Methadone increases intracellular calcium in SH-SY5Y and SH-EP1-halpha7 cells by activating neuronal nicotinic acetylcholine receptors.
(-)-Methadone acts as an agonist at opioid receptors. Both (+)- and (-)-enantiomers of methadone have been suggested to be potent non-competitive antagonists of alpha3beta4 neuronal nicotinic acetylcholine receptors (nAChRs). In the present study, we have examined interactions of methadone with nAChRs by using receptor binding assays, patch-clamp recording and calcium fluorometry imaging with SH-SY5Y cells naturally expressing alpha7 and alpha3* nAChR subtypes and SH-EP1-halpha7 cells heterologously expressing human alpha7 nAChRs. Methadone potently inhibited binding of [3H]methyllycaconitine to alpha7 nAChRs and that of [3H]epibatidine to alpha3* nAChRs. Methadone pretreatment induced up-regulation of epibatidine binding sites in SH-SY5Y cells. Using whole-cell patch-clamp recording, both isomers of methadone activated cation currents via mecamylamine-sensitive nAChRs in SH-SY5Y cells. Nicotine and both (+)- and (-)-methadone evoked increases in [Ca2+]i in both fluo-3AM loaded cell lines, and these effects were blocked by mecamylamine and by the alpha7 selective antagonist methyllycaconitine, suggesting effects of methadone as alpha7-nAChR agonist. Sensitivity of sustained nicotine and methadone effects to blockade by CdCl2, ryanodine and xestospongin-c implicates voltage-operated Ca2+ channels and intracellular Ca2+ stores as downstream modulators of elevated [Ca2+]i. Collectively, our results suggest that methadone engages in complex and potentially pharmacologically significant interactions with nAChRs.
Division of Pharmacology and Toxicology, University of Helsinki, Helsinki, Finland. jukka.s.pakkanen@helsinki.fi
Jukka S
JS
Pakkanen
Heli
H
Nousiainen
Jari
J
Yli-Kauhaluoma
Irene
I
Kylänlahti
Tommi
T
Möykkynen
Esa R
ER
Korpi
Jian-Hong
JH
Peng
Ronald J
RJ
Lukas
Liisa
L
Ahtee
Raimo K
RK
Tuominen
D000701:Analgesics, Opioid / Q000494:pharmacology*; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism*; D045744:Cell Line, Tumor; D005470:Fluorometry; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D008024:Ligands; D008691:Methadone / Q000494:pharmacology*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D018722:Nicotinic Agonists / Q000494:pharmacology*; D018408:Patch-Clamp Techniques; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000378:metabolism*; D013997:Time Factors; D015854:Up-Regulation
D000701:Analgesics, Opioid; D008024:Ligands; D018722:Nicotinic Agonists; D011978:Receptors, Nicotinic; D002118:Calcium; D008691:Methadone