1994 Fiscal Year Final Research Report Summary
Immunocytochemical study of vertebrate cone photoreceptors labeled by antibody against cone opsin.
| Project/Area Number |
05680730
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
神経・脳内生理学
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| Research Institution | Okazaki Naitonal Research Institutes |
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Principal Investigator |
OHTSUKA Teruya Okazaki National Research Institute, National Institute for Physiological Sciences, Associate Professor, 生理学研究所, 助教授 (10051814)
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| Project Period (FY) |
1993 – 1994
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| Keywords | retina / photoreceptor / opsin / antibody |
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| Research Abstract |
The outer-segment lengths of photoreceptor are dependent on the shedding of their apical ends and the continuous addition of newly formed membrane at their basal ends. I studied diurnal changes in the supply of opsin to goldfish blue-sensitive cones by immunocyto-chemistry and light-microscope autoradiography. Fish were maintained on a 12/12 hr L/D cycle. Rabbit polyclonal antibody (Ab) was raised against the N-terminus of goldfish blue cone opsin. Cells labeled by Ab and a FITC-conjugated secondary antibody were observed with a laser confocal microscope. Newly synthesized protein was measured in the following way : groups of goldfish were injected intraocularly with 50 muCi ^3H-leucine either at 06 : 00 (light-onset) or at 18 : 00 (light-offset). Individual fish were then sacrificed at hourly intervals. Radial retinal sections (10 mum thick) were made, and RI grains within cones labeled by Ab and ABC-DAB were counted. Ab labeled outer-segments of small single cones that contain a blue
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-sensitive visual pigment. It also labeled complicated infoldings in the perinuclear region that corrsponded to Golgi bodies and/or rough endoplasmic reticulum. There was labeling of numerous tiny vesicles in the inner segment as well. Labeled organelles were most evident in goldfish sacrificed during nighttime. Tissue from animals sacrificed during daytime hours showed far less labeling, less than 5 vesicles per cone. The number of labeled vesicles increased rapidly in the dark reaching a peak of about 50 vesicles per cone within 3 hours after light-offset. Production of labeled vesicles decreased thereafter to the daytime level before light-onset. This time course is similar to the shedding rates of goldfish cone outer-segments reported by O'Day & Young (1978). The number of RI grains incorporated into blud-sensitive cones during nighttime hours overall is about twice that of grains incorporated during daytime. The time course of labeled vesicle production suggests that blue-sensitive cones supply opsin mainly after light-offset. The small diurnal changes in protein synthesis indicates that the cones produce opsin mainly during nighttime hours, while other kinds of proteins are produced during daylight hours. Less
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