Introduction
The mechanisms of ammonia excretion and its associated toxicity have been extensively studied in freshwater fish; however, comparable investigations in anadromous species, particularly during early developmental stages of Atlantic salmon ( Salmo salar ), remain limited. Elevated environmental ammonia is highly toxic to fish, disrupting critical physiological processes such as osmoregulation, respiration, metabolism, neurotransmission, and cellular function, which can compromise health and even lead to mortality. However, exposure to high environmental ammonia during embryonic development may induce phenotypic plasticity, potentially conferring long-term benefits for stress resilience later in life. This study aimed to elucidate the patterns of ammonia transport in Atlantic salmon embryos and larvae up to 976 day-degrees (DD) post-fertilization. A secondary objective was to characterize the mRNA expression of key ammonia transporters (Rhag , Rhbg , and Rhcg), the urea transporter (UT), H+-ATPase, Na+/K+-ATPase, and sodium-hydrogen exchangers (NHE3 and NHE2) during early development under exposure to three sub-lethal ammonia concentrations . In addition, the study investigated whether ammonia exposure affects the cellular stress response by analyzing the mRNA expression of heat shock proteins HSP70 and HSP90.
Materials and methods
An ammonia exposure trial was conducted by fertilising a batch of Atlantic salmon eggs followed by the exposure to three levels of ammonia (low: 22 µg/L, medium: 75 µg/L, and high: 148 µg/L ) until the stage prior to start feeding (976 DD). Samples were taken from six developmental timepoints for mRNA expression analysis of important genes responsible for ammonia transport, urea excretion and cellular stress response . The spatial localization of ammonia transporters was also examined at two key developmental stages: hatching (525 DD) and near complete yolk absorption (900 DD), using immunohistochemistry.
Results and Discussion
The results demonstrated that Atlantic salmon embryos are primarily ammoniotelic throughout development, with ammonia serving as the predominant nitrogenous waste product. T he mRNA expression of Rhesus glycoprotein (Rh) genes, particularly Rhag and Rhcg, as well as the UT gene, showed strong alignment with the ontogeny of ammonia and urea excretion rates observed in other salmonid species. However, exposure to elevated ammonia levels significantly altered the expression patterns of these transporters suggesting that environmental ammonia influences the regulation of nitrogenous waste excretion.
Spatial localization revealed that Rhag was expressed in the developing gills, yolk sac, and operculum, while Rhbg was localized to the gills and oral cavity. Rhcg was detected in the sub-operculum and skin. These findings indicate that ammonia excretion is the primary mechanism of nitrogenous waste elimination during early development in Atlantic salmon, with Rhesus glycoproteins playing a pivotal role in ammonia transport via the gills and yolk sac. This process is likely facilitated by the coordinated activity of H+-ATPase, Na+/K+-ATPase, and sodium-hydrogen exchangers.
Furthermore, the upregulation of HSP70 and HSP90 in response to elevated ammonia concentrations suggests that ammonia exposure induces a cellular stress response during early development. This highlights the potential physiological impacts of environmental ammonia on developing salmon and underscores the importance of understanding how early exposure shapes stress-related molecular pathways.
Conclusion
This study reveals that elevated environmental ammonia regulates the early development of ammonia transport and excretion mechanisms in Atlantic salmon. Early exposure to ammonia alters the molecular responses of developing embryos, potentially influencing their ability to handle stress later in life. These findings provide valuable insights into the physiological and molecular adaptations of Atlantic salmon to environmental stressors during critical developmental stages.
Acknowledgements
This work was supported by the Norwegian Research Council No. 331892 (CandRAS).