Abstract
The contractile effect of okadaic acid (OA) and its derivatives was investigated in the rat uterus. OA (20 μM) induced a transient contraction which, after plateauing, slowly decreased. The structurally related compound okadanol (20 μM) failed to induce any significant contraction. Conversely, the synthetic compound methyl okadaate (20 μM) and the naturally occurring ester 7′-hydroxy-4′-methyl-2′-methylen-hept-4′(E)-enyl okadaate (20 μM) were as active as the free acid. The OA-induced contraction was unaffected in the presence of neomycin (5 mM), mepacrine (30 μM), 1-[N,O-bis(1,5-isoquinolinesulfonyl)-N-methyl-l-tyrosyl]-4-phenylpiperazine (10 μM), calphostin C (3 μM) and 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (30 μM). The calmodulin inhibitor N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (100 μM) did not modify the amplitude of the OA-induced contraction but significantly increased the rate of tension decay. The myosin light chain kinase inhibitor 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (1 mM) significantly reduced the peak amplitude of the contraction. Staurosporine (0.03–0.1 μM) did not modify the contractile component of the OA-induced response but inhibited the subsequent decrease in tension. In freshly dispersed myometrial cells loaded with the fluorescent Ca++ indicator indo 1, OA did not produce any significant increase in [Ca++]i. OA (5- to 90-min contact) also failed to modify the intracellular levels of arachidonic acid, compared with basal values. These data suggest that in the rat uterus 1) the contractile effect of OA (20 μM) is specifically mediated by inhibition of protein phosphatases type 1 and/or 2A and is related to a direct interaction with the contractile machinery; 2) the decreasing phase of the OA-induced mechanical response could be mediated by a staurosporine-sensitive protein kinase different from protein kinase C.
Footnotes
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Send reprint requests to: Jean-Pierre Savineau, Laboratoire de Physiologie, Faculté de Médecine Victor Pachon, Université de Bordeaux II, 146 rue Léo Saignat, 33076 Bordeaux Cédex, France.
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↵1 This research was supported by grants from Pôle Médicament Aquitaine (France), the UE (contract CI1-CT92–0049) and the Ministry of Education and Science (Spain, grants PB 92–0487 and ALI-95–1012-CO5–02).
- Abbreviations:
- OA
- okadaic acid
- KN-62
- 1-[N,O-bis(1,5-isoquinolinesulfonyl)-N-methyl-l-tyrosyl]-4-phenylpiperazine
- H-7
- 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine dihydrochloride
- W-7
- N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride
- ML-9
- 1-(5-chloronaphthalene-1-sulphonyl)-1H-hexahydro-1,4-diazepine hydrochloride
- PP1
- protein phosphatase type 1
- PP2A
- protein phosphatase type 2A
- PKC
- protein kinase C
- PKA
- protein kinase A
- PKG
- protein kinase G
- MLCK
- myosin light chain kinase
- MLCP
- myosin light chain phosphatase
- LC20
- 20 kDa myosin light chain
- [Ca++]i
- cytosolic Ca++concentration
- cAMP
- cyclic AMP
- cGMP
- cyclic GMP
- ATP
- adenosine 5′-triphosphate
- ACh
- acetylcholine
- Emax
- maximal contractile effect
- AM
- acetoxymethyl ester
- DIBAL-H
- diisobutylaluminum hydride
- PSS
- physiological salt solution
- ANOVA
- analysis of variance
- Received December 16, 1996.
- Accepted March 17, 1997.
- The American Society for Pharmacology and Experimental Therapeutics
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