Acute- and long-term flight stimulated antioxidant protection
(H1)
We found evidence that repeated bouts of flight initiated a hormetic
response that activated the antioxidant system to protect against the
accumulation of oxidative damage, consistent with H1. This concept of
hormesis, the mild exposure to reactive species and subsequent
activation of protective and repair mechanisms, has been demonstrated
mainly in the skeletal muscle of humans (McArdle et al. 2002, Rattan
2008). In the present study, the immediate, acute effects of a flight
included a reduction in both non-enzymatic antioxidant capacity as well
as oxidative damage (PF-AF; Fig. 2A, B), consistent with the
acute-effects of flight hypothesis (H1a). The coincident reduction in
both antioxidant capacity and oxidative damage suggests that birds
effectively avoided oxidative damage caused by short-term exercise at
least in part by using the non-enzymatic component of their antioxidant
system. Similarly, birds exposed to 15 days of flight-training and two
days of recovery were able to decrease circulating damage to lipids
(RC-BG; Fig. 2A) while untrained birds were not (long-term flight
effect, H1b). The apparent over-compensation of the antioxidant system
to enable a decrease in oxidative damage during a given flight (H1a)
seems novel to this study, as some flight-training studies report no
change in markers of circulating damage after an acute flight (Skrip et
al. 2016, Cooper-Mullin et al. 2019). Most studies report increases in
markers of circulating oxidative damage shortly after a migratory flight
in free-living birds (Jenni-Eiermann et al. 2014, Costantini et al.
2018), after a 200 km experimental flight in homing pigeons (Costantini
et al. 2008), or in the pectoralis after an experimental flight in a
wind tunnel (Dick and Guglielmo 2019). We found that the reduction in
both non-enzymatic antioxidant capacity as well as oxidative damage was
more apparent in birds that expended more energy per unit time during
their longest flight while flying at a prescribed pace (Fig. 3). Perhaps
flying birds that expend less energy per unit time are less
metabolically challenged, and thus do not need to deplete OXY as much to
prevent oxidative damage. This conclusion is similar in concept to (Dick
and Guglielmo 2019) who found less pectoralis damage in Yellow-rumped
warblers with lower flight energy expenditures.
Consistent with the flight effects proposed in H1b, regular daily flying
depleted non-enzymatic antioxidant capacity with an associated decrease
in oxidative damage immediately after a flight; although both
non-enzymatic antioxidant capacity and oxidative damage returned to
baseline levels after 48 hrs of rest (without wind tunnel flying). These
results are in accordance with flight-trained zebra finches that also
decreased OXY during 2-hrs of acute flight and then increased OXY after
birds had a reprieve from regular flying (Cooper-Mullin et al. 2019).
This consistent and relatively rapid (no more than a few days) depletion
and recovery of non-enzymatic antioxidant capacity suggests that
stopovers during migration may be important in allowing birds to
maintain their oxidative status over the course of the entire migration
(McWilliams et al. 2021). In support of this, plasma d-ROMs decreased
with increasing stopover duration (0-8 nights) in Garden warblersSylvia borin , and plasma non-enzymatic antioxidant capacity
increased with fat stores accumulated at a stopover site in Blackpoll
Warblers Setophaga striata and Red-eyed Vireos Vireo
olivaceus (Skrip et al. 2015). Importantly, after 15 days of exercise
training, European starlings exposed to an acute flight further depleted
their non-enzymatic antioxidant capacity without an associated increase
in oxidative damage. This provides additional evidence that birds during
migration seem able to rapidly adjust their antioxidant system to
maintain overall low levels of circulating damage.
We also found evidence in the liver and pectoralis that flight training
activated the antioxidant system so that after several weeks of daily
flying the non-enzymatic antioxidant capacity was maintained at levels
similar to those of untrained birds, and oxidative damage was prevented
in all tissues (H1c). Repeated bouts of flight apparently protected
birds against lipid damage in the liver and the pectoralis (Frawley et
al. 2021a), as lipid hydroperoxide concentration was lower or similar in
the liver and pectoralis, respectively, of flight-trained birds compared
to untrained birds. In our companion study (citation redacted for
initial review), the gene expression of CAT , SOD2 , andGPX1 were upregulated in the liver of flight-trained birds, while
only SOD2 was upregulated in the pectoralis, which likely
explains how lower liver damage levels in flight-trained birds was
achieved. This enzymatic upregulation combined with lower damage in the
liver suggests that the liver is preferentially protected compared to
the pectoralis, perhaps to preserve its crucial role in processing fatty
acids during flight or to protect those fatty acids from degradation.
There were also no differences detected in plasma OXY or liver or
pectoralis hydroxyl and peroxyl scavenging capacities between
flight-trained and untrained birds after 48 hrs of rest; thus birds were
able to maintain constant long-term levels of antioxidant capacity when
energetically challenged and avoid increases in oxidative damage.
Although long-term antioxidant capacity in the tissues was unchanged, it
is possible that flight-trained birds were able to prevent the
accumulation of oxidative damage by utilizing non-enzymatic antioxidant
capacity in tissues during flight, as also shown for OXY in the plasma,
and by increasing the gene expression of enzymatic antioxidants
(citation redacted for initial review).