response of these circadian pacemaker neurones was reversibly enhanced by 8-bromo-cGMP, a tissue permeable analogue of cyclic guanosine monophosphate, and reduced by L-NAME (Nω-nitro-L-arginine methyl ester), a specific but irreversible inhibitor of NO synthesis. Furthermore, isolated BRNs in primary cell culture responded to the NO-donor SNAP with an elevated intracellular calcium concentration as measured by fluorescent calcium imaging using fura-2. This effect was blocked in the presence of the calcium chelator ethylene glycol tetra acetic acid (EGTA), and mimicked by application of the phosphodiesterase blocker isobutylmethylxanthine (IBMX). We propose that NO released from neurons entering the eyes, modulates the light responsiveness of the circadian pacemaker cells in Bulla by activating cGMP-gated calcium channels.