the earliest mammals were shrew - like creatures who could search for food and hide in places that were too cool and dark for many of their lizard contemporaries ( 1 ) .
in addition to being warm blooded , mammals also were adept at performing adaptive thermogenesis , giving them the ability to further increase their metabolic rates when conditions would otherwise result in hypothermia . from the perspective of humans in 2015 , it is easy to forget that thermoregulation , in all of its dimensions , was a critical part of mammalian history
thermogenesis comes from chemical reactions in which the liberated free energy is not captured in other molecules ( e.g. , atp or creatine phosphate ) or used for work .
the best known example of a heat - generating pathway is the futile cycle of proton pumping in brown fat and beige fat through the actions of uncoupling protein 1 ( ucp1 ) ( 2,3 ) . in cells expressing ucp1 , the oxidation of lipids and carbohydrates results in the extraction of high - energy electrons , which flow down the electron transport chain ( etc ) as protons are pumped across the inner mitochondrial membrane ( fig .
the resulting electrochemical gradient across the inner mitochondrial membrane , which is usually coupled to atp synthesis via complex v of the etc , is dissipated by a  leak  of protons back across the inner membrane by ucp1 .
thus , much of the chemical energy generated by fuel oxidation in brown fat cells powers a futile proton cycle , which does no work and is instead liberated as heat .
this process is typically thought of as being indirectly activated by cold , via the sympathetic nervous system ( sns ) .
adaptive thermogenesis can also be activated directly by cold in beige adipocytes and by other stimuli that may signal independently from the -adrenergic receptors .
the reducing equivalents generated by the tricarboxylic acid ( tca ) cycle enter the etc .
this generates a proton gradient across the inner mitochondrial membrane . instead of linking this gradient to atp synthesis via complex v ,
it is also worth noting that , from a thermodynamic perspective , there is nothing special about proton cycling , and such a thermogenic machine could , in theory , be built from other components of cellular metabolism .
indeed , some deep - diving fish have a  heater organ ,  which is a specialized muscle that entirely lacks a myofibrillar apparatus .
this organ , found between the eye and brain in deep - diving marlin and certain tuna , uses a futile cycle of calcium leaking from the sarcoplasmic reticulum and atp - dependent calcium uptake to raise the local temperature near the brain of the fish ( 4 ) . in this case , the physiological goal is not to warm the whole body of the marlin , but to just make it a bit more astute than the other fish around it ! is ucp1 the only thermogenic pathway of importance in mammals ? while no other pathway of thermogenesis except shivering has been convincingly demonstrated , suggestive data from the scientific literature hints that such pathways probably exist . if mice lacking ucp1 are abruptly switched from ambient temperatures to cold ( 4c ) , they develop life - threatening hypothermia ( 5 ) . on the other hand ,
if they are gradually exposed to lower temperatures , they can survive quite normally at 4c ( 6 ) .
this suggests that there are compensatory thermogenic programs that can be activated as long as thermal stress is applied more gradually .
in addition , there are examples of experiments in mice where certain mutations have caused very robust increases in energy expenditure and thermogenesis , but where the levels of ucp1 mrna and protein are unchanged .
for example , deletion of par-1b / mark2 in mice results in increased energy expenditure and brown fat activation , although ucp1 protein levels were unchanged ( 7 ) .
ucp1 brown fat has long been known to occur in two distinct anatomical locations in rodents .
developmentally formed depots , best typified by the interscapular and perirenal regions , are composed of tightly packed brown fat cells of relatively uniform appearance .
these depots exist under most physiological conditions , although they may change in color and lipid content . on the other hand
, ucp1 cells can accumulate in small pockets in white fat depots , especially in the subcutaneous adipose tissues , particularly when mice are exposed to long - term cold or stimuli with hormones such as catecholamines and other -adrenergic agonists ( 8) . this pathway of adaptive thermogenesis is robustly activated by cold via an indirect pathway mediated by the sympathetic nervous system .
however , fat cells are also able to directly sense cold , though the mechanism is not well understood ( 9 ) . in both anatomical situations , brown fat cells have multiple small lipid droplets , numerous mitochondria , and rich innervation and vascularization . despite these similarities , it is now clear that the  classical  brown fat cells and the inducible  beige  fat cells come from different developmental lineages and are , in fact , distinct cell types .
the key finding in this regard was the observation that the classical developmentally formed brown fat arises from a myf5 lineage shared with skeletal muscle ( 10 ) .
consistent with this , primary cultures of classical brown fat express low levels of certain genes characteristic of skeletal muscle ( 11 ) .
subsequent work indicated that the brown fat / skeletal muscle decision occurs between 9.5 and 12.5 days of gestation in mice ( 12 ) .
conversely , the ucp1 cells induced in subcutaneous white fat depots by cold or -adrenergic agonists come from a myf5 lineage ( 10 ) . these beige fat cells have now been cloned and express a relatively low level of thermogenic genes , such as ucp1 and type 2 deiodinase in the basal state , but can induce these genes to levels essentially equivalent to classical brown fat cells when given hormonal stimuli ( 13 ) . moreover ,
the beige and brown fat cells have gene signatures that allow for distinction of these cells types ; however , their comparative physiological properties and overall roles in metabolism are not completely understood .
it is now clear that thermogenic adipocytes , taken as a whole , contribute very importantly to metabolic homeostasis in rodents .
the partial ablation of ucp1 cells through the transgenic expression of a toxigene led to these animals being more susceptible to obesity and diabetes ( 14 ) .
similarly , mice lacking ucp1 through targeted mutation had an increased body weight and fat content , at least when raised at thermoneutrality ( 15 ) .
excessive shivering apparently prevented obesity when these experiments were performed at ambient temperatures . the first transgenic model with increased brown and beige
fat was the adipose - selective expression of foxc2 , a transcription factor that activated camp metabolism in recipient cells ( 16 ) .
these mice , which were resistant to diet - induced obesity and diabetes , had expansions of both the classical brown and beige fat . deciphering the individual roles of brown and beige fat cells
prd1-bf-1-riz1 homologous domain - containing protein-16 ( prdm16 ) is an important transcriptional coregulator in both brown and beige fat , but when its expression was elevated through a promoter expressed in all fat depots ( ap2 ) , phenotypic changes were observed in the erstwhile white fat depots , which developed copious pockets of beige fat cells ( 17,18 ) .
the classical brown fat , which expresses very high levels of prdm16 in the basal state , showed few or no changes with a further elevation in prdm16 expression .
but this occurred only when the transgenic mice also received stimulation with a -adrenergic agonist . while studies in cultured cells had shown that classical brown fat cells require prdm16 to develop and maintain a thermogenic gene program , ablation of prdm16 from all fat cells in vivo was not sufficient to significantly alter the function of the classical brown fat . on the other hand ,
the development of beige fat cells was severely reduced , at both the histological and molecular level ( 19 ) .
these mice developed a moderate obesity , compared with control animals , with an unusual expansion of the subcutaneous fat depots .
these beige fat  deficient mice also showed a rather profound hepatic insulin resistance that was associated with liver steatosis .
one last point of interest is that the subcutaneous fat from the prdm16-ablated mice showed the presence of more inflammatory macrophages and increased the expression of proinflammatory genes .
the latter aspect was also observed with cultured subcutaneous cells lacking prdm16 , indicating that this transcriptional coregulator plays a role in the relative resistance of subcutaneous fat to inflammation and is an important determinant of subcutaneous versus visceral fat phenotype .
the role of prdm16 in the classical brown fat in vivo is interesting and somewhat more complex .
ablation of this factor with a myf5-driven cre recombinase causes a loss of brown fat function and gene expression , but only at 6 months of age ( 20 ) .
however , ablation of prdm16 along with its closest homolog , prdm3 , causes an early and severe loss of brown fat thermogenic gene expression and normal histology .
thus , it appears that both thermogenic cell types depend on prdm16 , but , at least in early life , the classical brown fat has a second factor that can support development , prdm3 .
harnessing the therapeutic potential of brown and beige fat requires a detailed understanding of the molecular mechanisms responsible for the determination and maintenance of each cell type .
the nuclear hormone receptor peroxisome proliferator  activated receptor  ( ppar ) is necessary and sufficient for the development of all fat cells ( 21 ) .
however , white and brown adipocytes have drastically different phenotypes , indicating that other transcriptional regulators must be involved .
a yeast two - hybrid screen identified ppar coactivator-1 ( pgc-1 ) as a cold - inducible binding partner of ppar ( 22 ) . in brown fat
pgc-1 is now appreciated to regulate numerous physiological processes in a variety of metabolically important tissues .
while brown and beige fat cells lacking pgc-1 have significantly blunted thermogenic gene expression , these cells still retain the molecular signature of brown fat cells ( 23,24 ) .
the transcriptional regulator prdm16 was found to be highly enriched in brown fat , whereas it is virtually undetectable in visceral white fat cells .
forced expression of prdm16 in cultured white fat cells induces pgc-1 and thermogenic genes , as well as mitochondrial genes and brown fat identity genes ( 17 ) .
in addition to activating these pathways , prdm16 also binds ccaat / enhancer binding protein  ( c / ebp ) and recruits the corepressor proteins ctbp1 and ctbp2 to repress white fat or muscle gene expression ( 25 ) .
importantly , the expression of prdm16 and its binding partner c / ebp in fibroblasts is sufficient to promote differentiation into functional brown fat cells , which can be detected by [ f]-2-fluoro-2-deoxy - d - glucose ( [ f]-fdg ) positron emission tomography ( pet ) when transplanted into mouse models ( 26 ) .
an increasing number of transcriptional regulators has been identified as important in brown and beige fat biology .
the transcription factor early b - cell factor-2 ( ebf2 ) is enriched in brown relative to beige adipocytes .
it functions upstream of prdm16 to promote binding of ppar to the promoters of brown - selective genes ( 27 ) .
ehmt1 is a histone lysine methyltransferase that purifies with the prdm16 transcriptional complex in brown fat .
ehmt1 is required for brown fat lineage specification and for thermogenesis ( 28 ) . presumably , there are also beige - selective factors upstream of prdm16 and beige - selective epigenetic regulators that modulate the phenotype of these cells .
it is able to compete with prdm16 for binding to ppar , and thereby modulates white versus brown / beige phenotype ( 29,30 ) . forced expression of tle3 in adipocytes results in impaired thermogenesis , while deletion enhances thermogenesis in brown and beige fat .
a number of other factors have also been shown to play roles in this biology , but a comprehensive review is beyond the scope of this article .
until fairly recently , brown fat was thought to be present in meaningful amounts only in human infants and small mammals .
the combination of insufficient hair , fur , and insulation , along with a high body surface area - to - mass ratio make babies and small mammals particularly susceptible to hypothermia . as a result
, they have developed significant interscapular brown fat , which defends body temperature by adaptive thermogenesis .
this brown fat was thought to regress by adulthood , unless exposed to catecholamine excess ( as in pheochromocytoma ) or long - term cold exposure ( as in outdoor workers in cold climates ) . in 2009 ,
these studies made use of [ f]-fdg pet to confirm that adults have glucose - avid tissue with imaging characteristics of adipose in deposits in the supraclavicular and spinal regions . as assayed by [ f]-fdg pet ,
the amount of  active  tissue is inversely associated with bmi , and this activity is increased by cold exposure .
follow - up studies ( 34 ) have shown that this tissue has the histological appearance of adipose tissue and expresses several molecular markers of brown fat including ucp1 .
current efforts are focused on developing optimal imaging modalities to detect this tissue and to quantify its absolute amounts and activity .
this is particularly important since brown and beige fat also oxidize lipids , which would not be detectable by [ f]-fdg pet technology .
research has increasingly suggested that adult human brown fat shares features of the inducible beige fat described in rodents .
brown and beige fat cell lines have now been cloned from mice , allowing for the identification of unique molecular markers of each cell type .
multiple studies ( 13,35,36 ) have shown that beige - selective markers are expressed more in human brown fat samples than are markers of the classical brown fat of rodents . a very recent study ( 37 ) characterized a human brown fat cell line and showed that it actually shares more molecular features with beige than brown adipocytes .
however , the brown fat in human infants , located in the interscapular region , seems most similar to the interscapular classical brown fat in rodents . in some adult humans , cells with both brown and beige characteristics
have been identified , with the depth in the neck seeming to be a determinant of relative brown versus beige character ( 36,38,39 ) . while further studies will clarify this question ,
this tissue may well contain both brown and beige cells in varying amounts in each individual .
as a deeper understanding of brown and beige fat has emerged , researchers have turned to strategies to induce the activity of these tissues as a possible therapy for obesity and metabolic diseases .
calculations have suggested that maximal cold - induced brown fat thermogenesis would be between 25 and 400 kcal / day in lean healthy volunteers ( 40 ) .
given the inducibility of beige fat , activating these cells could result in even more substantial effects .
however , the greatest benefits may not relate to increased energy expenditure per se , but rather might result from enhanced glucose and lipid disposal .
in addition to taking up and consuming glucose ( 3133 ) , brown adipose tissue takes up free fatty acids from triglyceride - rich lipoproteins .
cold exposure in mice upregulates this pathway , resulting in accelerated plasma triglyceride clearance ( 41 ) .
while compounds like dinitrophenol can result in impressive weight loss , they have also been associated with complications , including fatal hyperthermia . while this has understandably resulted in caution , it is possible that more specific uncouplers or targeted activators of ucp1 or other futile cycles may provide metabolic benefit with an acceptable safety profile . in that regard ,
a recent publication ( 42 ) described the development of controlled - release mitochondrial protonophores that can apparently safely uncouple in the liver , and in rats these compounds can improve insulin resistance , diabetes , hypertriglyceridemia , and hepatic steatosis .
brown and beige fat activity is robustly induced by thyroid hormone and catecholamines , but side effects would almost certainly limit their use . the 3-selective adrenergic agonist cl 316,243 potently activates brown and beige fat in rodents ( 43 ) .
this drug was recently shown to activate human brown fat , which may renew interest in this pathway ( 44 ) .
since cold robustly activates brown and beige fat , some investigators have suggested that moderate cold exposure could be used as a therapeutic approach . at least in healthy subjects , daily exposure to 19c for 2 h was sufficient to activate brown fat , resulting in weight loss ( 45 ) . in another small human study ( 46 )
while these studies suggest the potential of cold as a treatment modality , our societal preference for thermal comfort may make this unfeasible as a broad approach .
thiazolidinediones ( tzds ) cause browning of the white fat and may do so by stabilizing prdm16 , resulting in its accumulation ( 47 ) . however , these drugs have been associated with weight gain , fluid retention , and cardiovascular events , which have diminished enthusiasm for their use .
deacetylation of ppar by sirt1 promotes browning of the white fat , suggesting the possibility of developing compounds that selectively modulate this pathway ( 48 ) .
other regulators of interest include bone morphogenetic protein 7 ( bmp7 ) and bmp8b , cyclooxygenase-2 ( cox-2 ) , and natriuretic peptides , though their pleiotropic actions might limit their potential as drug targets .
fibroblast growth factor 21 ( fgf21 ) is being examined as a therapeutic agent , but it also has diverse actions and may be associated with bone loss . translating these discoveries into drug candidates will ultimately require a more complete understanding of brown / beige fat biology in humans .
mouse studies suggest that prdm16 is a key control point in brown and beige fat phenotype .
an increasing understanding of the relevant modifications of prdm16 and the upstream signaling pathways involved may provide new opportunities for therapeutic targeting of brown and beige fat .
 the epidemic of obesity and type 2 diabetes has increased interest in pathways that affect energy balance in mammalian systems . 
 brown fat , in all of its dimensions , can increase energy expenditure through the dissipation of chemical energy in the form of heat , using mitochondrial uncoupling and perhaps other pathways . 
 we discuss here some of the thermodynamic and cellular aspects of recent progress in brown fat research . 
 this includes studies of developmental lineages of ucp1 + adipocytes , including the discovery of beige fat cells , a new thermogenic cell type . 
 we also discuss the physiology and transcriptional control of brown and beige cells in rodents and the state of current knowledge about human brown fat . 
Role of Brown Fat From a Physiological Perspective: Thermodynamics
Discovery of Beige Fat and Its Physiology
Transcriptional Control
Rediscovery of Human Brown Fat and Its Brown Versus Beige Nature
Therapeutic Potential