Characterization of [3H]-diprenorphine binding in Rana pipiens: Observations of filter binding enhanced by naltrexone

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Initial studies were undertaken to examine the properties of [3H]-diprenorphine binding to Rana pipiens whole brain tissue using naltrexone for the definition of nonspecific binding. Saturation analysis demonstrated the binding of [3H]-diprenorphine to be saturable with a K(D) value of 0.65 nM and a B(max) value of 287.7 fmol/mg protein. Unlabeled diprenorphine dose-dependently displaced [3H]-diprenorphine from a single noninteractive site in competition studies which yielded a K(i) of 0.22 nM. However, control studies in the absence of tissue revealed significant binding of [3H]-diprenorphine to the filter alone. Interestingly, [3H]-diprenorphine in the presence of unlabeled naltrexone as well as with unlabeled naloxone showed significantly greater binding to the filter than did [3H]-diprenorphine alone. Given this observation of increased nonspecific binding, an artificially low B(max) value would be expected. It is our hypothesis that the unlabeled nonspecific drug forms a complex with [3H]-diprenorphine preventing it from being effectively washed through the filter or the unlabeled drug itself is blocking the flow of [3H]-diprenorphine through the filter. The latter is unlikely however as other binding studies done in our lab using the radioligand [3H]-naloxone with unlabeled naltrexone do not show significant binding to the filter. Copyright (C) 1999 Elsevier Science Ltd.

Original languageEnglish
Pages (from-to)43-48
Number of pages6
JournalJournal of Pharmacological and Toxicological Methods
Issue number1
StatePublished - Feb 1999


  • Naltrexone
  • Rana pipiens
  • [H]-Diprenorphine


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