Dynamic Imaging of Retinal Microglia

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Immune cells in the retina demonstrate significant dynamic motility that report on their physiology and function. We have examined the anatomy and behavior of ocular immune cells including 1. microglia in the retina, and 2. macrophages in the choroid. Our work employed ex vivo time-lapse confocal imaging techniques to visualize fluorescence-labeled microglia from transgenic CX3CR1+/GFP mice and follow dynamic microglia behavior in intact retinal and choroidal explants in real time. Immune and vascular alterations in the choroid are implicated in age-related macular degeneration (AMD). As choroidal immune cells are incompletely understood with regard to their physiology and interactions with choroidal vessels, we examined the associations between myeloid and vascular components of the choroid in young and aged mice. Albino CX3CR1(GFP/+) transgenic mice, whose choroidal myeloid cells possess green fluorescence, were perfused intraluminally with the vital dye DiI to label choroidal vessels. The distribution, morphology, behavior, and vascular associations of resident myeloid cells were examined using time-lapse live confocal imaging and immunohistochemical analysis. Dendritiform myeloid cells, comprising most of the resident immune cell population in the choroid, were widely distributed across the choroid and demonstrated close associations with choroidal vessels that varied with their position in the vascular tree. Notably, myeloid cells associated with choroidal arteries and arterioles appeared as elongated cells flanking the long axes of vessels, whereas those associated with the choriocapillaris were distributed as a layer of stellate cells on the scleral but not vitreal choriocapillaris surface. While stationary in position, dendritiform myeloid cells demonstrated the rapid process dynamism well suited to comprehensive immunosurveillance of the perivascular space. Myeloid cells also increased in density as a function of aging, correlating locally with greater choroidal vascular attenuation. Resident myeloid cells demonstrated close but dynamic physical interactions with choroidal vessels, indicative of constitutive immune-vascular interactions in the normal choroid. These interactions may alter progressively with aging, providing a basis for understanding age-related choroidal dysfunction underlying AMD.