| Project/Area Number |
15590283
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Pathological medical chemistry
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| Research Institution | Kansai Medical University |
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Principal Investigator |
NISHIZAWA Mikio Kansai Medical University, Faculty of Medicine, Lecturer, 医学部, 講師 (40192687)
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| Co-Investigator(Kenkyū-buntansha) |
ITO Seiji Kansai Medical University, Faculty of Medicine, Professor, 医学部, 教授 (80201325)
FURUTA Kyoji Gifu University, Graduate School of Medicine, Associate Professor, 大学院・医学研究科, 助教授 (40173538)
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| Project Period (FY) |
2003 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2004: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2003: ¥1,900,000 (Direct Cost: ¥1,900,000)
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| Keywords | acromelic acid / allodynia / prostaglandin F2alpha / antisense oligonucleotide / subtraction / neuron / intracellular free calcium ion / spinal cord / プロスタグランジンF_<2α> |
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| Research Abstract |
Acromelic acid (ACRO), a kainate analogue isolated from a poisonous mushroom Clitocybe acromelalga, produced tactile pain (mechanical allodynia), when intrathecally (i.t.) injected to mice. Induction of allodynia by prostaglandin (PG) F2alpha, as well as ACRO, was selectively lost one week after i.t. injection of a sublethal dose of ACRO, although no neuronal damage except slight gliosis was observed in the lower spinal cord. To isolate cDNA involving in allodynia, we established the method for i.t. administration of antisense oligonucleotides through tubing to mice. We identified an antisense oligonucleotide targeting FP mRNA, which caused disappearance of PGF2alpha-induced allodynia and decrease of FP mRNA in the spinal cord. PGF2alpha rapidly increased [Ca^<2+>]i of the cells in the deeper layer of the dorsal horn, when the spinal cord slices were prepared from the saline- administered mice. When the FP antisense oligonucleotide was administered, the population of PGF2alpha-responsi
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ve cells in the slices reduced, and PGF2alpha-induced [Ca^<2+>]i increase of these cells diminished. These data suggested that there are the FP-expressing cells involved in PGF2alpha-induced allodynia in the dorsal horn. As a biochemical probe for an ACRO receptor, we designed and synthesized a novel ACRO analog possessing an azido-functionalized phenyl group. Although the analog exerted a biological activity equivalent to ACRO, specific bands could not be identified by photoaffinity labeling experiments. To isolate candidates for an ACRO receptor, we screened a subtraction cDNA library using the spinal cord of the mice to which ACRO or saline was injected. Positive clones expressed specifically in spinal cord were isolated, and then the full-length cDNAs were isolated. Antisense oligonucleotides targeting these cDNAs were injected to mice by the above-mentioned method, and several suppressed the PGF2alpha-induced allodynia. Some cDNAs showed interaction with FP and other receptors, suggesting the involvement to the ACRO receptor. Less
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