Light-induced Mn2+ influx in Limulus ventral photoreceptors

Abstract

In contrast to insect species, light-activated influx of divalent ions into Limulus ventral photoreceptors has proven difficult to demonstrate. We used the quench of the fluorescent indicator dye, fura-2, to measure Mn²⁺ influx. Limulus ventral photoreceptors were injected with fura-2 and excited at 360 nm. When the photoreceptors were bathed in 1 mmol · l⁻¹ Mn²⁺, an approximately 1% per 10 s decline in the fura-2 fluorescence during intervals between 50-ms flashes was taken as a measure of Mn²⁺ entry in darkness. Fluorescence decline during 10-s flashes was used to monitor Mn²⁺ entry during the photoresponse. During the 10-s flashes we observed a small rapid decline of the fura-2 fluorescence even in the absence of Mn²⁺. This reflected a contamination of the fluorescence signal arising from light-induced release of intracellular calcium stores. A subsequent slower decline in fluorescence during the 10-s flash, amounting to approximately 9% per 10 s, was only observed in the presence of extracellular Mn²⁺ and was attributed to Mn²⁺ influx. This light-activated influx was not through voltage-gated calcium channels since it persisted under voltage clamp, was not stimulated by depolarizing current injections, nor blocked by NiCl₂. Depletion of internal calcium stores by cyclopiazonic acid treatment did not accelerate Mn²⁺ influx.