Tilapia (Oreochromis mossambicus) brain cells respond to hyperosmotic challenge by inducing myo-inositol biosynthesis

University of California, Davis, USA.
Journal of Experimental Biology (Impact Factor: 2.9). 09/2013; 216(24). DOI: 10.1242/jeb.088906
Source: PubMed


This study aimed to determine the regulation of the de novo myo-inositol biosynthetic (MIB) pathway in Mozambique tilapia (Oreochromis mossambicus) brain following acute (25 parts per thousand (ppt)) and chronic (30, 60, 90ppt) salinity acclimations. The MIB pathway plays an important role in cells for accumulating the compatible osmolyte, myo-inositol, in response to hyperosmotic challenge and consists of two enzymes, myo-inositol phosphate synthase and inositol monophosphatase. In tilapia brain, MIB enzyme transcriptional regulation was found to robustly increase in a time (acute acclimation) or dose (chronic acclimation) dependent manner. Blood plasma osmolality, Na(+), and Cl(-) concentrations were also measured and found to significantly increase in response to both acute and chronic salinity challenges. Interestingly, highly significant positive correlations were found between MIB enzyme mRNA and blood plasma osmolality in both acute and chronic salinity acclimations. Additionally, a mass spectrometry assay was established and used to quantify total myo-inositol concentration in tilapia brain, which closely mirrored the hyperosmotic MIB pathway induction. Thus, myo-inositol is a major compatible osmolyte that is accumulated in brain cells when exposed to acute and chronic hyperosmotic challenge. These data show that the MIB pathway is highly induced in response to environmental salinity challenge in tilapia brain and that this induction is likely prompted by increases in blood plasma osmolality. Because the MIB pathway uses glucose-6-phsosphate as a substrate and large amounts of myo-inositol are being synthesized, our data also illustrate that the MIB pathway likely contributes to the high energetic demand posed by salinity challenge.

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    • "way is strongly activated in multiple tissues of O . mossambicus and tilapia larvae already withstand severe osmotic stress immediately after hatching we investigated the effect of salinity acclimation on this pathway in tilapia larvae . Both transcript variants of MIPS ( MIPS160 , MIPS250 ) that were previously detected in adult tilapia tissues ( Gardell et al . , 2013 ; Sacchi et al . , 2013 ) are also present in larvae ."
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    ABSTRACT: The myo-inositol biosynthesis (MIB) pathway converts glucose-6-phosphate to the compatible osmolyte myo-inositol, which protects cells from salinity stress. We exposed tilapia larvae just after yolk sac resorption to various hypersaline environments and recorded robust induction of the enzymes that constitute the MIB pathway, myo-inositol-phosphate synthase (MIPS), and inositol monophosphatase 1 (IMPA1). Strong up-regulation of these enzymes is evident at both mRNA (quantitative real-time PCR) and protein (densitometric analysis of Western blots) levels. The highest level of induction of these enzymes occurs at the highest salinity that larvae were exposed to (90 ppt). Less severe salinity stress causes a proportionately reduced induction of the MIB pathway. Two distinct MIPS mRNA variants are present in tilapia larvae and both are induced at comparable levels for all the salinity challenges tested (34, 70, and 90 ppt). Immunohistochemical localization of IMPA1 protein in sagittal sections of salinity stressed and control larvae identified tissues that are particularly potent in inducing the MIB pathway. These tissues include the skin (epidermis), gills, eye (ciliary epithelium) and heart. In particular, the epidermis directly facing the external milieu showed a very strong induction of IMPA1 immunoreactivity. IMPA1 induction in response to salinity stress was not observed in other tissues suggesting that tilapia larvae may also utilize compatible organic osmolytes other than solely myo-inositol for osmoprotection. We conclude that the MIB pathway plays an important role in protecting multiple (but not all) tissues of tilapia larvae from hyperosmotic salinity stress. J. Exp. Zool. 9999A: XX–XX, 2014. © 2014 Wiley Periodicals, Inc.
    Journal of Experimental Zoology Part A Ecological Genetics and Physiology 10/2014; 321(8). DOI:10.1002/jez.1878 · 1.44 Impact Factor
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    • "Greater expression of IMPA1 relative to MIPS-160 is similar to what is reported in vivo [24], [27]. We found that the 700 mOsm/kg response of MIPS and IMPA mRNA abundances in the brain-derived OmB cell line was more similar to the in vivo response of brain tissue [27] than the 450 mOsm/kg, physiologically relevant treatment. However, the delay in MIB pathway induction in culture is likely a direct result of inhibition of critical cell functions by severe osmotic stress (see above). "
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    ABSTRACT: Fish cell cultures are becoming more widely used models for investigating molecular mechanisms of physiological response to environmental challenge. In this study, we derived two immortalized Mozambique tilapia (Oreochromis mossambicus) cell lines from brain (OmB) and lip epithelium (OmL), and compared them to a previously immortalized bulbus arteriosus (TmB) cell line. The OmB and OmL cell lines were generated without or with Rho-associated kinase (ROCK) inhibitor/3T3 feeder layer supplementation. Although both approaches were successful, ROCK inhibitor/feeder layer supplementation was found to offer the advantages of selecting for epithelial-like cell type and decreasing time to immortalization. After immortalization (≥ passage 5), we characterized the proteomes of the newly derived cell lines (OmB and OmL) using LCMS and identified several unique cell markers for each line. Subsequently, osmotolerance for each of the three cell lines following acute exposure to elevated sodium chloride was evaluated. The acute maximum osmotolerance of these tilapia cell lines (>700 mOsm/kg) was markedly higher than that of any other known vertebrate cell line, but was significantly higher in the epithelial-like OmL cell line. To validate the physiological relevance of these tilapia cell lines, we quantified the effects of acute hyperosmotic challenge (450 mOsm/kg and 700 mOsm/kg) on the transcriptional regulation of two enzymes involved in biosynthesis of the compatible organic osmolyte, myo-inositol. Both enzymes were found to be robustly upregulated in all three tilapia cell lines. Therefore, the newly established tilapia cells lines represent valuable tools for studying molecular mechanisms involved in the osmotic stress response of euryhaline fish.
    PLoS ONE 05/2014; 9(5):e95919. DOI:10.1371/journal.pone.0095919 · 3.23 Impact Factor
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    ABSTRACT: Through the experiments presented here we wanted to test whether egg production of the black-chinned tilapia Sarotherodon melanotheron heudelotii under experimental conditions varies as a function of ambient salinity (fresh waters vs. sea waters vs. hypersaline waters, 0, 35 and 70, respectively) and whether these responses differ between fish acclimated within a few weeks from fresh water to saline and hypersaline environments (experiments E1 and E2, monitoring over 10 and 18 weeks), and individuals born and raised all life long at the experimental salinities (E3, monitoring over 18 weeks). In total, 233 spawns were collected. In each of the three experiments, the reproductive investment (gram of egg per gram of female over 2 weeks) did not differ between salinities of 0 and 35, whereas it was 2-3 times lower at 70 than at 0-35, because of lower spawning frequency (E1-E3), smaller clutch size (E1) and lower spawn mass (E1-E3). Finally, fish acclimated to salinity from fresh water over a few weeks and those maintained at a particular salinity all life long showed similar reproductive traits, thereby emphasizing the remarkable physiological plasticity of this species.
    Aquatic Living Resources 04/2014; 27(2-2):63-72. DOI:10.1051/alr/2014008 · 1.01 Impact Factor
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