Characterization of rat brain opioid receptors by [Tyr-3,5-3H]1, d-Ala2, Leu5-enkephalin binding

S. Benyhe, Géza Tóth, Judit Kevei, M. Szücs, A. Borsodi, Katalin Di Gléria, Judit Szécsi, H. Süli-Vargha, Kálmán Medzihradszky

Research output: Contribution to journalArticle

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Abstract

[Tyr-3,5-3H]1, d-Ala2, Leu5-enkephalin ([3H]DALA) was used for labeling the opioid receptors of rat brain plasma membranes. The labeled ligand was prepared from [Tyr-3,5-diiodo]1, d-Ala2, Leu5-enkephalin by catalytic reductive dehalogenation in the presence of Pd catalyst. The resulting [Tyr-3,5-3H]1, d-Ala2, Leu5-enkephalin had a specific activity of 37.3 Ci/mmol. In the binding experiments steady-state level was reached at 24°C within 45 min. The pseudo first order association rate constant was 0.1 min-1. The dissociation of the receptor-ligand complex was biphasic with k-1-s of 0.009 and 0.025 min-1. The existence of two binding sites was proved by equilibrium studies. The high affinity site showed a KD=0.7 nM and Bmax=60 fmol/mg protein; the low affinity site had a KD=5 nM and Bmax=160 fmol/mg protein. A series of opioid peptides inhibited [3H]DALA binding more efficiently than morphine-like drugs suggesting that labeled ligand binds preferentially to the δ subtype of opioid receptors. Modification of the original peptides either at the C or N terminal ends of the molecules resulted in a decrease in their affinity.

Original languageEnglish
Pages (from-to)627-635
Number of pages9
JournalNeurochemical Research
Volume10
Issue number5
DOIs
Publication statusPublished - May 1985

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Enkephalins
Opioid Receptors
Rats
Brain
Ligands
Dehalogenation
Opioid Peptides
Cell membranes
Labeling
Morphine
Rate constants
Proteins
Binding Sites
Cell Membrane
Association reactions
Peptides
Catalysts
Molecules
Pharmaceutical Preparations
Experiments

ASJC Scopus subject areas

  • Neuroscience(all)
  • Biochemistry

Cite this

Characterization of rat brain opioid receptors by [Tyr-3,5-3H]1, d-Ala2, Leu5-enkephalin binding. / Benyhe, S.; Tóth, Géza; Kevei, Judit; Szücs, M.; Borsodi, A.; Di Gléria, Katalin; Szécsi, Judit; Süli-Vargha, H.; Medzihradszky, Kálmán.

In: Neurochemical Research, Vol. 10, No. 5, 05.1985, p. 627-635.

Research output: Contribution to journalArticle

Benyhe, S. ; Tóth, Géza ; Kevei, Judit ; Szücs, M. ; Borsodi, A. ; Di Gléria, Katalin ; Szécsi, Judit ; Süli-Vargha, H. ; Medzihradszky, Kálmán. / Characterization of rat brain opioid receptors by [Tyr-3,5-3H]1, d-Ala2, Leu5-enkephalin binding. In: Neurochemical Research. 1985 ; Vol. 10, No. 5. pp. 627-635.
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