1. Physiological basis and energetic consequences of vertebrate endothermy (2003-2007)
Project leader: Esa Hohtola
Support: Academy of Finland

2. Energy metabolism and thermal biology of endothermic animals in the North: Genotypic adaptation and phenotypic flexibility  (2006-2011)
Project leader: Esa Hohtola
Support: Thule-Institute

Group members

Seniors: Esa Hohtola, Seppo Saarela; Satu Mänttäri

Doctoral students (past & present):
1) Mirja Laurila see dissertation pics, May 20, 2005, supervisor Esa Hohtola
2) Juli Broggi (dissertation on Sept 1, 2006), see Juli with crested tit (Parus cristatus), supervisors Esa Hohtola & Markku Orell
3) Pirkko Sallinen, supervisor Seppo Saarela dissertation March 31, 2008

Pro gradu -students presently supervised (as main supervisor) by EH: Hanna Välimaa, Virpi Evesti, Maria Toivanen, Satu Timlin
 

Why study endothermy?

"Endothermy (the elevation of body temperature by metabolic heat production) represents one of the
most significant developments during vertebrate evolution." Hayes JP & Garland T Jr.: The evolution
of endothermy: testing the aerobic capacity model. - Evolution 49:839 (1995).

Endothermy is a novel phenomenon in the evolution of vertebrates (Ruben, 1995). In its strict form, it is limited to birds and mammals, whose adaptive radiation occurred after the mass extinction of dinosaurs. While all metabolic events produce heat as a byproduct, endothermic species are able to use such mechanisms in a controlled manner to maintain a thermal gradient towards the environment. Endothermy has thus a pivotal role in homeothermy.

The two known thermoregulatory mechanisms of endogenous heat production that birds and mammals use for endothermy are shivering thermogenesis in skeletal muscles and brown adipose tissues thermogenesis (also called non-shivering thermogenesis, NST). Shivering is used by both groups while brown fat-related NST is limited to mammals. In addition to thermoregulatory thermogenesis, the obligatory component of thermogenesis in non-thermoregulatory tissues is also significantly higher in endotherms than other vertebrates. This is the basis of endothermy in thermoneutral conditions (Hayes & Garland, 1995).

The study of the mechanisms of endothermic heat production has potential for shedding light on many biologically important phenomena:

1. Energy is the currency by which the proximate costs and benefits of many biological functions are measured. As endothermy is energetically very expensive, birds and mammals frequently encounter situations where various physiological functions compete for a limited energy supply and where survival necessitates special adaptive responses. Short-term hypothermia and hibernation in response to limited food supply are examples of such adaptations (Reinertsen, 1996). The study of endothermy thus promises advances in the understanding the controlling systems of animal bioenergetics.

2. The evolution of endothermy was based largely on utilisation of pre-existing physiological and anatomical structures (Satinoff, 1978). In particular, muscular thermogenesis is a classical example of harnessing a pre-existing system to a novel function. Shivering thermogenesis appears to have developed independently in birds and mammals to surprisingly analogous modes of function. Comparative studies of endothermic heat production thus promise advances in the understanding of the mechanisms by which physiological systems evolve.

3. Mammals and birds are characterised by a continuum of ontogenetic modes from altricial to precocial (Starck & Ricklefs, 1998). Altricial young rely on parental care during their development while precocial young are more independent. An important difference between altricial and precocial young is their capacity for endothermic heat production: precocial young are endothermic at birth while altricial species have to rely on external heat sources, usually their parents. This difference is reflected in almost all features of their development: growth rate, maturation of sensory systems and energetics. Utilisation of this continuum for comparative studies on endothermy is a potentially fertile field of developmental physiology.

4. Variations in the capacity for endothermic heat production are intimately related to the life history and distribution of many small mammals and birds in northern latitudes. In addition non-genetic adaptation (acclimatisation), genetic adaptation of populations to local conditions occurs. Maximal thermogenesis is an essential factor in the survival of many non-migrating species. Because basal metabolic rate increases with the maximal capacity for heat production, any increase in peak thermogenesis incurs additional costs in the form of increased basal energy consumption. Studying these parameters in northern birds and mammals thus combined ecophysiology to population biology.

Goals

We classify the objectives of the studies into 4 categories that address the questions posed in the title of this proposal at various level of biological organization. Accordingly, the objectives are listed below according to these levels from cellular and tissue level functions to organismal and population-level research plans.

1. To elucidate the biochemical, neuromuscular and thermal control of muscle thermogenesis with special reference to differences in mammals and birds. Also, the role of the recently discovered avian UCP in endothermy will be studied.

Specific aims include:

a) A comparative study of motor unit synchronization during shivering. Shivering thermogenesis has evolved independently in birds and mammals (Hohtola 1981) and there is some evidence indicating that these groups use slightly different neuronal control for regulating the level of muscle thermogenesis. Shivering comprises of a thermally induced activity of motor units leading to a sustained muscle tonus and sometimes tremor. It is adaptive to keep the level of synchronization low during shivering to avoid tremor that would increase convective heat loss. Preliminary data suggest that the level of synchronization is lower in birds than in mammals. Using signal analysis of electromyographic and accelerometer recordings it is possible to detect differences in motor unit functions during shivering and to shed light on how these mechanisms evolved during the phylogeny of birds and mammals.

b) The role of avian uncoupling protein (UCP) in the ontogeny and seasonal changes of thermogenesis. Precocial birds hatch with functional thermogenic mechanisms while in altricial hatchlings thermogenic capacity develops later. By measuring the expression of UCP-gene in skeletal muscle during the ontogenetic development of precocial and altricial birds, and correlating it with the development of thermogenic capacity (body temperature and metabolic rate during acute coldexposure), we can shed light on the role of avian UCP in thermogenesis.

2. The consequences of high rate of aerobic metabolism: damage by reactive oxygen species and mechanisms for protection. The high activity and large number of mitochondria in endotherms represent a source of free oxygen radicals from the respiratory chain. However, despite the higher aerobic metabolism in birds, their life span is 3 to 4 times longer than in mammals of similar body size. By studying respiratory chain function and protection by antioxidants in aged and coldacclimated birds and mammals we try to elucidate the senescence consequences of endothermy and explain why avian species live longer than mammals. This study will not encompass all aspects of antioxidant defence, but will concentrate on melatonin and respiratory chain function, which connect endothermy to endogenous diurnal rhythms and elevated metabolic rate. An interesting avenue in this context is the putative role of mitochondrial uncoupling proteins in suppressing production of reactive oxygen species (Brand 2000).

Specific aims:

a) In addition to the role of melatonin in mediating the diurnal rhythms of metabolic rate and its potential role in signaling energy balance, all these factors are connected by the potential direct and in direct (receptor-mediated) antioxidant functions of melatonin (Reiter 1995). This has important implications for studies of endotherm mitochondrial function and longevity. We plan to measure variations in melatonin levels in plasma and various tissues in birds and mammals in relation to age, food restriction and cold acclimation. In addition, indirect effects of melatonin will be studied by measuring the expression of melatonin membrane receptors in rat heart by q-RTPCR. The putative protective action of melatonin are necessarily time-dependent due to the diurnal cycle of melatonin secretion. Any indirect influence would thus be reflected as changes in receptor density. Interestingly, human heart failures are more frequent in the early morning when melatonin levels are at their minimum (Willich et al. 1992).

b) Oxidative capacity of tissues: affects of age, cold-acclimation and melatonin treatment. We will monitor the function of the respiratory chain by measuring cytochrome c -oxidase activity in various tissues after the experimental treatments to obtain an index of the level of mitochondrial deterioration. Different mitochondrial populations in muscle (intermyofibrillar vs. subsarcolemmal) will be studied separately.

3. To study how the energetic costs of endothermy can be alleviated by hypometabolic adaptations and optimization of obligatory thermogenesis related to feeding and digestion. Endotherms are an excellent model for studying the interactions of various homeostatic mechanisms that regulate energy consumption. This is due to the high energy costs of endothermy. Birds and mammals have a host of regulatory mechanisms that influence the amount of energy allocated to thermoregulation when they are confronted with restricted food availability.

Specific aims for experimental studies include:

a) Plasticity of fasting-induced hypometabolism. This will studied by measuring the effect on daylength and repeated fasts on fasting-induced nocturnal hypothermia. Most studies on fasting-induced hypothermia and hypometabolism have used only a single experimental fast or food restriction. In preliminary experiments with quails and pigeons, we have found that nocturnal hypothermia is rather inflexible as it does not influenced photoperiod or repeated fasts. Instead, a number of novel responses were found. These include a daytime decrease in body temperature and progressive increase in interfast body mass (Laurila et al. 2005). Further studies with a wider array of species will be performed to test the adaptive nature and flexibility of the hypothermic response and the physiological costs that limit its use only to situations of energetic stress. The experiments will include tests on the ability of birds to 'predict' food shortage from environmental cues. Quails and pigeons will be subjected to conditions where various visual stimuli will precede periods of food deprivation. The variables that will be recorded are body temperature, metabolic rate (oxygen consumption), body mass and motor activity.

b) Adaptive use of digestion-related thermogenesis to reduce the need for muscle thermogenesis. A significant amount of heat liberated during the processing of food in the gastrointestinal system. We have shown recently (Rashotte et al. 1999) that this thermogenesis can be effectively used to substitute for actual thermogenesis and thus to save energy. Additional savings could be accrued by the timing of digestion. This is possible in birds because feeding and actual digestion are 'uncoupled' due to the presence of a crop. Our preliminary work where accumulation of excreta was used as an index of digestion rate suggests that pigeons are able to retain food in the crop and postpone digestion to a time when the digestion-related heat production can be most effectively used. Future work will include more direct measurements of crop emptying and gastrointestinal function combined with measurements of metabolic rate.

4. Ecophysiological aspects and geographical variation of metabolic traits in small passerine birds.
Resident small passerines in winter face a combination of higher energetic demands (cold weather and long nights) and reduced food supply (shorter days in which to feed and reduced and more variable food supply). These conditions are more pronounced at high latitudes, where low temperatures and short photoperiod can last for several months. Birds have several physiological and behavioural strategies in order to survive the high energy demanding non-breeding period. Fat supplies the major source of metabolic fuel during winter fasts e.g. sudden harsh conditions and long cold nights. In order to minimise starvation risk birds should carry the maximum amount of fat that prevailing conditions would allow. Nevertheless they rarely do so, implying some costs in carrying fat reserves (Witter and Cuthill 1993). Instead they seem to actively manage their reserve levels taking into account food predictability (feeding conditions, food availability and food abundance), energy requirements and predation risk (Lima 1986). For the same purpose, some species may build up external food hoards during favourable periods and turn to repertoire of energy saving strategies such as special roosting places, better plumage insulation or nocturnal hypothermia.
Specific aims include:
 

a) Night and Day body reserves regulation. Most of theoretical and empirical research has been focused on energy regulation on a daytime basis, but birds not only manage daytime energy reserves but they also manage night-time energy expenditure. By decreasing night-time energy expenditure, birds might reduce starvation risk without increasing levels of fat reserves. Some birds are capable of reducing their body temperature during night (nocturnal hypothermia), by reducing their metabolic rate. Hypothermia has found to be regulated according to season, time-of-day, ambient temperature and energy reserves (Reinertsen 1996). This facultative use of hypothermia suggests that there must be some costs of using it. It has been hypothesised that hypothermic birds may suffer from an increased nocturnal predation risk, as their reaction time to external stimuli is increased and thus their ability to detect approaching predators is probably impaired. We test empirically the predictions made by a recent stochastic dynamic model by (Pravosudov and Lucas 2000). (1) Low body reserves, external caches, temperature and food predictability favour the use of hypothermia. (2) Nocturnal predation has an important effect on individual's decision to use hypothermia and (4) Optimal levels of body reserves are predicted to be smaller for birds employing nocturnal hypothermia. We expect birds to show a decreased hypothermia by a higher body mass at dusk and a higher mass loss during night (higher metabolism) when perceived night predation risk is increased. Alternatively, birds should decrease dusk body mass and increase hypothermia when daytime predation risk is increased. Birds can increase mass at dusk by increasing food intake or by diminishing energy expenditure throughout the day. This question can be resolved by measuring the amount of food consumed or by making estimates of the daily activity. By offering food permanently (ad libitum) we can assure that birds won't have any availability constraints in achieving the optimal body mass. For measuring the occurrence of nocturnal hypothermia we have developed a novel method that minimizes disturbance, which is necessary for experiments on wild birds. Briefly, the roosting box will consist of a wooden box outside the aviary connected to it through a hole. Birds will be able to roost in the box freely. When entering it they will fall into a cloth bag hanging from a load cell (an electronic mass-recording device). The load cell will register mass changes continuously. The deepness achieved of hypothermia has shown to be highly and negatively correlated with the change in body mass throughout the night (Reinertsen 1986). Birds will be exposed to first a three consecutive three-day experimental treatment and then a three-day control or the opposite. Birds are expected to start using the roosting boxes in the first day of captivity. Three birds will experience the same treatments for six days and after will be released. Following three birds will receive the alternate order of treatments.

b) Cold endurance differences between populations. Small birds wintering in boreal regions survive the non-breeding season by a process of winter acclimatization that improves cold resistance with respect to summer. Some species with wide distribution ranges may experience extremely different winter conditions ranging from mild winters to extremely cold ones. It is unclear how some species can deal with such different conditions. Winter acclimatization, at an individual level, is primarily a physiological and behavioural response but might include morphological differences at a population level. At a physiological level, winter acclimatized birds may show increased thermogenic capacity (maximum cold-induced metabolic rates), increased energy reserve levels, i.e winter fattening, or increased digestive efficiency. We intend to study basal and summit cold-induced metabolic rates and track those variables throughout the winter within individuals, populations and between populations differing in winter harshness. For this we have established collaboration with a study group in Lund with whom the methods have already been standardized. Metabolic rates will be measured using indirect calorimetry and a helium-air mixture will be used to induce summit metabolism.