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Whitney Marie Cleghorn, Michelle M Giarmarco, James B Hurley, Susan E. Brockerhoff; Calcium modulation and dynamics in adult zebrafish cone photoreceptors. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):5510.
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© ARVO (1962-2015); The Authors (2016-present)
Ca2+ regulates a variety of cellular processes including signal transduction, energy production, and mitochondrial function. Because photoreceptors (PRs) use significant quantities of energy, consuming more ATP/minute per gram tissue than even the brain, our goal is to understand how PRs respond to external fluctuations of Ca2+, and how intracellular Ca2+ dynamics influence energy metabolism. We have previously shown that larval zebrafish PRs display heterogeneous Ca2+ transients. In this study we investigate Ca2+ dynamics in adult zebrafish PRs.
Adult zebrafish retinas expressing the genetically encoded fluorescent Ca2+ sensor GCaMP in cone PRs were dissected, flat-mounted, and sliced into 400-µm sections. Retinas were stained with propidium iodide or BODIPY and slices were imaged in a temperature-controlled chamber. Retinas were perfused with 0 mM or 3 mM Ca2+ Ringer’s solution with and without 10 mM isridipine. GCaMP fluorescence was monitored by confocal microscopy.
Ca2+ transients heterogeneous in amplitude and duration are prevalent in both the synapse and cell body of adult PRs. When Ca2+ is removed from the media, intracellular GCaMP fluorescence initially decreases, but then transiently increases, suggesting a compensatory Ca2+ release from internal stores. When extracellular Ca2+ is restored, intracellular fluorescence increases, first in the synapse and then throughout the cell. To understand how PRs take up Ca2+ from extracellular stores, we treated retinas with isridipine to block Ca2+ entry through L-type Ca2+ channels in the synapse. Preliminary experiments show that isridipine slows recovery of GCaMP fluorescence, suggesting PRs take up Ca2+ predominately through the synapse. Further, repeated transients released from intracellular stores may compensate for this block in external Ca2+ uptake.
Transgenic adult zebrafish retinas with GCaMP in cone PRs display robust Ca2+ dynamics. Intracellular Ca2+ responds to changes in external Ca2+. PRs take up Ca2+ predominantly from the synapse and then distribute it to the cell body. Insufficient external Ca2+ or a block in Ca2+ uptake elicits compensatory release of Ca2+ into the cytoplasm from internal stores. Understanding how PRs regulate Ca2+ homeostasis will provide a framework for understanding how PRs use Ca2+ to modulate metabolic processes such as NADH and ATP production, to regulate energy production and maintain cell health.
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