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SL Facchina, A Verma MD PhD, W Watson MD PhD; Localization of Endoplasmic Reticulum Calcium Pools in the Porcine Eye . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2463.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Using a novel histochemical approach we have sought to determine the anatomical location of functional intracellular endoplasmic reticulum (ER) calcium pools within the porcine eye. Methods: Tissue Preparation: Eyes were harvested from euthanized animals, frozen rapidly, sectioned (18um frozen sections) along the horizontal axis and mounted onto Superfrost microscope slides. The slides were stored at -80 C until use in the calcium uptake assay. Calcium Uptake Assay: Each slide was allowed to equilibrate to room temperature for five minutes. The slides processed for in situ ATP-dependent endoplasmic reticulum 45Ca2+ transport activity as described previously (1). After incubation, slides were removed from uptake buffers, wash, dried and processed for 45Ca2+ autoradiography. Results: Prominent ER calcium pools were found not only in the retina, as expected but also in the trabecular meshwork and ciliary muscle. The optic nerve, sclera, chorioid and ciliary body have notably less calcium uptake. All calcium uptake was abolished by the calcium ionophore A23187 (10uM) and by the specific ER calcium pump inhibitor thapsigargin (1uM). Inositol-1,4,5-phosphate and caffeine were both able to release the accumulated calcium. These results demonstrate that functional calcium pools can be discretely localized in frozen sections of the eye. Conclusion: The endoplasmic reticulum calcium pools show definite heterogeneity within the structures of the eye and can be studied with a the simple and novel procedure described. Calcium plays a major role in ocular physiology and ocular pathophysiology. Our approach may be utilized for the development of pharmacotherapeutics in retinal diseases and glaucoma and may also be useful in studying the pathophysiological mechanisms underlying several eye diseases.
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