MRNA expression analysis of the physiological responses to ammonia infusion in rainbow trout

Abstract

We recently reported that tissue levels of Rhesus (Rh) mRNA in rainbow trout changed in response to high-external ammonia (HEA). To investigate whether or not these changes could be due to elevated plasma ammonia levels, we infused rainbow trout for 12 h with 140 mmol L(-1) NH(4)HCO(3), or with 140 mmol L(-1) NaCl as a control for the effects of infusion. We also analyzed the effects of dorsal aortic catheterization alone, without infusion. Catheterization alone resulted in an elevated ammonia excretion rate, a downregulation of Rhbg mRNA in the brain, and mRNA upregulations of Rhbg, Rhcg1, and Rhcg2 in the gill, Rhbg and Rhcg1 in the skin, and Rhag in the erythrocytes. In NH(4)HCO(3)-infused fish, plasma cortisol peaked at 6 h, erythrocyte Rhag mRNA was downregulated, gill Rhbg, Rhcg1, and Rhcg2 mRNA were upregulated, and skin Rhbg mRNA was also upregulated. NaCl infusion resulted in elevated plasma ammonia and ammonia excretion rates as well as gill mRNA upregulations of Rhbg, carbonic anhydrase, NHE2, H(+)-ATPase, Na(+)/K(+)-ATPase. Taken together, the results indicated that infusion of NH(4)HCO(3) induced a similar pattern of Rh transcript changes as that seen when fish were exposed to HEA. Second, catheterization alone, as well as isotonic NaCl infusion, significantly altered mRNA levels, highlighting the necessity for careful data interpretation and inclusion of appropriate controls for gene expression studies in fish that have undergone anaesthesia/surgery and infusion procedures. Finally, elevated plasma ammonia and cortisol may both be involved in the signaling mechanism for Rh gene regulation.