title: Comparison of MEK/ERK pathway inhibitors on the upregulation of vascular G-protein coupled receptors in rat cerebral arteries.
abstract: Organ culture is an in vitro method for investigating cellular mechanisms involved in upregulation of vasocontractile G-protein coupled receptors. We hypothesize that mitogen-activated-protein kinase (MEK) and/or extracellular-signal-regulated kinase (ERK) specific inhibitors will attenuate the G-protein coupled receptor expression following organ culture. Rat cerebral arteries were incubated 48h in the presence of MEK/ERK specific inhibitors U0126, PD98059, SL327, or AG126 for different time periods. Contractile responses by activation of endothelin receptor type A and type B, serotonin receptor 5-HT(1B), prostanoid TP receptor, and angiotensin II receptor type 1 and type 2 were investigated. Results were verified by measurement of mRNA with real time PCR and by protein immunohistochemistry. Organ culture induced transcriptional upregulation of endothelin ET(B) receptor and of serotonin 5-HT(1B) receptor on translational level and increased respective contractions. The prostanoid TP receptor mediated contraction curve was left-wards shifted by organ culture. Organ culture was associated with elevated pERK1/2 in the vascular smooth muscle cells: the MEK1/2 inhibitor U0126 attenuated the endothelin ET(B) receptor mediated contraction at post-translational level or by changing the receptor affinities. The serotonin 5-HT(1B) receptor and prostanoid TP receptor mediated contractions were abolished by U0126. Administration of U0126 6h after start of incubation blocked the receptor upregulation. In conclusion, MEK specific inhibitor U0126 is a potent inhibitor of G-protein coupled receptor alteration seen during organ culture. Given the ability to inhibit G-protein coupled receptor alteration at the clinically relevant time-point 6h post incubation makes it an attractive therapeutic agent for in vivo studies.
authors.affiliation: Department of Clinical Experimental Research, Glostrup Research Institute, Glostrup University Hospital, Denmark. sandhu.hardip@gmail.com
authors.forename: Hardip
authors.initials: H
authors.lastname: Sandhu
authors.forename: Saema
authors.initials: S
authors.lastname: Ansar
authors.forename: Lars
authors.initials: L
authors.lastname: Edvinsson
mesh_terms: D000818:Animals; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors*; D008297:Male; D020768:Middle Cerebral Artery / Q000187:drug effects / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D009924:Organ Culture Techniques; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D044023:Receptor, Endothelin B / Q000187:drug effects* / Q000235:genetics; D044302:Receptor, Serotonin, 5-HT1B / Q000187:drug effects* / Q000235:genetics; D017482:Receptors, Thromboxane / Q000187:drug effects / Q000378:metabolism; D013997:Time Factors; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D012333:RNA, Messenger; D044023:Receptor, Endothelin B; D044302:Receptor, Serotonin, 5-HT1B; D017482:Receptors, Thromboxane; D048049:Extracellular Signal-Regulated MAP Kinases; D020928:Mitogen-Activated Protein Kinases