Detailed localization of photoreceptor guanylate cyclase activating protein-1 and -2 in mammalian retinas using light and electron microscopy

Guanylate cyclase activating proteins, GCAP-1 and GCAP-2, have a pivotal ro le in the activation of guanylate cyclase in phototransduction. Previous st udies on the localization of GCAP-1 and GCAP-2 are contradictory. In this s tudy, we tried to avoid possible artifacts accompanied by immunocytochemist ry. Immunolabeling of a GCAP was carried out using antibodies pre-adsorbed with a different type of GCAP. In addition, immunolabeling was performed us ing three different animal species under different fixation and embedding. Electron microscopic immunocytochemistry was also performed to reveal subce llular localization of GCAPs as well as confirming data obtained by light m icroscopy. All data indicate that anti-GCAP-1 antibody binding sites were f ound predominantly in cone outer segments, in particular, in disk membrane regions. Sparse labeling was observed in rod outer segments, but the labeli ng was much lower than that seen in cone outer segments. Less labeling is a lso found in synaptic regions and inner segments of cones. No labeling was detected in connecting cilia and its cytoplasmic extensions. Such labeling patterns were similar among human. monkey and bovine retinas. The localizat ion of GCAP-1 is consistent with the pattern of a recently reported human c one-specific degeneration. Anti-GCAP-2 antibody binding sites were detected in both inner and outer segments of rods and cones of all three animals al though the labeling density was slightly different among species. Cryo-immu no-labeling of GCAP-2 in bovine retinas revealed that labeling sites were m ore concentrated in rods than those of cones. and that synaptic regions wer e also labeled. The different localization of GCAPs suggest that roles of G CAP-1 and GCAP-2 may be different. (C) 1999 Academic Press.