BRAIN-TYPE CREATINE-KINASE IN PHOTORECEPTOR CELL OUTER SEGMENTS - ROLE OF A PHOSPHOCREATINE CIRCUIT IN OUTER SEGMENT ENERGY-METABOLISM AND PHOTOTRANSDUCTION

Different isoforms of creatine kinase, an important enzyme of vertebra te energy metabolism, were localized in bovine photoreceptor cells, wi th particular emphasis on the identification and quantification of the brain-type isoform within the outer segment compartment. Using immuno fluorescence and immunoelectron microscopy, brain-type creatine kinase was shown to be present in bovine photoreceptor cell outer and inner segments. The presence of this isoenzyme in rod outer segments was add itionally confirmed by immunoblotting and immunolabeling of isolated r od outer segments. The content of creatine kinase in rod outer segment s was quantified by measuring creatine kinase activity after membrane disruption with detergent. The ATP regeneration potential provided by the creatine kinase in isolated, washed bovine rod outer segments was 1.2 +/- (0.4) i.u. . mg-1 rhodopsin. This value was calculated to be a t least an order of magnitude larger than that necessary to replenish the energy required for cGMP resynthesis in rod outer segments, and hi gh enough to regenerate the entire ATP pool of rod outer segments with in the time span of a photic cycle. A mitochondrial creatine kinase is oenzyme was located within the ellipsoid portions of bovine rod and co ne inner segments by immunofluorescence microscopy and, using immunogo ld staining, was specifically localized in the mitochondria clustered within bovine rod and cone inner segments. These results suggest that vertebrate photoreceptor cells contain a functional phosphocreatine ci rcuit. Outer segment creatine kinase may play an important role in pho totransduction by providing energy for the visual cycle, maintaining h igh local ATP/ADP ratios and consuming protons produced by enzymes loc ated in the outer segment.