Evidence that SLC26 anion transporters mediate branchial chloride uptake in adult zebrafish (Danio rerio)

2009 ◽  
Vol 297 (4) ◽  
pp. R988-R997 ◽  
Author(s):  
S. F. Perry ◽  
B. Vulesevic ◽  
M. Grosell ◽  
M. Bayaa
Keyword(s):  
Mrna Expression ◽  
Danio Rerio ◽  
Whole Body ◽  
Mrna Levels ◽  
Adult Zebrafish ◽  
Chloride Uptake ◽  
Anion Transporters ◽  
Anion Transporter ◽  
Normal Water

Experiments were performed to test the hypothesis that three members of the SLC26 anion transporter gene family (SLC26a3, A4, and A6; hereafter termed za3, za4, and za6) mediate branchial Cl−/HCO3− exchange in adult zebrafish ( Danio rerio). Real-time RT-PCR demonstrated that the gill expressed relatively high levels of za6 mRNA; za3 and za4 mRNA, while present, were less abundant. Also, za4 and za6 were expressed at relatively high levels in the kidney. The results of in situ hybridization or immunocytochemistry (za3 only) experiments performed on gill sections revealed that the SLC26 transporters were predominantly expressed on the filament epithelium (especially within the interlamellar regions) and to a lesser extent on the lamellar epithelium at the base of lamellae. This distribution pattern suggests that the SLC26 anion transporters are localized to mitochondrion-rich cells (ionocytes). Transferring fish to water containing low [Cl−] (0.02 mmol/l) resulted in significant increases in branchial SLC26 mRNA expression after 5–10 days of exposure relative to fish raised in normal water [Cl−] (0.4 mmol/l); transferring fish to Cl−-enriched water (2.0 mmol/l) was without effect on mRNA levels. Transferring fish to water containing elevated levels of NaHCO3 (10–12.5 mmol/l) caused marked increases in branchial SLC26 mRNA expression between 3 and 10 days of transfer that was associated with a significant 40% increase in Cl− uptake (as measured upon return to normal water after 7 days). A decrease in whole body net acid excretion (equivalent to an increase in net base excretion) in fish previously maintained in high [NaHCO3] water, concurrent with increases in Cl− uptake and SLC26 mRNA levels, suggests a role for these anion transporters in Cl− uptake and acid-base regulation owing to their Cl−/HCO3− exchange activities.

scholarly journals Involvement of the calcium-sensing receptor in calcium homeostasis in larval zebrafish exposed to low environmental calcium

2014 ◽  
Vol 306 (4) ◽  
pp. R211-R221 ◽  
Author(s):  
Raymond W. M. Kwong ◽  
Dan Auprix ◽  
Steve F. Perry
Keyword(s):  
In Situ Hybridization ◽  
Confocal Microscopy ◽  
Mrna Expression ◽  
Danio Rerio ◽  
Whole Body ◽  
Calcium Sensing Receptor ◽  
Larval Zebrafish ◽  
Calcium Sensing ◽  
Normal Water

The involvement of the calcium-sensing receptor (CaSR) in Ca2+ homeostasis was investigated in larval zebrafish, Danio rerio. The expression of CaSR mRNA was first observed at 3 h posfertilization (hpf) and increased with development until plateauing at ∼48 hpf. At 4 dpf, CaSR mRNA was increased in fish acclimated to low Ca2+ water (25 μM vs. 250 μM in normal water). Using immunohistochemistry and confocal microscopy, we demonstrated that the CaSR is expressed in the olfactory epithelium, neuromasts, ionocytes on the yolk sac epithelium, and corpuscles of Stannius. Results of double immunohistochemistry and/or in situ hybridization indicated that the CaSR is localized to a subset of mitochondrion-rich ionocytes enriched with Na+/K+-ATPase and epithelial Ca2+ channel ( ecac). Translational knockdown of the CaSR prevented 4 dpf larvae from regulating whole body Ca2+ levels when exposed to a low Ca2+ environment. Further, the increases in ecac mRNA expression and Ca2+ influx, normally associated with exposure to low-Ca2+ water, were prevented by CaSR knockdown. These findings demonstrate that larval zebrafish lacking the CaSR lose their ability to regulate Ca2+ when confronted with a low-Ca2+ environment. Results from real-time PCR suggested that the mRNA expression of the hypocalcemic hormone stanniocalcin ( stc-1) remained elevated in the CaSR morphants following acclimation to low-Ca2+ water. Overall, the results suggest that the CaSR is critical for Ca2+ homeostasis in larval zebrafish exposed to low environmental Ca2+ levels, possibly owing to its modulation of stanniocalcin mRNA expression.

scholarly journals Selenium status affects selenoprotein expression, reproduction, and F1 generation locomotor activity in zebrafish (Danio rerio)

2014 ◽  
Vol 111 (11) ◽  
pp. 1918-1931 ◽  
Author(s):  
Sam Penglase ◽  
Kristin Hamre ◽  
Josef D. Rasinger ◽  
Staale Ellingsen
Keyword(s):  
Locomotor Activity ◽  
Reproductive Success ◽  
Mrna Expression ◽  
Danio Rerio ◽  
Expression Patterns ◽  
Maximum Growth ◽  
Cellular Growth ◽  
Mrna Levels ◽  
Gpx Activity ◽  
Se Status

Se is an essential trace element, and is incorporated into selenoproteins which play important roles in human health. Mammalian selenoprotein-coding genes are often present as paralogues in teleost fish, and it is unclear whether the expression patterns or functions of these fish paralogues reflect their mammalian orthologues. Using the model species zebrafish (Danio rerio; ZF), we aimed to assess how dietary Se affects key parameters in Se metabolism and utilisation including glutathione peroxidase (GPX) activity, the mRNA expression of key Se-dependent proteins (gpx1a, gpx1b, sepp1a and sepp1b), oxidative status, reproductive success and F1 generation locomotor activity. From 27 d until 254 d post-fertilisation, ZF were fed diets with graded levels of Se ranging from deficient ( < 0·10 mg/kg) to toxic (30 mg/kg). The mRNA expression of gpx1a and gpx1b and GPX activity responded in a similar manner to changes in Se status. GPX activity and mRNA levels were lowest when dietary Se levels (0·3 mg/kg) resulted in the maximum growth of ZF, and a proposed bimodal mechanism in response to Se status below and above this dietary Se level was identified. The expression of the sepp1 paralogues differed, with only sepp1a responding to Se status. High dietary Se supplementation (30 mg/kg) decreased reproductive success, while the offspring of ZF fed above 0·3 mg Se/kg diet had lower locomotor activity than the other groups. Overall, the novel finding of low selenoprotein expression and activity coinciding with maximum body growth suggests that even small Se-induced variations in redox status may influence cellular growth rates.

Estrogen-related receptor γ2 controls NaCl uptake to maintain ionic homeostasis

2021 ◽  
Author(s):  
Shang-Wu Shih ◽  
Jia-Jiun Yan ◽  
Yi-Hsing Wang ◽  
Yi-Ling Tsou ◽  
Ling Chiu ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Body Fluid ◽  
Whole Body ◽  
Ionic Homeostasis ◽  
Ion Regulation ◽  
Expression Levels ◽  
Morpholino Knockdown ◽  
Oryzias Melastigma ◽  
Euryhaline Teleost

Estrogen-related receptors (ERRs) are known to function in mammalian kidney as key regulators of ion transport-related genes; however, a comprehensive understanding of the physiological functions of ERRs in vertebrate body fluid ionic homeostasis is still elusive. Here, we used medaka (Oryzias melastigma), a euryhaline teleost, to investigate how ERRs are involved in ion regulation. After transferring medaka from hypertonic seawater to hypotonic freshwater (FW), the mRNA expression levels of errγ2 were highly upregulated, suggesting that ERRγ2 may play a crucial role in ion uptake. In situ hybridization and immunofluorescence staining showed that errγ2 was specifically expressed in ionocytes, the cells responsible for Na+/Cl- transport. In normal FW, ERRγ2 morpholino knockdown caused reductions in the mRNA expression of Na+/Cl- cotransporter (NCC), the number of NCC ionocytes, Na+/Cl- influxes of ionocytes, and whole-body Na+/Cl- contents. In FW with low Na+ and low Cl-, the expression levels of mRNA for Na+/H+ exchanger 3 (NHE3) and NCC were both decreased in ERRγ2 morphants. Treating embryos with DY131, an agonist of ERRγ, increased the whole-body Na+/Cl- contents and ncc mRNA expression in ERRγ2 morphants. As such, medaka ERRγ2 may control Na+/Cl- uptake by regulating ncc and/or nhe3 mRNA expression and ionocyte number, and these regulatory actions may be subtly adjusted depending on internal and external ion concentrations. These findings not only provide new insights into the underpinning mechanism of actions of ERRs, but also enhance our understanding of their roles in body fluid ionic homeostasis for adaptation to changing environments during vertebrate evolution.

scholarly journals Upregulation of the angiogenic factor heparin affin regulatory peptide by progesterone in rat uterus

1998 ◽  
Vol 158 (3) ◽  
pp. 389-399 ◽  
Author(s):  
PE Milhiet ◽  
F Vacherot ◽  
JP Caruelle ◽  
D Barritault ◽  
D Caruelle ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Hormonal Regulation ◽  
Angiogenic Factor ◽  
Mrna Levels ◽  
Heparin Binding ◽  
Regulatory Peptide ◽  
Rat Uterus ◽  
New Family ◽  
Immunohistochemical Studies

Heparin affin regulatory peptide (HARP), also named pleiotropin, is a secreted polypeptide that belongs to a new family of heparin-binding growth/differentiation factors. In this study, we investigated the expression and distribution of HARP mRNA and protein in rat uterus. Semi-quantitative reverse transcriptase PCR experiments showed variations in HARP mRNA levels throughout the estrous cycle, with a maximum during diestrus, pointing to hormonal regulation of HARP mRNA expression. Uterine expression of HARP mRNA was studied in ovariectomized animals treated with 17 beta-estradiol, progesterone alone or progesterone and RU486. In these experiments, progesterone upregulated HARP mRNA expression. Induction was observed 6 h after progesterone injection and was inhibited by RU486 treatment. In contrast, after 17 beta-estradiol injection, a slight decrease in HARP mRNA expression was observed. In situ hybridization studies with digoxigenin-labeled DNA probe revealed that HARP mRNA was present in smooth muscle cells of both myometrium and blood vessels and also in endothelial cells from endometrium. Immunohistochemical studies showed that HARP expression was not limited to cells that expressed HARP mRNA, but also occurred in both the luminal and glandular epithelium even though its transcript was never detected. We conclude that HARP may mediate the effects of progesterone on the homeostasis and vascularization of uterine tissue.

Somatostatin-28 inhibitory action on somatolactin-α and -β gene expression in goldfish

2014 ◽  
Vol 307 (6) ◽  
pp. R755-R768 ◽  
Author(s):  
Quan Jiang ◽  
Anderson O. L. Wong
Keyword(s):  
Gene Expression ◽  
Mrna Expression ◽  
Primary Cultures ◽  
Inhibitory Effect ◽  
Pituitary Cells ◽  
Mrna Levels ◽  
Structural Identity ◽  
Rapid Amplification ◽  
Differential Coupling

Somatostain (SS) is known to inhibit growth hormone (GH) and prolactin (PRL) secretion. Somatolactin (SL) is a member of the GH/PRL family, but its regulation by goldfish brain somatostatin-28 (gbSS-28) has not been examined. To this end, the structural identity of goldfish SLα was established by 5′/3′-rapid amplification of cDNA ends. As revealed by in situ hybridization and immunohistochemical staining, the expression of SL isoforms was detected in pituitary cells located in the neurointermediate lobe (NIL). The transcripts of goldfish SS receptor 5a (Sst5a) but not Sst1b, Sst2, or Sst3a were detected in the goldfish NIL cells by RT-PCR. In goldfish pituitary cells, gbSS-28 not only had an inhibitory effect on basal SLα and SLβ mRNA levels but also could abolish insulin-like growth factor-stimulated SL gene expression. In primary cultures of goldfish NIL cells, gbSS-28 reduced forskolin-stimulated total cAMP production. With the use of a pharmacological approach, the adenylate cyclase (AC)/cAMP and phospholipase C (PLC)/inositol trisphosphate (IP3)/protein kinase C (PKC) cascades were shown to be involved in gbSS-28-inhibited SLα mRNA expression. Similar postreceptor signaling cascades were also observed for gbSS-28-reduced SLβ mRNA expression, except that PKC coupling to PLC was not involved. These results provide evidence that gbSS-28 can inhibit SLα and SLβ gene expression at the goldfish pituitary level via Sst5 through differential coupling of AC/cAMP and PLC/IP3/PKC cascades.

Renal expression of organic anion transporter OAT2 in rats and mice is regulated by sex hormones

2007 ◽  
Vol 292 (1) ◽  
pp. F361-F372 ◽  
Author(s):  
Marija Ljubojević ◽  
Daniela Balen ◽  
Davorka Breljak ◽  
Marija Kušan ◽  
Naohiko Anzai ◽  
...  
Keyword(s):  
Gender Differences ◽  
Mrna Expression ◽  
Rat Kidney ◽  
Organic Anion ◽  
Staining Intensity ◽  
Protein Band ◽  
Organic Anion Transporter ◽  
Adult Rats ◽  
Anion Transporters ◽  
Anion Transporter

The renal reabsorption and/or excretion of various organic anions is mediated by specific organic anion transporters (OATs). OAT2 (Slc22a7) has been identified in rat kidney, where its mRNA expression exhibits gender differences [females (F) > males (M)]. The exact localization of OAT2 protein in the mammalian kidney has not been reported. Here we studied the expression of OAT2 mRNA by RT-PCR and its protein by Western blotting (WB) and immunocytochemistry (IC) in kidneys of adult intact and gonadectomized M and F, sex hormone-treated castrated M, and prepubertal M and F rats, and the protein in adult M and F mice. In adult rats, the expression of OAT2 mRNA was predominant in the outer stripe (OS) tissue, exhibiting 1) gender dependency (F > M), 2) upregulation by castration and downregulation by ovariectomy, and 3) strong downregulation by testosterone and weak upregulation by estradiol and progesterone treatment. A polyclonal antibody against rat OAT2 on WB of isolated renal membranes labeled a ∼66-kDa protein band that was stronger in F. By IC, the antibody exclusively stained brush border (BB) of the proximal tubule S3 segment (S3) in the OS and medullary rays (F > M). In variously treated rats, the pattern of 66-kDa band density in the OS membranes and the staining intensity of BB in S3 matched the mRNA expression. The expression of OAT2 protein in prepubertal rats was low and gender independent. In mice, the expression pattern largely resembled that in rats. Therefore, OAT2 in rat (and mouse) kidney is localized to the BB of S3, exhibiting gender differences (F > M) that appear in puberty and are caused by strong androgen inhibition and weak estrogen and progesterone stimulation.

scholarly journals Progestin increases the expression of gonadotropins in pituitaries of male zebrafish

2016 ◽  
Vol 230 (1) ◽  
pp. 143-156 ◽  
Author(s):  
Cuili Wang ◽  
Dongteng Liu ◽  
Weiting Chen ◽  
Wei Ge ◽  
Wanshu Hong ◽  
...  
Keyword(s):  
Time Course ◽  
Ex Vivo ◽  
Mrna Levels ◽  
Adult Zebrafish ◽  
Male Adult ◽  
Positive Effects ◽  
Time Course Experiment ◽  
Nuclear Progesterone Receptor

Our previous study showed that the in vivo positive effects of 17α,20β-dihydroxy-4-pregnen-3-one (DHP), the major progestin in zebrafish, on early spermatogenesis was much stronger than the ex vivo ones, which may suggest an effect of DHP on the expression of gonadotropins. In our present study, we first observed that fshb and lhb mRNA levels in the pituitary of male adult zebrafish were greatly inhibited by 3 weeks exposure to 10nM estradiol (E2). However, an additional 24h 100nM DHP exposure not only reversed the E2-induced inhibition, but also significantly increased the expression of fshb and lhb mRNA. These stimulatory effects were also observed in male adult fish without E2 pretreatment, and a time course experiment showed that it took 24h for fshb and 12h for lhb to respond significantly. Because these stimulatory activities were partially antagonized by a nuclear progesterone receptor (Pgr) antagonist mifepristone, we generated a Pgr-knockout (pgr–/–) model using the TALEN technique. With and without DHP in vivo treatment, fshb and lhb mRNA levels of pgr–/– were significantly lower than those of pgr+/+. Furthermore, ex vivo treatment of pituitary fragments of pgr–/– with DHP stimulated lhb, but not fshb mRNA expression. Results from double-colored fluorescent in situ hybridization showed that pgr mRNA was expressed only in fshb-expressing cells. Taken together, our results indicated that DHP participated in the regulation of neuroendocrine control of reproduction in male zebrafish, and exerted a Pgr-mediated direct stimulatory effect on fshb mRNA at pituitary level.

scholarly journals Distribution and kinetics of superantigen-induced cytokine gene expression in mouse spleen.

1993 ◽  
Vol 178 (5) ◽  
pp. 1531-1539 ◽  
Author(s):  
M Bette ◽  
M K Schäfer ◽  
N van Rooijen ◽  
E Weihe ◽  
B Fleischer
Keyword(s):  
T Cell ◽  
Mrna Expression ◽  
Interleukin 2 ◽  
Cytokine Gene Expression ◽  
Tnf Alpha ◽  
Second Phase ◽  
Mrna Levels ◽  
Messenger Rnas ◽  
Kinetics Of

The polyclonal stimulation of T cells by bacterial superantigens is involved in the pathogenesis of the toxic shock syndrome in certain staphylococcal and streptococcal infections. Here we describe the onset and kinetics of superantigen-induced cytokine production in situ in spleens of normal BALB/c mice monitored at the level of cytokine mRNA expression by in situ hybridization. Messenger RNAs for interleukin 2 (IL-2), interferon gamma, and tumor necrosis factors (TNF) alpha and beta were not expressed at detectable levels in spleens of unstimulated animals but became visible already 30 min after intraperitoneal application of 50 micrograms staphylococcal enterotoxin B. All mRNA levels showed peak expression approximately 3 h after injection and a slow decrease up to 24 h after injection. Expression of the mRNAs was restricted to the T cell-dependent area of the periarteriolar lymphatic sheets of the spleen. Interestingly, TNF-alpha mRNA showed a biphasic response, the early appearing mRNA had the same localization as the other mRNAs, whereas after 3 h TNF-alpha mRNA showed a broader distribution indicating a second cell population producing TNF-alpha. The expression of IL-2 and TNF proteins in the serum increased in parallel to the observed mRNA changes with a slight delay. The presence of macrophages was not required for the expression of the cytokine mRNAs in the spleen as the expression was unchanged in macrophage-depleted mice. Only the second phase of TNF-alpha mRNA expression was abrogated in such animals. The expression of all mRNAs was completely suppressed by prior administration of cyclosporin A. These data show that nonphagocytic cells are the essential superantigen-presenting cells in vivo and indicate that at least part of the pathogenetic TNF-alpha is T cell derived.

scholarly journals Gene expression and secretion of LH and FSH in relation to gene expression of GnRH receptors in the brushtail possum (Trichosurus vulpecula) demonstrates highly conserved mechanisms

Reproduction ◽  
2009 ◽  
Vol 137 (1) ◽  
pp. 129-140 ◽  
Author(s):  
J L Crawford ◽  
D A Heath ◽  
L J Haydon ◽  
B P Thomson ◽  
D C Eckery
Keyword(s):  
Gene Expression ◽  
Mrna Expression ◽  
Oestrous Cycle ◽  
Brushtail Possum ◽  
Mrna Levels ◽  
Gonadotrophin Secretion ◽  
Pituitary Glands ◽  
Sampling Regime ◽  
And Storage

In eutherian mammals, the gonadotrophins (LH and FSH) are synthesized and stored in gonadotroph cells under the regulation of multiple mechanisms including GnRH. Very little is known about the regulation of gonadotrophin secretion and storage in pituitary glands of marsupials. This study revealed, using quantitative PCR and heterologous RIA techniques, thatLHBmRNA expression levels remained constant over the oestrous cycle, regardless of the presence of a preovulatory LH surge, which is characteristic of a hormone secreted under regulation. Our sampling regime was unable to detect pulses of LH during the follicular phase, althoughGNRHRmRNA levels had increased at this time. Pulses of LH were, however, detected in the luteal phase of cycling females, in anoestrus females and in males. There was a positive correlation between gene expression ofFSHBand plasma levels of FSH at different stages of the oestrous cycle and no pulses of FSH were detected at any time; all characteristics of a hormone secreted via the constitutive pathway. Usingin situhybridisation and immunohistochemistry methods, we determined that mRNA expression ofLHBandFSHB, and protein storage of gonadotrophins exhibited a similar pattern of localisation within the pituitary gland. Additionally, sexual dimorphism of gonadotroph populations was evident. In summary, these findings are similar to that reported in eutherians and considering that marsupial evolution diverged from eutherians over 100 million years ago suggests that the regulation of gonadotrophins is highly conserved indeed.

scholarly journals Characterization of rat epimorphin/syntaxin 2 expression suggests a role in crypt-villus morphogenesis

1998 ◽  
Vol 275 (1) ◽  
pp. G114-G124 ◽  
Author(s):  
Alka Goyal ◽  
Renu Singh ◽  
Elzbieta A. Swietlicki ◽  
Marc S. Levin ◽  
Deborah C. Rubin
Keyword(s):  
Small Bowel ◽  
Mrna Expression ◽  
Bowel Resection ◽  
Intestinal Adaptation ◽  
Critical Role ◽  
Small Bowel Resection ◽  
Mrna Levels ◽  
Skin Morphogenesis

The rodent intestinal mucosa undergoes a remarkable morphogenesis as the crypt-villus axis is formed. Endoderm-mesenchymal interactions play a critical role in this process. Epimorphin is a mesenchymal protein postulated to play a role in lung and skin morphogenesis. The rat homologue, syntaxin 2, belongs to a family of integral membrane proteins that function in vesicle docking and fusion. To clarify its role in fetal gut morphogenesis, epimorphin expression was examined during ontogeny, in an isograft model of ischemic injury and mucosal repair, and during intestinal adaptation after small bowel resection. Epimorphin/syntaxin 2 mRNA levels were increased in fetal gut during lumen formation and villus morphogenesis. mRNA levels remained elevated in the first 2 wk after birth and then declined at weaning. In situ hybridization showed epimorphin/syntaxin 2 mRNA in gestational day 14( G14) and G15 intestinal mesenchymal cells and in the mucosal lamina propria during villus formation. Epimorphin/syntaxin 2 mRNA expression increased during villus repair in the isograft. In contrast, in the early stages of intestinal adaptation after small bowel resection, epimorphin/syntaxin 2 mRNA expression was suppressed in the adapting gut. We conclude the cell-specific and temporal patterns of epimorphin expression in the models used in this study suggest a role in the morphogenesis of the crypt-villus axis.

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