Relevance and significance
The multifaceted antioxidant system consists of numerous components that collectively provide individuals with a tool-box of ‘antioxidant strategies’ they can use to protect against oxidative damage (Halliwell and Gutteridge 2007, Costantini 2014). Migratory birds can use a combination of non-enzymatic antioxidants gained from their diets and a suite of endogenous antioxidant enzymes and sacrificial molecules to protect against excess reactive species produced by relevant oxidative challenges, including repeated bouts of flight and high-PUFA diets (Skrip and McWilliams 2016, McWilliams et al. 2021). This study demonstrated the ability of a songbird to use OXY during an acute flight and to recover OXY levels within 2 days of rest (Fig. 2), revealing the recovery timeline of antioxidant capacity for the first time (McWilliams et al. 2021). We also found that such rapid adjustments to the antioxidant system allowed birds to maintain overall low levels of circulating damage, although the extent was tissue-dependent. Researchers should remain mindful of the tissue-dependency of antioxidant capacity and oxidative damage demonstrated here and elsewhere (Frawley et al. 2021a) and that future studies that assess oxidative status select biologically relevant tissues to measure that directly relate to the hypotheses of interest.
Individual variation in the ability to utilize antioxidants must exist for phenotypic flexibility of the antioxidant system to persist (Piersma and Van Gils 2011) and we found evidence of such individual-level, condition-dependent modulation of OXY and uric acid during long-duration flight (Fig. S1). A subset of birds that expended energy in the upper 50% range (>0.51 kJ/min) seemed to either deplete OXY or increase levels of uric acid, a potent antioxidant (Fig. 3B and Fig. S1C), whereas the remaining birds did not change these circulating measures (OXY, uric acid) from the averages. Studies, such as those done with mosquitofish (Loughland and Seebacher 2020), that elucidate the antioxidant strategies used by individuals to maintain their oxidative balance under oxidatively challenging conditions and its effects on performance (e.g. temperature acclimation, running speed, breeding success, flight efficiency) would be particularly revealing for migratory songbirds.
The study described here was part of a larger integrated research project that also compared metabolic and antioxidant gene expression and enzyme activities in multiple tissues within the same individuals (citations redacted for review). This allows us to compare how flight training, dietary antioxidants, and dietary PUFA affect the antioxidant system across the traditional scales of the Central Dogma (DNA, to RNA, to protein). All measures except oxidative status in tissues responded strongly to flight training. In the plasma, proteins are subject to posttranslational modifications and so better able to optimal to detect individual strategies to combat oxidative damage in response to flight exercise. In contrast, diet effects on oxidative status were most evident in tissues (i.e., relative gene expression and antioxidant assays) compared to functional assays in the plasma (i.e., markers for antioxidant capacity and oxidative damage) perhaps because the former is more tightly regulated. Our studies also suggest that plasma oxidative capacity is not a good predictor of oxidative damage in tissues, whereas antioxidant and metabolic gene expression and antioxidant enzyme activities are significant predictors of oxidative damage in the pectoralis and liver (citation redacted for initial review). Thus, future studies must carefully consider the appropriate scale of measurement(s) to best reveal how the antioxidant system of wild animals responds to ecologically-relevant oxidative challenges such as migration.