General and Comparative Endocrinology 206 (2014) 43–50
Contents lists available at ScienceDirect
General and Comparative Endocrinology journal homepage: www.elsevier.com/locate/ygcen
Interplay among nocturnal activity, melatonin, corticosterone and performance in the invasive cane toad (Rhinella marinus) Tim S. Jessop a,⇑, Tim Dempster a, Mike Letnic b, Jonathan K. Webb c a
Department of Zoology, University of Melbourne, Victoria 3010, Australia School of Biological, Earth and Environmental Sciences, The University of New South Wales, NSW 2052, Australia c School of the Environment, University of Technology, Broadway, NSW 2007, Australia b
a r t i c l e
i n f o
Article history: Received 7 August 2013 Revised 26 June 2014 Accepted 14 July 2014 Available online 22 July 2014 Keywords: Sleep–wake cycles Ecology Environmental variation Behavioral flexibility Hormonal plasticity Organismal performance
a b s t r a c t Most animals conduct daily activities exclusively either during the day or at night. Here, hormones such as melatonin and corticosterone, greatly influence the synchronization or regulation of physiological and behavioral cycles needed for daily activity. How then do species that exhibit more flexible daily activity patterns, responses to ecological, environmental or life-history processes, regulate daily hormone profiles important to daily performance? This study examined the consequences of (1) nocturnal activity on diel profiles of melatonin and corticosterone and (2) the effects of experimentally increased acute melatonin levels on physiological and metabolic performance in the cane toad (Rhinella marinus). Unlike inactive captive toads that had a distinct nocturnal melatonin profile, nocturnally active toads sampled under field and captive conditions, exhibited decreased nocturnal melatonin profiles with no evidence for any phase shift. Nocturnal corticosterone levels were significantly higher in field active toads than captive toads. In toads with experimentally increased melatonin levels, plasma lactate and glucose responses following recovery post exercise were significantly different from control toads. However, exogenously increased melatonin did not affect resting metabolism in toads. These results suggest that toads could adjust daily hormone profiles to match nocturnal activity requirements, thereby avoiding performance costs induced by high nocturnal melatonin levels. The ability of toads to exhibit plasticity in daily hormone cycles, could have broad implications for how they and other animals utilize behavioral flexibility to optimize daily activities in response to natural and increasingly human mediated environmental variation. Ó 2014 Elsevier Inc. All rights reserved.
1. Introduction Animals typically exhibit daily sleep–wake cycles with segregated periods of active (e.g. foraging and social interactions) and inactive (e.g. sleep, rest) behavior. The conserved evolution of discreet behavioral partitioning within 24 h activity cycles, suggests animals respond to strong temporal selection from daily variation in environmental and ecological processes to maximise fitness (Daan, 1981). However, despite the generality of discreet daily patterns of activity, animal may also exhibit phase shifts of varying durations in daily activity schedules in response to temporal variation in resources, to accommodate life-history requirements, or because environmental or ecological constraints limit daily activity. The capacity for animals to mediate flexibility (i.e. plasticity) in the timing of daily activity patterns is likely to confer higher ⇑ Corresponding author. E-mail address: [email protected] (T.S. Jessop). http://dx.doi.org/10.1016/j.ygcen.2014.07.013 0016-6480/Ó 2014 Elsevier Inc. All rights reserved.
fitness in temporally variable or rapidly changing environments (Daan, 1981; Cotton and Parker, 2000; Kronfeld-Schor and Dayan, 2003; Webb et al., 2014). The endocrine regulation of sleep wake cycles in diurnal vertebrates (particularly under captive conditions or using laboratory model species) is well understood (Underwood, 1990; Reiter, 1991). Here two hormones, melatonin (peak levels at night) and corticosterone (peak levels pre awakening or during the day) often exhibit opposing daily profiles that regulates the interplay between daily cycles of behavior (e.g. activity), physiology (e.g. metabolism/digestion) and entrainment with environmental timing cues such daily as photo- and thermal-cycles (Firth et al., 1989; Underwood, 1990; Reiter, 1991). In some diurnal species, melatonin sets the phase of the daily activity cycle, but also reduces arousal state that causes inactivity or sleepiness (Underwood, 1990; Hyde and Underwood, 2000; Zhdanova et al., 2001; Azpeleta et al., 2010) and can even inhibit activity (Chiba et al., 1985; Phol, 2000). Diel variation in basal corticosterone
44
T.S. Jessop et al. / General and Comparative Endocrinology 206 (2014) 43–50
levels typically peak before or during the active phase of an animal daily sleep–awake cycle (Breuner et al., 1999; Jessop et al., 2002). The peak in corticosterone up regulates multiple physiological and behavioral processes consistent with increased arousal, activity and metabolism associated with the active period of the sleep– wake cycle (Dallman et al., 1993; Breuner et al., 1999). How then does temporal flexibility in daily activity schedules of animals influence these physiological processes? Some nocturnal species (e.g. rodents) maintain high melatonin and corticosterone levels during periods of peak nocturnal activity (Mendelson et al., 1980; Tobler et al., 1994), whilst others, including nocturnal birds exhibit limited increase in night time plasma melatonin levels (Taniguchi et al., 1993; Wikelski et al., 2006). By contrast, diurnal species that exploit nocturnal activity for seasonal life-history events (e.g. vernal migration in diurnal birds: Gwinner et al., 1993; Gwinner, 1996; Fusani and Gwinner, 2001, nocturnal nesting in green sea turtles: Jessop et al., 2002) facultatively reduce the nocturnal peak in melatonin, nor exhibit any commensurate phase shift in the hormone’s profile. These examples intuitively suggest some capacity for physiological plasticity in regulation of the daily melatonin cycle outside the typical nocturnal cycle exhibited by diurnal species. Because phase shifts in these activities are often highly predictable (e.g. due to seasonal environmental cues) and important, it suggests that selection may act on individuals to resynchronize their endocrine cycles with phase shifts in daily activity to maximise organismal performance (Gwinner et al., 1993; Fusani and Gwinner, 2004, 2005). Otherwise, such animals might, as suggested by human shift work research, be exposed to the broad scale negative health effects that arise from uncoupling between daily behavioral and physiological cycles (Knutsson, 2003; Schernhammer et al., 2003). However, some animals display even greater flexibility, and adjust daily activity in response to short term fluctuations in environmental (e.g. rainfall, temperature) or ecological processes (e.g. food pulses, breeding activities or predation risk). Both diurnal and nocturnal species may vary the onset of daily activities to accommodate local conditions. For example, under field conditions nocturnal animals such as many anurans, bats, and rodents exhibit intermittent periods of nocturnal activity interspersed by nightly bouts of inactivity (Zug and Zug, 1979; Halle and Stenseth, 2000). How these animals regulate their daily melatonin and corticosterone profiles to accommodate highly flexible daily activity cycles is not well understood. Presumably, such animals also require compensation among activity cycles, hormones and performance (Kronfeld-Schor and Dayan, 2003; Fusani et al., 2011). Using field and laboratory experiments we investigated: (1) if animals that utilize flexible activity cycles alter daily hormone profiles of melatonin and corticosterone; and (2) if melatonin affects physiological aspects of exercise and metabolic performance which under ecological conditions could infer fitness costs. We used the cane toad (Rhinella marina), to investigate the interplay among daily hormone profiles, variable activity cycles and performance. Toads are an interesting model for such studies as they exhibit ontogenetic shifts from diurnal to nocturnal activity (Pizzatto et al., 2008). Once nocturnally active, toads exhibit highly variable daily activity patterns, alternating between days with and without nocturnal activity (Zug and Zug, 1979). These activity shifts arise because of inter-daily variation in temperature and rainfall. However, even when conditions are favorable, toads can be inactive to reconcile time lags associated with digestion and gut clearance (Zug and Zug, 1979). Simply toads may alter between days of nocturnal activity and inactivity and such schedules are likely to be highly variably due to dynamic variation in biophysical constraints (e.g. rainfall) and individual foraging success. We predicted that if toads compensate hormone cycles due to daily variation in
nocturnal activity, daily melatonin and corticosterone profiles should match periods of low and high activity, respectively. Further if daily melatonin profiles are indeed altered, does this suggest nocturnal performance compensation to prevent the potentially inhibitory effects of melatonin on activity? Here we tested the effects of exogenous melatonin on two measures or whole organism physiological performance. First we examined the effects of exogenous melatonin on post exercise plasma lactate (a metabolic waste product causing fatigue) and glucose recovery as markers of melatonin’s potential performance costs for toads conducting nocturnal activity. Similar to most vertebrates, toads need to utilize both anaerobic and aerobic metabolisms to facilitate different behaviors appropriate to specific ecological contexts. Typically high performance and intense physical activities associated with escape or social conflict involve anaerobic metabolism (Pough, 1989). Clearance and release of plasma lactate and glucose during such behaviors can have profound effects on animal performance and in turn have important fitness consequences (Pough, 1989). Melatonin has been reported in some studies to have an inhibitory effect on the regulation of intermediate metabolites such as lactate and glucose (Soengas et al., 1996; Prunet-Marcassus et al., 2003). Similarly there is evidence that melatonin can inhibit glucocorticoid and thyroid hormones that also regulate intermediate metabolites necessary for optimal physical performance (Appa-Rao et al., 2001; Saito et al., 2005; Azpeleta et al., 2010). Second, we investigated the effects of melatonin on resting metabolism in toads. Putative effects of melatonin suggest that it has a regulatory role related to energy metabolism through inhibition of thyroid hormones (John et al., 1990; Krotewicz and Lewinski, 1994) or glucocorticoid hormones (Azpeleta et al., 2010). These hormones can have an important stimulatory effect on resting metabolic rate in ecotherms (Chiu and Tong, 1979; John-Alder, 1983, 1990; Durant et al., 2008). Any reduction in metabolic rate through melatonin inhibiting these hormones could again have important performance implications for toads undertaking nocturnal activity. 2. Materials and methods 2.1. Ethics statement All animals were maintained and tested in accordance with the Institutional Animal Care and Use Committee of the University of Queensland (zoo/225/96/urg). 2.2. Study animals For all experiments we captured adults cane toads (130 g), a large and now wide spread terrestrial anuran introduced intro Australia from South America (via Hawaii) in the 1930’s (Phillips et al., 2007). Animals used in our experiments were predominantly males (80%) reflecting the natural sex bias in Australian toad populations. Thus we did not consider toad sex as a potential effect in our statistical analyses. Furthermore, we did not collect animals exhibiting reproductive behavior (e.g. males calling or male/female toads in amplexus) since reproduction can elevate corticosterone levels in Bufoniid toads (e.g. Orchinik et al., 1988; Leary et al., 2004) and could confound measures in our study. Cane toads were studied under field and short term captive conditions and we used individuals sourced from a large population inhabiting the alumni forest located on the University of Queensland campus (Brisbane, Australia; 27°290 S, 153°80 E). This habitat comprises tall dense and closed canopy mesic forest that provides good habitat for toads to forage at night and seek shelter during the day. Immediately adjacent to this forest are large ponds that toads use to mate and rehydrate. The alumni forest is disjunct from the
T.S. Jessop et al. / General and Comparative Endocrinology 206 (2014) 43–50
campus and not influenced by artificial light from university buildings or street lights. The study was conducted in summer during February, where monthly temperature averaged 29.0 °C and 20.6 °C for maximum and minimum daily temperature. The natural photoperiod during our study was 13 h light:11 dark with sun rise at 5:20 am and sun set at 6:40 pm. We conducted the study in summer, when toads are nocturnally active (to facilitate capture and performance experiments), and when daily cycles in plasma melatonin and corticosterone levels are near annual maxima (to facilitate assay measurements; Firth et al., 1989; Moore and Jessop, 2003). 2.3. Field studies To record daily melatonin and corticosterone profiles from toads under natural field conditions we selected a very dark (
Contents lists available at ScienceDirect
General and Comparative Endocrinology journal homepage: www.elsevier.com/locate/ygcen
Interplay among nocturnal activity, melatonin, corticosterone and performance in the invasive cane toad (Rhinella marinus) Tim S. Jessop a,⇑, Tim Dempster a, Mike Letnic b, Jonathan K. Webb c a
Department of Zoology, University of Melbourne, Victoria 3010, Australia School of Biological, Earth and Environmental Sciences, The University of New South Wales, NSW 2052, Australia c School of the Environment, University of Technology, Broadway, NSW 2007, Australia b
a r t i c l e
i n f o
Article history: Received 7 August 2013 Revised 26 June 2014 Accepted 14 July 2014 Available online 22 July 2014 Keywords: Sleep–wake cycles Ecology Environmental variation Behavioral flexibility Hormonal plasticity Organismal performance
a b s t r a c t Most animals conduct daily activities exclusively either during the day or at night. Here, hormones such as melatonin and corticosterone, greatly influence the synchronization or regulation of physiological and behavioral cycles needed for daily activity. How then do species that exhibit more flexible daily activity patterns, responses to ecological, environmental or life-history processes, regulate daily hormone profiles important to daily performance? This study examined the consequences of (1) nocturnal activity on diel profiles of melatonin and corticosterone and (2) the effects of experimentally increased acute melatonin levels on physiological and metabolic performance in the cane toad (Rhinella marinus). Unlike inactive captive toads that had a distinct nocturnal melatonin profile, nocturnally active toads sampled under field and captive conditions, exhibited decreased nocturnal melatonin profiles with no evidence for any phase shift. Nocturnal corticosterone levels were significantly higher in field active toads than captive toads. In toads with experimentally increased melatonin levels, plasma lactate and glucose responses following recovery post exercise were significantly different from control toads. However, exogenously increased melatonin did not affect resting metabolism in toads. These results suggest that toads could adjust daily hormone profiles to match nocturnal activity requirements, thereby avoiding performance costs induced by high nocturnal melatonin levels. The ability of toads to exhibit plasticity in daily hormone cycles, could have broad implications for how they and other animals utilize behavioral flexibility to optimize daily activities in response to natural and increasingly human mediated environmental variation. Ó 2014 Elsevier Inc. All rights reserved.
1. Introduction Animals typically exhibit daily sleep–wake cycles with segregated periods of active (e.g. foraging and social interactions) and inactive (e.g. sleep, rest) behavior. The conserved evolution of discreet behavioral partitioning within 24 h activity cycles, suggests animals respond to strong temporal selection from daily variation in environmental and ecological processes to maximise fitness (Daan, 1981). However, despite the generality of discreet daily patterns of activity, animal may also exhibit phase shifts of varying durations in daily activity schedules in response to temporal variation in resources, to accommodate life-history requirements, or because environmental or ecological constraints limit daily activity. The capacity for animals to mediate flexibility (i.e. plasticity) in the timing of daily activity patterns is likely to confer higher ⇑ Corresponding author. E-mail address: [email protected] (T.S. Jessop). http://dx.doi.org/10.1016/j.ygcen.2014.07.013 0016-6480/Ó 2014 Elsevier Inc. All rights reserved.
fitness in temporally variable or rapidly changing environments (Daan, 1981; Cotton and Parker, 2000; Kronfeld-Schor and Dayan, 2003; Webb et al., 2014). The endocrine regulation of sleep wake cycles in diurnal vertebrates (particularly under captive conditions or using laboratory model species) is well understood (Underwood, 1990; Reiter, 1991). Here two hormones, melatonin (peak levels at night) and corticosterone (peak levels pre awakening or during the day) often exhibit opposing daily profiles that regulates the interplay between daily cycles of behavior (e.g. activity), physiology (e.g. metabolism/digestion) and entrainment with environmental timing cues such daily as photo- and thermal-cycles (Firth et al., 1989; Underwood, 1990; Reiter, 1991). In some diurnal species, melatonin sets the phase of the daily activity cycle, but also reduces arousal state that causes inactivity or sleepiness (Underwood, 1990; Hyde and Underwood, 2000; Zhdanova et al., 2001; Azpeleta et al., 2010) and can even inhibit activity (Chiba et al., 1985; Phol, 2000). Diel variation in basal corticosterone
44
T.S. Jessop et al. / General and Comparative Endocrinology 206 (2014) 43–50
levels typically peak before or during the active phase of an animal daily sleep–awake cycle (Breuner et al., 1999; Jessop et al., 2002). The peak in corticosterone up regulates multiple physiological and behavioral processes consistent with increased arousal, activity and metabolism associated with the active period of the sleep– wake cycle (Dallman et al., 1993; Breuner et al., 1999). How then does temporal flexibility in daily activity schedules of animals influence these physiological processes? Some nocturnal species (e.g. rodents) maintain high melatonin and corticosterone levels during periods of peak nocturnal activity (Mendelson et al., 1980; Tobler et al., 1994), whilst others, including nocturnal birds exhibit limited increase in night time plasma melatonin levels (Taniguchi et al., 1993; Wikelski et al., 2006). By contrast, diurnal species that exploit nocturnal activity for seasonal life-history events (e.g. vernal migration in diurnal birds: Gwinner et al., 1993; Gwinner, 1996; Fusani and Gwinner, 2001, nocturnal nesting in green sea turtles: Jessop et al., 2002) facultatively reduce the nocturnal peak in melatonin, nor exhibit any commensurate phase shift in the hormone’s profile. These examples intuitively suggest some capacity for physiological plasticity in regulation of the daily melatonin cycle outside the typical nocturnal cycle exhibited by diurnal species. Because phase shifts in these activities are often highly predictable (e.g. due to seasonal environmental cues) and important, it suggests that selection may act on individuals to resynchronize their endocrine cycles with phase shifts in daily activity to maximise organismal performance (Gwinner et al., 1993; Fusani and Gwinner, 2004, 2005). Otherwise, such animals might, as suggested by human shift work research, be exposed to the broad scale negative health effects that arise from uncoupling between daily behavioral and physiological cycles (Knutsson, 2003; Schernhammer et al., 2003). However, some animals display even greater flexibility, and adjust daily activity in response to short term fluctuations in environmental (e.g. rainfall, temperature) or ecological processes (e.g. food pulses, breeding activities or predation risk). Both diurnal and nocturnal species may vary the onset of daily activities to accommodate local conditions. For example, under field conditions nocturnal animals such as many anurans, bats, and rodents exhibit intermittent periods of nocturnal activity interspersed by nightly bouts of inactivity (Zug and Zug, 1979; Halle and Stenseth, 2000). How these animals regulate their daily melatonin and corticosterone profiles to accommodate highly flexible daily activity cycles is not well understood. Presumably, such animals also require compensation among activity cycles, hormones and performance (Kronfeld-Schor and Dayan, 2003; Fusani et al., 2011). Using field and laboratory experiments we investigated: (1) if animals that utilize flexible activity cycles alter daily hormone profiles of melatonin and corticosterone; and (2) if melatonin affects physiological aspects of exercise and metabolic performance which under ecological conditions could infer fitness costs. We used the cane toad (Rhinella marina), to investigate the interplay among daily hormone profiles, variable activity cycles and performance. Toads are an interesting model for such studies as they exhibit ontogenetic shifts from diurnal to nocturnal activity (Pizzatto et al., 2008). Once nocturnally active, toads exhibit highly variable daily activity patterns, alternating between days with and without nocturnal activity (Zug and Zug, 1979). These activity shifts arise because of inter-daily variation in temperature and rainfall. However, even when conditions are favorable, toads can be inactive to reconcile time lags associated with digestion and gut clearance (Zug and Zug, 1979). Simply toads may alter between days of nocturnal activity and inactivity and such schedules are likely to be highly variably due to dynamic variation in biophysical constraints (e.g. rainfall) and individual foraging success. We predicted that if toads compensate hormone cycles due to daily variation in
nocturnal activity, daily melatonin and corticosterone profiles should match periods of low and high activity, respectively. Further if daily melatonin profiles are indeed altered, does this suggest nocturnal performance compensation to prevent the potentially inhibitory effects of melatonin on activity? Here we tested the effects of exogenous melatonin on two measures or whole organism physiological performance. First we examined the effects of exogenous melatonin on post exercise plasma lactate (a metabolic waste product causing fatigue) and glucose recovery as markers of melatonin’s potential performance costs for toads conducting nocturnal activity. Similar to most vertebrates, toads need to utilize both anaerobic and aerobic metabolisms to facilitate different behaviors appropriate to specific ecological contexts. Typically high performance and intense physical activities associated with escape or social conflict involve anaerobic metabolism (Pough, 1989). Clearance and release of plasma lactate and glucose during such behaviors can have profound effects on animal performance and in turn have important fitness consequences (Pough, 1989). Melatonin has been reported in some studies to have an inhibitory effect on the regulation of intermediate metabolites such as lactate and glucose (Soengas et al., 1996; Prunet-Marcassus et al., 2003). Similarly there is evidence that melatonin can inhibit glucocorticoid and thyroid hormones that also regulate intermediate metabolites necessary for optimal physical performance (Appa-Rao et al., 2001; Saito et al., 2005; Azpeleta et al., 2010). Second, we investigated the effects of melatonin on resting metabolism in toads. Putative effects of melatonin suggest that it has a regulatory role related to energy metabolism through inhibition of thyroid hormones (John et al., 1990; Krotewicz and Lewinski, 1994) or glucocorticoid hormones (Azpeleta et al., 2010). These hormones can have an important stimulatory effect on resting metabolic rate in ecotherms (Chiu and Tong, 1979; John-Alder, 1983, 1990; Durant et al., 2008). Any reduction in metabolic rate through melatonin inhibiting these hormones could again have important performance implications for toads undertaking nocturnal activity. 2. Materials and methods 2.1. Ethics statement All animals were maintained and tested in accordance with the Institutional Animal Care and Use Committee of the University of Queensland (zoo/225/96/urg). 2.2. Study animals For all experiments we captured adults cane toads (130 g), a large and now wide spread terrestrial anuran introduced intro Australia from South America (via Hawaii) in the 1930’s (Phillips et al., 2007). Animals used in our experiments were predominantly males (80%) reflecting the natural sex bias in Australian toad populations. Thus we did not consider toad sex as a potential effect in our statistical analyses. Furthermore, we did not collect animals exhibiting reproductive behavior (e.g. males calling or male/female toads in amplexus) since reproduction can elevate corticosterone levels in Bufoniid toads (e.g. Orchinik et al., 1988; Leary et al., 2004) and could confound measures in our study. Cane toads were studied under field and short term captive conditions and we used individuals sourced from a large population inhabiting the alumni forest located on the University of Queensland campus (Brisbane, Australia; 27°290 S, 153°80 E). This habitat comprises tall dense and closed canopy mesic forest that provides good habitat for toads to forage at night and seek shelter during the day. Immediately adjacent to this forest are large ponds that toads use to mate and rehydrate. The alumni forest is disjunct from the
T.S. Jessop et al. / General and Comparative Endocrinology 206 (2014) 43–50
campus and not influenced by artificial light from university buildings or street lights. The study was conducted in summer during February, where monthly temperature averaged 29.0 °C and 20.6 °C for maximum and minimum daily temperature. The natural photoperiod during our study was 13 h light:11 dark with sun rise at 5:20 am and sun set at 6:40 pm. We conducted the study in summer, when toads are nocturnally active (to facilitate capture and performance experiments), and when daily cycles in plasma melatonin and corticosterone levels are near annual maxima (to facilitate assay measurements; Firth et al., 1989; Moore and Jessop, 2003). 2.3. Field studies To record daily melatonin and corticosterone profiles from toads under natural field conditions we selected a very dark (
