Sensitivity of norepinephrine-evoked vasoconstriction to pertussis toxin in the old rat

1998 ◽  
Vol 274 (6) ◽  
pp. R1604-R1612 ◽  
Author(s):  
Alain Robert ◽  
Nguyen N. P. Tran ◽  
Philippe Giummelly ◽  
Jeffrey Atkinson ◽  
Christine Capdeville-Atkinson
Keyword(s):  
Intracellular Calcium ◽  
G Protein ◽  
Pertussis Toxin ◽  
Receptor Activation ◽  
Plasma Norepinephrine ◽  
Adrenoceptor Antagonist ◽  
Vasoconstrictor Response ◽  
High K ◽  
Male Wistar Rats

In male Wistar rats, the in vitro vasoconstrictor response of the perfused tail artery elicited by norepinephrine or serotonin decreased with age (24 mo old vs. 3 mo old), whereas the fluorescent signal (fura 2) produced by intracellular calcium ([Formula: see text]) mobilization increased. Both vasoconstriction and the increase in intracellular calcium concentration elicited by a high-K+, depolarizing solution were unaffected by aging. Pertussis toxin, a G protein inhibitor, had no effect on vasoconstriction induced by high K+ but diminished vasoconstrictor responses to norepinephrine in 3- and 12-mo-old animals but not in 24-mo-old animals. Pertussis toxin had no effect on[Formula: see text] mobilization. The sensitivity of receptor activation to pertussis toxin in tail arteries from 24-mo-old animals was restored by pretreatment with the α-adrenoceptor antagonist nicergoline. Nicergoline had no effect on vasoconstriction induced by high K+. Plasma norepinephrine concentration rose with age; nicergoline had no effect on this rise. We suggest that aging leads to a decrease in the intracellular G protein-modulated amplification of vasoconstriction produced by receptor activation and that this could be linked to the hyperadrenergic state. Ca2+sensitivity can be restored by chronic treatment with an α-adrenoceptor antagonist.

Effects of 5-day hypoxia on cardiac adrenergic neurotransmission in rats

1998 ◽  
Vol 85 (3) ◽  
pp. 890-897 ◽  
Author(s):  
Karine Mardon ◽  
Pascal Merlet ◽  
André Syrota ◽  
Bernard Mazière
Keyword(s):  
G Protein ◽  
Plasma Norepinephrine ◽  
Protein Subunit ◽  
Α Subunit ◽  
In Vitro Binding ◽  
Vitro Binding ◽  
Male Wistar Rats ◽  
Inhibitory G Protein ◽  
Protein Density

Chronic hypoxia induces an overall sympathetic hyperactivation associated with a myocardial β-receptor desensitization. The mechanisms involved in this desensitization were evaluated in 32 male Wistar rats kept in a hypobaric pressure chamber ([Formula: see text] = 40 Torr, atmospheric pressure = 450 Torr) for 5 days. In hypoxic compared with normoxic conditions, plasma norepinephrine (NE) levels were higher (2.1 ± 0.7 vs. 0.6 ± 0.2 ng/ml) with no difference in the plasma epinephrine levels (2.2 ± 0.7 vs. 1.8 ± 0.3 ng/ml). In hypoxia neuronal NE uptake measured by [3H]NE was decreased by 32% in the right ventricle (RV) and by 35% in the left ventricle (LV), and [3H]mazindol in vitro binding showed a decrease in uptake-1 carrier protein density by 38% in the RV and by 41% in the LV. In vitro binding assays with [3H]CGP-12177 indicate β-adrenoceptor density reduced by 40% in the RV and by 32% in the LV, and this was due to reduced β1-subtype fraction (competition binding experiments with practolol). Hypoxia reduced the production of cAMP induced by isoproterenol (36% decrease in the RV and 41% decrease in the LV), 5′-guanylylimododiphosphate (40% decrease in the RV and 42% decrease in the LV), and forskolin (39% decrease in the RV and 41% decrease in the LV) but did not alter the effect of MnCl2 and NaF. Quantitation of inhibitory G-protein α-subunit by immunochemical analysis showed a 46% increase in the cardiac-specific isoform[Formula: see text] in hypoxic hearts. The present data demonstrate that in rats 5-day hypoxia leads to changes in pre- and postsynaptic myocardial adrenergic function. The myocardial desensitization associated with both a reduction in externalized β1-adrenoceptor and an increase in inhibitory G-protein subunit may be caused by increased synaptic NE levels due to impaired uptake-1 system.

scholarly journals A Blunted Sympathetic Function and an Enhanced Nitrergic Activity Contribute to Reduce Mesenteric Resistance in Hyperthyroidism

2021 ◽  
Vol 22 (2) ◽  
pp. 570
Author(s):  
Laia Cros-Brunsó ◽  
Laura Camacho-Rodríguez ◽  
Ángel Martínez-González ◽  
Pablo Llévenes ◽  
Mercedes Salaices ◽  
...  
Keyword(s):  
Citrate Synthase ◽  
Purinergic Receptor ◽  
Body Weight Gain ◽  
Electrical Field Stimulation ◽  
Alpha Adrenoceptor ◽  
Adrenoceptor Antagonist ◽  
Sympathetic Function ◽  
Vasoconstrictor Response ◽  
Relevant Role ◽  
Male Wistar Rats

We aimed to determine whether an experimental model of hyperthyroidism could alter the function of sympathetic and nitrergic components of mesenteric innervation. For this purpose, male Wistar rats were divided into (1) control rats (CT) and (2) rats infused with L-Thyroxine (HT). Body weight gain and adipose tissue accumulation were lower in HT rats, while systolic blood pressure and citrate synthase activity in the soleus muscle were increased by HT. In segments from the superior mesenteric artery, the application of an electrical field stimulation (EFS) induced a vasoconstrictor response, which was lower in arteries from HT animals. The alpha-adrenoceptor antagonist phentolamine diminished EFS-induced vasoconstriction to a lower extent in HT arteries, while the purinergic receptor antagonist suramin reduced contractile response to EFS only in segments from CT. In line with this, noradrenaline release, tyrosine hydroxylase expression and activation and dopamine β hydroxylase expression were diminished in HT. The unspecific nitric oxide synthase (NOS) inhibitor L-NAME increased EFS-induced vasoconstriction more markedly in segments from HT rats. NO release was enhanced in HT, probably due to an enhancement in neuronal NOS activity, in which a hyperactivation of both PKC and PI3K-AKT signaling pathways might play a relevant role. In conclusion, perivascular mesenteric innervation might contribute to reduce the vascular resistance observed in hyperthyroidism.

ATP induces contraction of cultured brain capillary pericytes, via activation of P2Y type purinergic receptors

2020 ◽  
Author(s):  
Sofie Hørlyck ◽  
Changsi Cai ◽  
Hans C Helms ◽  
Martin Lauritzen ◽  
Birger Brodin
Keyword(s):  
Intracellular Calcium ◽  
Purinergic Receptors ◽  
Calcium Release ◽  
Purinergic Receptor ◽  
Cytosolic Calcium ◽  
Receptor Activation ◽  
Confocal Imaging ◽  
Inositol Triphosphate ◽  
Brain Capillary

Brain capillary pericytes have been suggested to play a role in the regulation of cerebral blood-flow under physiological and pathophysiological conditions. ATP has been shown to cause constriction of capillaries under ischemic conditions and suggested to be involved in the "no-reflow" phenomenon. In order to investigate the effects of extracellular ATP on pericyte cell contraction, we studied purinergic receptor activation of cultured bovine brain capillary pericytes. We measured [Ca2+]i-responses to purinergic agonists with the fluorescent indicators fura-2 and Cal-520 and estimated contraction of pericytes as relative change in cell area, using real-time confocal imaging. Addition of ATP caused an increase in cytosolic calcium and contraction of the brain capillary pericytes, both reversible and inhibited by a purinergic receptor antagonist PPADS. Furthermore, we demonstrated that ATP-induced contraction could be eliminated by intracellular calcium-chelation with BAPTA, indicating that the contraction was mediated via purinergic P2 -type receptor-mediated [Ca2+]i-signaling. ATP stimulation induced inositol triphosphate signaling, consistent with the notion of P2Y receptor activation. Receptor profiling studies demonstrated presence of P2Y1 and P2Y2 receptors, using ATP, UTP, ADP and the subtype specific agonists MRS2365 (P2Y1) and 2-thio-UTP (P2Y2)). Addition of specific P2X agonists only caused a [Ca2+]i increase at high concentrations, attributed to activation of inositol triphosphate signaling. Our results suggest that contraction of brain capillary pericytes in vitro by activation of P2Y type purinergic receptors is caused by intracellular calcium release. This adds more mechanistic understanding to the role of pericytes in vessel constriction, and points towards P2Y receptors as potential therapeutic targets.

scholarly journals Specific oxylipins enhance vertebrate hematopoiesis via the receptor GPR132

2018 ◽  
Vol 115 (37) ◽  
pp. 9252-9257 ◽  
Author(s):  
Jamie L. Lahvic ◽  
Michelle Ammerman ◽  
Pulin Li ◽  
Megan C. Blair ◽  
Emma R. Stillman ◽  
...  
Keyword(s):  
Fatty Acids ◽  
G Protein ◽  
Cell Lines ◽  
Saturated Fatty Acids ◽  
Receptor Activation ◽  
G Protein Coupled Receptors ◽  
Hydroxy Fatty Acids ◽  
High Affinity ◽  
G Protein Coupled

Epoxyeicosatrienoic acids (EETs) are lipid-derived signaling molecules with cardioprotective and vasodilatory actions. We recently showed that 11,12-EET enhances hematopoietic induction and engraftment in mice and zebrafish. EETs are known to signal via G protein-coupled receptors, with evidence supporting the existence of a specific high-affinity receptor. Identification of a hematopoietic-specific EET receptor would enable genetic interrogation of EET signaling pathways, and perhaps clinical use of this molecule. We developed a bioinformatic approach to identify an EET receptor based on the expression of G protein-coupled receptors in cell lines with differential responses to EETs. We found 10 candidate EET receptors that are expressed in three EET-responsive cell lines, but not expressed in an EET-unresponsive line. Of these, only recombinant GPR132 showed EET-responsiveness in vitro, using a luminescence-based β-arrestin recruitment assay. Knockdown of zebrafish gpr132b prevented EET-induced hematopoiesis, and marrow from GPR132 knockout mice showed decreased long-term engraftment capability. In contrast to high-affinity EET receptors, GPR132 is reported to respond to additional hydroxy-fatty acids in vitro, and we found that these same hydroxy-fatty acids enhance hematopoiesis in the zebrafish. We conducted structure–activity relationship analyses using both cell culture and zebrafish assays on diverse medium-chain fatty acids. Certain oxygenated, unsaturated free fatty acids showed high activation of GPR132, whereas unoxygenated or saturated fatty acids had lower activity. Absence of the carbon-1 position carboxylic acid prevented activity, suggesting that this moiety is required for receptor activation. GPR132 responds to a select panel of oxygenated polyunsaturated fatty acids to enhance both embryonic and adult hematopoiesis.

Vanadate causes synthesis of endothelium-derived NO via pertussis toxin-sensitive G protein in pigs

1996 ◽  
Vol 271 (1) ◽  
pp. H296-H302 ◽  
Author(s):  
R. Nakaike ◽  
H. Shimokawa ◽  
M. K. Owada ◽  
O. Tokunaga ◽  
H. Yasutake ◽  
...  
Keyword(s):  
G Protein ◽  
Pertussis Toxin ◽  
Tyrosine Kinases ◽  
Kinase Inhibitors ◽  
Active Form ◽  
Protein Tyrosine Phosphatases ◽  
Jugular Veins ◽  
Aortic Endothelial Cells ◽  
Endothelial Nitric Oxide

The effects of sodium orthovanadate, an inhibitor of protein tyrosine phosphatases, on the endothelial nitric oxide (NO) pathway were studied in vitro. Vanadate caused endothelium-dependent relaxations in isolated porcine coronary arteries, which were abolished by N omega-nitro-L-arginine methyl ester. The relaxations were also abolished by pertussis toxin, an inhibitor of certain G proteins. Tyrosine kinase inhibitors, genistein and alpha-cyano-3-ethoxy-4-hydroxy-5-phenyl-methylcinnamamide (ST-638), significantly attenuated the vanadate-induced relaxations. Vanadate also caused pertussis toxin-sensitive, endothelium-dependent relaxations in isolated porcine renal and femoral arteries and jugular veins. Immunoblots, using an antibody to phosphotyrosines and to c-Src in native porcine aortic endothelial cells, respectively, showed that vanadate induced an elevation of phosphotyrosine proteins and a decrease in the amount of the active form of c-Src family kinases; both changes were markedly suppressed by cotreatment with ST-638. These results indicate that in porcine blood vessels, vanadate causes a synthesis of endothelium-derived NO for which endothelial tyrosine kinases and pertussis toxin-sensitive G protein are considered to be closely involved.

Somatostatin inhibits the norepinephrine-activated calcium channels in rMTC 6–23 cells: possible involvement of a pertussis toxin-sensitive G-protein

1992 ◽  
Vol 127 (4) ◽  
pp. 378-384 ◽  
Author(s):  
A Zink ◽  
F Raue
Keyword(s):  
Calcium Channels ◽  
Intracellular Calcium ◽  
G Protein ◽  
Pertussis Toxin ◽  
Single Cells ◽  
Medullary Carcinoma ◽  
Inhibitory Effect ◽  
Somatostatin Analogue ◽  
Intracellular Camp ◽  
Long Acting

The mechanisms by which somatostatin inhibits hormone release are complex and involve, among other things, reduction of both intracellular cAMP and intracellular calcium. We studied the influence of the long-acting somatostatin analogue octreotide on norepinephrine (NE)-induced changes in intracellular calcium ([Ca2+]i) in fura-2 loaded single cells of a rat medullary carcinoma cell line, rMTC 6–23. Increases in the extracellular calcium concentration ([Ca2+]e) induced a sudden rise in [Ca2+]i which could be blocked by EGTA or the calcium channel blocker verapamil. NE evoked a similar increase in [Ca2+]i, which also could be blocked by the addition of EGTA or verapamil. Octreotide prevented or reversed the NE-induced increase in [Ca2+ ]i. Pretreatment of the cells with pertussis toxin abolished the inhibitory effect of octreotide. Thus we conclude that the NE-induced rise in [Ca2+]i is due to an influx of [Ca2+ ]e, most probably through voltage-dependent calcium channels. Octreotide inhibits the NE-stimulated rise in [Ca2+ ]i by a pertussis toxin-sensitive G-protein, most probably through a direct effect on NE-activated calcium channels.

Pertussis toxin inhibition of T-cell hybridoma invasion is reversed by manganese-induced activation of LFA-1

1994 ◽  
Vol 107 (3) ◽  
pp. 551-559
Author(s):  
G. La Riviere ◽  
J.W. Klein Gebbinck ◽  
M.H. Driessens ◽  
E. Roos
Keyword(s):  
T Cell ◽  
G Protein ◽  
Pertussis Toxin ◽  
Hybridoma Cells ◽  
Autocrine Motility Factor ◽  
Motility Factor ◽  
Serum Component ◽  
Extracellular Factor

Pertussis toxin (PT) inhibits invasiveness of T-cell hybridomas in vitro and metastasis formation in vivo. We present evidence for the hypothesis that PT interferes with functional activation of LFA-1. Invasion by TAM2D2 T-cell hybridoma cells of fibroblast monolayers was completely blocked by PT pretreatment, but the cells regained invasiveness in the presence of Mn2+, which activates LFA-1. This invasion was blocked by anti-LFA-1 mAb, and Mn2+ did not stimulate invasiveness of LFA-1-deficient TAM2D2 mutants. TAM2D2 cells did not adhere to surfaces coated with the LFA-1 counterstructure ICAM-1, but Mn2+ induced adhesion. Hence, LFA-1 on TAM2D2 cells requires activation before it can participate in the invasion process. The hypothesis is further supported by the slightly different results obtained with the TAM8C4 T-cell hybridoma. PT inhibited invasion strongly but not completely. This reduced invasion was increased by Mn2+. TAM8C4 cells did adhere to ICAM-1, but Mn2+ enhanced adhesion. Thus, part of LFA-1 on TAM8C4 cells is constitutively active, allowing for some PT-insensitive invasion, but further activation is required for optimal adhesion and invasion. PT blocks G-protein-mediated signals, suggesting that an extracellular factor is involved. This is not a serum component or an autocrine motility factor, since the PT effect was serum-independent, and PT did not inhibit motility. Therefore, it is probably produced by the fibroblasts, and either secreted or associated with the cell surface. These results are in line with the hypothesis that a fibroblast constituent activates LFA-1 via a PT-sensitive G-protein and thus stimulates invasion of T-cell hybridomas into the fibroblast monolayer.

scholarly journals Distinct Roles for Gαi2 and Gβγ in Signaling to DNA Synthesis and Gαi3 in Cellular Transformation by Dopamine D2S Receptor Activation in BALB/c 3T3 Cells

2000 ◽  
Vol 20 (5) ◽  
pp. 1497-1506 ◽  
Author(s):  
Mohammad H. Ghahremani ◽  
Christine Forget ◽  
Paul R. Albert
Keyword(s):  
Cell Proliferation ◽  
Dna Synthesis ◽  
G Protein ◽  
Pertussis Toxin ◽  
Cellular Response ◽  
Short Form ◽  
Ectopic Expression ◽  
Cellular Transformation ◽  
Receptor Activation ◽  
Neuroendocrine Cells

ABSTRACT Control of cell proliferation depends on intracellular mediators that determine the cellular response to external cues. In neuroendocrine cells, the dopamine D2 receptor short form (D2S receptor) inhibits cell proliferation, whereas in mesenchymal cells the same receptor enhances cell proliferation. Nontransformed BALB/c 3T3 fibroblast cells were stably transfected with the D2S receptor cDNA to study the G proteins that direct D2S signaling to stimulate cell proliferation. Pertussis toxin inactivates Gi and Go proteins and blocks signaling of the D2S receptor in these cells. D2S receptor signaling was reconstituted by individually transfecting pertussis toxin-resistant Gαi/o subunit mutants and measuring D2-induced responses in pertussis toxin-treated cells. This approach identified Gαi2 and Gαi3 as mediators of the D2S receptor-mediated inhibition of forskolin-stimulated adenylyl cyclase activity; Gαi2-mediated D2S-induced stimulation of p42 and p44 mitogen-activated kinase (MAPK) and DNA synthesis, whereas Gαi3 was required for formation of transformed foci. Transfection of toxin-resistant Gαi1 cDNA induced abnormal cell growth independent of D2S receptor activation, while Gαo inhibited dopamine-induced transformation. The role of Gβγ subunits was assessed by ectopic expression of the carboxyl-terminal domain of G protein receptor kinase to selectively antagonize Gβγ activity. Mobilization of Gβγ subunits was required for D2S-induced calcium mobilization, MAPK activation, and DNA synthesis. These findings reveal a remarkable and distinct G protein specificity for D2S receptor-mediated signaling to initiate DNA synthesis (Gαi2 and Gβγ) and oncogenic transformation (Gαi3), and they indicate that acute activation of MAPK correlates with enhanced DNA synthesis but not with transformation.

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