CELLULAR RETINALDEHYDE-BINDING PROTEIN IN DEVELOPING RETINAL ASTROCYTES

Cellular retinaldehyde binding protein (CRALBP) is present in Muller g lia and in cells of the retinal pigment epithelium, but we have recent ly observed CRALBP-like immunoreactivity near the inner limiting membr ane in the newborn mouse retina, The present study has examined whethe r this protein is present in developing retinal astrocytes. Retinal ti ssue was collected at various embryonic and postnatal ages and in adul thood. Tissue for immunohistochemistry was fixed by immersion in 4% pa raformaldehyde and immunostained using rabbit polyclonal antisera to C RALBP or glial fibrillary acidic protein (GFAP), while fresh tissue wa s homogenized for Western analysis. Specificity of the antiserum for t he 33 kDa protein was shown in retinal homogenates by immunoblotting, with expression of the protein increasing steadily from E15.5 through adulthood. Immunostaining of sections from fetal eye-cups revealed fai nt labeling of cells in the optic nerve, with progressive migration of CRALBP-immunoreactive cells into the retina at the inner limiting mem brane during the perinatal period. By the day of birth, these cells we re intensely immunoreactive, showing a morphology characteristic of mi grating astrocytes. These CRALBP-immunoreactive cells mimicked the pro gressive infiltration of GFAP-positive astrocytes which are known to m igrate into the retina from the optic nerve head, many of which were d ouble-labeled with GFAP. Their distribution across the retina is disti nct from that of the lighter-staining Muller glial somata during these stages, and they are not misidentified Muller glial endfeet, Astrocyt es are only transiently CRALBP-immunoreactive, no longer containing th e protein after the second post-natal week, Preincubation of the antis erum with purified CRALBP abolished all staining of astrocytes. Couple d with the fact that only a single (similar to 33 kDa) molecular weigh t protein is labeled by the antiserum, it was concluded that retinal a strocytes contain CRALBP during a limited period of development. (C) 1 997 Academic Press Limited.