Abstract
The effects of N-methyl-d-aspartate (NMDA) on opioid receptor-mediated G protein activation were explored in neuroblastoma X glioma hybrid (NG108–15) cells. Treatment of the cells with NMDA resulted in a remarkable attenuation of [35S]guanosine-5′-O-(3-thio)triphosphate binding stimulated by [d-Pen2,d-Pen5]-enkephalin (DPDPE), a δ-opioid receptor agonist. The effects of NMDA were dose and time dependent with an IC50 value of 5 nmand could be blocked by NMDA receptor antagonists. After NMDA treatment, the DPDPE dose-response curve shifted to the right (EC50 value increased ∼7-fold, from 6 to 40 nm), and the maximal response induced by DPDPE was reduced by ∼60%. The effects of NMDA were reversible, and the DPDPE response could recover within 60 min. The functional responses of δ-, μ-, and κ-opioid receptors in primarily cultured neurons also were attenuated significantly by NMDA treatment. The inhibitory effects of NMDA on opioid receptor-mediated G protein activation could be blocked by coadministration of the protein kinase C (PKC) inhibitors or by elimination of the extracellular Ca2+. Correspondingly, NMDA treatment of NG108 cells significantly elevated cellular PKC activity and stimulated Giα2 phosphorylation. Transient transfection into NG108–15 cells of the wild-type Giα2and a mutated Giα2 (Ser144Ala) resulted in a 2-fold increase in DPDPE-stimulated G protein activation. The DPDPE responses were greatly inhibited by NMDA treatment in the wild-type Giα2-transfected cells but much less affected in the mutant Giα2-transfected cells. In summary, NMDA attenuates opioid receptor/G protein coupling, and this process requires activation of PKC.
Footnotes
- Received July 18, 1997.
- Accepted January 12, 1998.
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Send reprint requests to: Dr. Gang Pei, Shanghai Institute of Cell Biology, Chinese Academy of Sciences, 320 Yue Yang Road, Shanghai 200031, People’s Republic of China. E-mail:gangpei{at}sunm.shcnc.ac.cn
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This work was supported by research grants from National Natural Science foundation of China (39630130 and 39625015), Chinese Academy of Sciences, Shanghai Research Center of Life Sciences, Shanghai Educational Development Foundation and German Max-Planck Society.
- The American Society for Pharmacology and Experimental Therapeutics
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