| Project/Area Number |
60480101
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
General anatomy (including Histology/Embryology)
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| Research Institution | Jichi Medical School |
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Principal Investigator |
SATITO T. Department of Anatomy, jichi Medical School, 医学部, 教授 (40090419)
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| Co-Investigator(Kenkyū-buntansha) |
HANIHARA T. Department of Anatomy, Jichi Medical School, 医学部, 助手 (00180919)
ARAKI M. Department of Anatomy, Jichi Medical School, 医学部, 助手 (00118449)
MATSUSHITA M. Department of Anatomy, Jichi Medical School, 医学部, 助手 (00118457)
YAMAKADO M. Department of Anatomy, Jichi Medical School, 医学部, 助教授 (80010114)
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| Project Period (FY) |
1985 – 1987
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| Project Status |
Completed (Fiscal Year 1987)
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| Budget Amount *help |
¥6,700,000 (Direct Cost: ¥6,700,000)
Fiscal Year 1987: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1986: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 1985: ¥4,600,000 (Direct Cost: ¥4,600,000)
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| Keywords | Papid freeze / enzyme histochemistry / phosphodiesterase / guanylate cycle / GTPase / 5'-nucleotidase / retina / 視制帽外節 / 急速凍結 / 酵素 / 組織化学 / 細胞化学 |
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| Research Abstract |
The development of the rapid freeze substitution method enables us to see excellent morphology in electron microscopy. Therefore, the possibility of enzyme histochemistry of cyclic nucleotide metabolizing enzymes on the quick feeze substituted rat retina was studied.
The ultrastructure of the freeze substituted samples was excellent in showing the fine parallel arrangement of the disk membranes in the rod outer segnt.
In spite of a long substitution fixation of 3 days, using 0.5% glutraldehyde and 6% tannic acid in acetone, guanylate cyclase, phosphodiesterase, CTPst, and 5'-nucleotidase were demonstrably retained. With the contrast enhancemen on biogical membranes by the 0.5-4% glutaraldehyde and 6% tannic acid in aceton, the site of phosphodiesterase activity was clearly visualized on the disk membranes of rod outer segment sepecially at cytoplasmic side. The low phosphodiesterase activity was observed in the rod outer segment from the dark adapted retina showing as some disk membranes were positive but the others were not. All disk membranes were appleared positive and enhanced at light conditions.
The present investigation reveals that the freeze substitution technique can preserve not only fine morphology but also enzyme activity in cells and tissues at cerain physiological conditions.
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