Confocal microscopy and computer-assisted image reconstruction of astrocytes in the mammalian retina

The distribution of astrocytes in the vascularized retina of pigs, rats and cats was investigated by confocal microscopy and computer-assisted image processing. In whole mounts, immunocytochemical identification was done by staining astrocytes for glial fibrillary acidic protein (GFAP), and blood vessels for alpha-smooth muscle actin or collagen IV. Double-staining was followed through consecutive optical sections and made it possible to precisely align the two markers in the inner retina. The resulting computer-assisted image reconstructions revealed asymmetric ensheathment of blood vessels by GFAP-positive fibres. The ultrastructural basis for this asymmetry, as studied by electron microscopy, was found to be different in pigs and cats. In the pig, astrocytes firmly ensheathed the vessel circumference, but glial filaments were much more abundant on the vitreal and lateral than on the scleral side. By contrast, in the cat astrocytes were generally confined to regions occupied by axonal bundles and constituted only part of the vascular glia limitans, else formed by Müller cells. Moreover, our observations unambiguously showed that individual astrocytes maintained simultaneous contact with axons and blood vessels and lined the vitreous body. The physical links provided by astrocytes suggest that they are able to function as central communicating elements between ganglion cells, the vasculature and the vitreous body.