Methadone but not Morphine Inhibits Lubiprostone-Stimulated Cl− Currents in T84 Intestinal Cells and Recombinant Human ClC-2, but not CFTR Cl− Currents
Open Access
- 24 August 2012
- journal article
- research article
- Published by Springer Science and Business Media LLC in Cell Biochemistry and Biophysics
- Vol. 66 (1), 53-63
- https://doi.org/10.1007/s12013-012-9406-6
Abstract
In clinical trials, methadone, but not morphine, appeared to prevent beneficial effects of lubiprostone, a ClC-2 Cl− channel activator, on opioid-induced constipation. Effects of methadone and morphine on lubiprostone-stimulated Cl− currents were measured by short circuit current (Isc) across T84 cells. Whole cell patch clamp of human ClC-2 (hClC-2) stably expressed in HEK293 cells and in a high expression cell line (HEK293EBNA) as well as human CFTR (hCFTR) stably expressed in HEK293 cells was used to study methadone and morphine effects on recombinant hClC-2 and hCFTR Cl− currents. Methadone but not morphine inhibited lubiprostone-stimulated Isc in T84 cells with half-maximal inhibition at 100 nM. Naloxone did not affect lubiprostone stimulation or methadone inhibition of Isc. Lubiprostone-stimulated Cl− currents in hClC-2/HEK293 cells, but not forskolin/IBMX-stimulated Cl− currents in hCFTR/HEK293 cells, were inhibited by methadone, but not morphine. HEK293EBNA cells expressing hClC-2 showed time-dependent, voltage-activated, CdCl2-inhibited Cl− currents in the absence (control) and the presence of lubiprostone. Methadone, but not morphine, inhibited control and lubiprostone-stimulated hClC-2 Cl− currents with half-maximal inhibition at 100 and 200–230 nM, respectively. Forskolin/IBMX-stimulated hClC-2 Cl− currents were also inhibited by methadone. Myristoylated protein kinase inhibitor (a specific PKA inhibitor) inhibited forskolin/IBMX- but not lubiprostone-stimulated hClC-2 Cl− currents. Methadone caused greater inhibition of lubiprostone-stimulated currents added before patching (66.1 %) compared with after patching (28.7 %). Methadone caused inhibition of lubiprostone-stimulated Cl− currents in T84 cells and control; lubiprostone- and forskolin/IBMX-stimulated recombinant hClC-2 Cl− currents may be the basis for reduced efficacy of lubiprostone in methadone-treated patients.Keywords
This publication has 43 references indexed in Scilit:
- Lubiprostone for the treatment of opioid-induced bowel dysfunctionExpert Opinion on Pharmacotherapy, 2011
- Lubiprostone Activates Cl− Secretion via cAMP Signaling and Increases Membrane CFTR in the Human Colon Carcinoma Cell Line, T84Digestive Diseases and Sciences, 2010
- Gating of human ClC‐2 chloride channels and regulation by carboxy‐terminal domainsThe Journal of Physiology, 2008
- A synthetic prostone activates apical chloride channels in A6 epithelial cellsAmerican Journal of Physiology-Gastrointestinal and Liver Physiology, 2008
- Development of a generic transient transfection process at 100 L scaleCytotechnology, 2008
- Functional Expression of μ-Opioid Receptors in the Human Colon Cancer Cell Line, HT-29, and their Localization in Human ColonDigestive Diseases and Sciences, 2007
- Large-scale Transient Transfection of Mammalian Cells: A Newly Emerging Attractive Option for Recombinant Protein ProductionJournal of Structural and Functional Genomics, 2005
- Thiazolidinone CFTR inhibitor identified by high-throughput screening blocks cholera toxin–induced intestinal fluid secretionJCI Insight, 2002
- Differential Coupling of μ‐, δ‐, and κ‐Opioid Receptors to Gα16‐Mediated Stimulation of Phospholipase CJournal of Neurochemistry, 1998
- Methadone block of K+ current in squid giant fiber lobe neurons.The Journal of general physiology, 1996