Abstract
The air-breathing singhi catfish (Heteropneustes fossilis) is frequently being challenged by bacterial contaminants, and different environmental insults like osmotic, hyper-ammonia, dehydration and oxidative stresses in its natural habitats throughout the year. The main objectives of the present investigation were to determine (a) the possible induction of inducible nitric oxide synthase (iNOS) gene with enhanced production of nitric oxide (NO) by intra-peritoneal injection of lipopolysaccharide (LPS) (a bacterial endotoxin), and (b) to determine the effects of hepatic cell volume changes due to anisotonicity or by infusion of certain metabolites, stress hormones and by induction of oxidative stress on production of NO from the iNOS-induced perfused liver of singhi catfish. Intra-peritoneal injection of LPS led to induction of iNOS gene and localized tissue specific expression of iNOS enzyme with more production and accumulation of NO in different tissues of singhi catfish. Further, changes of hydration status/cell volume, caused either by anisotonicity or by infusion of certain metabolites such as glutamine plus glycine and adenosine, affected the NO production from the perfused liver of iNOS-induced singhi catfish. In general, increase of hydration status/cell swelling due to hypotonicity caused decrease, and decrease of hydration status/cell shrinkage due to hypertonicity caused increase of NO efflux from the perfused liver, thus suggesting that changes in hydration status/cell volume of hepatic cells serve as a potent modulator for regulating the NO production. Significant increase of NO efflux from the perfused liver was also observed while infusing the liver with stress hormones like epinephrine and norepinephrine, accompanied with decrease of hydration status/cell volume of hepatic cells. Further, oxidative stress, caused due to infusion of t-butyl hydroperoxide and hydrogen peroxide separately, in the perfused liver of singhi catfish, resulted in significant increase of NO efflux accompanied with decrease of hydration status/cell volume of hepatic cells. However, the reasons for these cell volume-sensitive changes of NO efflux from the liver of singhi catfish are not fully understood with the available data. Nonetheless, enhanced or decreased production of NO from the perfused liver under osmotic stress, in presence of stress hormones and oxidative stress reflected its potential role in cellular homeostasis and also for better adaptations under environmental challenges. This is the first report of osmosensitive and oxidative stress-induced changes of NO production and efflux from the liver of any teleosts. Further, the level of expression of iNOS in this singhi catfish could also serve as an important indicator to determine the pathological status of the external environment.
Highlights
Nitric oxide (NO), one of the smallest known bioactive products of cells, is known to act as an intra- and extracellular mediator of various cell functions [1,2]
The inducible nitric oxide synthase (iNOS) activity, which could not be detected in control fish by the assay method used, was able to detect in fish after 12 and 24 h of LPS injection (Fig 2)
In addition to reports on various biochemical adaptations related to nitrogen metabolism in singhi catfish under environmental stresses [40], the role of NO has been emphasized in this fish under ammonia stress during exposure to high external ammonia and during desiccation stress by our group [45,55]
Summary
Nitric oxide (NO), one of the smallest known bioactive products of cells, is known to act as an intra- and extracellular mediator of various cell functions [1,2]. Presence of very efficient volume regulatory mechanisms have been demonstrated in hepatocytes of air-breathing catfish (Clarias batrachus) but the hepatocytes were found to remain in partly swollen or shrunken state as long as they were exposed to anisotonicity [15]. These minute changes of cell volume owing to anisotonicity have been reported to cause changes in carbohydrate metabolism [16,17,18], autophagic proteolysis [19] and protein synthesis [20] in this air-breathing catfish. Fishes represent excellent models to identify and understand elements and mechanisms controlling the physiological and behavioral changes that occur in response to osmotic and oxidative stresses [22,24]
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