Pharmacologic and molecular characterization of the vascular ETA receptor in the venomous snake Bothrops jararaca.
Endothelins (ETs) and sarafotoxins (SRTXs) are active isopeptides that have very similar structures and functions. All isoforms interact with two specific G-protein-coupled receptors, ET(A) and ET(B). To characterize functional vascular ET receptors in the poisonous snake, Bothrops jararaca, cumulative concentration-response curves to ETs and SRTXs were performed in isolated aortic rings, in the absence and presence of selective ET receptor antagonists. Vascular expression of ET receptor messenger RNA (mRNA) was evaluated by reverse transcriptase (RT) polymerase chain reaction (PCR) analysis, and a fragment of the ET(A) receptor was cloned and sequenced. In vivo, ET-1 induced a dose-dependent biphasic response on anesthetized B. jararaca snakes. In vitro, ET-1, SRTX-b, ET-3, SRTX-c, and IRL-1620 induced concentration-dependent vasoconstriction, with a potency order suggesting the presence of typical ET(A) receptors. BQ-123, a selective ET(A) antagonist, inhibited contractions induced by ET-1 and SRTX-b with expected negative log of the dissociation constant, K(B), (pK(B)) values for mixed ET(A)/ET(B) receptor populations. The nonselective ET(A)/ET(B) receptors antagonist, PD-142893, produced similar inhibition. The ET(B) antagonist, IRL-1038, potentiated contractile responses to SRTX-c. ET-1 and SRTX-c responses were also potentiated when aortic rings were pretreated with N(omega)-nitro-L-arginine methyl ester (L-NAME) plus indomethacin. Processing of the B. jararaca aortic first-strand complementary DNA, by RT-PCR with primers designed from the Gallus gallus ET(A) receptor sequence, enabled isolation, purification, cloning, and sequencing of a single band. The partial sequence of the B. jararaca ET(A) receptor showed a very high sequence similarity with ET(A) receptor sequences from chicken, rat, human, and Xenopus. In conclusion, vascular responses to SRTXs/ETs in the B. jararaca aorta are mediated predominantly, but not exclusively, by typical ET(A) receptors.
Borgheresi
Rosa A M B
RA
Laboratory of Pharmacology, Butantan Institute, Av. Vital 1500, 05503-900 Sao Paulo, Brazil. rosabbor-gheresi@butantan.gov.br
Leroy
Janine M G
JM
Yogi
Alvaro
A
DosSantos
Rosangela A
RA
Breno
Maria C
MC
Tostes
Rita C
RC
D000595:Amino Acid Sequence; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects; D001483:Base Sequence; D001794:Blood Pressure / Q000187:drug effects; D017837:Bothrops; D017124:Conserved Sequence; D003435:Crotalid Venoms / Q000494:pharmacology*; D018076:DNA, Complementary / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D065130:Endothelin A Receptor Antagonists; D005260:Female; D007262:Infusions, Intravenous; D008297:Male; D008969:Molecular Sequence Data; D009119:Muscle Contraction / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects; D010477:Perfusion; D012333:RNA, Messenger / Q000032:analysis; D044022:Receptor, Endothelin A / Q000819:agonists / Q000737:chemistry* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D014661:Vasoconstriction / Q000187:drug effects; D014662:Vasoconstrictor Agents / Q000494:pharmacology
D003435:Crotalid Venoms; D018076:DNA, Complementary; D065130:Endothelin A Receptor Antagonists; D012333:RNA, Messenger; D044022:Receptor, Endothelin A; D014662:Vasoconstrictor Agents