Acute aggressive behavior perturbates the oxidative status of a wild bird independently of testosterone and progesterone
Introduction
Animals frequently engage in aggressive behaviors to obtain and secure limited resources that can enhance their reproductive success, such as a breeding territory (Beckman and Ames, 1998; Duckworth, 2006; Demas et al., 2007; Rosvall, 2008; Smith and Blumstein, 2008; Clutton-Brock, 2009). However, aggressive behaviors entail costs (e.g., territory loss or injuries), which could partially explain why individuals differ in their degree of aggressiveness. A particularly relevant but understudied cost associated with this behavior concerns the oxidative state of an individual (Costantini et al., 2008; Isaksson et al., 2011). The oxidative state of an individual is determined by the concentration of pro-oxidants (i.e., reactive oxygen species) and antioxidants (i.e., non-enzymatic and enzymatic compounds) present in cells and tissues (reviewed by Costantini, 2019). A change in any of these molecular components in favor of pro-oxidants can lead to damage of biomolecules such as lipids, proteins and DNA (reviewed by Costantini, 2008; Monaghan et al., 2009). This damage can shape life-history decisions of individuals as well as life history traits such as reproduction and longevity (e.g., Finkel and Holbrook, 2000; Costantini, 2008; Monaghan et al., 2009), potentially translating into ecological and evolutionary consequences.
Aggressive behaviors are energetically demanding activities that increase the metabolic rate, exposing an individual to an elevated concentration of pro-oxidants (Fig. 1a; Costantini, 2008; Powers and Jackson, 2008; Skrip and McWilliams, 2016; Cooper-Mullin and McWilliams, 2016; but see Salin et al., 2015). Such perturbations in the oxidative status of animals are expected since more than 90% of the cellular energy is generated by the mitochondria (Bottje, 2015) and natural by-products of aerobic respiration are reactive oxygen species (ROS). However, our understanding of aggression altering the oxidative state is still in its early stages. In selected lines of mice (Mus musculus domesticus), aggressive males had lower levels of antioxidants than non-aggressive males (Costantini et al., 2008). In contrast, aggressive males of wild-caught white skinks (Egernia whitii) had higher levels of non-enzymatic antioxidants than less aggressive males, whereas in females no such relationship was apparent (Isaksson et al., 2011). Acute aggressive behaviors, such as territorial fights, that represent an increase in energy expenditure, could also incur an oxidative challenge; yet it remains unknown whether this is the case.
The oxidative state of an individual can be indirectly altered by aggressive behaviors through an increase in steroid hormone concentrations (Fig. 1b; Schantz et al., 1999; Alonso-Alvarez et al., 2007). The sex steroid testosterone is assumed to be the key hormone related to resource-defense aggression and can increase during aggressive interactions in males (Wingfield et al., 1990; Hirschenhauser and Oliveira, 2006; Goymann et al., 2007; Hau, 2007). Testosterone has been proposed to be a cause of increased ROS production (e.g., Alonso-Alvarez et al., 2007; Koch et al., 2016), which can occur either because testosterone enhances the metabolic rate of individuals (e.g., Marler and Moore, 1989; Welle et al., 1992; Wikelski et al., 1999; Buchanan et al., 2001; Koch et al., 2016) and/or because testosterone has intrinsic oxidative properties (e.g., Zhu et al., 1997; Chainy et al., 2009; Casagrande et al., 2011). An increase in aggressive behavior after a territorial intrusion can, therefore, potentially lead to an increase in testosterone concentrations (e.g., Wingfield and Wada, 1989; Wingfield and Hahn, 1994; McGlothlin et al., 2008), with testosterone levels influencing the concentrations of pro-oxidant and antioxidant compounds and resulting in an overall perturbation of the oxidative state. To date, only one study has looked at the relationship between aggression, oxidative status and testosterone concentrations in a wild-caught vertebrate (Isaksson et al., 2011). Isaksson et al. (2011) found that the aggressive phenotype of male white skinks, but not testosterone concentrations, was positively related to the oxidative condition of individuals. Aggression can also be associated with another steroid hormone: progesterone. Progesterone has been mainly proposed as a mediator of aggressive behavior in females (e.g., Goymann et al., 2008; but see e.g., Adreani et al., 2018). As with testosterone, progesterone can increase the metabolic rate of organisms (Gavrilova-Jordan and Price, 2007) and the concentration of pro-oxidants (only from studies done in vitro e.g., Zhu et al., 1997; Itagaki et al., 2005). Investigating the relationships between aggressive behavior, oxidative status, testosterone and progesterone in females and males is fundamental if we aim to understand the sex-specificity of these physiological pathways.
The main goal of our study was to test the hypothesis that an acute increase in conspecific aggressive behavior can impair the oxidative status of wild rufous horneros (Aves: Furnarius rufus, hereafter termed hornero), and to test whether this was related to testosterone and progesterone concentrations. To elicit an aggressive response, we challenged female and male birds with 20 min of simulated conspecific territorial intrusions (STI) during the nest-building period. Immediately after the STI we collected blood samples to measure the concentrations of three oxidative status markers, testosterone and progesterone. We then compared these concentrations with the ones of birds that did not engage in aggressive behaviors. The hornero represents an excellent model system to study the effects of acute aggression across sexes. It is a seasonal breeder, and both females and males are involved in territorial defense throughout the year (Fraga, 1980; Diniz et al., 2016; Mentesana et al. 2020). Further, horneros are sexually monomorphic in plumage coloration and body size (i.e., there is no difference in body condition between sexes; Diniz et al., 2016), and all breeding behaviors studied so far, except for aggressive behaviors, are shared and coordinated between the sexes (Massoni et al., 2012; Diniz et al., 2018; Mentesana et al. 2020).
Section snippets
Field site and simulated territorial intrusion (STI)
The study was conducted between August 22nd and September 28th of 2016 on the campus of INIA ‘Las Brujas’ (National Institute of Agricultural Research), Department of Canelones, Uruguay (34°40′ S, 56°20′ W; 0–35 m a.s.l.). During this period, horneros were building their nests and females were close to egg laying. In total, we collected behavioral and physiological data from birds defending 51 territories that were subjected to the experimental treatments between 7 am and 1 pm.
We experimentally
Sex differences in the concentrations of oxidative markers, testosterone and progesterone of control birds
Control females had a lower non-enzymatic antioxidant capacity (OXY) than control males (Fig. 2A; p(dif.) > 99.99%, Table S3). Both sexes had similar enzymatic antioxidant concentrations (GPX; Fig. 2B; p(dif.) = 79.03%, Table S3). We found a trend for control males having lower levels of pro-oxidants than females (ROMs; Fig. 2C; p(dif.) = 94.67%, Table S3). Control females had lower testosterone concentrations than control males (Fig. 2D; p(dif.) = 99.86%, Table S3), but we found no difference
Discussion
We experimentally tested if conspecific aggressive behavior induced an acute change in the oxidative state of female and male wild birds, and whether this effect was mediated by testosterone or progesterone. Our results provide the first evidence that an acute increase in conspecific aggressive behavior (i.e., for 20 min) perturbates the oxidative status of an individual by decreasing the concentration of non-enzymatic antioxidants in plasma. This perturbation in oxidative condition was not
Conclusions
Our study provides the first evidence that an acute increase in aggressive behavior perturbates the oxidative state of female and male wild birds. Our study also suggests that the mechanisms underlying these changes differ between sexes. Until now, the effect of aggression on the oxidative state was investigated in the framework of personality traits (Costantini et al., 2008; Isaksson et al., 2011). Altogether, the existing studies suggest that aggressive behaviors performed over both short and
Author Contributions
NMA and LM contributed equally in the study. Both conceived and designed the study, secured funding and analysed the data. LM took lead in writing the initial version of the mauscript and NMA provided critical feedback and helped shape the manuscript.
Ethical statement
The experimental procedures were approved by the Ethics Committee of Animal Experimentation (CEUA) of the Facultad de Ciencias, Universidad de la República, Uruguay (University of the Republic of Uruguay). Protocol number 186, file 2400-11000090-16.
Funding
We acknowledge funding and support from the International Max Planck Research School (IMPRS) for Organismal Biology (Max Planck Institute, Germany), and by Idea Wild (USA) that provided field equipment.
Declaration of competing interest
We declare no competing interests.
Acknowledgements
We thank Enzo Cavalli and Ernesto Guedes for their assistance in the field, and Monika Trappschuh for conducting the hormone analysis. We also thank the members from the Ethology Lab at the Universidad de la República in Montevideo and especially to Bettina Tassino for their support. Also, we thank all the “INIA Las Brujas” staff for providing us with accommodation and equipment during fieldwork. Furthermore, we are thankful to Juan Carlos Reboreda and the Ecology and Behaviour lab (LEyCA) from
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