Coupling of the expressed alpha 1B-adrenergic receptor to the phospholipase C pathway in Xenopus oocytes. The role of Go

• Published in: The Journal of biological chemistry Vol. 268; no. 10; pp. 7532 - 7537
• Main Authors: Blitzer, R D, Omri, G, De Vivo, M, Carty, D J, Premont, R T, Codina, J, Birnbaumer, L, Cotecchia, S, Caron, M G, Lefkowitz, R J
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 05-04-1993
• Subjects: Animals; Cattle; Egtazic Acid - metabolism; Female; GTP-Binding Proteins - metabolism; Humans; In Vitro Techniques; Inositol 1,4,5-Trisphosphate - metabolism; Kinetics; Oocytes - enzymology; Pertussis Toxin; Receptors, Adrenergic, alpha - metabolism; Signal Transduction; Type C Phospholipases - metabolism; Virulence Factors, Bordetella - pharmacology; Xenopus laevis
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/s0021-9258(18)53208-7

alpha 1B-Adrenergic receptor mRNA was injected into Xenopus oocytes, resulting in a norepinephrine-evoked Cl- current. The
response was proportional to norepinephrine concentration, blocked by prazosin, and dependent on intracellular Ca2+ derived
from inositol trisphosphate-sensitive stores. Oocytes treated with 2 micrograms/ml pertussis toxin showed a time-dependent
decrease of the norepinephrine response, taking up to 72 h to show an 80% decrease. Overnight treatment with 10 micrograms/ml
pertussis toxin also resulted in 80% reduction. Responses to two other cloned receptors (M1-muscarinic and serotonin-1c) expressed
in oocytes were also reduced 50% or more by 72 h of pertussis toxin treatment. Pertussis toxin labeling of the cloned Xenopus
alpha o-subunit translated in vitro showed that it was a significantly poorer substrate for pertussis toxin than the two mammalian
alpha o-subunits expressed and assayed under identical conditions. This unexpected biochemical behavior of the Xenopus alpha
o-subunit is in agreement with the rather unusual treatment conditions required to observe the effects of pertussis toxin
on the receptor-evoked Cl- current in the oocyte. Injection of mammalian heterotrimeric G(o) but not Gi3 significantly enhanced
the norepinephrine-evoked Cl- current in oocytes. Injection of mixtures of anti-sense oligonucleotides to the Xenopus alpha
o-subunit reduced the norepinephrine-evoked Cl- current by 60% within 24 h, compared with oocytes injected with the oligonucleotides
encoding sense sequences. These studies indicate that the expressed alpha 1B-adrenergic receptor, like the native muscarinic
receptor, utilizes G(o) to couple to the phospholipase C-mediated Cl- current in Xenopus oocytes.