In:
Pharmacology, S. Karger AG, Vol. 72, No. 3 ( 2004), p. 205-212
Abstract:
Metabotropic G protein-coupled receptors have recently been recognized as targets for anesthetics and analgesics. In particular, G 〈 sub 〉 q 〈 /sub 〉 -coupled receptors such as muscarinic M 〈 sub 〉 1 〈 /sub 〉 receptors (M 〈 sub 〉 1 〈 /sub 〉 R) and 5-hydroxytryptamine (5-HT) type 2A receptors have been reported to be targets for anesthetics. Much less is known, however, about the effects of anesthetics on G 〈 sub 〉 i 〈 /sub 〉 -coupled receptors. Here we report a method to analyze functions of G 〈 sub 〉 i 〈 /sub 〉 -coupled receptors in 〈 i 〉 Xenopus 〈 /i 〉 oocytes expressing a chimeric Gα protein. A chimeric Gα 〈 sub 〉 q 〈 /sub 〉 protein Gα 〈 sub 〉 qi5 〈 /sub 〉 , which contains carboxy-terminus five amino acids of Gα 〈 sub 〉 i 〈 /sub 〉 , enables G 〈 sub 〉 i 〈 /sub 〉 -coupled receptors to couple to Gq-coupled receptor-mediated downstream pathways such as activation of phospholipase C. We determined acetylcholine (ACh)-induced Ca 〈 sup 〉 2+ 〈 /sup 〉 -activated Cl 〈 sup 〉 – 〈 /sup 〉 currents in 〈 i 〉 Xenopus 〈 /i 〉 oocytes coexpressing G 〈 sub 〉 i 〈 /sub 〉 -coupled muscarinic M 〈 sub 〉 2 〈 /sub 〉 receptors (M 〈 sub 〉 2 〈 /sub 〉 R) with the chimeric Gα 〈 sub 〉 qi5 〈 /sub 〉 . Although ACh did not induce any currents in oocytes expressing M 〈 sub 〉 2 〈 /sub 〉 R alone, it caused robust Cl 〈 sup 〉 – 〈 /sup 〉 currents in oocytes coexpressing M 〈 sub 〉 2 〈 /sub 〉 R with Gα 〈 sub 〉 qi5 〈 /sub 〉 . The EC 〈 sub 〉 50 〈 /sub 〉 of the ACh-induced Cl 〈 sup 〉 – 〈 /sup 〉 current mediated through Gα 〈 sub 〉 qi5 〈 /sub 〉 was 0.2 µmol/l, which was 2.2 times higher than that of the ACh-induced G protein-activated inwardly rectifying K 〈 sup 〉 + 〈 /sup 〉 currents activated by Gβγ subunits liberated from endogenously expressed Gα 〈 sub 〉 i 〈 /sub 〉 in 〈 i 〉 Xenopus 〈 /i 〉 oocytes. Other G 〈 sub 〉 i 〈 /sub 〉 -coupled somatostatin type 2, 5-HT 〈 sub 〉 1A 〈 /sub 〉 and δ-opioid receptors, when coexpressed with Gα 〈 sub 〉 qi5 〈 /sub 〉 in oocytes, also caused robust Ca 〈 sup 〉 2+ 〈 /sup 〉 -activated Cl 〈 sup 〉 – 〈 /sup 〉 currents. In oocytes coexpressing M 〈 sub 〉 2 〈 /sub 〉 R and Gα 〈 sub 〉 qi5 〈 /sub 〉 , a volatile anesthetic halothane inhibited M 〈 sub 〉 2 〈 /sub 〉 R-induced Cl 〈 sup 〉 – 〈 /sup 〉 currents in a concentration-dependent manner with the IC 〈 sub 〉 50 〈 /sub 〉 of 1.1 mmol/l, suggesting that halothane inhibits M 〈 sub 〉 2 〈 /sub 〉 R-induced cellular responses at clinically relevant concentrations. Treatment with the protein kinase C inhibitor GF109203X produced a 3.5-fold enhancement of the initial Cl 〈 sup 〉 – 〈 /sup 〉 currents induced by 1 µmol/l ACh in oocytes expressing M 〈 sub 〉 2 〈 /sub 〉 R and G 〈 sub 〉 qi5 〈 /sub 〉 . The rate of halothane-induced inhibition of Cl 〈 sup 〉 – 〈 /sup 〉 currents elicited by ACh, however, was not changed in such oocytes pretreated with GF109203X. These findings suggest that halothane inhibits the M 〈 sub 〉 2 〈 /sub 〉 R-induced signaling by acting at sites other than PKC activity. Collectively these findings suggest that the use of oocyte expressing Gα 〈 sub 〉 qi5 〈 /sub 〉 would be helpful to examine the effects of anesthetics or analgesics on the function of G 〈 sub 〉 i 〈 /sub 〉 -coupled receptors in the 〈 i 〉 Xenopus 〈 /i 〉 oocyte expression system.
Type of Medium:
Online Resource
ISSN:
0031-7012
,
1423-0313
Language:
English
Publisher:
S. Karger AG
Publication Date:
2004
detail.hit.zdb_id:
1483550-2
SSG:
15,3
