The visual arrestins in rhabdomeral photoreceptors are multifunctional phosphoproteins. They are rapidly phosphorylated in response to light, but the functional relevance of this phosphorylation is not yet fully understood. The phosphorylation of Limulus visual arrestin is particularly complex in that it becomes phosphorylated on three sites, and one or more of these site are phosphorylated even in the dark. The purpose of this study was to examine in detail the light-stimulated phosphorylation of each of the three sites in Limulus visual arrestin in intact photoreceptors. We found that light increased the phosphorylation of all three sites (S377, S381, and S396), that S381 is a preferred phosphorylation site, and that S377 and S381 are highly phosphorylated in the dark. The major effect of light was to increase the phosphorylation of S396, the site located closest to the C-terminal and very close to the adaptin binding motif. We speculate that the phosphorylation of this site may be particularly important for regulating the light-driven endocytosis of rhabdomeral membrane.