Regulation of Rhesus glycoprotein-related genes in large-scale loach Paramisgurnus dabryanus during ammonia loading.

Abstract

Waterborne ammonia is one of the crucial issues that limited production and animal health in aquaculture. Ammonia-tolerant varieties are highly desired in intensive fish farming. Screening for the key regulatory genes of ammonia tolerance is essential for variety breeding. According to the previous hypothesis, Rh glycoproteins play an important role in ammonia excretion in teleosts. However, the ammonia defensive mechanisms are not well described at present for large-scale loach (Paramisgurnus dabryanus), a typical air-breathing and commercially important fish in East Asia. Here we show that the transcription of Rh glycoprotein-related genes was significantly affected by ammonia exposure in this species. Probit analysis showed that 96h-LC50 of NH4Cl at 23℃ and pH 7.2 was 92.64mmol/L. A significant increase of Rhcg expression in gills was observed after 48h of 60mmol/L and 36h of 80mmol/L NH4Cl exposure, suggesting that Rhcg present on the apical side of the branchial epithelium facilitates NH3 excretion out of gills. A high concentration of acute ammonia exposure induced elevated Rhbg transcript in the gills of large-scale loaches, while a slight change in Rhbg expression was observed in response to lower ammonia, suggesting that transcriptions of Rhbg genes are activated by a considerably high level of ambient ammonia to eliminate excessive endogenous nitrogen. The Rhag mRNA level in gills of large-scale loaches increased markedly with the prolonging of exposure time from 0 to 36h of ammonia loading, suggesting Rhag localized in gills may be primarily associated with ammonia handling. During 7-21 days of ammonia exposure, the expression of most Rh glycoproteins-related genes in the gills decreased, indicating that the functional role of Rh glycoproteins is not primarily associated with ammonia defense over a long period (more than 7 days). Although a significant transcript of Rhbg was found in the skin of a large-scale loach, the lack of Rhcg and down-regulation of Rhag may indicate that the skin is not an essential location of ammonia excretion, at least when submerged to high levels of ammonia in the environment. In conclusion, Rh glycoproteins localized in gills as ammonia transporters play a momentous role in ammonia detoxification in this species during acute ammonia loading. However, it does not show a positive function during long-term ammonia exposure. Furthermore, the physiological function of Rh glycoproteins localized in the skin is still unclear and deserves further study.