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A. B. Condren, P. Mettu, K. J. Liang, R. N. Fariss, J.-Y. Tsai, W. T. Wong; In situ Regulation of Choroidal Bloodflow by Smooth Muscle Cells and Pericytes: An ex vivo Confocal Time-Lapse Imaging Approach in Sclerochoroidal Explants. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4279.
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© ARVO (1962-2015); The Authors (2016-present)
Investigations of choroidal blood vessel regulation have been largely performed in dissociated and fragmented tissue. Here, we aim to study the regulation of choroidal bloodflow using an in situ model system of choroidal vessel behavior in intact sclerochoroidal explants using live imaging techniques.
Sclerochoroidal explants, containing intact sclera, choroid, and RPE, were acutely isolated from smooth muscle type -actin (SMA) promoter-driven green fluorescent protein (GFP) transgenic mice. Dynamic behavior of living smooth muscle cells and pericytes labeled with GFP was followed in situ using 3-dimensional time-lapse confocal imaging in response to vasoactive agonists.
GFP-labeled, SMA-positive perivascular cells had two distinct morphologies and distributions: (1) a band-like morphology that surrounds the circumference of vessels in a dense, ladder-like pattern, and (2) a pericyte-like morphology with sprawling processes extending from a central soma that sparsely envelope choroidal vessels. Time-lapse imaging revealed that both cell morphologies exhibit dynamic structural changes that resulted in vessel constriction when vasoactive agonists were applied. SMA-positive cells produced reversible vessel constriction in a dose-dependent manner to endothelin-1. Cellular behavior is also likely regulated by intracellular calcium as calcium entry, induced by A23187, a calcium-ionophore, produced vessel constriction, while calcium efflux, induced by transfer to a calcium-free medium, produced vessel dilation. The degree of vasoconstriction in imaged vessels increased as a function of increasing perivascular cell coverage, and is lower in vessels covered with low-density pericyte-like cells, compared to the vessels of similar diameter that are covered at a higher density by band-like cells.
Vital GFP-labeling of pericytes in transgenic mice presents a good in situ model (and a potential in vivo model) for studying regulatory changes in choroidal vasculature using live imaging. Perivascular SMA-positive cells in the choroid are associated with distinct morphologies and distributions that may help confer functional diversity in the vasoregulation of the choroidal bloodflow.
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