Sedation and slaughter of Atlantic salmon (Salmo salar, L.) with carbon monoxide, and a possible regulatory role of neuroglobin

Abstract

Until recently CO2 has been utilized as an anesthetic method before gill cutting of Norwegian Atlantic salmon farmed fish. From 1. July 2012 this is no longer permitted in Norway since it causes strong aversive reactions and severe stress to the animal. In the search for alternative methods for sedation and anesthesia carbon monoxide (CO) has gained interest. In our project, studies demonstrate that CO treatment has a positive effect on product quality and animal welfare. The use of CO in slaughtering, processing and packaging of muscle foods has been reviewed. The present thesis focuses on how sedation of Atlantic salmon with CO affects animal welfare and quality of fish when used as an alternative method prior to anaesthetization. CO is widely used in meat and fish industry for smoking of fish or packaging of meat to maintain product quality. In this study CO was used to sedate salmon by diffusion of the gas in seawater tanks with fish. The level of CO in the seawater was estimated on the basis of oxygen level measured in the seawater and the length of time of diffusion. The methodology to measure CO in water by use of GC-chromatography was not until recently available in our laboratories. Behavior analysis and cortisol levels indicate that CO treatment may result in a more calm fish, not showing increased cortisol values compared to other treatments but rather a tendency of the opposite. However, at high levels of CO a more rapid swimming activity was observed. Another effect was a more rapid onset of rigor mortis and a faster decrease in pH observed in CO exposed salmon compared to the control group. Early onset of rigor mortis could indicate stressed fish, but in this case a more likely explanation is the anaerobic metabolism as a secondary response of CO treatment. Moreover, fillet from CO treated salmon showed a higher degree of red color. Mackerel fillets exposed to CO also showed a perceptible more red color. Both fillets of salmon and mackerel are rich in Mb compared to e.g. cod fishes. In the search for factors with a controlling effect on CO we focused on a recently discovered globin located in nerve tissue, neuroglobin (Ngb). Ngb has been shown to protect the brain during stress and hypoxia in fish. Ngb was detected in salmon brain tissue, but only in small amounts, 1-10 μg/g, after immune prespitation and western analysis. Thus in order to study salmon neuroglobin (sNgb) in more detail, we cloned and expressed the protein in E. coli. Recombinant sNgb showed a strong response using a salmon Ngb specific antibody, raised against the C-terminal peptide of sNgb. Homology modeling of 3D structures of sNgb and mouse neuroglobin indicated that sNgb has a more flexible structure compared to the mammalian neuroglobins that have typical psychrophilic characteristics.