Solving the conundrum of intra-specific variation in metabolic rate: A multidisciplinary conceptual and methodological toolkit: New technical developments are opening the door to an understanding of why metabolic rate varies among individual animals of a species: New technical developments are opening the door to an understanding of why metabolic rate varies among individual animals of a species.

Researchers from diverse disciplines, including organismal and cellular physiology, sports science, human nutrition, evolution and ecology, have sought to understand the causes and consequences of the surprising variation in metabolic rate found among and within individual animals of the same species. Research in this area has been hampered by differences in approach, terminology and methodology, and the context in which measurements are made. Recent advances provide important opportunities to identify and address the key questions in the field. By bringing together researchers from different areas of biology and biomedicine, we describe and evaluate these developments and the insights they could yield, highlighting the need for more standardisation across disciplines. We conclude with a list of important questions that can now be addressed by developing a common conceptual and methodological toolkit for studies on metabolic variation in animals.

Is torpor a water conservation strategy? Heterothermic responses to acute water and food deprivation are repeatable among individuals of Phodopus sungorus.

Energy conservation is a clear function of torpor. Although many studies imply that torpor is also a water-saving strategy, the experimental evidence linking water availability with torpor is inconclusive. We tested the relative roles of water and energy shortages in driving torpor, using the Siberian hamster Phodopus sungorus as a model species. To account for the seasonal development of spontaneous heterothermy, we used male hamsters acclimated to short (8L:16D, SP; n = 40) and long (16L:8D, LP; n = 36) photoperiods. We continuously measured body temperature (Tb) during consecutive 32 h of complete removal of water, food, or both, separated by 7.5 d recovery periods. We predicted that all deprivation types would increase the frequency of spontaneous torpor in SP, and induce torpor in LP-acclimated hamsters. Individuals underwent each deprivation type twice in random orders. Food and water deprivation did not induce torpor in LP-acclimated P. sungorus. Patterns of torpor expression varied among deprivation types in SP individuals. Torpor frequency was significantly lower, but bouts were ∼2 h longer and 2.5 °C deeper, during water deprivation compared to food and food-and-water deprivation. Heterothermic responses to all deprivation types were repeatable among individuals. Different torpor patterns during water and food deprivation suggest that water and energy shortages are distinct physiological challenges. Deeper and longer bouts during water deprivation likely led to higher energy and water savings, while shorter and shallower bouts during fasting may reflect a trade-off between energy conservation and food-seeking activity. The lack of a difference between food- and food-and-water-deprived hamsters suggests a higher sensitivity to food than water shortage. This supports the traditional view that energy conservation is the major function of torpor, but suggests that water shortages may also modulate torpor use. The high repeatability of thermoregulatory responses to resource deprivation suggests that these may be heritable traits subject to natural selection.

Seasonal variation in body composition in an Afrotropical passerine bird: increases in pectoral muscle mass are, unexpectedly, associated with lower thermogenic capacity.

Phenotypic flexibility in avian metabolic rates and body composition have been well-studied in high-latitude species, which typically increase basal metabolic rate (BMR) and summit metabolism (Msum) when acclimatized to winter conditions. Patterns of seasonal metabolic acclimatization are more variable in lower-latitude birds that experience milder winters, with fewer studies investigating adjustments in avian organ and muscle masses in the context of metabolic flexibility in these regions. We quantified seasonal variation (summer vs winter) in the masses of organs and muscles frequently associated with changes in BMR (gizzard, intestines and liver) and Msum (heart and pectoral muscles), in white-browed sparrow-weavers (Plocepasser mahali). We also measured pectoral muscle thickness using a portable ultrasound system to determine whether we could non-lethally estimate muscle size. A concurrent study measured seasonal changes in BMR and Msum in the same population of sparrow-weavers, but different individuals. There was no seasonal variation in the dry masses of the gizzard, intestines or liver of sparrow-weavers, and during the same period, BMR did not vary seasonally. We found significantly higher heart (~ 18% higher) and pectoral muscle (~ 9% higher) dry mass during winter, although ultrasound measurements did not detect seasonal changes in pectoral muscle size. Despite winter increases in pectoral muscle mass, Msum was ~ 26% lower in winter compared to summer. To the best of our knowledge, this is the first study to report an increase in avian pectoral muscle mass but a concomitant decrease in thermogenic capacity.

Phenotypic flexibility of metabolic rate and evaporative water loss does not vary across a climatic gradient in an Afrotropical passerine bird.

Small birds inhabiting northern temperate and boreal latitudes typically increase metabolic rates during cold winters or acclimation to low air temperatures (Taccl). Recent studies suggest considerable variation in patterns of seasonal metabolic acclimatization in birds from subtropical and tropical regions with milder winters, but there remains a dearth of acclimation studies investigating metabolic flexibility among lower-latitude birds. We used short-term thermal acclimation experiments to investigate phenotypic flexibility in basal metabolic rate (BMR), thermoneutral evaporative water loss (EWL) and summit metabolism (Msum) in three populations of white-browed sparrow-weavers (Plocepasser mahali) along a climatic and aridity gradient. We allocated individuals to one of three Taccl treatments (5, 20 and 35°C; n=11 per population per Taccl) for 28 days, and measured post-acclimation BMR, EWL and Msum using flow-through respirometry. Our data reveal the expected pattern of lower BMR and EWL (∼12% and 25% lower, respectively) in birds at Taccl=35°C compared with cooler Taccl treatments, as observed in previous acclimation studies on subtropical birds. We found no variation in the reaction norms of BMR and EWL among populations in response to acclimation, suggesting previously documented differences in seasonal BMR acclimatization are the result of phenotypic flexibility. In contrast to higher-latitude species, Msum did not significantly vary in response to thermal acclimation. These findings support the idea that factors other than enhancing cold tolerance may be driving patterns of metabolic variation in subtropical birds.

Seasonal Metabolic Acclimatization Varies in Direction and Magnitude among Years in Two Arid-Zone Passerines.

Adaptive plasticity in avian thermal physiology is increasingly apparent, with a well-studied example being metabolic upregulation during cold winters in small birds inhabiting temperate and boreal latitudes. Recent studies have revealed greater variation in the direction and magnitude of seasonal metabolic adjustments among subtropical/tropical birds experiencing milder winters compared with higher-latitude counterparts, suggesting that patterns could vary among years within populations. We quantified seasonal metabolic variation (summer vs. winter) in Kalahari Desert populations of two Afrotropical passerines, the white-browed sparrow-weaver (WBSW; Plocepasser mahali; ∼40 g) and the scaly-feathered weaver (SFW; Sporopipes squamifrons; ∼10 g) over subsequent years (2014-2017). We used flow-through respirometry to measure basal metabolic rate (BMR) and summit metabolism (Msum; maximum cold-induced resting metabolic rate) and quantified seasonal fluctuations in air temperature (Ta) and food abundance (arthropod and grass seed abundance) at the study site. Our data reveal that the direction and magnitude of seasonal metabolic acclimatization vary among years in both species, with the winter BMR of WBSWs ranging from ∼20% lower to 68% higher compared with the summer BMR. In contrast to higher-latitude species, Msum was not related to the cold-limit temperature of birds or to winter minimum Ta at the study site, but interannual variation in BMR and Msum was significantly lower in seasons with lower food abundance in both WBSWs and SFWs. Our data support the idea that patterns of seasonal acclimatization are more variable in birds from lower latitudes and that there is considerable phenotypic flexibility in avian thermal physiology.

Reaction norms for heat tolerance and evaporative cooling capacity do not vary across a climatic gradient in a passerine bird.

There is increasing evidence for considerable phenotypic flexibility in endotherm thermal physiology, a phenomenon with far-reaching implications for the evolution of traits related to heat tolerance. Numerous studies have documented intraspecific variation in avian thermoregulatory traits, but few have revealed the shapes of thermoregulatory reaction norms or how these might vary among populations. We investigated phenotypic flexibility in the ability of a model Afrotropical passerine bird (the white-browed sparrow-weaver, Plocepasser mahali) to handle high air temperatures (Ta). We allocated birds from three sites varying by ~ 11 °C in mean daily summer maximum Ta to three acclimation temperature (Taccl) treatments (daytime Taccl ≈ 30 °C, 36 °C or 42 °C respectively; n ≈ 10 per site per Taccl). After an acclimation period of 30 days, heat tolerance and evaporative cooling capacity was quantified by exposing birds to progressively higher Ta until they approached severe hyperthermia (body temperature [Tb] = 44.5 °C; Ta range: 38-54 °C). We measured metabolic rate and evaporative water loss using open flow-through respirometry, and Tb using temperature-sensitive passive-integrated transponder tags. Hyperthermia threshold Ta (Ta,HT) was significantly higher and Tb significantly lower in birds acclimated to the hottest Taccl compared to those from milder acclimation treatments. Population (i.e., site of capture) was not a significant predictor of any thermoregulatory variables or hyperthermia threshold Ta (Ta,HT) after acclimation, revealing that the shape of reaction norms for heat tolerance and evaporative cooling capacity does not vary among these three populations.

Interactions between humidity and evaporative heat dissipation in a passerine bird.

Environmental humidity is thought to be a major determinant of evaporative cooling capacity at high air temperatures (Ta), but the technical challenges of experimentally manipulating humidity in respirometry chambers have resulted in far less being known about the effects of humidity compared to those of Ta. We tested the prediction that at Ta approaching and exceeding normothermic body temperature (Tb), high humidity would result in higher Tb, lower evaporative water loss (EWL) and/or higher resting metabolic rate (RMR) in a passerine bird, the white-browed sparrow-weaver (Plocepasser mahali). We used open-system flow-through respirometry to measure EWL, RMR and Tb in sparrow-weavers experiencing Ta = 36-44 °C and chamber humidities of 6, 13, 19 or 25 g m- 3. Increasing humidity was associated with significantly higher Tb. The strongest effect of humidity, however, involved significant increases in RMR; at Ta = 40 °C, RMR at a humidity of 25 g m- 3 was ~ 40% higher compared to a humidity of 6 g m- 3. Moreover, the interaction between Ta and humidity exerted a significant effect on the ratio of evaporative heat loss (EHL) to metabolic heat production (MHP), evident as an increasing effect of humidity with increasing Ta. Our results, when compared with those of the limited number of previous studies that involved similar ranges of Ta and humidity, reveal that the relative effects of humidity on EWL and RMR vary among avian taxa, and support the notion that the overall effect of high humidity is a reduction in maximum EHL/MHP.

Seasonal Metabolic Acclimatization Varies in Direction and Magnitude among Populations of an Afrotropical Passerine Bird.

Avian metabolic responses demonstrate considerable diversity under fluctuating environmental conditions, a well-studied example being the seasonal upregulation of basal metabolic rate (BMR) and summit metabolism (Msum) in temperate species experiencing harsh winters. Fewer studies have examined seasonal metabolic acclimatization in subtropical or tropical species. We investigated seasonal metabolic variation in an Afrotropical ploceid passerine, the white-browed sparrow-weaver (Plocepasser mahali; ∼47 g), at three sites along a climatic gradient of approximately 7°C in winter minimum air temperature (Ta). We measured Msum (n ≥ 10 per site per season) in a helox atmosphere, BMR of the same birds at thermoneutrality (Ta ≈ 30°C), and resting metabolic rates at 5°C ≤ Ta ≤ 20°C. Patterns of seasonal adjustments in BMR varied among populations in a manner not solely related to variation in seasonal Ta extremes, ranging from BMR ∼52% higher in winter than in summer to no seasonal difference. Greater cold tolerance was found in a population at a colder desert site, manifested as higher Msum (∼25% higher) and lower helox temperature at cold limit values compared with a milder, mesic site. Our results lend support to the idea that greater variance in the pattern of seasonal metabolic responses occurs in subtropical and tropical species compared with their temperate-zone counterparts and that factors other than Ta extremes (e.g., food availability) may be important in determining the magnitude and direction of seasonal metabolic adjustments in subtropical birds.

Avian thermoregulation in the heat: evaporative cooling capacity in an archetypal desert specialist, Burchell's sandgrouse (Pterocles burchelli).

Sandgrouse (Pterocliformes) are quintessential examples of avian adaptation to desert environments, but relatively little is known about the limits to their heat tolerance and evaporative cooling capacity. We predicted that evaporative cooling in Burchell's sandgrouse (Pterocles burchelli) is highly efficient and provides the basis for tolerance of very high air temperature (Ta). We measured body temperature (Tb), resting metabolic rate (RMR) and evaporative water loss (EWL) at Ta between 25°C and ∼58°C in birds exposed to successive increments in Ta Normothermic Tb averaged 39.0°C, lower than typical avian values. At Ta>34.5°C, Tb increased linearly to a maximum of 43.6°C at Ta=56°C. The upper critical limit of thermoneutrality (Tuc) was Ta=43.8°C, closely coinciding with the onset of panting and gular flutter. Above the Tuc, RMR increased 2.5-fold to 2.89 W at Ta=56°C, a fractional increase far exceeding that of many other species under comparable conditions. Rates of EWL increased rapidly at Ta>42.9°C to 7.84±0.90 g h(-1) at Ta=56°C, an 11-fold increase above minimal levels. Maximum evaporative cooling efficiency (ratio of evaporative heat loss to metabolic heat production) was 2.03, but could be as high as 2.70 if our assumption that the birds were metabolising lipids is incorrect. Thermoregulation at very high Ta in P. burchelli was characterised by large increases in RMR and EWL, and is much less efficient than in taxa such as columbids and caprimulgids.

Seasonal and geographical variation in heat tolerance and evaporative cooling capacity in a passerine bird.

Intraspecific variation in avian thermoregulatory responses to heat stress has received little attention, despite increasing evidence that endothermic animals show considerable physiological variation among populations. We investigated seasonal (summer versus winter) variation in heat tolerance and evaporative cooling in an Afrotropical ploceid passerine, the white-browed sparrow-weaver (Plocepasser mahali; ∼ 47 g) at three sites along a climatic gradient with more than 10 °C variation in mid-summer maximum air temperature (Ta). We measured resting metabolic rate (RMR) and total evaporative water loss (TEWL) using open flow-through respirometry, and core body temperature (Tb) using passive integrated transponder tags. Sparrow-weavers were exposed to a ramped profile of progressively higher Ta between 30 and 52 °C to elicit maximum evaporative cooling capacity (N=10 per site per season); the maximum Ta birds tolerated before the onset of severe hyperthermia (Tb ≈ 44 °C) was considered to be their hyperthermia threshold Ta (Ta,HT). Our data reveal significant seasonal acclimatisation of heat tolerance, with a desert population of sparrow-weavers reaching significantly higher Ta in summer (49.5 ± 1.4 °C, i.e. higher Ta,HT) than in winter (46.8 ± 0.9 °C), reflecting enhanced evaporative cooling during summer. Moreover, desert sparrow-weavers had significantly higher heat tolerance and evaporative cooling capacity during summer compared with populations from more mesic sites (Ta,HT=47.3 ± 1.5 and 47.6 ± 1.3 °C). A better understanding of the contributions of local adaptation versus phenotypic plasticity to intraspecific variation in avian heat tolerance and evaporative cooling capacity is needed for modelling species' responses to changing climates.

Thermoregulation in African Green Pigeons (Treron calvus) and a re-analysis of insular effects on basal metabolic rate and heterothermy in columbid birds.

Columbid birds represent a useful model taxon for examining adaptation in metabolic and thermal traits, including the effects of insularity. To test predictions concerning the role of insularity and low predation risk as factors selecting for the use of torpor, and the evolution of low basal metabolic rate in island species, we examined thermoregulation under laboratory and semi-natural conditions in a mainland species, the African Green Pigeon (Treron calvus). Under laboratory conditions, rest-phase body temperature (T b) was significantly and positively correlated with air temperature (T a) between 0 and 35 °C, and the relationship between resting metabolic rate (RMR) and T a differed from typical endothermic patterns. The minimum RMR, which we interpret as basal metabolic rate (BMR), was 0.825 ± 0.090 W. Green pigeons responded to food restriction by significantly decreasing rest-phase T b, but the reductions were small (at most ~5 °C below normothermic values), with a minimum T b of 33.1 °C recorded in a food-deprived bird. We found no evidence of the large reductions in T b and metabolic rate and the lethargic state characteristic of torpor. The absence of torpor in T. calvus lends support to the idea that species restricted to islands that are free of predators are more likely to use torpor than mainland species that face the risk of predation during the rest-phase. We also analysed interspecific variation in columbid BMR in a phylogenetically informed framework and verified the conclusions of an earlier study which found that BMR is significantly lower in island species compared to those that occur on mainlands.