A subtype of kappa-opioid receptor mediates inhibition of high-affinity GTPase inherent in Gi1 in guinea pig cerebellar membranes.
The kappa-opioid receptor agonists including U-50,488H and dynorphin A (1-17) in ranges of 0.1-100 nM inhibited the hydrolysis of GTP to GDP (P(i) release) inherent in GTP-binding proteins (G proteins) in guinea pig cerebellar membranes. U-50,488H inhibited only high-affinity GTPase activity, not low-affinity activity. The action of this agonist was found to be biphasic, and there was no inhibition at concentrations > 1 microM. The inhibition was abolished by pretreatment with preactivated pertussis toxin (PTX) at concentrations > 1 micrograms/ml but not with preactivated cholera toxin (30 micrograms/ml). Similar blockade of kappa-receptor-mediated inhibition was also observed when membranes were pretreated with a low concentration (8 microM) of N-ethylmaleimide (NEM) at low temperature (4 degrees C), which alkylates the cysteine residue to be ADP-ribosylated by PTX; but this treatment caused no significant change in kappa-agonist binding. When purified Gi1, but not G(o), was reconstituted into membranes pretreated with NEM, the kappa-receptor-mediated inhibition was recovered. These findings suggest that a subtype of kappa-opioid receptor is coupled to inhibition of intrinsic activity of Gi1.
Ueda
H
H
Department of Pharmacology, Yokohama City University School of Medicine, Japan.
Misawa
H
H
Fukushima
N
N
Katada
T
T
Ui
M
M
Satoh
M
M
D000818:Animals; D002531:Cerebellum / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000037:antagonists & inhibitors*; D019204:GTP-Binding Proteins / Q000378:metabolism* / Q000502:physiology; D006168:Guinea Pigs; D008297:Male; D008566:Membranes / Q000378:metabolism; D037342:Pertussis Toxin; D017473:Receptors, Opioid, kappa / Q000819:agonists / Q000502:physiology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D004791:Enzyme Inhibitors; D017473:Receptors, Opioid, kappa; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D005033:Ethylmaleimide