Muscarinic blockade of beta-adrenoceptor-stimulated adenylyl cyclase: the role of stimulatory and inhibitory guanine-nucleotide binding regulatory proteins (Gs and Gi)

N J Pyne, M W Grady, D Shehnaz, P A Stevens, S Pyne, I W Rodger

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

The functional antagonism that exists between muscarinic and beta-adrenoceptor function in guinea-pig tracheal smooth muscle was investigated by assessing Gs and Gi regulated adenylyl cyclase activity in isolated membranes. 2. Membranes from guinea-pig tracheal smooth muscle contain both Gi alpha and Gs alpha as assessed by Western blots with anti-G-protein antibodies. 3. GppNHp, a non-hydrolysable analogue of guanosine 5'-triphosphate (GTP), was shown to stimulate adenylyl cyclase activity at high concentrations (10(-6)-10(-4) M). GppNHp also produced a concentration-dependent reduction in pertussis toxin-catalysed adenosine diphosphate (ADP)-ribosylation of Gi alpha. 4. Pretreatment of tracheal smooth muscle slices with methacholine (10(-6) M) provoked a blockade of isoprenaline plus GTP, GppNHp- and GTP-stimulated adenylyl cyclase. 5. Addition of methacholine to membranes did not trigger inhibition of GTP-stimulated adenylyl cyclase activity but did block the isoprenaline-mediated augmentation of GTP-stimulated adenylyl cyclase activity. 6. Pretreatment of tracheal smooth muscle with methacholine (10(-6) M) provoked a blockade of cholera toxin-catalysed NAD(+)-dependent ADP-ribosylation of Gs alpha. 7. Phorbol-12-myristate 13-acetate (PMA)-treatment of tracheal smooth muscle slices actually enhanced GppNHp-stimulated adenylyl cyclase activity in subsequently prepared membranes. 8. We suggest that methacholine in addition to inhibiting adenylyl cyclase via a Gi-dependent mechanism induces a functional inactivation of Gs activity. These results together may explain the functional antagonism that exists between increased muscarinic tone and the ability of beta-adrenoceptor agonists to provoke excitation-contraction uncoupling.
Original languageEnglish
Pages (from-to)881-7
Number of pages7
JournalBritish Journal of Pharmacology
Volume107
Issue number3
Publication statusPublished - 1992

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GTP-Binding Proteins
Adenylyl Cyclases
Adrenergic Receptors
Cholinergic Agents
Carrier Proteins
Guanosine Triphosphate
Methacholine Chloride
Smooth Muscle
Membranes
Isoproterenol
Adenosine Diphosphate
Guinea Pigs
Gs GTP-Binding Protein alpha Subunits
Cholera Toxin
Pertussis Toxin
NAD
Acetates
Western Blotting
Antibodies

Keywords

  • adenylate cyclase
  • adrenergic beta-antagonists
  • animals
  • cholera toxin
  • GTP-binding proteins
  • guanylyl imidodiphosphate
  • guinea pigs
  • isoproterenol
  • membranes
  • methacholine compounds
  • muscle
  • parasympathomimetics
  • tetradecanoylphorbol acetate
  • trachea

Cite this

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title = "Muscarinic blockade of beta-adrenoceptor-stimulated adenylyl cyclase: the role of stimulatory and inhibitory guanine-nucleotide binding regulatory proteins (Gs and Gi)",
abstract = "The functional antagonism that exists between muscarinic and beta-adrenoceptor function in guinea-pig tracheal smooth muscle was investigated by assessing Gs and Gi regulated adenylyl cyclase activity in isolated membranes. 2. Membranes from guinea-pig tracheal smooth muscle contain both Gi alpha and Gs alpha as assessed by Western blots with anti-G-protein antibodies. 3. GppNHp, a non-hydrolysable analogue of guanosine 5'-triphosphate (GTP), was shown to stimulate adenylyl cyclase activity at high concentrations (10(-6)-10(-4) M). GppNHp also produced a concentration-dependent reduction in pertussis toxin-catalysed adenosine diphosphate (ADP)-ribosylation of Gi alpha. 4. Pretreatment of tracheal smooth muscle slices with methacholine (10(-6) M) provoked a blockade of isoprenaline plus GTP, GppNHp- and GTP-stimulated adenylyl cyclase. 5. Addition of methacholine to membranes did not trigger inhibition of GTP-stimulated adenylyl cyclase activity but did block the isoprenaline-mediated augmentation of GTP-stimulated adenylyl cyclase activity. 6. Pretreatment of tracheal smooth muscle with methacholine (10(-6) M) provoked a blockade of cholera toxin-catalysed NAD(+)-dependent ADP-ribosylation of Gs alpha. 7. Phorbol-12-myristate 13-acetate (PMA)-treatment of tracheal smooth muscle slices actually enhanced GppNHp-stimulated adenylyl cyclase activity in subsequently prepared membranes. 8. We suggest that methacholine in addition to inhibiting adenylyl cyclase via a Gi-dependent mechanism induces a functional inactivation of Gs activity. These results together may explain the functional antagonism that exists between increased muscarinic tone and the ability of beta-adrenoceptor agonists to provoke excitation-contraction uncoupling.",
keywords = "adenylate cyclase, adrenergic beta-antagonists, animals, cholera toxin, GTP-binding proteins, guanylyl imidodiphosphate, guinea pigs, isoproterenol, membranes, methacholine compounds, muscle, parasympathomimetics, tetradecanoylphorbol acetate, trachea",
author = "Pyne, {N J} and Grady, {M W} and D Shehnaz and Stevens, {P A} and S Pyne and Rodger, {I W}",
year = "1992",
language = "English",
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Muscarinic blockade of beta-adrenoceptor-stimulated adenylyl cyclase: the role of stimulatory and inhibitory guanine-nucleotide binding regulatory proteins (Gs and Gi). / Pyne, N J; Grady, M W; Shehnaz, D; Stevens, P A; Pyne, S; Rodger, I W.

In: British Journal of Pharmacology, Vol. 107, No. 3, 1992, p. 881-7.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Muscarinic blockade of beta-adrenoceptor-stimulated adenylyl cyclase: the role of stimulatory and inhibitory guanine-nucleotide binding regulatory proteins (Gs and Gi)

AU - Pyne, N J

AU - Grady, M W

AU - Shehnaz, D

AU - Stevens, P A

AU - Pyne, S

AU - Rodger, I W

PY - 1992

Y1 - 1992

N2 - The functional antagonism that exists between muscarinic and beta-adrenoceptor function in guinea-pig tracheal smooth muscle was investigated by assessing Gs and Gi regulated adenylyl cyclase activity in isolated membranes. 2. Membranes from guinea-pig tracheal smooth muscle contain both Gi alpha and Gs alpha as assessed by Western blots with anti-G-protein antibodies. 3. GppNHp, a non-hydrolysable analogue of guanosine 5'-triphosphate (GTP), was shown to stimulate adenylyl cyclase activity at high concentrations (10(-6)-10(-4) M). GppNHp also produced a concentration-dependent reduction in pertussis toxin-catalysed adenosine diphosphate (ADP)-ribosylation of Gi alpha. 4. Pretreatment of tracheal smooth muscle slices with methacholine (10(-6) M) provoked a blockade of isoprenaline plus GTP, GppNHp- and GTP-stimulated adenylyl cyclase. 5. Addition of methacholine to membranes did not trigger inhibition of GTP-stimulated adenylyl cyclase activity but did block the isoprenaline-mediated augmentation of GTP-stimulated adenylyl cyclase activity. 6. Pretreatment of tracheal smooth muscle with methacholine (10(-6) M) provoked a blockade of cholera toxin-catalysed NAD(+)-dependent ADP-ribosylation of Gs alpha. 7. Phorbol-12-myristate 13-acetate (PMA)-treatment of tracheal smooth muscle slices actually enhanced GppNHp-stimulated adenylyl cyclase activity in subsequently prepared membranes. 8. We suggest that methacholine in addition to inhibiting adenylyl cyclase via a Gi-dependent mechanism induces a functional inactivation of Gs activity. These results together may explain the functional antagonism that exists between increased muscarinic tone and the ability of beta-adrenoceptor agonists to provoke excitation-contraction uncoupling.

AB - The functional antagonism that exists between muscarinic and beta-adrenoceptor function in guinea-pig tracheal smooth muscle was investigated by assessing Gs and Gi regulated adenylyl cyclase activity in isolated membranes. 2. Membranes from guinea-pig tracheal smooth muscle contain both Gi alpha and Gs alpha as assessed by Western blots with anti-G-protein antibodies. 3. GppNHp, a non-hydrolysable analogue of guanosine 5'-triphosphate (GTP), was shown to stimulate adenylyl cyclase activity at high concentrations (10(-6)-10(-4) M). GppNHp also produced a concentration-dependent reduction in pertussis toxin-catalysed adenosine diphosphate (ADP)-ribosylation of Gi alpha. 4. Pretreatment of tracheal smooth muscle slices with methacholine (10(-6) M) provoked a blockade of isoprenaline plus GTP, GppNHp- and GTP-stimulated adenylyl cyclase. 5. Addition of methacholine to membranes did not trigger inhibition of GTP-stimulated adenylyl cyclase activity but did block the isoprenaline-mediated augmentation of GTP-stimulated adenylyl cyclase activity. 6. Pretreatment of tracheal smooth muscle with methacholine (10(-6) M) provoked a blockade of cholera toxin-catalysed NAD(+)-dependent ADP-ribosylation of Gs alpha. 7. Phorbol-12-myristate 13-acetate (PMA)-treatment of tracheal smooth muscle slices actually enhanced GppNHp-stimulated adenylyl cyclase activity in subsequently prepared membranes. 8. We suggest that methacholine in addition to inhibiting adenylyl cyclase via a Gi-dependent mechanism induces a functional inactivation of Gs activity. These results together may explain the functional antagonism that exists between increased muscarinic tone and the ability of beta-adrenoceptor agonists to provoke excitation-contraction uncoupling.

KW - adenylate cyclase

KW - adrenergic beta-antagonists

KW - animals

KW - cholera toxin

KW - GTP-binding proteins

KW - guanylyl imidodiphosphate

KW - guinea pigs

KW - isoproterenol

KW - membranes

KW - methacholine compounds

KW - muscle

KW - parasympathomimetics

KW - tetradecanoylphorbol acetate

KW - trachea

M3 - Article

VL - 107

SP - 881

EP - 887

JO - British Journal of Pharmacology

JF - British Journal of Pharmacology

SN - 0007-1188

IS - 3

ER -