The relationship between mitochondrial respiration, resting metabolic rate and blood cell count in great tits.

Although mitochondrial respiration is believed to explain a substantial part of the variation in resting metabolic rate (RMR), few studies have empirically studied the relationship between organismal and cellular metabolism. We therefore investigated the relationship between RMR and mitochondrial respiration of permeabilized blood cells in wild great tits (Parus major L.). We also studied the correlation between mitochondrial respiration traits and blood cell count, as normalizing mitochondrial respiration by the cell count is a method commonly used to study blood metabolism. In contrast to previous studies, our results show that there was no relationship between RMR and mitochondrial respiration in intact blood cells (i.e. with the ROUTINE respiration). However, when cells were permeabilized and interrelation re-assessed under saturating substrate availability, we found that RMR was positively related to phosphorylating respiration rates through complexes I and II (i.e. OXPHOS respiration) and to the mitochondrial efficiency to produce energy (i.e. net phosphorylation efficiency), though variation explained by the models was low (i.e. linear model: R2=0.14 to 0.21). However, unlike studies in mammals, LEAK respiration without [i.e. L(n)] and with [i.e. L(Omy)] adenylates was not significantly related to RMR. These results suggest that phosphorylating respiration in blood cells can potentially be used to predict RMR in wild birds, but that this relationship may have to be addressed in standardized conditions (permeabilized cells) and that the prediction risks being imprecise. We also showed that, in our conditions, there was no relationship between any mitochondrial respiration trait and blood cell count. Hence, we caution against normalising respiration rates using this parameter as is sometimes done. Future work should address the functional explanations for the observed relationships, and determine why these appear labile across space, time, taxon, and physiological state.

Thermoregulatory consequences of growing up during a heatwave or a cold snap in Japanese quail.

Changes in environmental temperature during development can affect growth, metabolism and temperature tolerance of the offspring. We know little about whether such changes remain to adulthood, which is important to understand the links between climate change, development and fitness. We investigated whether phenotypic consequences of the thermal environment in early life remained in adulthood in two studies on Japanese quail (Coturnix japonica). Birds were raised under simulated heatwave, cold snap or control conditions, from hatching until halfway through the growth period, and then in common garden conditions until reproductively mature. We measured biometric and thermoregulatory [metabolic heat production (MHP), evaporative water and heat loss (EWL, EHL) and body temperature] responses to variation in submaximal air temperature at the end of the thermal acclimation period and in adulthood. Warm birds had lower MHP than control birds at the end of the thermal acclimation period and, in the warmest temperature studied (40°C), also had higher evaporative cooling capacity compared with controls. No analogous responses were recorded in cold birds, although they had higher EWL than controls in all but the highest test temperature. None of the effects found at the end of the heatwave or cold snap period remained until adulthood. This implies that chicks exposed to higher temperatures could be more prepared to counter heat stress as juveniles but that they do not enjoy any advantages of such developmental conditions when facing high temperatures as adults. Conversely, cold temperature does not seem to confer any priming effects in adolescence.

Measuring body temperature in birds - the effects of sensor type and placement on estimated temperature and metabolic rate.

Several methods are routinely used to measure avian body temperature, but different methods vary in invasiveness. This may cause stress-induced increases in temperature and/or metabolic rate and, hence, overestimation of both parameters. Choosing an adequate temperature measurement method is therefore key to accurately characterizing an animal's thermal and metabolic phenotype. Using great tits (Parus major) and four common methods with different levels of invasiveness (intraperitoneal, cloacal, subcutaneous, cutaneous), we evaluated the preciseness of body temperature measurements and effects on resting metabolic rate (RMR) over a 40°C range of ambient temperatures. None of the methods caused overestimation or underestimation of RMR compared with un-instrumented birds, and body or skin temperature estimates did not differ between methods in thermoneutrality. However, skin temperature was lower compared with all other methods below thermoneutrality. These results provide empirical guidance for future research that aims to measure body temperature and metabolic rate in small bird models.

Temperature-dependent Developmental Plasticity and Its Effects on Allen's and Bergmann's Rules in Endotherms.

Ecogeographical rules, describing common trends in animal form across space and time, have provided key insights into the primary factors driving species diversity on our planet. Among the most well-known ecogeographical rules are Bergmann's rule and Allen's rule, with each correlating ambient temperature to the size and shape of endotherms within a species. In recent years, these two rules have attracted renewed research attention, largely with the goal of understanding how they emerge (e.g., via natural selection or phenotypic plasticity) and, thus, whether they may emerge quickly enough to aid adaptations to a warming world. Yet despite this attention, the precise proximate and ultimate drivers of Bergmann's and Allen's rules remain unresolved. In this conceptual paper, we articulate novel and classic hypotheses for understanding whether and how plastic responses to developmental temperatures might contributed to each rule. Next, we compare over a century of empirical literature surrounding Bergmann's and Allen's rules against our hypotheses to uncover likely avenues by which developmental plasticity might drive temperature-phenotype correlations. Across birds and mammals, studies strongly support developmental plasticity as a driver of Bergmann's and Allen's rules, particularly with regards to Allen's rule. However, plastic contributions toward each rule appear largely non-linear and dependent upon: (1) efficiency of energy use (Bergmann's rule) and (2) thermal advantages (Allen's rule) at given ambient temperatures. These findings suggest that, among endotherms, rapid changes in body shape and size will continue to co-occur with our changing climate, but generalizing the direction of responses across populations is likely naive.

Plasticity of mitochondrial function safeguards phosphorylating respiration during in vitro simulation of rest-phase hypothermia.

Many animals downregulate body temperature to save energy when resting (rest-phase hypothermia). Small birds that winter at high latitudes have comparatively limited capacity for hypothermia and so pay large energy costs for thermoregulation during cold nights. Available evidence suggests this process is fueled by adenosine triphosphate (ATP)-dependent mechanisms. Most ATP is produced by oxidative phosphorylation in the mitochondria, but mitochondrial respiration may be lower during hypothermia because of the temperature dependence of biological processes. This can create conflict between increased organismal ATP demand and a lower mitochondrial capacity to provide it. We studied this in blood cell mitochondria of wild great tits (Parus major) by simulating rest-phase hypothermia via a 6°C reduction in assay temperature in vitro. The birds had spent the night preceding the experiment in thermoneutrality or in temperatures representing mild or very cold winter nights, but night temperatures never affected mitochondrial respiration. However, across temperature groups, endogenous respiration was 14% lower in hypothermia. This did not reflect general thermal suppression of mitochondrial function because phosphorylating respiration was unaffected by thermal state. Instead, hypothermia was associated with a threefold reduction of leak respiration, from 17% in normothermia to 4% in hypothermia. Thus, the coupling of total respiration to ATP production was 96% in hypothermia, compared to 83% in normothermia. Our study shows that the thermal insensitivity of phosphorylation combined with short-term plasticity of leak respiration may safeguard ATP production when endogenous respiration is suppressed. This casts new light on the process by which small birds endure harsh winter cold and warrants future tests across tissues in vivo.

A whole blood approach improves speed and accuracy when measuring mitochondrial respiration in intact avian blood cells.

Understanding mitochondrial biology and pathology is key to understanding the evolution of animal form and function. However, mitochondrial measurement often involves invasive, or even terminal, sampling, which can be difficult to reconcile in wild models or longitudinal studies. Non-mammal vertebrates contain mitochondria in their red blood cells, which can be exploited for minimally invasive mitochondrial measurement. Several recent bird studies have measured mitochondrial function using isolated blood cells. Isolation adds time in the laboratory and might be associated with physiological complications. We developed and validated a protocol to measure mitochondrial respiration in bird whole blood. Endogenous respiration was comparable between isolated blood cells and whole blood. However, respiration towards oxidative phosphorylation was higher in whole blood, and whole blood mitochondria were better coupled and had higher maximum working capacity. Whole blood measurement was also more reproducible than measurement on isolated cells for all traits considered. Measurements were feasible over a 10-fold range of sample volumes, although both small and large volumes were associated with changes to respiratory traits. The protocol was compatible with long-term storage: after 24 h at 5°C without agitation, all respiration traits but maximum working capacity remained unchanged, the latter decreasing by 14%. Our study suggests that whole blood measurement provides faster, more reproducible, and more biologically and physiologically relevant (mitochondrial integrity) assessment of mitochondrial respiration. We recommend future studies to take a whole blood approach unless specific circumstances require the use of isolated blood cells.

Variation in breeding phenology in response to climate change in two passerine species.

Increasingly warmer springs have caused phenological shifts in both plants and animals. In birds, it is well established that mean laying date has advanced to match the earlier food peak. We know less about changes in the distribution of egg-laying dates within a population and the environmental variables that determine this variation. This could be an important component of how populations respond to climate change. We, therefore, used laying date and environmental data from 39 years (1983-2021) to determine how climate change affected laying date variation in blue tits (Cyanistes caeruleus) and marsh tits (Poecile palustris), two sympatric passerines with different life histories. Both species advanced mean laying date (0.19-0.24 days per year) and mean laying date showed a negative relationship with maximum spring temperature in both blue and marsh tits. In springs with no clear temperature increase during the critical time window (the time-window in which mean laying date was most sensitive to temperature) start of breeding in blue tits was distributed over a longer part of the season. However, there was no such pattern in marsh tits. Our findings suggest that temperature change, and not necessarily absolute temperature, can shape the variation in breeding phenology in a species-specific manner, possibly linked to variation in life-history strategies. This is an important consideration when predicting how climate change affects timing of breeding within a population.

Experimental warming during incubation improves cold tolerance of blue tit (Cyanistes caeruleus) chicks.

Climate change and increasing air temperature may alter environmental conditions for developing birds, with a range of phenotypic consequences for offspring. The thermal environment during incubation may affect the trade-off between growth and thermoregulation, but the effects of temperature on the ontogeny of endothermy are not fully understood. Therefore, we experimentally tested whether heating the nest cup of Eurasian blue tits (Cyanistes caeruleus) during incubation would influence cold tolerance of the chicks after hatching. Chicks from both heated and control nests showed a decrease in cooling rate with age as they became increasingly endothermic and homeothermic. However, chicks from previously heated nests cooled at a lower rate per unit surface area and from across the whole body. These chicks also had a greater body mass during the first 12 days of life compared with chicks from control nests. Lower cooling rates in heated chicks may reflect greater thermogenic capacity or a reduced surface area to volume ratio owing to a greater body mass. Future projections for climate change predict rising air temperature and increased likelihood of heatwaves, even in temperate regions. Our results indicate that nest microclimate can affect thermoregulation in offspring, and thus may be used to predict some of the future physiological responses of birds to climate change during breeding.

Major population splits coincide with episodes of rapid climate change in a forest-dependent bird.

Climate change influences population demography by altering patterns of gene flow and reproductive isolation. Direct mutation rates offer the possibility for accurate dating on the within-species level but are currently only available for a handful of vertebrate species. Here, we use the first directly estimated mutation rate in birds to study the evolutionary history of pied flycatchers (Ficedula hypoleuca). Using a combination of demographic inference and species distribution modelling, we show that all major population splits in this forest-dependent system occurred during periods of increased climate instability and rapid global temperature change. We show that the divergent Spanish subspecies originated during the Eemian-Weichselian transition 115-104 thousand years ago (kya), and not during the last glacial maximum (26.5-19 kya), as previously suggested. The magnitude and rates of climate change during the glacial-interglacial transitions that preceded population splits in pied flycatchers were similar to, or exceeded, those predicted to occur in the course of the current, human-induced climate crisis. As such, our results provide a timely reminder of the strong impact that episodes of climate instability and rapid temperature changes can have on species' evolutionary trajectories, with important implications for the natural world in the Anthropocene.

Variation in reproductive investment increases body temperature amplitude in a temperate passerine.

Many birds and mammals show substantial circadian variation in body temperature, which has been attributed to fluctuations in ambient temperature and energy reserves. However, to fully understand the variation in body temperature over the course of the day, we also need to consider effects of variation in work rate. We made use of a dataset on body temperature during the resting and active periods in female marsh tits (Poecile palustris) that bred in a temperate area and were subjected to experimental changes in reproductive investment through brood size manipulations. Furthermore, the amplitude increased with daytime, but were unaffected by nighttime, ambient temperature. Amplitudes in females with manipulated broods were 44% above predictions based on inter-specific allometric relationships. In extreme cases, amplitudes were > 100% above predicted values. However, no individual female realised the maximum potential amplitude (8.5 °C, i.e. the difference between the highest and lowest body temperature within the population) but seemed to prioritise either a reduction in body temperature at night or an increase in body temperature in the day. This suggests that body temperature amplitude might be constrained by costs that preclude extensive use of both low nighttime and high daytime body temperatures within the same individual. Amplitudes in the range found here (0.5-6.7 °C) have previously mostly been reported from sub-tropical and/or arid habitats. We show that comparable values can also be found amongst birds in relatively cool, temperate regions, partly due to a pronounced increase in body temperature during periods with high work rate.

Extreme altitudes during diurnal flights in a nocturnal songbird migrant.

Billions of nocturnally migrating songbirds fly across oceans and deserts on their annual journeys. Using multisensor data loggers, we show that great reed warblers (Acrocephalus arundinaceus) regularly prolong their otherwise strictly nocturnal flights into daytime when crossing the Mediterranean Sea and the Sahara Desert. Unexpectedly, when prolonging their flights, they climbed steeply at dawn, from a mean of 2394 meters above sea level to reach extreme cruising altitudes (mean 5367 and maximum 6267 meters above sea level) during daytime flights. This previously unknown behavior of using exceedingly high flight altitudes when migrating during daytime could be caused by diel variation in ambient temperature, winds, predation, vision range, and solar radiation. Our finding of this notable behavior provides new perspectives on constraints in bird flight and might help to explain the evolution of nocturnal migration.

Plumage development and environmental factors influence surface temperature gradients and heat loss in wandering albatross chicks.

Young birds in cold environments face a range of age-specific thermal challenges. Studying the thermal biology of young birds throughout ontogeny may further our understanding of how such challenges are met. We investigated how age and environmental parameters influenced surface temperature gradients across various body regions of wandering albatross (Diomedea exulans) chicks on Bird Island, South Georgia. This study was carried out over a 200 d period during the austral winter, from the end of the brood-guard period until fledging, bridging a gap in knowledge of surface temperature variation and heat loss in developing birds with a long nestling stage in severe climatic conditions. We found that variation in surface temperature gradients (i.e. the difference between surface and environmental temperature) was strongly influenced by chick age effects for insulated body regions (trunk), with an increase in the surface temperature gradient that followed the progression of plumage development, from the second set of down (mesoptiles), to final chick feathers (teleoptiles). Environmental conditions (primarily wind speed and relative humidity) had a stronger influence on the gradients in uninsulated areas (eye, bill) than insulated regions, which we interpret as a reflection of the relative degree of homeothermy exhibited by chicks of a given age. Based on biophysical modelling, total heat loss of chicks was estimated to increase linearly with age. However, mass specific heat loss decreased during the early stages of growth and then subsequently increased. This was attributed to age-related changes in feather growth and activity that increased surface temperature and, hence, metabolic heat loss. These results provide a foundation for further work on the effects of environmental stressors on developing chicks, which are key to understanding the physiological responses of animals to changes in climate in polar regions.

Avian red blood cell mitochondria produce more heat in winter than in autumn.

Endotherms in cold regions improve heat-producing capacity when preparing for winter. We know comparatively little about how this change is fueled by seasonal adaptation in cellular respiration. Thus, we studied the changes of mitochondrial function in red blood cells in sympatric Coal (Periparus ater), Blue (Cyanistes caeruleus), and Great (Parus major) tits between autumn and winter. These species differ more than twofold in body mass and in several aspects of their foraging ecology and social dominance, which could require differential seasonal adaptation of energy expenditure. Coal and Great tits in particular upregulated the mitochondrial respiration rate and mitochondrial volume in winter. This was not directed toward ATP synthesis, instead reflecting increased uncoupling of electron transport from ATP production. Because uncoupling is exothermic, this increased heat-producing capacity at the sub-cellular level in winter. This previously unexplored the route of thermogenesis in birds should be addressed in future work.

Body Temperature and Activity Rhythms Under Different Photoperiods in High Arctic Svalbard ptarmigan (Lagopus muta hyperborea).

Organisms use circadian rhythms to anticipate and exploit daily environmental oscillations. While circadian rhythms are of clear importance for inhabitants of tropic and temperate latitudes, its role for permanent residents of the polar regions is less well understood. The high Arctic Svalbard ptarmigan shows behavioral rhythmicity in presence of light-dark cycles but is arrhythmic during the polar day and polar night. This has been suggested to be an adaptation to the unique light environment of the Arctic. In this study, we examined regulatory aspects of the circadian control system in the Svalbard ptarmigan by recording core body temperature (T b) alongside locomotor activity in captive birds under different photoperiods. We show that T b and activity are rhythmic with a 24-h period under short (SP; L:D 6:18) and long photoperiod (LP; L:D 16:8). Under constant light and constant darkness, rhythmicity in T b attenuates and activity shows signs of ultradian rhythmicity. Birds under SP also showed a rise in T b preceding the light-on signal and any rise in activity, which proves that the light-on signal can be anticipated, most likely by a circadian system.

Low incubation temperature slows the development of cold tolerance in a precocial bird.

Incubating birds trade off self-maintenance for keeping eggs warm. This causes lower incubation temperature in more challenging conditions, with consequences for a range of offspring traits. It is not yet clear how low developmental temperature affects cold tolerance early in life. This is ecologically important because before full thermoregulatory capacity is attained, precocial chicks must switch between foraging and being brooded when their body temperature declines. Hence, we studied how cold tolerance during conditions similar to a feeding bout in the wild was affected by incubation temperature in Japanese quail (Coturnix japonica). Cold-incubated (35.5°C) chicks took the longest to develop, hatched at a smaller size, and remained smaller during their first week of life compared with chicks incubated at higher temperatures (37.0 and 38.5°C). This was reflected in increased cooling rate and reduced homeothermy, probably on account of reductions in both heat-producing capacity and insulation. Lower cold tolerance could exacerbate other temperature-linked phenotypic effects and, hence, also the trade-off between future and current reproduction from the perspective of the incubating parent.

Endocrinology of thermoregulation in birds in a changing climate.

The ability to maintain a (relatively) stable body temperature in a wide range of thermal environments by use of endogenous heat production is a unique feature of endotherms such as birds. Endothermy is acquired and regulated via various endocrine and molecular pathways, and ultimately allows wide aerial, aquatic, and terrestrial distribution in variable environments. However, due to our changing climate, birds are faced with potential new challenges for thermoregulation, such as more frequent extreme weather events, lower predictability of climate, and increasing mean temperature. We provide an overview on thermoregulation in birds and its endocrine and molecular mechanisms, pinpointing gaps in current knowledge and recent developments, focusing especially on non-model species to understand the generality of, and variation in, mechanisms. We highlight plasticity of thermoregulation and underlying endocrine regulation, because thorough understanding of plasticity is key to predicting responses to changing environmental conditions. To this end, we discuss how changing climate is likely to affect avian thermoregulation and associated endocrine traits, and how the interplay between these physiological processes may play a role in facilitating or constraining adaptation to a changing climate. We conclude that while the general patterns of endocrine regulation of thermogenesis are quite well understood, at least in poultry, the molecular and endocrine mechanisms that regulate, e.g. mitochondrial function and plasticity of thermoregulation over different time scales (from transgenerational to daily variation), need to be unveiled. Plasticity may ameliorate climate change effects on thermoregulation to some extent, but the increased frequency of extreme weather events, and associated changes in resource availability, may be beyond the scope and/or speed for plastic responses. This could lead to selection for more tolerant phenotypes, if the underlying physiological traits harbour genetic and individual variation for selection to act on - a key question for future research.

Deep body and surface temperature responses to hot and cold environments in the zebra finch.

Global warming increasingly challenges thermoregulation in endothermic animals, particularly in hot and dry environments where low water availability and high temperature increase the risk of hyperthermia. In birds, un-feathered body parts such as the head and bill work as 'thermal windows', because heat flux is higher compared to more insulated body regions. We studied how such structures were used in different thermal environments, and if heat flux properties change with time in a given temperature. We acclimated zebra finches (Taeniopygia guttata) to two different ambient temperatures, 'cold' (5 °C) and 'hot' (35 °C), and measured the response in core body temperature using a thermometer, and head surface temperature using thermal imaging. Birds in the hot treatment had 10.3 °C higher head temperature than those in the cold treatment. Thermal acclimation also resulted in heat storage in the hot group: core body temperature was 1.1 °C higher in the 35 °C group compared to the 5 °C group. Hence, the thermal gradient from core to shell was 9.03 °C smaller in the hot treatment. Dry heat transfer rate from the head was significantly lower in the hot compared to the cold treatment after four weeks of thermal acclimation. This reflects constraints on changes to peripheral circulation and maximum body temperature. Heat dissipation capacity from the head region increased with acclimation time in the hot treatment, perhaps because angiogenesis was required to reach peak heat transfer rate. We have shown that zebra finches meet high environmental temperature by heat storage, which saves water and energy, and by peripheral vasodilation in the head, which facilitates dry heat loss. These responses will not exclude the need for evaporative cooling, but will lessen the amount of energy expend on body temperature reduction in hot environments.