The control by feedback of the actions of an intracellular messenger will be investigated in a living photoreceptor. Inositol trisphosphate (InsP3) mobilizes intracellular calcium. In the field of eye research. InsP3 has been proposed as a messenger that mediates contraction of iris smooth muscle in response to acetylcholine as well as a mediator or modulator of phototransduction in both vertebrate and invertebrate photoreceptors. Elsewhere, inositol trisphosphate is thought to mediate events as diverse as secretion, fertilization, platelet activation and synaptic transmission. In the nervous system, the InsP3 messenger pathway has been suggested as a target for the anti-depressive actions of lithium. Despite this interest in InsP3, the small size of mammalian cells has limited knowledge about the dynamics of action of InsP3 in living cells. The cells chosen for the study are the giant photoreceptors of Limulus, in which InsP3 is thought to mediate light-induced calcium-release. The InsP3-sensitive store of calcium has been localized within the photoreceptor and methods for the rapid delivery of solutions into the living photoreceptor are available. The roles of diffusion of InsP3 to the calcium stores and of depletion and refilling of those stores will be assessed. The effects on calcium release by InsP3 of temperature, metabolites of InsP3, of pH, and of calcium ions will be determined. The specific hypothesis of a feedback loop by which calcium ions previously released by InsP3 can prevent InsP3 from releasing more calcium will be tested. If present, this feedback limits calcium release and may be responsible for the oscillations in free calcium frequently associated with the action of InsP3 in these and in other cells. Possible biochemical sites for this feedback will be investigated by measuring the calcium dependence of the breakdown of InsP3 and of the binding of radiolabelled InsP3 to photoreceptor membranes.
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