Regulation of the CHAPS-solubilized muscarinic receptors by an inhibitory GTP binding protein (Gi) in the brain of neonatal and adult rats

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Abstract

CHAPS-solubilized muscarinic receptors (mAChRs) retain selective agonist or antagonist binding and sensitivity to GTP and ADP-ribosylation by pertussis toxin that ribosylates a group of G protein called Gi. This suggests that CHAPS solubilizes the mAChR-Gi complex both from neonatal and adult neurons. The gradient centrifugation of solubilized samples indicated that the neonatal samples contain mostly Gi-coupled mAChR (mAChRn,) exhibiting a high affinity for [3H]OxM, but the adult samples contain a comparable quantity of Gi coupled mAChR (mAChRa1) and uncoupled mAChRs (mAChRa2) exhibiting low affinity for the agonist. The binding of [3H]OxM to or the dissociation of [3H]OxM from mAChRn and mAChRa1 was sensitive to GTP. However, the effects of GTP were sensitive to Na+ for mAChRn but not for mAChRa1. ADP-ribosylation by pertussis toxin but not by cholera toxin abolished the effects of Na+ on agonist binding to solubilized mAChRn, suggesting that NaCl affects mAChRn not by interacting with the receptor but by interacting with the mAChR-Gi complex. The subtype composition of mAChRn, mAChRa1 and mAChRa2 was studied by determining the ic50 and the Ki values for the inhibition by subtype-selective antagonists in the binding of [3H]OxM to the receptors. The neonatal and adult samples exhibited differences in the distribution of the mAChR subtypes, the neonatal samples containing mostly the M1-mAChR subspecies but the adult samples containing M1-, M2- and M3-mAChR subspecies. The pirenzepine-induced inhibition in the binding of [3H]OxM both to M1-mAChRn and M1-mAChRa1 was sensitive to GTP. The GTP-induced effects on mAChRn were modulated by Na+, but the effect on M1-mAChRa1 was not. This suggests functional differences between Gi coupled M1-mAChRn and M1-mAChRa1.

Original languageEnglish (US)
Pages (from-to)255-267
Number of pages13
JournalComparative Biochemistry and Physiology -- Part A: Physiology
Volume109
Issue number2
DOIs
StatePublished - Oct 1994

Keywords

  • Adult
  • Brain
  • CHAPS
  • G
  • Muscarinic
  • Neonatal
  • Receptors
  • Subspecies

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