Essential role of rho kinase in the Ca2+ sensitization of prostaglandin F(2alpha)-induced contraction of rabbit aortae.
Inhibition of dephosphorylation of the 20 kDa myosin light chain (MLC(20)) is an important mechanism for the Ca(2+)-induced sensitization of vascular smooth muscle contraction. We investigated whether this mechanism operates in prostaglandin F(2alpha) (PGF(2alpha))-induced contraction of rabbit aortic smooth muscle and, if so, whether protein kinase C (PKC) or rho-associated kinase (rho kinase) contribute to the inhibition of dephosphorylation. In normal medium, PGF(2alpha) (10 microM) increased the phosphorylation of MLC(20) and developed tension. The rho-kinase inhibitors fasudil and hydroxyfasudil inhibited these changes, despite having no effect on a phorbol-ester-induced MLC(20) phosphorylation. After treatment with verapamil or chelation of external Ca(2+) with EGTA, PGF(2alpha) increased the MLC(20) phosphorylation and tension without an increase in [Ca(2+)](i), all of which were sensitive to fasudil and hydroxyfasudil. ML-9, a MLC kinase inhibitor, quickly reversed the KCl-induced MLC(20) phosphorylation and contraction to the resting level. However, fractions of PGF(2alpha)-induced contraction and MLC(20) phosphorylation were resistant to ML-9 but were sensitive to fasudil. Ro31-8220 (10 microM), a PKC inhibitor, did not affect the phosphorylation of MLC(20) and the tension caused by PGF(2alpha), thus excluding the possibility of the involvement of PKC in the PGF(2alpha)-induced MLC(20) phosphorylation. PGF(2alpha) increased phosphorylation at Thr654 of the myosin binding subunit (MBS) of myosin phosphatase, which is a target of rho kinase, and fasudil decreased the phosphorylation. These data suggest that the PGF(2alpha)-induced contraction is accompanied by the inhibition of MLC(20) dephosphorylation through rho kinase-induced MBS phosphorylation, leading to Ca(2+) sensitization of contraction. An actin-associated mechanism may also be involved in the PGF(2alpha)-induced sensitization.
Ito
Katsuaki
K
Department of Veterinary Pharmacology, Faculty of Agriculture, Miyazaki University, Miyazaki 889-2192, Japan.
Shimomura
Erika
E
Iwanaga
Takahiro
T
Shiraishi
Mitsuya
M
Shindo
Kazutoshi
K
Nakamura
Junji
J
Nagumo
Hiromitsu
H
Seto
Minoru
M
Sasaki
Yasuharu
Y
Takuwa
Yoh
Y
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D001381:Azepines / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015237:Dinoprost / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066298:In Vitro Techniques; D007211:Indoles / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes / Q000378:metabolism; D008970:Molecular Weight; D018994:Myosin Light Chains / Q000737:chemistry / Q000378:metabolism; D043263:Myosin-Light-Chain Phosphatase; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D011817:Rabbits; D014661:Vasoconstriction / Q000187:drug effects / Q000502:physiology*; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology; D054460:rho-Associated Kinases
D001381:Azepines; D004791:Enzyme Inhibitors; D007211:Indoles; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; D018994:Myosin Light Chains; D014662:Vasoconstrictor Agents; D014665:Vasodilator Agents; C431085:hydroxyfasudil; C056218:ML 9; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D015237:Dinoprost; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases; D043263:Myosin-Light-Chain Phosphatase; C049347:fasudil; D002118:Calcium; C064758:Ro 31-8220