Coincident signalling between the Gi/Go-coupled delta-opioid receptor and the Gq-coupled m3 muscarinic receptor at the level of intracellular free calcium in SH-SY5Y cells.
In SH-SY5Y cells, activation of delta-opioid receptors with [D-Pen(2,5)]-enkephalin (DPDPE; 1 microM) did not alter the intracellular free Ca(2+) concentration [Ca(2+)](i). However, when DPDPE was applied during concomitant Gq-coupled m3 muscarinic receptor stimulation by carbachol or oxotremorine-M, it produced an elevation of [Ca(2+)](i). The DPDPE-evoked increase in [Ca(2+)](i) was abolished when the carbachol-sensitive intracellular Ca(2+) store was emptied. There was a marked difference between the concentration-response relationship for the elevation of [Ca(2+)](i) by carbachol (EC(50) 13 microM, Hill slope 1) and the concentration-response relationship for carbachol's permissive action in revealing the delta-opioid receptor-mediated elevation of [Ca(2+)] (EC(50) 0.7 mM; Hill slope 1.8). Sequestration of free G protein beta gamma dimers by transient transfection of cells with a beta gamma binding protein (residues 495-689 of the C terminal tail of G protein-coupled receptor kinase 2) reduced the ability of delta opioid receptor activation to elevate [Ca(2+)](i). However, DPDPE did not elevate either basal or oxotremorine-M-evoked inositol phosphate production indicating that delta-opioid receptor activation did not stimulate phospholipase C. Furthermore, delta-opioid receptor activation did not result in the reversal of muscarinic receptor desensitization, membrane hyperpolarization or stimulation of sphingosine kinase. There was no coincident signalling between the delta-opioid receptor and the lysophosphatidic acid receptor which couples to elevation of [Ca(2+)](i) in SH-SY5Y cells by a PLC-independent mechanism. In SH-SY5Y cells the coincident signalling between the endogenously expressed delta-opioid and m3 muscarinic receptors appears to occur in the receptor activation-Ca(2+) release signalling pathway at a step after the activation of phospholipase C.
Department of Pharmacology, University of Bristol, Bristol, UK.
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Yeo
D S
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Samways
C E
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Fowler
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Gunn-Moore
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Henderson
D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000502:physiology*; D002217:Carbachol / Q000494:pharmacology*; D006224:Cricetinae; D020881:Enkephalin, D-Penicillamine (2,5)- / Q000494:pharmacology*; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D009447:Neuroblastoma; D010095:Oxotremorine / Q000494:pharmacology*; D020239:Receptor Cross-Talk / Q000187:drug effects / Q000502:physiology*; D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000502:physiology*; D017465:Receptors, Opioid, delta / Q000187:drug effects / Q000502:physiology*; D014162:Transfection; D014407:Tumor Cells, Cultured
D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic; D017465:Receptors, Opioid, delta; D010095:Oxotremorine; D020881:Enkephalin, D-Penicillamine (2,5)-; D002217:Carbachol; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins; D002118:Calcium