Molecular and Cellular Biochemistry

, Volume 140, Issue 1, pp 65–71 | Cite as

Signal transduction pathways of muscarinic receptors in circular smooth muscle from the rabbit caecum

  • Pierre Cuq
  • Richard Magous
  • Jean-Pierre Bali
Article

Abstract

The effects of muscarinic acetylcholine receptor stimulation on phosphoinositides breakdown and adenylate cyclase activity were examined in the circular smooth muscle of the rabbit caecum. InMyo-[3H]inositol-labeled circular smooth muscle cells, carbachol caused a concentration-dependent increase in [3H]inositol phosphates ([3H]IPs) accumulation (EC50 of 3±1 μM). The M1-selective antagonist pirenzepine (PRZ), the M2-selective AF-DX 116 (11-2[[2-[(diethyl-amino)methyl]-1-piperidinyl]acetyl]-5, 11-dihydro-6Hpyrido[2,3-b][1,4]benzodiazepin-6-one) and the M3-selective para-fluoro-hexahydrosiladifenidol (p-F-HHSiD) inhibited the carbachol-induced [3]inositol phosphates accumulation with the following order of potency: p-F-HHSiD>PRZ>AF-DX 116. In saponin-permeabilized circular smooth muscle cells, carbachol and GTPγ[S] elicited a concentration-dependent increase in [3H]inositol phosphates accumulation. The concentration-response curve for GTPγ[S] was shifted to the left when cells were incubated with 1 μM carbachol. The [3H]inositol phosphates accumulation elicited by simultaneous addition of 0.1 μM GTPγ[S] and 1 μM carbachol to permeabilized cells was significantly decreased (78.28±18.23% inhibition) when cells were preincubated for 5 min with 0.1 mM GDPβ[S]. In nonpermeabilized cells, pertussis toxin did not alter the carbachol-induced increase in [3H]inositol phosphates accumulation. On the other hand, the 0.1 mM carbachol-induced inhibition of forskolin-stimulated adenylate cyclase activity in circular smooth muscle homogenates was significantly reversed by atropine and AF-DX 116, whereas PRZ and p-F-HHSiD were ineffective (muscarinic antagonists were used at 1 μM final concentration). Moreover, the carbachol-induced inhibition of the cyclic AMP accumulation elicited by 10 μM isoproterenol was abolished by pertussis toxin pretreatment of isolated circular smooth muscle cells. In conclusion, our data suggest that in circular smooth muscle of rabbit caecum, the muscarinic receptor stimulation of [3H]inositol phsophates accumulation is mediated by M3 subtype receptors coupled to a pertussis toxin-insensitive G protein, whereas inhibition of adenylate cyclase activity is mediated by M2 subtype receptors coupled to a pertussis toxin-sensitive GTP-binding protein Gi.

Key words

muscarinic receptors GTP-binding proteins phosphoinositides adenylate cyclase smooth muscle caecum 

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Copyright information

© Kluwer Academic Publishers 1994

Authors and Affiliations

  • Pierre Cuq
    • 1
  • Richard Magous
    • 1
  • Jean-Pierre Bali
    • 1
  1. 1.Laboratoire de Biochimie des Membranes, INSERM CJF 92-07Faculté de PharmacieMontpellier CédexFrance

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