Intracellular pathways involved in upregulation of vascular endothelin type B receptors in cerebral arteries of the rat.
OBJECTIVE
Previous studies have shown that contractile endothelin type B (ETB) receptors are upregulated in cerebral arteries after experimental focal cerebral ischemia. The aim of this study was to examine the upregulation of contractile ETB receptors in cerebral arteries after organ culture and to elucidate the intracellular pathways involved.


METHODS
Rat middle cerebral arteries (MCAs) were incubated with or without inhibitors. The vessels were mounted in myographs, and the contractile responses to endothelin-1 (ET-1) (ETA and ETB receptor agonist) and sarafotoxin 6c (ETB receptor agonist) were measured. Levels of ETB receptor mRNA were measured with real-time polymerase chain reaction.


RESULTS
In fresh MCA, sarafotoxin 6c had no contractile effect. However, after organ culture, a strong concentration-dependent contraction was induced. ET-1 produced a strong contraction, in which the Emax was unaffected by organ culture but the EC50 was decreased with time. The sarafotoxin 6c-induced contraction after 24 hours of organ culture was attenuated by the transcriptional inhibitor actinomycin D and the translational inhibitor cycloheximide as well as the protein kinase C inhibitor Ro-31-8220. Real-time polymerase chain reaction revealed that the mRNA levels of the ETB receptor were increased after organ culture compared with fresh vessels. Actinomycin D and Ro-31-8220 diminished the enhanced mRNA levels considerably.


CONCLUSIONS
The results suggest that, in fresh MCA, the ETA receptor is the most prominent subtype, while after organ culture ETB receptors also contribute to the contraction. This upregulation is due to de novo transcription of receptors. Protein kinase C is involved in the upregulation as Ro-31-8220 attenuates the contraction and the mRNA increase.
Division of Experimental Vascular Research, Lund University Hospital, Wallenberg Neurocentrum, BMC A13, 221 84 Lund, Sweden. Marie.Henriksson@med.lu.se
Marie
M
Henriksson
Emelie
E
Stenman
Lars
L
Edvinsson
D000818:Animals; D004305:Dose-Response Relationship, Drug; D019332:Endothelin-1 / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D020768:Middle Cerebral Artery / Q000187:drug effects / Q000502:physiology*; D009213:Myography; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology*; D014661:Vasoconstriction / Q000187:drug effects; D014757:Viper Venoms / Q000494:pharmacology
D019332:Endothelin-1; D004791:Enzyme Inhibitors; D019384:Nucleic Acid Synthesis Inhibitors; D012333:RNA, Messenger; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin; D014757:Viper Venoms; C057354:sarafotoxins s6; D011493:Protein Kinase C; D011188:Potassium