Characterization of the Binding of the GABA Agonist [3H]Piperidine‐4‐Sulphonic Acid to Bovine Brain Synaptic Membranes

P. Krogsgaard‐Larsen, A. Snowman, S. C. Lummis, R. W. Olsen

Research output: Contribution to journalArticlepeer-review

Abstract

Abstract: The binding of radioactive piperidine‐4‐sulphonic acid ([3H]P4S) to thoroughly washed, frozen, and thawed membranes isolated from cow and rat brains has been studied. Quantitative computer analysis of the binding curves for four regions of bovine brain revealed the general presence of two binding sites. In these brain regions less satisfactory computer fits were obtained for receptor models showing one or three binding sites or negative cooperativity. With the use of Tris‐citrate buffer at 0°C the two affinity classes for P4S in bovine cortex membranes revealed the following binding parameters: KD= 17 ± 7 nM (Bmax= 0.15 ± 0.07 pmol/mg protein) and KD= 237 ± 100 nM (Bmax= 0.80 ± 0.20 pmol/mg protein). Heterogeneity was also observed for association and dissociation rates of [3H]P4S. The slow binding component (kon= 5.6 × 107 or 8.8 × 107 M‐1 min‐1, kOff= 0.83 min‐1, and KD= 14.7 or 9.4 nM, determined by two different methods in phosphate buffer containing potassium chloride) corresponds to the high‐affinity component of the equilibrium binding curve (KD= 11 nM, Bmax= 0.12 pmol/mg protein in the same buffer system). The association and dissociation rates for the subpopulation of rapidly dissociating sites, apparently corresponding to the low‐affinity sites, were too rapid to be measured accurately. The binding of [3H]P4S appears to involve the same two populations of sites with Bmax values similar to those for [3H]GABA binding to the same tissue, although the kinetic parameters for the two ligands are somewhat different. Furthermore, comparative studies on the inhibition of [3H]P4S and [3H]GABA binding by various GABA analogues, strongly suggest that P4S binds to the GABA receptors. The different effects of P4S and GABA on benzodiazepine binding are discussed.

Original languageEnglish (US)
Pages (from-to)401-409
Number of pages9
JournalJournal of Neurochemistry
Volume37
Issue number2
DOIs
StatePublished - Aug 1981
Externally publishedYes

Keywords

  • GABA agonist
  • GABA receptor
  • GABA‐benzodiazepine interaction
  • Piperidine‐4‐sulphonic acid
  • Receptor binding

ASJC Scopus subject areas

  • Biochemistry
  • Cellular and Molecular Neuroscience

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