S(+)-ketamine suppresses desensitization of γ-aminobutyric acid type B receptor-mediated signaling by inhibition of the interaction of γ-aminobutyric acid type B receptors with G protein-coupled receptor kinase 4 or 5.
BACKGROUND
Intrathecal baclofen therapy is an established treatment for severe spasticity. However, long-term management occasionally results in the development of tolerance. One of the mechanisms of tolerance is desensitization of γ-aminobutyric acid type B receptor (GABABR) because of the complex formation of the GABAB2 subunit (GB2R) and G protein-coupled receptor kinase (GRK) 4 or 5. The current study focused on S(+)-ketamine, which reduces the development of morphine tolerance. This study was designed to investigate whether S(+)-ketamine affects the GABABR desensitization processes by baclofen.


METHODS
The G protein-activated inwardly rectifying K channel currents induced by baclofen were recorded using Xenopus oocytes coexpressing G protein-activated inwardly rectifying K channel 1/2, GABAB1a receptor subunit, GB2R, and GRK. Translocation of GRKs 4 and 5 and protein complex formation of GB2R with GRKs were analyzed by confocal microscopy and fluorescence resonance energy transfer analysis in baby hamster kidney cells coexpressing GABAB1a receptor subunit, fluorescent protein-tagged GB2R, and GRKs. The formation of protein complexes of GB2R with GRKs was also determined by coimmunoprecipitation and Western blot analysis.


RESULTS
Desensitization of GABABR-mediated signaling was suppressed by S(+)-ketamine in a concentration-dependent manner in the electrophysiologic assay. Confocal microscopy revealed that S(+)-ketamine inhibited translocation of GRKs 4 and 5 to the plasma membranes and protein complex formation of GB2R with the GRKs. Western blot analysis also showed that S(+)-ketamine inhibited the protein complex formation of GB2R with the GRKs.


CONCLUSIONS
S(+)-Ketamine suppressed the desensitization of GABABR-mediated signaling at least in part through inhibition of formation of protein complexes of GB2R with GRK 4 or 5.
Department of Anesthesiology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan.
Yuko
Y
Ando
Minoru
M
Hojo
Masato
M
Kanaide
Masafumi
M
Takada
Yuka
Y
Sudo
Seiji
S
Shiraishi
Koji
K
Sumikawa
Yasuhito
Y
Uezono
D000700:Analgesics / Q000494:pharmacology*; D000818:Animals; D001418:Baclofen / Q000494:pharmacology; D015153:Blotting, Western; D002478:Cells, Cultured; D006224:Cricetinae; D004347:Drug Interactions; D004361:Drug Tolerance; D055724:Electrophysiological Phenomena; D031541:Fluorescence Resonance Energy Transfer; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels / Q000187:drug effects / Q000378:metabolism; D054768:G-Protein-Coupled Receptor Kinases / Q000187:drug effects / Q000378:metabolism*; D058786:GABA-B Receptor Agonists / Q000494:pharmacology; D007649:Ketamine / Q000494:pharmacology*; D007668:Kidney / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D051381:Rats; D018080:Receptors, GABA-B / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D014981:Xenopus
D000700:Analgesics; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D058786:GABA-B Receptor Agonists; D018080:Receptors, GABA-B; D007649:Ketamine; D054768:G-Protein-Coupled Receptor Kinases; D001418:Baclofen