Abstract

The development of decompression sickness in humans is linked to inflammatory responses to nitrogen bubbles which form during ascent (decompression). Bubble formation does not always lead to symptoms however, and activity level prior to or during a dive may alter the susceptibility of an individual by priming immune responses. Observations of gas bubble injury in stranded and by-caught marine mammals have renewed questions concerning bubble formation. Should bubbles be more common than previously thought, it has been suggested that a less-reactive immune response is one mechanism by which marine mammals avoid injury under normal circumstances. In 2017, our lab reported that various in vitro exposures to nitrogen bubbles did not result in significant activation of inflammatory responses, measured as an increase in C5a, an important protein in the complement cascade, in belugas (Delphinapterus leucas).¹ As a continuation of that work, this project measures in vivo changes in C5a related to activity level during diving, and evaluated the potential for activity to modulate the complement response to nitrogen bubbles in vitro. A beluga at Mystic Aquarium was trained to perform two contrasting dive behaviors for durations of 3 minutes each: 1) active diving; submerged swimming, with active fluking, and 2) stationary diving; submergence with minimal body movement. Each behavior was repeated 3 times. Blow samples were collected before, immediately following and 1 hour following each dive behavior. Blood samples were only obtained immediately following each dive. Baseline blood samples were obtained during routine monthly health checks. Plasma catecholamines were quantified using high performance liquid chromatography, and C5a was measured in both serum and blow using a commercial harbor seal (Phoca vitulina) EIA kit (MyBiosource.com) which we previously validated for use in belugas. Due to a sample size of one, statistical analyses were deemed inappropriate, yet interesting patterns were observed. Norepinephrine was higher following both dive behaviors as compared with monthly baseline values. Dopamine was detectable only following active dives, and was higher than in baseline months. Serum C5a was higher following both dive behaviors as compared with monthly baseline values, with the largest difference observed following stationary dives. Most notably, large increases in serum C5a were detected following exposure to nitrogen bubbles following both dive behaviors that were not seen in baseline samples previously tested (n=7). Minimal changes were observed in blow C5a for either dive behavior; however, similar to blood results, the effect of stationary dives seemed to be slightly greater than active dives. While the small scope of this study limits definitive conclusions, it is, to our knowledge, the first report of catecholamine measures in a beluga following specific dive behaviors. That dopamine was detectable only following active dives may suggest activation of “reward” systems in the nervous system during the performance of this behavior. The observed changes in C5a suggest that breath hold and/or activity level during a dive may play roles in modulating immune responsiveness. Such information is relevant to understanding how anthropogenic stressors may alter susceptibility of marine mammals to dive related injury.

Acknowledgements

The authors especially thank the beluga husbandry staff at Mystic Aquarium for training of the dive behaviors, and participation in this study, particularly Carey Richard, Lindsey Nelson and Kathryn Justice. Thank you also to the veterinary team at Mystic Aquarium for carrying out blood sampling after each dive. This work was conducted under Mystic Aquarium IACUC protocol #15004. Funding for this work was provided by the Office of Naval Research (ONR) award #N000141512203.

* Presenting author
+ Student presenter

Literature Cited

1.  Thompson LA, Goertz CEC, Hobbs R, Romano, TA. 2017. The complement cascade in marine mammals as a mechanism to avoid damage from nitrogen bubbles during diving. IAAAM 48th Annual Conference Proceedings, Cancun, Mexico.