Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 28
Filtrar
Mais filtros

Base de dados
País/Região como assunto
Tipo de documento
Intervalo de ano de publicação
1.
J Evol Biol ; 35(5): 763-771, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35324039

RESUMO

Selection acting across environmental gradients, such as latitudes, can cause spatial structuring of genomic variants even within panmictic populations. In this study, we focused on the within-generation latitudinal selection between northernmost and southernmost individuals of the North Pacific population of a tropical eel Anguilla marmorata, which shares its northernmost distribution with a temperate eel Anguilla japonica. Whole-genome sequencing data indicated that the northernmost and southernmost individuals of A. marmorata belong to a single panmictic population, as suggested by previous studies. On the contrary, parts of genomic regions across multiple chromosomes exhibited significant genetic differentiation between the northernmost and southernmost individuals, and in these genomic regions, the genotypes of the northernmost individuals were similar to those of A. japonica. These findings suggested within-generation latitudinal selection of A. marmorata, which might have led to genetic closeness between northernmost A. marmorata and A. japonica.


Assuntos
Anguilla , Anguilla/genética , Animais , Genômica , Genótipo , Humanos
2.
Artigo em Inglês | MEDLINE | ID: mdl-33059022

RESUMO

The effects of high external ammonia (HEA) exposure on breathing and the potential involvement of ammonia transporting Rh proteins in ammonia sensing were assessed in larval and adult zebrafish. Acute exposure of adults to either 250 or 500 µM (NH4)2SO4 caused increases in ventilation amplitude (AVENT) without affecting frequency (fVENT), resembling the ventilatory response to hypercapnia rather than hypoxia, during which fVENT was increased exclusively. The hyperventilatory response to HEA was prevented by hyperoxia, indicating that control of breathing through ammonia sensing is likely secondary to O2 chemoreception. Neuroepithelial cells (NECs) isolated from gill filaments exhibited a significant increase of intracellular [Ca2+] in response to 1 mM NH4Cl but this response was small (roughly 30%) compared to the response to hypercapnia (37.5 mmHg; ~800% increase). Immunohistochemistry (IHC) failed to reveal the presence of Rh proteins (Rhcgb, Rhbg or Rhag) in gill filament NECs. Knockout of rhcgb did not affect the ventilatory response of adults to HEA. Larvae at 4 days post fertilization (dpf) responded to HEA with increases in fVENT (AVENT was not measured). The hyperventilatory response of larvae to HEA was attenuated (60% reduction) after treatment from 0 to 4 dpf with the sympathetic neurotoxin 6-hydroxydopamine. In larvae, Rhcgb, Rhbg and Rhag were undetectable by IHC in cutaneous NECs yet the fVENT to HEA following Rhbg knockdown was slightly (22%) attenuated. Thus, the hyperventilatory response to external ammonia in adult zebrafish, while apparently initiated by activation of NECs, does not require Rhcgb, nor is the entry of ammonia into NECs reliant on other Rh proteins. The lack of colocalization of Rh proteins with NECs suggests that the entry of ammonia into NECs in larvae, also is not facilitated by this family of ammonia channels.


Assuntos
Amônia/farmacologia , Hiperventilação/fisiopatologia , Fenômenos Fisiológicos Respiratórios/efeitos dos fármacos , Peixe-Zebra/fisiologia , Amônia/metabolismo , Animais , Proteínas Sanguíneas/metabolismo , Cálcio/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Brânquias/citologia , Brânquias/efeitos dos fármacos , Brânquias/metabolismo , Imuno-Histoquímica , Larva/citologia , Larva/efeitos dos fármacos , Larva/metabolismo , Glicoproteínas de Membrana/metabolismo , Células Neuroepiteliais/efeitos dos fármacos , Células Neuroepiteliais/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/metabolismo
3.
J Fish Biol ; 99(1): 288-292, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33651431

RESUMO

Water-choice trial experiments revealed that Anguilla japonica glass eels collected in southern Japan possess strong preferences for fresh water and agricultural water. Their locomotor activity and preference for fresh water were higher and stronger, respectively, in this study when compared to previous studies conducted at lower temperatures. These results suggest that their locomotor activity and preference for fresh water is influenced by water temperature. The attraction to agricultural water indicates their upstream migration and habitat selection could be influenced by agricultural water.


Assuntos
Anguilla , Animais , Água Doce , Japão , Salinidade , Água
4.
Kidney Int ; 94(3): 514-523, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30146013

RESUMO

Pseudohypoaldosteronism type II (PHAII) is a genetic disease characterized by association of hyperkalemia, hyperchloremic metabolic acidosis, hypertension, low renin, and high sensitivity to thiazide diuretics. It is caused by mutations in the WNK1, WNK4, KLHL3 or CUL3 gene. There is strong evidence that excessive sodium chloride reabsorption by the sodium chloride cotransporter NCC in the distal convoluted tubule is involved. WNK4 is expressed not only in distal convoluted tubule cells but also in ß-intercalated cells of the cortical collecting duct. These latter cells exchange intracellular bicarbonate for external chloride through pendrin, and therefore, account for renal base excretion. However, these cells can also mediate thiazide-sensitive sodium chloride absorption when the pendrin-dependent apical chloride influx is coupled to apical sodium influx by the sodium-driven chloride/bicarbonate exchanger. Here we determine whether this system is involved in the pathogenesis of PHAII. Renal pendrin activity was markedly increased in a mouse model carrying a WNK4 missense mutation (Q562E) previously identified in patients with PHAII. The upregulation of pendrin led to an increase in thiazide-sensitive sodium chloride absorption by the cortical collecting duct, and it caused metabolic acidosis. The function of apical potassium channels was altered in this model, and hyperkalemia was fully corrected by pendrin genetic ablation. Thus, we demonstrate an important contribution of pendrin in renal regulation of sodium chloride, potassium and acid-base homeostasis and in the pathophysiology of PHAII. Furthermore, we identify renal distal bicarbonate secretion as a novel mechanism of renal tubular acidosis.


Assuntos
Acidose Tubular Renal/fisiopatologia , Túbulos Renais Coletores/fisiopatologia , Proteínas Serina-Treonina Quinases/genética , Pseudo-Hipoaldosteronismo/complicações , Transportadores de Sulfato/metabolismo , Acidose Tubular Renal/sangue , Acidose Tubular Renal/etiologia , Animais , Modelos Animais de Doenças , Técnicas de Inativação de Genes , Humanos , Túbulos Renais Coletores/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação de Sentido Incorreto , Potássio/sangue , Potássio/metabolismo , Pseudo-Hipoaldosteronismo/genética , Pseudo-Hipoaldosteronismo/fisiopatologia , Eliminação Renal , Cloreto de Sódio/metabolismo , Simportadores de Sódio-Bicarbonato/metabolismo , Transportadores de Sulfato/genética , Regulação para Cima
5.
Artigo em Inglês | MEDLINE | ID: mdl-29913320

RESUMO

Na+ uptake in larval zebrafish (Danio rerio) is coordinated by three mechanisms: Na+/H+-exchanger 3b (NHE3b) expressed in H+-ATPase-rich (HR) cells, an unidentified Na+ channel coupled to electrogenic H+-ATPase expressed in HR cells, and Na+-Cl--cotransporter (NCC) expressed in NCC cells. Recently, acid-sensing ion channels (ASICs) were proposed to be the putative Na+ channel involved in H+-ATPase-mediated Na+ uptake in adult zebrafish and rainbow trout. In the present study, we hypothesized that ASICs also play this role in Na+ uptake in larval zebrafish. In support of this hypothesis, immunohistochemical analyses revealed that ASIC4b was expressed in HR cells on the yolk sac skin at 4 days post-fertilization (dpf). However, neither treatment with the ASIC-specific blocker 4,6-diamidino-2-phenylindole (DAPI) nor morpholino knockdown of ASIC4b reduced Na+ uptake in circumneutral conditions at 4 dpf. However, because ASIC4b knockdown led to significant increases in the mRNA expression of nhe3b and ncc and a significant increase in HR cell density, it is possible that Na+ influx was sustained by increased participation of non-ASIC4b pathways. Moreover, when fish were reared in acidic water (pH = 4), ASIC4b knockdown led to a stimulation of Na+ uptake at 3 and 4 dpf, results which also were inconsistent with an essential role for ASIC-mediated Na+ uptake, even under conditions known to constrain Na+ uptake via NHE3b. Thus, while ASIC4b clearly is expressed in HR cells, the current functional experiments cannot confirm its involvement in Na+ uptake in larval zebrafish.


Assuntos
Canais Iônicos Sensíveis a Ácido/fisiologia , Larva/metabolismo , Sódio/metabolismo , Proteínas de Peixe-Zebra/fisiologia , Peixe-Zebra/crescimento & desenvolvimento , Canais Iônicos Sensíveis a Ácido/efeitos dos fármacos , Canais Iônicos Sensíveis a Ácido/metabolismo , Animais , Feminino , Indóis/farmacologia , Transporte de Íons , Masculino , Morfolinos/farmacologia , Proteínas de Peixe-Zebra/metabolismo
6.
Nephrol Dial Transplant ; 32(7): 1137-1145, 2017 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-28064162

RESUMO

BACKGROUND: Pendrin, the chloride/bicarbonate exchanger of ß-intercalated cells of the renal connecting tubule and the collecting duct, plays a key role in NaCl reabsorption by the distal nephron. Therefore, pendrin may be important for the control of extracellular fluid volume and blood pressure. METHODS: Here, we have used a genetic mouse model in which the expression of pendrin can be switched-on in vivo by the administration of doxycycline. Pendrin can also be rapidly removed when doxycycline administration is discontinued. Therefore, our genetic strategy allows us to test selectively the acute effects of loss of pendrin function. RESULTS: We show that acute loss of pendrin leads to a significant decrease of blood pressure. In addition, acute ablation of pendrin did not alter significantly the acid-base status or blood K + concentration. CONCLUSION: By using a transgenic mouse model, avoiding off-target effects related to pharmacological compounds, this study suggests that pendrin could be a novel target to treat hypertension.


Assuntos
Proteínas de Transporte de Ânions/fisiologia , Pressão Sanguínea/fisiologia , Hipertensão/etiologia , Animais , Hipertensão/metabolismo , Hipertensão/patologia , Masculino , Camundongos , Camundongos Transgênicos , Transportadores de Sulfato
7.
J Exp Biol ; 219(Pt 24): 3988-3995, 2016 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-27802147

RESUMO

The present study investigated the potential role of hypoxia-inducible factor (HIF) in calcium homeostasis in developing zebrafish (Danio rerio). It was demonstrated that zebrafish raised in hypoxic water (30 mmHg; control, 155 mmHg PO2 ) until 4 days post-fertilization exhibited a substantial reduction in whole-body Ca2+ levels and Ca2+ uptake. Ca2+ uptake in hypoxia-treated fish did not return to pre-hypoxia (control) levels within 2 h of transfer back to normoxic water. Results from real-time PCR showed that hypoxia decreased the whole-body mRNA expression levels of the epithelial Ca2+ channel (ecac), but not plasma membrane Ca2+-ATPase (pmca2) or Na+/Ca2+-exchanger (ncx1b). Whole-mount in situ hybridization revealed that the number of ecac-expressing ionocytes was reduced in fish raised in hypoxic water. These findings suggested that hypoxic treatment suppressed the expression of ecac, thereby reducing Ca2+ influx. To further evaluate the potential mechanisms for the effects of hypoxia on Ca2+ regulation, a functional gene knockdown approach was employed to prevent the expression of HIF-1αb during hypoxic treatment. Consistent with a role for HIF-1αb in regulating Ca2+ balance during hypoxia, the results demonstrated that the reduction of Ca2+ uptake associated with hypoxic exposure was not observed in fish experiencing HIF-1αb knockdown. Additionally, the effects of hypoxia on reducing the number of ecac-expressing ionocytes was less pronounced in HIF-1αb-deficient fish. Overall, the current study revealed that hypoxic exposure inhibited Ca2+ uptake in developing zebrafish, probably owing to HIF-1αb-mediated suppression of ecac expression.


Assuntos
Cálcio/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Hipóxia/metabolismo , Peixe-Zebra/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Transporte Biológico/genética , Contagem de Células , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Íons , Morfolinos/farmacologia , Estabilidade Proteica/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
8.
Gen Comp Endocrinol ; 234: 40-6, 2016 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-27179885

RESUMO

Zebrafish (Danio rerio) is an emerging model for integrative physiological research. In this mini-review, we discuss recent advances in the neuroendocrine control of ionic balance in this species, and identify current knowledge gaps and issues that would benefit from further investigation. Zebrafish inhabit a hypo-ionic environment and therefore are challenged by a continual loss of ions to the water. To maintain ionic homeostasis, they must actively take up ions from the water and reduce passive ion loss. The adult gill or the skin of larvae are the primary sites of ionic regulation. Current models for the uptake of major ions in zebrafish incorporate at least three types of ion transporting cells (also called ionocytes); H(+)-ATPase-rich cells for Na(+) uptake, Na(+)/K(+)-ATPase-rich cells for Ca(2+) uptake, and Na(+)/Cl(-)-cotransporter expressing cells for both Na(+) and Cl(-) uptake. The precise molecular mechanisms regulating the paracellular loss of ions remain largely unknown. However, epithelial tight junction proteins, including claudins, are thought to play a critical role in reducing ion losses to the surrounding water. Using the zebrafish model, several key neuroendocrine factors were identified as regulators of epithelial ion movement, including the catecholamines (adrenaline and noradrenaline), cortisol, the renin-angiotensin system, parathyroid hormone and prolactin. Increasing evidence also suggests that gasotransmitters, such as H2S, are involved in regulating ion uptake.


Assuntos
Transporte de Íons/fisiologia , Sistemas Neurossecretores , Proteínas de Peixe-Zebra/metabolismo , Animais , Equilíbrio Hidroeletrolítico , Peixe-Zebra
9.
Pflugers Arch ; 467(8): 1623-41, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25515081

RESUMO

Because majority of biological processes are dependent on pH, maintaining systemic acid-base balance is critical. The kidney contributes to systemic acid-base regulation, by reabsorbing HCO3 (-) (both filtered by glomeruli and generated within a nephron) and acidifying urine. Abnormalities in those processes will eventually lead to a disruption in systemic acid-base balance and provoke metabolic acid-base disorders. Research over the past 30 years advanced our understanding on cellular and molecular mechanisms responsible for those processes. In particular, a variety of transgenic animal models, where target genes are deleted either globally or conditionally, provided significant insights into how specific transporters are contributing to the renal acid-base regulation. Here, we broadly overview the mechanisms of renal ion transport participating to acid-base regulation, with emphasis on data obtained from transgenic mice models.


Assuntos
Rim/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Equilíbrio Hidroeletrolítico , Desequilíbrio Hidroeletrolítico/metabolismo , Animais , Modelos Animais de Doenças , Genótipo , Humanos , Rim/fisiopatologia , Proteínas de Membrana Transportadoras/genética , Camundongos Transgênicos , Modelos Biológicos , Fenótipo , Eliminação Renal , Reabsorção Renal , Desequilíbrio Hidroeletrolítico/genética , Desequilíbrio Hidroeletrolítico/fisiopatologia
10.
Pflugers Arch ; 467(4): 753-65, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24893788

RESUMO

The present study investigated the role of the transcription factor, glial cell missing 2 (gcm2), in Ca(2+) regulation in zebrafish larvae. Translational gene knockdown of gcm2 decreased Ca(2+) uptake and the density of ionocytes expressing the epithelial Ca(2+) channel (ecac), and disrupted the overall Ca(2+) balance. Ca(2+) uptake and the expression of gcm2 messenger RNA (mRNA) were significantly elevated in larvae acclimated to low Ca(2+) water (25 µM); the stimulation of Ca(2+) uptake was not observed in fish experiencing gcm2 knockdown. Acclimation to acidic water (pH 4) significantly reduced whole-body Ca(2+) content owing to reduced Ca(2+) uptake and increased Ca(2+) efflux. However, ecac mRNA levels and the density of ecac-expressing ionocytes were increased in fish acclimated to acidic water, and maximal Ca(2+) uptake capacity (J MAX) was significantly increased when measured in control water (pH ~7.4). Acclimation of larvae to acidic water significantly increased gcm2 mRNA expression, and in gcm2 morphants, no such stimulation in Ca(2+) uptake was observed after their return to control water. Overexpression of gcm2 mRNA resulted in a significant increase in the numbers of ecac-expressing ionocytes and Ca(2+) uptake. These observations reveal a critical role for gcm2 in Ca(2+) homeostasis in zebrafish larvae.


Assuntos
Cálcio/metabolismo , Proteínas de Ligação a DNA/metabolismo , Homeostase , Fatores de Transcrição/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Aclimatação , Animais , Proteínas de Ligação a DNA/genética , Células Epiteliais/metabolismo , Concentração de Íons de Hidrogênio , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo , Fatores de Transcrição/genética , Peixe-Zebra , Proteínas de Peixe-Zebra/genética
11.
Pflugers Arch ; 467(4): 651-64, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24939700

RESUMO

The present study investigated the role of hydrogen sulfide (H2S) in regulating Na(+) uptake in larval zebrafish, Danio rerio. Waterborne treatment of larvae at 4 days post-fertilization (dpf) with Na2S or GYY-4137 (chemicals known to generate H2S) significantly reduced Na(+) uptake. Exposure of larvae to water enriched with NaCl (1 mM NaCl) caused a pronounced reduction in Na(+) uptake which was prevented by pharmacological inhibition of cystathionine ß-synthase (CBS) or cystathionine γ-lyase (CSE), two key enzymes involved in the endogenous synthesis of H2S. Furthermore, translational gene knockdown of CSE and CBSb significantly increased the basal rate of Na(+) uptake. Waterborne treatment with Na2S significantly decreased whole-body acid excretion and reduced Na(+) uptake in larval zebrafish preexposed to acidic (pH 4.0) water (a condition shown to promote Na(+) uptake via Na(+)-H(+)-exchanger 3b, NHE3b). However, Na2S did not affect Na(+) uptake in larvae depleted of NHE3b-containing ionocytes (HR cells) after knockdown of transcription factor glial cell missing 2 (gcm2) in which Na(+) uptake occurs predominantly via Na(+)-Cl(-) co-transporter (NCC)-containing cells. These observations suggest that Na(+) uptake via NHE3b, but not NCC, is regulated by H2S. Whole-mount immunohistochemistry demonstrated that ionocytes expressing NHE3b also express CSE. These data suggests a physiologically relevant role of H2S as a mechanism to lower Na(+) uptake in zebrafish larvae, probably through its inhibitory action on NHE3b.


Assuntos
Sulfeto de Hidrogênio/farmacologia , Absorção Cutânea , Sódio/metabolismo , Animais , Cistationina beta-Sintase/antagonistas & inibidores , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Larva/efeitos dos fármacos , Larva/metabolismo , Liases/antagonistas & inibidores , Liases/genética , Liases/metabolismo , Simportadores de Cloreto de Sódio/metabolismo , Trocador 3 de Sódio-Hidrogênio , Trocadores de Sódio-Hidrogênio/genética , Trocadores de Sódio-Hidrogênio/metabolismo , Sulfitos/farmacologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
12.
J Exp Biol ; 218(Pt 23): 3746-53, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26486367

RESUMO

Nitric oxide (NO) is a gaseous neurotransmitter, which, in adult mammals, modulates the acute hypoxic ventilatory response; its role in the control of breathing in fish during development is unknown. We addressed the interactive effects of developmental age and NO in the control of piscine breathing by measuring the ventilatory response of zebrafish (Danio rerio) adults and larvae to NO donors and by inhibiting endogenous production of NO. In adults, sodium nitroprusside (SNP), a NO donor, inhibited ventilation; the extent of the ventilatory inhibition was related to the pre-existing ventilatory drive, with the greatest inhibition exhibited during exposure to hypoxia (PO2=5.6 kPa). Inhibition of endogenous NO production using L-NAME suppressed the hypoventilatory response to hyperoxia, supporting an inhibitory role of NO in adult zebrafish. Neuroepithelial cells (NECs), the putative oxygen chemoreceptors of fish, contain neuronal nitric oxide synthase (nNOS). In zebrafish larvae at 4 days post-fertilization, SNP increased ventilation in a concentration-dependent manner. Inhibition of NOS activity with L-NAME or knockdown of nNOS inhibited the hypoxic (PO2=3.5 kPa) ventilatory response. Immunohistochemistry revealed the presence of nNOS in the NECs of larvae. Taken together, these data suggest that NO plays an inhibitory role in the control of ventilation in adult zebrafish, but an excitatory role in larvae.


Assuntos
Óxido Nítrico/fisiologia , Oxigênio/metabolismo , Peixe-Zebra/fisiologia , Animais , Hipóxia Celular , Células Quimiorreceptoras/fisiologia , Brânquias/fisiologia , Larva/fisiologia , NG-Nitroarginina Metil Éster/farmacologia , Células Neuroepiteliais/efeitos dos fármacos , Células Neuroepiteliais/fisiologia , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico Sintase Tipo I/análise , Nitroprussiato/farmacologia
13.
J Physiol ; 592(14): 3075-88, 2014 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-24756639

RESUMO

The current study investigated the role of hydrogen sulphide (H2S) in oxygen sensing, intracellular signalling and promotion of ventilatory responses to hypoxia in adult and larval zebrafish (Danio rerio). Both larval and adult zebrafish exhibited a dose-dependent increase in ventilation to sodium sulphide (Na2S), an H2S donor. In vertebrates, cystathionine ß-synthase (CBS) and cystathionine γ-lyase (CSE) are enzymes that catalyse the endogenous production of H2S. In adult zebrafish, inhibition of both CBS and CSE with aminooxyacetate (AOA) and propargyl glycine (PPG) blunted or abolished the hypoxic hyperventilation, and the addition of Na2S to the water partially rescued the effects of inhibiting endogenous H2S production. In zebrafish larvae (4 days post-fertilization), gene knockdown of either CBS or CSE using morpholinos attenuated the hypoxic ventilatory response. Furthermore, the intracellular calcium concentration of isolated neuroepithelial cells (NECs), which are putative oxygen chemoreceptors, increased significantly when these cells were exposed to 50 µm Na2S, supporting a role for H2S in Ca(2+)-evoked neurotransmitter release in these cells. Finally, immunohistochemical labelling showed that NECs dissociated from adult gill contained CBS and CSE, whereas cutaneous NECs in larval zebrafish expressed only CSE. Taken together, these data show that H2S can be produced in the putative oxygen-sensing cells of zebrafish, the NECs, in which it appears to play a pivotal role in promoting the hypoxic ventilatory response.


Assuntos
Sulfeto de Hidrogênio , Hipóxia/fisiopatologia , Respiração , Alcinos/farmacologia , Ácido Amino-Oxiacético/farmacologia , Animais , Cistationina beta-Sintase/antagonistas & inibidores , Cistationina beta-Sintase/fisiologia , Cistationina gama-Liase/antagonistas & inibidores , Cistationina gama-Liase/fisiologia , Glicina/análogos & derivados , Glicina/farmacologia , Células Neuroepiteliais/fisiologia , Oxigênio/fisiologia , Sulfetos/farmacologia , Peixe-Zebra
14.
Am J Physiol Regul Integr Comp Physiol ; 306(1): R51-60, 2014 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-24259461

RESUMO

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 ß-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.


Assuntos
AMP Cíclico/metabolismo , Inibidores de Proteínas Quinases/metabolismo , Transdução de Sinais/fisiologia , Sódio/metabolismo , Peixe-Zebra/metabolismo , Animais , Espaço Intracelular/metabolismo , Larva/metabolismo , Receptores Adrenérgicos beta/metabolismo , Trocador 3 de Sódio-Hidrogênio , Trocadores de Sódio-Hidrogênio/genética , Trocadores de Sódio-Hidrogênio/metabolismo , Proteínas de Peixe-Zebra/metabolismo
15.
J Exp Biol ; 217(Pt 5): 651-62, 2014 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-24574381

RESUMO

Ionic regulation and acid-base balance are fundamental to the physiology of vertebrates including fish. Acidification of freshwater ecosystems is recognized as a global environmental problem, and the physiological responses to acid exposure in a few fish species are well characterized. However, the underlying mechanisms promoting ionic and acid-base balance for most fish species that have been investigated remain unclear. Zebrafish (Danio rerio) has emerged as a powerful model system to elucidate the molecular basis of ionic and acid-base regulation. The utility of zebrafish is related to the ease with which it can be genetically manipulated, its suitability for state-of-the-art molecular and cellular approaches, and its tolerance to diverse environmental conditions. Recent studies have identified several key regulatory mechanisms enabling acclimation of zebrafish to acidic environments, including activation of the sodium/hydrogen exchanger (NHE) and H(+)-ATPase for acid secretion and Na(+) uptake, cortisol-mediated regulation of transcellular and paracellular Na(+) movements, and ionocyte proliferation controlled by specific cell-fate transcription factors. These integrated physiological responses ultimately contribute to ionic and acid-base homeostasis in zebrafish exposed to acidic water. In the present review, we provide an overview of the general effects of acid exposure on freshwater fish, the adaptive mechanisms promoting extreme acid tolerance in fishes native to acidic environments, and the mechanisms regulating ionic and acid-base balance during acid exposure in zebrafish.


Assuntos
Equilíbrio Ácido-Base/efeitos dos fármacos , Ácidos/toxicidade , Adaptação Fisiológica , Peixes/fisiologia , Poluentes Químicos da Água/toxicidade , Animais , Água Doce/química , Concentração de Íons de Hidrogênio , Transporte de Íons/efeitos dos fármacos , Peixe-Zebra/fisiologia
16.
J Exp Biol ; 217(Pt 19): 3569-78, 2014 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-25063853

RESUMO

The ontogeny of carbon dioxide (CO2) sensing in zebrafish (Danio rerio) has not been examined. In this study, CO2-mediated increases in heart rate were used to gauge the capacity of zebrafish larvae to sense CO2. CO2 is thought to be detected via neuroepithelial cells (NECs), which are homologous to mammalian carotid body glomus cells. Larvae at 5 days post-fertilization (d.p.f.) exhibited tachycardia when exposed for 30 min to 0.75% CO2 (~5.63 mmHg); at 7 d.p.f., tachycardia was elicited by 0.5% CO2 (~3.75 mmHg). Based on pharmacological evidence using ß-adrenergic receptor (ß-AR) antagonists, and confirmed by ß1-AR translational gene knockdown using morpholinos, the reflex tachycardia accompanying hypercapnia was probably mediated by the interaction of catecholamines with cardiac ß1 receptors. Because the cardiac response to hypercapnia was abolished by the ganglionic blocker hexamethonium, it is probable that the reflex cardio-acceleration was mediated by catecholamines derived from sympathetic adrenergic neurons. Owing to its likely role in facilitating intracellular acidification during exposure to hypercapnia, it was hypothesized that carbonic anhydrase (CA) is involved in CO2 sensing, and that inhibition of CA activity would blunt the downstream responses. Indeed, the cardiac response to hypercapnia (0.75% CO2) was reduced in fish at 5 d.p.f. exposed to acetazolamide, a CA inhibitor, and in fish experiencing zCAc (CA2-like a) knockdown. Successful knockdown of zCAc was confirmed by CA activity measurements, western blotting and immunocytochemistry. Co-injection of embryos with zCAc morpholino and mRNA modified at the morpholino binding site restored normal levels of CA activity and protein levels, and restored (rescued) the usual cardiac responses to hypercapnia. These data, combined with the finding that zCAc is expressed in NECs located on the skin, suggest that the afferent limb of the CO2-induced cardiac reflex in zebrafish larvae is initiated by coetaneous CO2-sensing neuroepithelial cells.


Assuntos
Dióxido de Carbono/metabolismo , Anidrases Carbônicas/metabolismo , Catecolaminas/metabolismo , Frequência Cardíaca/fisiologia , Larva/crescimento & desenvolvimento , Larva/metabolismo , Peixe-Zebra/crescimento & desenvolvimento , Peixe-Zebra/metabolismo , Fatores Etários , Animais , Hipercapnia , Células Neuroepiteliais/metabolismo , Receptores Adrenérgicos beta
17.
Am J Physiol Regul Integr Comp Physiol ; 303(10): R1031-41, 2012 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-23019213

RESUMO

The potential role of adrenergic systems in regulating Na(+) uptake in zebrafish (Danio rerio) larvae was investigated. Treatment with isoproterenol (a generic ß-adrenergic receptor agonist) stimulated Na(+) uptake, whereas treatment with phenylephrine (an α(1)-adrenergic receptor agonist) as well as clonidine (an α(2)-adrenergic receptor agonist) significantly reduced Na(+) uptake, suggesting opposing roles of α- and ß-adrenergic receptors in Na(+) uptake regulation. The increase in Na(+) uptake associated with exposure to acidic water (pH = 4.0) was attenuated in the presence of the nonselective ß-receptor antagonist propranolol or the ß(1)-receptor blocker atenolol; the ß(2)-receptor antagonist ICI-118551 was without effect. The stimulation of Na(+) uptake associated with ion-poor water (32-fold dilution of Ottawa tapwater) was unaffected by ß-receptor blockade. Translational gene knockdown of ß-receptors using antisense oligonucleotide morpholinos was used as a second method to assess the role of adrenergic systems in the regulation of Na(+) uptake. Whereas ß(1)- or ß(2B)-receptor knockdown led to significant decreases in Na(+) uptake during exposure to acidic water, only ß(2A)-receptor morphants failed to increase Na(+) uptake in response to ion-poor water. In support of the pharmacology and knockdown experiments that demonstrated an involvement of ß-adrenergic systems in the control of Na(+) uptake, we showed that the H(+)-ATPase-rich (HR) cell, a subtype of ionocyte known to be a site of Na(+) uptake, is innervated and appears to express ß-adrenergic receptors (propranolol binding sites) at 4 days postfertilization. These data indicate an important role of adrenergic systems in regulating Na(+) uptake in developing zebrafish.


Assuntos
Receptores Adrenérgicos beta/metabolismo , Sódio/metabolismo , Agonistas Adrenérgicos beta/farmacologia , Antagonistas Adrenérgicos beta/farmacologia , Animais , Catecolaminas , Hidrocortisona , Concentração de Íons de Hidrogênio , Íons , Larva , Água/química , Peixe-Zebra
18.
Am J Physiol Regul Integr Comp Physiol ; 301(5): R1517-28, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21832207

RESUMO

The involvement of a Na(+)/H(+) exchanger (NHE) in mediating Na(+) uptake by freshwater fish is currently debated. Although supported indirectly by empirical molecular and pharmacological data, theoretically its operation should be constrained thermodynamically, owing to unfavorable chemical gradients. Recently, there has been an increasing focus on ammonia channels (Rh proteins) as potentially contributing to Na(+) uptake across the freshwater fish gill. In this study, we tested the hypothesis that Rhcg1, a specific apical isoform of Rh protein, is critically important in facilitating Na(+) uptake in zebrafish larvae via its interaction with NHE. Treating larvae (4 days postfertilization) with 5-(N-ethyl-N-isopropyl) amiloride (EIPA), an inhibitor of NHE, caused a significant reduction in Na(+) uptake in fish reared in acidic water (pH ∼ 4.0). A role for NHE in Na(+) uptake was further confirmed by translational knockdown of NHE3b, an isoform of NHE thought to be responsible for Na(+)/H(+) exchange in zebrafish larvae. Exposing the larvae reared in acidic water to 5 mM external ammonium sulfate or increasing the buffering capacity of the water with 10 mM HEPES caused concurrent reductions in ammonia excretion and Na(+) uptake. Furthermore, translational knockdown of Rhcg1 significantly reduced ammonia excretion and Na(+) uptake in larvae chronically (4 days) or acutely (24 h) exposed to acidic water. Unlike in sham-injected larvae, EIPA did not affect Na(+) uptake in fish experiencing Rhcg1 knockdown. Additionally, exposure of larvae to bafilomycin A1 (an inhibitor of H(+)-ATPase) significantly reduced Na(+) uptake in fish reared in acidic water. These observations suggest the existence of multiple mechanisms of Na(+) uptake in larval zebrafish in acidic water: one in which Na(+) uptake via NHE3b is linked to ammonia excretion via Rhcg1, and another facilitated by H(+)-ATPase.


Assuntos
Amônia/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Brânquias/metabolismo , Sódio/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Amilorida/análogos & derivados , Amilorida/farmacologia , Animais , Transporte Biológico , Soluções Tampão , Proteínas de Transporte de Cátions/genética , Técnicas de Silenciamento de Genes , Brânquias/efeitos dos fármacos , Brânquias/embriologia , Concentração de Íons de Hidrogênio , Cinética , Larva/metabolismo , Macrolídeos/farmacologia , ATPases Translocadoras de Prótons/antagonistas & inibidores , ATPases Translocadoras de Prótons/metabolismo , Trocador 3 de Sódio-Hidrogênio , Trocadores de Sódio-Hidrogênio/antagonistas & inibidores , Trocadores de Sódio-Hidrogênio/genética , Trocadores de Sódio-Hidrogênio/metabolismo , Peixe-Zebra , Proteínas de Peixe-Zebra/genética
19.
Artigo em Inglês | MEDLINE | ID: mdl-21600298

RESUMO

The objective of the present study was to characterize the capacity of zebrafish (Danio rerio) to regulate whole body Na⁺ levels during exposure to acidic (pH 3.8-4.0) water. Exposure to acidic water significantly affected the mRNA levels of 14 claudin and two occludin isoforms, tight junction proteins thought to be involved in regulating paracellular efflux. Despite these changes, Na⁺ efflux as well as uptake of polyethylene glycol (PEG), a marker for paracellular pathway, was persistently elevated during the 2-week period of acid exposure, although there was a transient recovery between 12- and 72-h. Pre-exposing fish to acidic water for 2 weeks failed to attenuate the increase in Na⁺ efflux associated with acute exposure to acidic water of low [Ca²âº]. However, during recovery in water of circumneutral pH following exposure to acidic water, normal rates of Na⁺ efflux were restored within 5h. The rate of Na⁺ uptake was significantly elevated between 4 and 7 days of exposure to acidic water; the increase was associated with significant increases in maximal Na⁺ uptake capacity (J(MAX)Na⁺) and affinity constant (K(M)). These results demonstrate that in acidic water, zebrafish maintain their whole body Na⁺ balance primarily by regulating Na⁺ uptake, rather than Na⁺ efflux.


Assuntos
Ácidos/farmacologia , Homeostase , Sódio/metabolismo , Junções Íntimas/metabolismo , Peixe-Zebra/metabolismo , Aclimatação , Ácidos/metabolismo , Análise de Variância , Animais , Cálcio/metabolismo , Permeabilidade da Membrana Celular , Claudinas/metabolismo , Concentração de Íons de Hidrogênio , Proteínas de Membrana/metabolismo , Ocludina , Polietilenoglicóis/metabolismo , Isoformas de Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Tempo , Água/química , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
20.
J Exp Biol ; 213(Pt 21): 3656-65, 2010 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-20952613

RESUMO

Goldfish acclimated to cold water (e.g. 7°C) experience a marked reduction in functional lamellar surface area owing to the proliferation of an interlamellar cell mass (ILCM), a phenomenon termed gill remodelling. The goal of the present study was to assess the consequences of the reduced functional surface area on the capacity of goldfish to excrete ammonia. Despite the expected impact of ambient temperature on functional surface area, fish acclimated to 7°C and 25°C exhibited similar rates of ammonia excretion (J(net,amm)); the Q10 values for fed and starved fish were 1.07 and 1.20, respectively. To control for possible temperature-related differences in rates of endogenous ammonia production, J(net,amm) was determined at the two acclimation temperatures after loading fish with 1.12 µmol g₋1 of NH4Cl. In the 3 h post-injection period, J(net,amm) was elevated to a greater extent in the 25°C fish. To estimate the potential contribution of increased ventilation and cardiac output to ammonia clearance in the warmer fish, the ammonia loading experiment was repeated on the 7°C fish immediately after they were exercised to exhaustion. The rate of excretion of ammonia was significantly increased in the exercised 7°C fish (presumably experiencing increased ventilation and cardiac output for at least some of the measurement period) suggesting that differences in external and internal convection may at least partially explain the enhanced capacity of the 25°C fish to clear the ammonia load. To more specifically assess the contribution of the different functional surface areas on the differing rates of ammonia clearance at the two acclimation temperatures, the 7°C fish were exposed for 7 days to hypoxia (P(O2)=10 mmHg=1.33 kPa), a treatment known to cause the disappearance of the ILCM. The results demonstrated that the hypoxia-associated loss of the ILCM was accompanied by a significant increase in the rate of ammonia clearance in the 7°C fish when returned to normoxic conditions. To determine whether compensatory changes in the ammonia transporting proteins might be contributing to sustaining J(net,amm) under conditions of reduced functional lamellar surface area, the relative expression and branchial distribution of four Rh proteins were assessed by western blotting and immunocytochemistry. Although the relative expression of the Rh proteins was unaffected by acclimation temperature, there did appear to be a change in the spatial distribution of Rhag, Rhbg and Rhcg1. Specifically, these three Rh proteins (and to a lesser extent Rhcg2) appeared to localize in cells on the outer edge of the ILCM that were enriched with Na(+)/K(+)-ATPase. Thus, we suggest that despite the impediment to ammonia excretion imposed by the ILCM, goldfish acclimated to 7°C are able to sustain normal rates of excretion owing to the redistribution of ammonia transporting cells.


Assuntos
Amônia/metabolismo , Brânquias/metabolismo , Carpa Dourada/metabolismo , Aclimatação/efeitos dos fármacos , Cloreto de Amônio/farmacologia , Animais , Western Blotting , Proteínas de Peixes/metabolismo , Imunofluorescência , Brânquias/citologia , Brânquias/efeitos dos fármacos , Hipóxia/metabolismo , Injeções Intraperitoneais , Estado Nutricional/efeitos dos fármacos , Condicionamento Físico Animal , Cloreto de Sódio/farmacologia , Temperatura
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA