1998 Fiscal Year Final Research Report Summary
Elucidation of cellular mechanism of mu-opioid receptor agonist-induced analgesia by employing antisense oligodeoxynucleotide
| Project/Area Number |
09470026
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General pharmacology
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| Research Institution | Tokai University |
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Principal Investigator |
OKA Tetsuo Tokai University, School of Medicine, Professor, 医学部, 教授 (40055976)
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| Co-Investigator(Kenkyū-buntansha) |
YOSHIKAWA Masanobu Tokai University, School of Medicine, Assistant Researcher, 医学部, 助手 (90276791)
HASHIMOTO Atsushi Tokai University, School of Medicine, Assistant Professor, 医学部, 講師 (80271592)
KOBAYASHI Hiroyuki Tokai University, School of Medicine, Assistant Professor, 医学部, 講師 (60195807)
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| Project Period (FY) |
1997 – 1998
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| Keywords | mu-opioid receptor / G protein / antisense oligodeoxynucleotides / RT-PCR / western blot / in situ hybridization / opioid peptides / morphine-induced analgesia |
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| Research Abstract |
Antisense oligodeoxynucleotide (AS ODN) targeted against mu-opioid receptor (MOR) significantly decreased MOR mRNA content but did not significantly change mRNA contents of 6- and K-opioid receptors (DOR and KOR). Significant decrease in MOR mRNA by MOR AS ODN was also shown by in situ hybridization. In contrast to MOR AS ODN.MOR sense ODN.and DOR and KOR AS ODNs did not significantly change MOR mRNA contents. Effects of AS ODNs against five G protein cx subunits (Gialpha1, Gialpha2, Gialpha3, Goalpha and Gsalpha) were then examined by RT-PCR and western blot. Since specificities of reported AS ODNs were found to be low, we designed new AS ODNs. Among these, specificities of Giod and Gocx AS ODNs found to be high. Two these AS ODNs significantly decreased targeted proteins, their mRNA contents, and morphine-induced analgesia. Results indicate that both Gil and Go were involved in morphine-induced analgesia. Since AS ODNs targeted against Gialpha2, Gialpha 3 and Gsalpha reduced mRNA con
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tents of not only targeted G protein cc. subunits but also other alpha subunits, the involvement of three these G proteins in morphine-induced analgesia remained to be elucidated.
In contrast to morphine, endogenous MOR agonists such as Met^5- and Leu^5-enkephalin (met-enk and leu-enk) are known to be rapidly inactivated. Thus, their analgesic potencies had been difficult to be estimated. Therefore, we explored methods to protect these peptides completely from hydrolytic inactivation, and found that the mixture of three peptidase inhibitors (Pis). Amastatin, captopril and phosphoramidon, almost completely prevented the hydrolysis of five peptides, met-enk. leu-enk met-enk-Arg^6Phe^7. Met-enk-Arg^6Gly^7-Leu^8 and dynorphin A- (1-8) in cerebral membrane preparations. Furthermore, we found that analgesic potency of met-enk in rats pretreated with three PIs was approximately tenfold higher than that of morphine. We plan to examine the effect of each AS ODN on analgesia induced by these endogenous opioid peptides. Less
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Research Products
(12 results)
