Methadone but not morphine inhibits lubiprostone-stimulated Cl- currents in T84 intestinal cells and recombinant human ClC-2, but not CFTR Cl- currents.
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.
Department of Molecular and Cellular Physiology, University of Cincinnati College of Medicine, 231 Albert Sabin Way, Cincinnati, OH 45267-0576, USA. John.Cuppoletti@uc.edu
John
J
Cuppoletti
Jayati
J
Chakrabarti
Kirti
K
Tewari
Danuta H
DH
Malinowska
D000527:Alprostadil / Q000031:analogs & derivatives* / Q000494:pharmacology; D001692:Biological Transport; D019256:Cadmium Chloride / Q000494:pharmacology; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000187:drug effects*; D005576:Colforsin / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D004353:Drug Evaluation, Preclinical; D057809:HEK293 Cells; D006801:Humans; D000068238:Lubiprostone; D008691:Methadone / Q000494:pharmacology*; D009020:Morphine / Q000494:pharmacology*; D009270:Naloxone / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
C505032:CFTR protein, human; D018118:Chloride Channels; D011994:Recombinant Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D009270:Naloxone; D000068238:Lubiprostone; D009020:Morphine; D000527:Alprostadil; D019256:Cadmium Chloride; D008691:Methadone