scholarly journals The role of cAMP-mediated intracellular signaling in regulating Na+ uptake in zebrafish larvae

2014 ◽  
Vol 306 (1) ◽  
pp. R51-R60 ◽  
Author(s):  
Yusuke Kumai ◽  
Raymond W. M. Kwong ◽  
Steve F. Perry
Keyword(s):  
Protein Trafficking ◽  
Intracellular Signaling ◽  
Whole Body ◽  
Larval Zebrafish ◽  
Zebrafish Larvae ◽  
Β Adrenergic Receptors ◽  
Camp Levels ◽  
Na Uptake ◽  
Stimulation Of

In the current study, the role of cAMP in stimulating Na+ uptake in larval zebrafish was investigated. Treating larvae at 4 days postfertilization (dpf) with 10 μM forskolin or 1 μM 8-bromo cAMP significantly increased Na+ uptake by three-fold and twofold, respectively. The cAMP-dependent stimulation of Na+ uptake was probably unrelated to protein trafficking via microtubules because pretreatment with 200 μM colchicine or 30 μM nocodazole did not attenuate the magnitude of the response. Na+ uptake was stimulated markedly following acute (2 h) exposure to acidic water. The acid-induced increase in Na+ uptake was accompanied by a twofold elevation in whole body cAMP levels and attenuated by inhibiting PKA with 10 μM H-89. Knockdown of Na+-H+ exchanger 3b (NHE3b) attenuated, but did not abolish, the stimulation of Na+ uptake during forskolin treatment. In glial cell missing 2 morphants, in which the role of NHE3b in Na+ uptake is diminished and the Na+-Cl− cotransporter (NCC) becomes the predominant route of Na+ entry, forskolin treatment continued to increase Na+ uptake. These data suggest that at least NHE3b and NCC are targeted by cAMP in zebrafish larvae. Staining of larvae with fluorescent forskolin and propranolol revealed the presence of transmembrane adenylyl cyclase within multiple subtypes of ionocytes expressing β-adrenergic receptors. Taken together, results of the present study demonstrate that cAMP-mediated intracellular signaling may regulate multiple Na+ transporters and plays an important role in regulating Na+ uptake in zebrafish larvae during acute exposure to an acidic environment.

Loss of M2 muscarinic receptor function inhibits development of hypoxic bradycardia and alters cardiac β-adrenergic sensitivity in larval zebrafish (Danio rerio)

2009 ◽  
Vol 297 (2) ◽  
pp. R412-R420 ◽  
Author(s):  
Shelby L. Steele ◽  
Kwok Hong Andy Lo ◽  
Vincent Wai Tsun Li ◽  
Shuk Han Cheng ◽  
Marc Ekker ◽  
...  
Keyword(s):  
Heart Rate ◽  
Muscarinic Receptor ◽  
Danio Rerio ◽  
Antisense Oligonucleotides ◽  
Adrenergic Receptor ◽  
Larval Zebrafish ◽  
M2 Muscarinic Receptor ◽  
Β Adrenergic Receptors ◽  
Relative Mrna Expression

Fish exposed to hypoxia develop decreased heart rate, or bradycardia, the physiological significance of which remains unknown. The general muscarinic receptor antagonist atropine abolishes the development of this hypoxic bradycardia, suggesting the involvement of muscarinic receptors. In this study, we tested the hypothesis that the hypoxic bradycardia is mediated specifically by stimulation of the M2 muscarinic receptor, the most abundant subtype in the vertebrate heart. Zebrafish ( Danio rerio) were reared at two levels of hypoxia (30 and 40 Torr Po2) from the point of fertilization. In hypoxic fish, the heart rate was significantly lower than in normoxic controls from 2 to 10 days postfertilization (dpf). At the more severe level of hypoxia (30 Torr Po2), there were significant increases in the relative mRNA expression of M 2 and the cardiac type β-adrenergic receptors ( β1AR, β2aAR, and β2bAR) at 4 dpf. The hypoxic bradycardia was abolished (at 40 Torr Po2) or significantly attenuated (at 30 Torr Po2) in larvae experiencing M2 receptor knockdown (using morpholino antisense oligonucleotides). Sham-injected larvae exhibited typical hypoxic bradycardia in both hypoxic regimens. The expression of β1AR, β2aAR, β2bAR, and M 2 mRNA was altered at various stages between 1 and 4 dpf in larvae experiencing M2 receptor knockdown. Interestingly, M2 receptor knockdown revealed a cardioinhibitory role for the β2-adrenergic receptor. This is the first study to demonstrate a specific role of the M2 muscarinic receptor in the initiation of hypoxic bradycardia in fish.

The Regulatory Role Of Camp In Induction Of Human Platelet Receptor For Fibrinogen

1981 ◽  
Author(s):  
S E Graber ◽  
J Hawiger
Keyword(s):  
Human Platelets ◽  
Membrane Receptor ◽  
Fibrinogen Receptor ◽  
Fibrinogen Binding ◽  
Platelet Receptor ◽  
The Relationship ◽  
Camp Levels ◽  
Dose Dependent ◽  
Stimulation Of

Membrane receptor for fibrinogen plays an essential role in adhesion and aggregation of human platelets by allowing fibrinogen to bridge two or more platelets together. Whereas in normal, unstimulated platelets fibrinogen receptor is not available, it becomes mobilized upon stimulation of platelets with thrombin, ADP, and other stimuli. The mechanism(s) regulating availability of membrane receptor for fibrinogen remains unknown. Following our recent demonstration that prostacyclin (PGI2) prevents mobilization of fibrinogen receptor by thrombin and ADP (Nature 1980, 283,195), we investigated the relationship between cAMP levels and fibrinogen receptor availability. Platelets separated from plasma proteins were briefly exposed to a low thrombin concentration (0.05 U/ml) followed by hirudin to inactivate free thrombin. Binding of 125I-fi- brinogen and cAMP levels were determined in parallel samples. A dose-dependent rise in platelet cAMP levels from 3.3 pM to 10.3 pM/108 platelets in response to PGI2 (3×10-9M - 3×108M) was accompanied by a corresponding inhibition of 125I-fibrinogen binding. The degree of the cAMP increment correlated with binding inhibition (r=0.96). The inhibition of 125I-fibrinogen binding by PGI2 was sustained up to 120 min and was paralleled by a persistent rise in cAMP level. Stimulation of platelet cAMP synthesis “from within” by a ribosylation of the nucleotide regulatory component with subunit A1 of cholera toxin also increased cAMP levels and inhibited fibrinogen receptor mobilization.These results provide evidence that “up and down” regulation of fibrinogen receptor in platelets is linked to changes in cAMP levels induced by different types of adenyl cyclase antagonists and agonists.

Ventromedial hypothalamic lesions impair glucoregulation in response to endotoxin

1997 ◽  
Vol 272 (5) ◽  
pp. R1525-R1531 ◽  
Author(s):  
J. P. Lynch ◽  
M. M. Wojnar ◽  
C. H. Lang
Keyword(s):  
Metabolic Response ◽  
Plasma Concentrations ◽  
Ventromedial Hypothalamus ◽  
Whole Body ◽  
Tnf Alpha ◽  
Hormone Release ◽  
Glucose Flux ◽  
Bilateral Lesions ◽  
Stimulation Of

The purpose of the present study was to determine the role of the ventromedial hypothalamus (VMH) in regulating counter-regulatory hormone release and the increase in glucose flux that is observed after injection of endotoxin [lipopolysaccharide (LPS)]. Bilateral lesions of the VMH were produced electrolytically 2 wk before the experiment; sham-operated rats served as controls. [3-3H]glucose was infused to assess whole body glucose flux before and for 4 h after intravenous injection of Escherichia coli LPS. In control rats, LPS increased the plasma concentrations of glucose and lactate and the rates of glucose appearance and disappearance. In these animals, LPS also produced sustained elevations in corticosterone, glucagon, and catecholamines. In contrast, the glucose metabolic response to LPS was attenuated by > 50% in VMH-lesioned rats. These changes were associated with a blunted increase in the plasma concentration of glucagon, epinephrine, and norepinephrine in VMH-lesioned rats compared with control animals. There was no difference in the plasma concentrations of corticosterone or TNF-alpha between the two groups after LPS or the responsiveness of sham- and VMH-lesioned rats to an infusion of either glucagon or epinephrine. These data indicate that the VMH plays a central role in regulating the secretion of glucagon and catecholamines and the stimulation of glucose flux after LPS.

Stimulation of prolactin release by dopamine withdrawal: role of calcium influx

1994 ◽  
Vol 267 (5) ◽  
pp. E789-E794 ◽  
Author(s):  
K. A. Gregerson ◽  
R. Chuknyiska ◽  
N. Golesorkhi
Keyword(s):  
Channel Blocker ◽  
Inositol Phosphates ◽  
Calcium Influx ◽  
Pituitary Cells ◽  
Induced Changes ◽  
Camp Levels ◽  
Ca2 Channels ◽  
Stimulation Of ◽  
Spiking Activity

Withdrawal of dopamine (DA), a neurotransmitter that inhibits prolactin (PRL) release from the anterior pituitary, stimulates PRL release with transient (30- to 45-min) secretory rates that exceed those observed before application of DA ("PRL rebound"). Using patch-clamp methods on identified rat lactotropes, we have demonstrated that a period of increased Ca(2+)-spiking activity follows recovery from the DA-induced hyperpolarization. The present experiments used dissociated pituitary cells to identify the relative roles of adenosine 3',5'-cyclic monophosphate (cAMP), inositol phosphates, and the enhanced influx of Ca2+ in the rebound secretion of PRL. Rebound secretion of PRL after DA withdrawal was completely blocked by the Ca2+ channel blocker verapamil (20 microM), which also inhibited spontaneous Ca(2+)-spiking activity. DA-induced changes in cAMP levels could be completely dissociated from the PRL rebound. Production of inositol phosphates rose after DA withdrawal but was secondary to the influx of Ca2+. These data demonstrate that influx of extracellular Ca2+ through verapamil-sensitive channels is a critical step in inducing PRL release after DA withdrawal. This finding supports our theory that DA-induced hyperpolarization recruits previously inactivated Ca2+ channels and upon DA washout the enhanced influx of Ca2+ through these voltage-regulated channels supports the rebound release of PRL.

Role of alpha- and beta-adrenoceptors in sympathetically mediated thermogenesis

1993 ◽  
Vol 264 (1) ◽  
pp. E11-E17 ◽  
Author(s):  
E. E. Blaak ◽  
M. A. van Baak ◽  
K. P. Kempen ◽  
W. H. Saris
Keyword(s):  
Blood Pressure ◽  
Heart Rate ◽  
Energy Expenditure ◽  
Whole Body ◽  
Healthy Male ◽  
Beta Adrenoceptor ◽  
Beta 2 ◽  
Body Energy ◽  
Stimulation Of

This study was intended to investigate the role of alpha- and beta-adrenoceptor populations in the sympathetically mediated thermogenesis in healthy lean males. In the first study, the beta 1-, beta 2-, and beta 3-agonist isoprenaline was infused in increasing doses with and without simultaneous infusion of the beta 1-blocker atenolol (Iso and Iso+AT, respectively). There was an increase in whole body energy expenditure (EE) after infusing Iso+AT (P < 0.001) and an almost twofold higher increase after infusion of Iso only (P < 0.001). Stimulation of the beta 2-adrenoceptors by a specific agonist (salbutamol) resulted in a significant increase in EE (P < 0.001). The effect of stimulation of alpha 1-adrenoceptors on EE was measured by infusing increasing doses of the alpha 1-agonist phenylephrine. EE did not change, whereas blood pressure (BP) increased (P < 0.001) and heart rate decreased (P < 0.01). In addition to this study, the alpha 1-, alpha 2-, beta 1-, beta 2-, and beta 3-agonists norepinephrine and epinephrine were infused with simultaneous infusion of the beta 1- and beta 2-blocker propranolol. In both studies, there was no effect on EE, whereas BP increased (P < 0.01). In conclusion, in healthy male lean volunteers both beta 1- and beta 2-adrenoceptors are involved in the sympathetically mediated thermogenesis, whereas the alpha 1-, alpha 2-, and beta 3-adrenoceptors do not play a role.

scholarly journals Angiotensin-II promotes Na+ uptake in larval zebrafish, Danio rerio, in acidic and ion-poor water

2013 ◽  
Vol 220 (3) ◽  
pp. 195-205 ◽  
Author(s):  
Yusuke Kumai ◽  
Nicholas J Bernier ◽  
Steve F Perry
Keyword(s):  
Angiotensin Ii ◽  
Danio Rerio ◽  
Glucocorticoid Receptors ◽  
Enzyme Inhibitor ◽  
Renin Angiotensin System ◽  
Whole Body ◽  
Ang Ii ◽  
Larval Zebrafish ◽  
Rapid Activation ◽  
Stimulation Of

The contribution of the renin–angiotensin system (RAS) to Na+uptake was investigated in larval zebrafish (Danio rerio). At 4 days post fertilization (dpf), the level of whole-body angiotensin-II (ANG-II) was significantly increased after 1- or 3-h exposure to acidic (pH=4.0) or ion-poor water (20-fold dilution of Ottawa tapwater), suggesting rapid activation of the RAS. Long-term (24 h) treatment of 3 dpf larvae with ANG-I or ANG-II significantly increased Na+uptake which was accompanied by an increase in mRNA expression of the Na+-Cl−cotransporter (zslc12a10.2). Induction of Na+uptake by exposure to ANG-I was blocked by simultaneously treating larvae with lisinopril (an angiotensin-converting enzyme inhibitor). Acute (2 h) exposure to acidic water or ion-poor water led to significant increase in Na+uptake which was partially blocked by the ANG-II receptor antagonist, telmisartan. Consistent with these data, translational knockdown of renin prevented the stimulation of Na+uptake following exposure to acidic or ion-poor water. The lack of any effects of pharmacological inhibition (using RU486), or knockdown of glucocorticoid receptors on the stimulation of Na+uptake during acute exposure to acidic or ion-poor environments, indicates that the acute effects of RAS occur independently of cortisol signaling. The results of this study demonstrate that the RAS is involved in Na+homeostasis in larval zebrafish.

scholarly journals A role for sodium-chloride cotransporters in the rapid regulation of ion uptake following acute environmental acidosis: new insights from the zebrafish model

2016 ◽  
Vol 311 (6) ◽  
pp. C931-C941 ◽  
Author(s):  
Raymond W. M. Kwong ◽  
Steve F. Perry
Keyword(s):  
Acid Exposure ◽  
Whole Body ◽  
Cell Type ◽  
Acidic Water ◽  
Zebrafish Model ◽  
Larval Zebrafish ◽  
Water Ph ◽  
Control Water ◽  
Compensatory Regulation

The effects of acute exposure to acidic water on Na+ and Cl− homeostasis, and the mechanisms underlying their compensatory regulation, were investigated in the larval zebrafish Danio rerio. Exposure to acidic water (pH 4.0; control pH 7.6) for 2 h significantly reduced Na+ uptake and whole body Na+ content. Nevertheless, the capacity for Na+ uptake was substantially increased in fish preexposed to acidic water but measured in control water. Based on the accumulation of the Na+-selective dye, Sodium Green, two ionocyte subtypes exhibited intracellular Na+ enrichment after preexposure to acidic water: H+-ATPase rich (HR) cells, which coexpress the Na+/H+ exchanger isoform 3b (NHE3b), and a non-HR cell population. In fish experiencing Na+-Cl− cotransporter (NCC) knockdown, we observed no Sodium Green accumulation in the latter cell type, suggesting the non-HR cells were NCC cells. Elimination of NHE3b-expressing HR cells did not prevent the increased Na+ uptake following acid exposure. On the other hand, the increased Na+ uptake was abolished when the acidic water was enriched with Na+ and Cl−, but not with Na+ only, indicating that the elevated Na+ uptake after acid exposure was associated with the compensatory regulation of Cl−. Further examinations demonstrated that acute acid exposure also reduced whole body Cl− levels and increased the capacity for Cl− uptake. Moreover, knockdown of NCC prevented the increased uptake of both Na+ and Cl− after exposure to acidic water. Together, the results of the present study revealed a novel role of NCC in the compensatory regulation of Na+ and Cl− uptake following acute acidosis.

scholarly journals Characteristics of Protooncogene Expression in A5 Cells

1993 ◽  
Vol 4 (3) ◽  
pp. 531-535
Author(s):  
Eleni Kousvelari ◽  
Chih-Ko Yeh
Keyword(s):  
Cell Cycle ◽  
Adrenergic Receptors ◽  
Early Response ◽  
Mrna Levels ◽  
Intracellular Pathways ◽  
Early Response Genes ◽  
Β Adrenergic Receptors ◽  
Jun B ◽  
Stimulation Of

Stimulation of β-adrenergic receptors by isoproterenol or addition of 8-BrcAMP rapidly and transiently induces the expression of the protooncogenes, c-fos, and jun B, but not that of c-jun in A5 cells. These results indicate that different intracellular pathways may operate within the same cell for the induction of this group of early response genes. The inducibility of c-fos and jun B genes by either isoproterenol of 8-BrcAMP is transcriptionally regulated and accompanied by increases in their respective products. Furthermore, both c-fos and jun B mRNA levels are elevated at G0/G 1 phase of the A5 cell cycle and are inducible by isoproterenol or 8-BrcAMP at the different phases of the cell cycle. These data further suggest a possible role of c-fos and jun B in A5 cell cycle.

Enhanced airway reactivity and inflammation in A2A adenosine receptor-deficient allergic mice

2007 ◽  
Vol 292 (6) ◽  
pp. L1335-L1344 ◽  
Author(s):  
Ahmed Nadeem ◽  
Ming Fan ◽  
Habib R. Ansari ◽  
Catherine Ledent ◽  
S. Jamal Mustafa
Keyword(s):  
Adenosine Receptor ◽  
Nitrosative Stress ◽  
Allergen Challenge ◽  
Inflammatory Cells ◽  
Whole Body ◽  
Airway Reactivity ◽  
A2a Adenosine Receptor ◽  
Group A ◽  
Camp Levels

A2A adenosine receptor (A2AAR) has potent anti-inflammatory properties, which may be important in the regulation of airway reactivity and inflammation. Inflammatory cells that possess A2AAR also produce nitrosative stress, which is associated with pathophysiology of asthma, so we hypothesized that A2AAR deficiency may lead to increased airway reactivity and inflammation through nitrosative stress. Thus the present study was carried out to investigate the role of A2AAR on airway reactivity, inflammation, NF-κB signaling, and nitrosative stress in A2AAR knockout (KO) and wild-type (WT) mice using our murine model of asthma. Animals were sensitized intraperitoneally on days 1 and 6 with 200 μg of ragweed, followed by aerosolized challenges with 0.5% ragweed on days 11, 12, and 13, twice a day. On day 14, airway reactivity to methacholine was assessed as enhanced pause (Penh) using whole body plethysmography followed by bronchoalveolar lavage (BAL) and lung collection for various analyses. Allergen challenge caused a significant decrease in expression of A2AAR in A2A WT sensitized mice, with A2AAR expression being undetected in A2A KO sensitized group leading to decreased lung cAMP levels in both groups. A2AAR deletion/downregulation led to an increase in Penh to methacholine and influx of total cells, eosinophils, lymphocytes, and neutrophils in BAL with highest values in A2A KO sensitized group. A2A KO sensitized group further had increased NF-κB expression and nitrosative stress compared with WT sensitized group. These data suggest that A2AAR deficiency leads to airway inflammation and airway hyperresponsiveness, possibly via involvement of nitrosative stress in this model of asthma.

Forskolin and thyrotrophin stimulation of rat FRTL-5 thyroid cell growth: the role of cyclic AMP

1987 ◽  
Vol 114 (2) ◽  
pp. 199-205 ◽  
Author(s):  
P. A. Ealey ◽  
C. A. Ahene ◽  
J. M. Emmerson ◽  
N. J. Marshall
Keyword(s):  
Cyclic Amp ◽  
Dose Range ◽  
Phosphodiesterase Inhibitor ◽  
Lag Phase ◽  
Thyroid Cell ◽  
Intracellular Camp ◽  
Camp Levels ◽  
Tsh Stimulation ◽  
Stimulation Of

ABSTRACT The adenylate cyclase stimulator forskolin increases intracellular cyclic AMP (cAMP) in rat FRTL-5 cells within minutes and, after a lag phase of 20–24 h, an increase of cells in metaphase is seen. The dose– response relationships were similar in both systems, with significant increases in the number of metaphases observed at ∼0·1 μmol/l and a doubling of cAMP levels at 1 μmol/l, whilst doses of 0·1 mmol/l and above proved cytotoxic. An involvement of intracellular cAMP as a positive intermediate in cell division was further suggested by the finding that a low dose of forskolin (0·1 μmol/l) potentiated TSH stimulation of mitosis. Isobutyl methyl xanthine (IBMX), a phosphodiesterase inhibitor, also acted as a mitogen and potentiated TSH action. Moreover, the simultaneous inclusion of low doses of IBMX and forskolin additionally potentiated TSH stimulation of mitosis. An analogue of cAMP, dibutyryl cAMP, also stimulated mitosis and acted over a restricted dose range, with maximal stimulation at 1 mmol/l. We conclude that cAMP may act as a positive signal for FRTL-5 thyroid cell proliferation. J. Endocr. (1987) 114, 199–205

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