title: Rho-kinase inhibition acutely augments blood flow in focal cerebral ischemia via endothelial mechanisms.
abstract: Rho-kinase is a serine threonine kinase that increases vasomotor tone via its effects on both endothelium and smooth muscle. Rho-kinase inhibition reduces cerebral infarct size in wild type, but not endothelial nitric oxide synthase deficient (eNOS-/-) mice. The mechanism may be related to Rho-kinase activation under hypoxic/ischemic conditions and impaired vasodilation because of downregulation of eNOS activity. To further implicate Rho-kinase in impaired vascular relaxation during hypoxia/ischemia, we exposed isolated vessels from rat and mouse to 60 mins of hypoxia, and showed that hypoxia reversibly abolished acetylcholine-induced eNOS-dependent relaxation, and that Rho-kinase inhibitor hydroxyfasudil partially preserved this relaxation during hypoxia. We, therefore, hypothesized that if hypoxia-induced Rho-kinase activation acutely impairs vasodilation in ischemic cortex, in vivo, then Rho-kinase inhibitors would acutely augment cerebral blood flow (CBF) as a mechanism by which they reduce infarct size. To test this, we studied the acute cerebral hemodynamic effects of Rho-kinase inhibitors in ischemic core and penumbra during distal middle cerebral artery occlusion (dMCAO) in wild-type and eNOS-/- mice using laser speckle flowmetry. When administered 60 mins before or immediately after dMCAO, Rho-kinase inhibitors hydroxyfasudil and Y-27632 reduced the area of severely ischemic cortex. However, hydroxyfasudil did not reduce the area of CBF deficit in eNOS-/- mice, suggesting that its effect on CBF within the ischemic cortex is primarily endothelium-dependent, and not mediated by its direct vasodilator effect on vascular smooth muscle. Our results suggest that Rho-kinase negatively regulates eNOS activity in acutely ischemic brain, thereby worsening the CBF deficit. Therefore, rapid nontranscriptional upregulation of eNOS activity by small molecule inhibitors of Rho-kinase may be a viable therapeutic approach in acute stroke.
authors.affiliation: Department of Radiology, Stroke and Neurovascular Regulation Laboratory, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts 02129, USA.
authors.forename: Hwa Kyoung
authors.initials: HK
authors.lastname: Shin
authors.forename: Salvatore
authors.initials: S
authors.lastname: Salomone
authors.forename: E Michelle
authors.initials: EM
authors.lastname: Potts
authors.forename: Sae-Won
authors.initials: SW
authors.lastname: Lee
authors.forename: Eric
authors.initials: E
authors.lastname: Millican
authors.forename: Kensuke
authors.initials: K
authors.lastname: Noma
authors.forename: Paul L
authors.initials: PL
authors.lastname: Huang
authors.forename: David A
authors.initials: DA
authors.lastname: Boas
authors.forename: James K
authors.initials: JK
authors.lastname: Liao
authors.forename: Michael A
authors.initials: MA
authors.lastname: Moskowitz
authors.forename: Cenk
authors.initials: C
authors.lastname: Ayata
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002545:Brain Ischemia / Q000188:drug therapy / Q000503:physiopathology*; D002560:Cerebrovascular Circulation / Q000187:drug effects*; D015536:Down-Regulation / Q000187:drug effects; D004730:Endothelium, Vascular / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D002534:Hypoxia, Brain / Q000201:enzymology / Q000378:metabolism; D020244:Infarction, Middle Cerebral Artery / Q000473:pathology; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors*; D017078:Laser-Doppler Flowmetry; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D052250:Nitric Oxide Synthase Type III / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors*; D051381:Rats; D013268:Stimulation, Chemical; D014665:Vasodilator Agents / Q000494:pharmacology; D054460:rho-Associated Kinases
chemical_list: D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D014665:Vasodilator Agents; C431085:hydroxyfasudil; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D052250:Nitric Oxide Synthase Type III; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D000109:Acetylcholine; D019331:NG-Nitroarginine Methyl Ester