scholarly journals Pertussis toxin does not inhibit muscarinic-receptor-mediated phosphoinositide hydrolysis or calcium mobilization

1985 ◽  
Vol 227 (3) ◽  
pp. 933-937 ◽  
Author(s):  
S B Masters ◽  
M W Martin ◽  
T K Harden ◽  
J H Brown
Keyword(s):  
Muscarinic Receptor ◽  
Muscarinic Receptors ◽  
Pertussis Toxin ◽  
Inositol Phosphate ◽  
Regulatory Protein ◽  
Calcium Mobilization ◽  
Phosphoinositide Hydrolysis ◽  
Heart Cells ◽  
Chick Heart ◽  
Stimulation Of

Pertussis toxin was used to examine the role of the inhibitory guanine nucleotide regulatory protein, Ni, in muscarinic-receptor-mediated stimulation of phosphoinositide turnover and calcium mobilization. In cultured chick heart cells, pertussis-toxin treatment inhibited muscarinic-receptor-mediated attenuation of isoprenaline-stimulated cyclic AMP accumulation. This finding is consistent with the proposal that pertussis toxin blocks the capacity of Ni to couple muscarinic receptors to adenylate cyclase. In contrast, treatment of chick heart cells or 1321N1 human astrocytoma cells with pertussis toxin did not block muscarinic-receptor-mediated stimulation of phosphoinositide hydrolysis, as measured by [3H]inositol phosphate accumulation in the presence of Li+. Pertussis-toxin treatment also had little effect on basal and muscarinic-receptor-stimulated phosphatidylinositol synthesis, as measured by the incorporation of [3H]inositol into phosphatidylinositol. Activation of muscarinic receptors also enhances the rate of unidirectional 45Ca2+ efflux in 1321N1 cells; this response, like phosphoinositide hydrolysis, was not prevented by pertussis-toxin treatment. Our data suggest that muscarinic receptors are not coupled to phosphoinositide hydrolysis or calcium mobilization through Ni.

scholarly journals Effects of pertussis toxin on growth factor-stimulated inositol phosphate formation and DNA synthesis in Swiss 3T3 cells

1988 ◽  
Vol 249 (3) ◽  
pp. 917-920 ◽  
Author(s):  
C W Taylor ◽  
D M Blakeley ◽  
A N Corps ◽  
M J Berridge ◽  
K D Brown
Keyword(s):  
Growth Factor ◽  
Dna Synthesis ◽  
Pertussis Toxin ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Phosphoinositide Hydrolysis ◽  
Polypeptide Growth Factors ◽  
Swiss 3T3 ◽  
Swiss 3T3 Cell ◽  
Stimulation Of

We have compared the effects of pretreatment of Swiss 3T3 cell with pertussis toxin on the stimulation of DNA synthesis and phosphoinositide hydrolysis in response to a wide variety of mitogens. The toxin substantially inhibited the stimulation of DNA synthesis in response to a phorbol ester or various peptide and polypeptide growth factors irrespective of their ability to activate phosphoinositidase C. Production of inositol phosphates in response to platelet-derived growth factor, fibroblast growth factor and prostaglandin F2 alpha were unaffected by the toxin while bombesin- and vasopressin-stimulated formation of inositol phosphates were inhibited by only 27 and 23% respectively. These results argue against a major role for a pertussis toxin-sensitive G protein in coupling any of these mitogen receptors to activation of a phosphoinositidase C. Furthermore, the results suggest that the widespread inhibitory effects of pertussis toxin on mitogen-stimulated DNA synthesis may be unrelated to the toxin's limited actions on phosphoinositide hydrolysis.

scholarly journals Guanine nucleotide-dependent pertussis-toxin-insensitive stimulation of inositol phosphate formation by carbachol in a membrane preparation from human astrocytoma cells

1986 ◽  
Vol 239 (1) ◽  
pp. 141-146 ◽  
Author(s):  
J R Hepler ◽  
T K Harden
Keyword(s):  
Pertussis Toxin ◽  
Inositol Phosphate ◽  
Regulatory Protein ◽  
Dependent Manner ◽  
Guanine Nucleotide ◽  
Astrocytoma Cells ◽  
Human Astrocytoma ◽  
Guanine Nucleotide Regulatory Protein ◽  
Human Astrocytoma Cells ◽  
Stimulation Of

The efficacy of muscarinic-receptor agonists for stimulation of inositol phosphate formation and Ca2+ mobilization in intact 1321N1 human astrocytoma cells is correlated with their capacity for formation of a GTP-sensitive high-affinity binding complex in membranes from these cells [Evans, Hepler, Masters, Brown & Harden (1985) Biochem. J. 232, 751-757]. These observations prompted the proposal that a guanine nucleotide regulatory protein serves to couple muscarinic receptors to the phospholipase C involved in phosphoinositide hydrolysis in 1321N1 cells. Inositol phosphate (InsP) formation was measured in a cell-free preparation from 1321N1 cells to provide direct support for this idea. The formation of InsP3, InsP2 and InsP1 was increased in a concentration-dependent manner (K0.5 approximately 5 microM) by guanosine 5′-[gamma-thio]triphosphate (GTP[S]) in washed membranes prepared from myo-[3H]inositol-prelabelled 1321N1 cells. Both GTP[S] and guanosine 5′-[beta gamma-imido]triphosphate (p[NH]ppG) stimulated InsP formation by 2-3-fold over control; GTP, GDP and GMP were much less efficacious. Millimolar concentrations of NaF also stimulated the formation of inositol phosphates in membrane preparations from 1321N1 cells. In the presence of 10 microM-GTP[S], the muscarinic cholinergic-receptor agonist carbachol stimulated (K0.5 approximately 10 microM) the formation of InsP above that achieved with GTP[S] alone. The effect of carbachol was completely blocked by atropine. The order of potency of nucleotides for stimulation of InsP formation in the presence of 500 microM-carbachol was GTP[S] greater than p[NH]ppG greater than GTP = GDP. Pertussis toxin, at concentrations that fully ADP-ribosylate and functionally inactivate Gi (the inhibitory guanine nucleotide regulatory protein), had no effect on InsP formation in the presence of GTP[S] or GTP[S] plus carbachol. These data are consistent with the idea that a guanine nucleotide regulatory protein that is not Gi is involved in receptor-mediated stimulation of InsP formation in 1321N1 human astrocytoma cells.

scholarly journals Characterization of Prejunctional Muscarinic Receptors: Effects on the Release of VIP and Functional Responses and Receptor Expression in the Ovine Submandibular Gland

2009 ◽  
Vol 2009 ◽  
pp. 1-6
Author(s):  
Anders T. Ryberg ◽  
Ondrej Soukup ◽  
Gunnar Tobin
Keyword(s):  
Electrical Stimulation ◽  
Submandibular Gland ◽  
Muscarinic Receptor ◽  
Muscarinic Receptors ◽  
Receptor Expression ◽  
Receptor Subtypes ◽  
Chorda Tympani Nerve ◽  
Functional Responses ◽  
Stimulation Of

In the in vivo experiments on anaesthetized sheep, it was presently examined whether muscarinic receptor antagonists with diverse selectivity affect the release of VIP in response to electrical stimulation of the parasympathetic chorda tympanic nerve differently, and if the changes in the release could be associated to altered secretory and vasodilator responses. The location of the muscarinic receptor subtypes was examined also. In the experiments, blood was collected out of the submandibular venous drainage before and during electrical stimulation of chorda tympani nerve in the absence and presence either of pirenzepine or methoctramine. While metchoctramine increased the output of protein, pirenzepine inhibited flow of saliva and increased protein output, vasodilatation, and VIP output. In morphological examinations, the inhibitory muscarinic M4 receptor occurred interacinarily in the gland. It is concluded that prejunctional muscarinic receptors, most likely of the M4 subtype, exert inhibitory modulation of the parasympathetic release of VIP in the ovine submandibular gland.

Functionally distinct muscarinic receptors on gastric somatostatin cells

1990 ◽  
Vol 258 (6) ◽  
pp. G982-G987 ◽  
Author(s):  
M. L. Schubert ◽  
J. Hightower
Keyword(s):  
Muscarinic Receptors ◽  
Pertussis Toxin ◽  
Inhibitory Effect ◽  
Tetraacetic Acid ◽  
Muscarinic Agonists ◽  
Rat Stomach ◽  
Dominant Effect ◽  
Somatostatin Secretion ◽  
Somatostatin Cells ◽  
Stimulation Of

The present study was designed to examine the mode of action of muscarinic agonists on somatostatin secretion in intact gastric tissues, i.e., mucosal segments from the fundus and antrum of rat and the isolated luminally perfused mouse stomach. Methacholine caused similar decreases in somatostatin secretion in segments from the fundus (35 +/- 3%; P less than 0.001) and antrum (35 +/- 2%; P less than 0.001) of rat stomach, and in whole mouse stomach (43 +/- 3%; P less than 0.001). The decrease was the net effect of a dominant inhibition and a lesser stimulation of somatostatin secretion. Pretreatment with the permeant derivative of the acetomethoxy ester form of the calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA/AM, 15 microM) caused a further decrease in methacholine-induced somatostatin secretion, implying that a stimulatory component existed that was mediated by intracellular calcium. Pretreatment with pertussis toxin (125 ng/ml) for 60 min converted the decrease in somatostatin secretion to an increase above basal levels. The increase induced by pretreatment with pertussis toxin was abolished by additional pretreatment with BAPTA/AM. Procaine (5 mM), which blocks release of calcium from intracellular stores, produced an effect on somatostatin secretion similar to that of BAPTA/AM. The results indicate that 1) methacholine exerts dual inhibitory and stimulatory effects on somatostatin cells of rat and mouse stomach, 2) the dominant effect is inhibitory and sensitive to pertussis toxin, and 3) a concurrent stimulatory effect, mediated by calcium, is unmasked after blockade of the inhibitory effect with pertussis toxin.

scholarly journals Accumulation and metabolism of Ins(1,4,5)P3 and Ins(1,3,4,5)P4 in muscarinic-receptor-stimulated SH-SY5Y neuroblastoma cells

1991 ◽  
Vol 273 (3) ◽  
pp. 791-794 ◽  
Author(s):  
D G Lambert ◽  
R A J Challiss ◽  
S R Nahorski
Keyword(s):  
Muscarinic Receptor ◽  
Muscarinic Receptors ◽  
Neuroblastoma Cells ◽  
Stimulation Of ◽  
Related Increase

Stimulation of M3 muscarinic receptors expressed by SH-SY5Y cells induced a dose- and time-related increase in the mass of Ins(1,4,5)P3 (basal 38.3 +/- 5.8 pmol/mg of protein) and Ins(1,3,4,5)P4 (basal 6.1 +/- 1.2 pmol/mg of protein). Comparison of radioreceptor mass assays with [3H]inositol labelling showed higher-fold stimulations with the former protocol. The later accumulation of Ins(1,4,5)P3 and Ins(1,3,4,5)P4 mass was dependent upon extracellular Ca2+.

Rapacuronium Augments Acetylcholine-induced Bronchoconstriction via  Positive Allosteric Interactions at the M3 Muscarinic Receptor

2005 ◽  
Vol 103 (6) ◽  
pp. 1195-1203 ◽  
Author(s):  
Edmund H. Jooste ◽  
Amit Sharma ◽  
Yi Zhang ◽  
Charles W. Emala
Keyword(s):  
Clinical Practice ◽  
Muscarinic Receptor ◽  
Muscarinic Receptors ◽  
Inositol Phosphate ◽  
Neuromuscular Blocking ◽  
Muscle Relaxants ◽  
Neuromuscular Blocking Agents ◽  
Clinical Use ◽  
Allosteric Interactions ◽  
M3 Muscarinic Receptor

Background Neuromuscular blocking agents' detrimental airway effects may occur as a result of interactions with muscarinic receptors, allergic reactions, or histamine release. Rapacuronium, a nondepolarizing muscle relaxant, was withdrawn from clinical use because of its association with fatal bronchospasm. Despite its withdrawal from clinical use, it is imperative that the mechanism by which bronchospasm occurred is understood so that new muscle relaxants introduced to clinical practice do not share these same detrimental airway effects. Methods Airway smooth muscle force was measured in guinea pig tracheal rings in organ baths exposed to muscle relaxants with or without subthreshold concentrations of acetylcholine. Antagonism of muscarinic, histamine, neurokinin, leukotriene receptors, or blockade of L-type calcium channels or depletion of nonadrenergic, noncholinergic neurotransmitters was performed. Muscle relaxants' potentiation of acetylcholine-stimulated inositol phosphate synthesis and allosteric interactions on the kinetics of atropine-induced [3H]N-methylscopolamine dissociation were measured in cells expressing recombinant human M3 muscarinic receptors. Results Rapacuronium, within clinically achieved concentrations, contracted tracheal rings in the presence but not in the absence of subthreshold concentrations of acetylcholine. This effect was prevented or reversed only by atropine. The allosteric action of rapacuronium was demonstrated by the slowing of atropine-induced dissociation of [3H]N-methylscopolamine, and positive cooperativity was demonstrated by potentiation of acetylcholine-induced inositol phosphate synthesis. Conclusion Many muscle relaxants have allosteric properties at muscarinic receptors; however, positive cooperativity at the M3 muscarinic receptor within clinically relevant concentrations is unique to rapacuronium. These findings establish novel parameters that should be considered in the evaluation of airway safety of any newly synthesized neuromuscular blocking agents considered for clinical practice.

Field Stimulation of Isolated Chick Heart Cells.

1999 ◽  
Vol 10 (1) ◽  
pp. 92-107 ◽  
Author(s):  
BRADLEY A. STONE ◽  
MELVYN LIEBERMAN ◽  
WANDA KRASSOWSKA
Keyword(s):  
Heart Cells ◽  
Field Stimulation ◽  
Chick Heart ◽  
Stimulation Of
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