Dysregulated pancreatic lipid phenotype, inflammation and cellular injury in a chronic ethanol feeding model of hepatic alcohol dehydrogenase-deficient deer mice.

AIMS: Dysregulation of pancreatic fat and lipotoxic inflammation are common clinical findings in alcoholic chronic pancreatitis (ACP). In this study, we investigated a relationship between dysregulated pancreatic lipid metabolism and the development of injury in a chronic ethanol (EtOH) feeding model of hepatic alcohol dehydrogenase 1- deficient (ADH-) deer mice. METHODS: ADH- and hepatic ADH normal (ADH+) deer mice were fed a liquid diet containing 3 % EtOH for three months and received a single gavage of binge EtOH with/without fatty acid ethyl esters (FAEEs) one week before the euthanasia. Plasma and pancreatic tissue were analyzed for lipids including FAEEs, inflammatory markers and adipokines using GC-MS, bioassays/kits, and immunostaining, respectively. Pancreatic morphology and proteins involved in lipogenesis were determined by the H & E staining, electron microscopy and Western blot analysis. KEY FINDINGS: Chronic EtOH feeding in ADH- vs. ADH+ deer mice resulted in a significant increase in the levels of pancreatic lipids including FAEEs, adipokines (leptin and resistin), fat infiltration with inflammatory cells and lipid droplet deposition along with the proteins involved in lipogenesis. The changes exacerbated by an administration of binge EtOH with/without FAEEs in the pancreas of ADH- vs. ADH+ deer mice fed chronic EtOH suggest a metabolic basis for ACP. SIGNIFICANCE: These findings suggest that the liver-pancreatic axis plays a crucial role in etiopathogenesis of ACP, as the increased body burden of EtOH due to hepatic ADH deficiency exacerbates pancreatic injury.

Exposure to binge ethanol and fatty acid ethyl esters exacerbates chronic ethanol-induced pancreatic injury in hepatic alcohol dehydrogenase-deficient deer mice.

Alcoholic chronic pancreatitis (ACP) is a fibroinflammatory disease of the pancreas. However, metabolic basis of ACP is not clearly understood. In this study, we evaluated differential pancreatic injury in hepatic alcohol dehydrogenase-deficient (ADH-) deer mice fed chronic ethanol (EtOH), chronic plus binge EtOH, and chronic plus binge EtOH and fatty acid ethyl esters (FAEEs, nonoxidative metabolites of EtOH) to understand the metabolic basis of ACP. Hepatic ADH- and ADH normal (ADH+) deer mice were fed Lieber-DeCarli liquid diet containing 3% (wt/vol) EtOH for 3 mo. One week before the euthanization, chronic EtOH-fed mice were further administered with an oral gavage of binge EtOH with/without FAEEs. Blood alcohol concentration (BAC), pancreatic injury, and inflammatory markers were measured. Pancreatic morphology, ultrastructural changes, and endoplasmic reticulum (ER)/oxidative stress were examined using H&E staining, electron microscopy, immunostaining, and/or Western blot, respectively. Overall, BAC was substantially increased in chronic EtOH-fed groups of ADH- versus ADH+ deer mice. A significant change in pancreatic acinar cell morphology, with mild to moderate fibrosis and ultrastructural changes evident by dilatations and disruption of ER cisternae, ER/oxidative stress along with increased levels of inflammatory markers were observed in the pancreas of chronic EtOH-fed groups of ADH- versus ADH+ deer mice. Furthermore, chronic plus binge EtOH and FAEEs exposure elevated BAC, enhanced ER/oxidative stress, and exacerbated chronic EtOH-induced pancreatic injury in ADH- deer mice suggesting a role of increased body burden of EtOH and its metabolism under reduced hepatic ADH in initiation and progression of ACP.NEW & NOTEWORTHY We established a chronic EtOH feeding model of hepatic alcohol dehydrogenase-deficient (ADH-) deer mice, which mimics several fibroinflammatory features of human alcoholic chronic pancreatitis (ACP). The fibroinflammatory and morphological features exacerbated by chronic plus binge EtOH and FAEEs exposure provide a strong case for metabolic basis of ACP. Most importantly, several pathological and molecular targets identified in this study provide a much broader understanding of the mechanism and avenues to develop therapeutics for ACP.

Pulmonary hypertension is attenuated and ventilation-perfusion matching is maintained during chronic hypoxia in deer mice native to high altitude.

Hypoxia at high altitude can constrain metabolism and performance and can elicit physiological adjustments that are deleterious to health and fitness. Hypoxic pulmonary hypertension is a particularly serious and maladaptive response to chronic hypoxia, which results from vasoconstriction and pathological remodeling of pulmonary arteries, and can lead to pulmonary edema and right ventricle hypertrophy. We investigated whether deer mice (Peromyscus maniculatus) native to high altitude have attenuated this maladaptive response to chronic hypoxia and whether evolved changes or hypoxia-induced plasticity in pulmonary vasculature might impact ventilation-perfusion (V-Q) matching in chronic hypoxia. Deer mouse populations from both high and low altitudes were born and raised to adulthood in captivity at sea level, and various aspects of lung function were measured before and after exposure to chronic hypoxia (12 kPa O2, simulating the O2 pressure at 4,300 m) for 6-8 wk. In lowlanders, chronic hypoxia increased right ventricle systolic pressure (RVSP) from 14 to 19 mmHg (P = 0.001), in association with thickening of smooth muscle in pulmonary arteries and right ventricle hypertrophy. Chronic hypoxia also impaired V-Q matching in lowlanders (measured at rest using SPECT-CT imaging), as reflected by increased log SD of the perfusion distribution (log SDQ) from 0.55 to 0.86 (P = 0.031). In highlanders, chronic hypoxia had attenuated effects on RVSP and no effects on smooth muscle thickness, right ventricle mass, or V-Q matching. Therefore, evolved changes in lung function help attenuate maladaptive plasticity and contribute to hypoxia tolerance in high-altitude deer mice.

Extrahypothalamic oxytocin neurons drive stress-induced social vigilance and avoidance.

Oxytocin increases the salience of both positive and negative social contexts and it is thought that these diverse actions on behavior are mediated in part through circuit-specific action. This hypothesis is based primarily on manipulations of oxytocin receptor function, leaving open the question of whether different populations of oxytocin neurons mediate different effects on behavior. Here we inhibited oxytocin synthesis in a stress-sensitive population of oxytocin neurons specifically within the medioventral bed nucleus of the stria terminalis (BNSTmv). Oxytocin knockdown prevented social stress-induced increases in social vigilance and decreases in social approach. Viral tracing of BNSTmv oxytocin neurons revealed fibers in regions controlling defensive behaviors, including lateral hypothalamus, anterior hypothalamus, and anteromedial BNST (BNSTam). Oxytocin infusion into BNSTam in stress naïve mice increased social vigilance and reduced social approach. These results show that a population of extrahypothalamic oxytocin neurons plays a key role in controlling stress-induced social anxiety behaviors.

Developmental exposure of California mice to endocrine disrupting chemicals and potential effects on the microbiome-gut-brain axis at adulthood.

Xenoestrogens are chemicals found in plant products, such as genistein (GEN), and in industrial chemicals, e.g., bisphenol A (BPA), present in plastics and other products that are prevalent in the environment. Early exposure to such endocrine disrupting chemicals (EDC) may affect brain development by directly disrupting neural programming and/or through the microbiome-gut-brain axis. To test this hypothesis, California mice (Peromyscus californicus) offspring were exposed through the maternal diet to GEN (250 mg/kg feed weight) or BPA (5 mg/kg feed weight, low dose- LD or 50 mg/kg, upper dose-UD), and dams were placed on these diets two weeks prior to breeding, throughout gestation, and lactation. Various behaviors, gut microbiota, and fecal metabolome were assessed at 90 days of age. The LD but not UD of BPA exposure resulted in individuals spending more time engaging in repetitive behaviors. GEN exposed individuals were more likely to exhibit such behaviors and showed socio-communicative disturbances. BPA and GEN exposed females had increased number of metabolites involved in carbohydrate metabolism and synthesis. Males exposed to BPA or GEN showed alterations in lysine degradation and phenylalanine and tyrosine metabolism. Current findings indicate cause for concern that developmental exposure to BPA or GEN might affect the microbiome-gut-brain axis.

Adaptive Shifts in Gene Regulation Underlie a Developmental Delay in Thermogenesis in High-Altitude Deer Mice.

Aerobic performance is tied to fitness as it influences an animal's ability to find food, escape predators, or survive extreme conditions. At high altitude, where low O2 availability and persistent cold prevail, maximum metabolic heat production (thermogenesis) is an aerobic performance trait that is closely linked to survival. Understanding how thermogenesis evolves to enhance survival at high altitude will yield insight into the links between physiology, performance, and fitness. Recent work in deer mice (Peromyscus maniculatus) has shown that adult mice native to high altitude have higher thermogenic capacities under hypoxia compared with lowland conspecifics, but that developing high-altitude pups delay the onset of thermogenesis. This finding suggests that natural selection on thermogenic capacity varies across life stages. To determine the mechanistic cause of this ontogenetic delay, we analyzed the transcriptomes of thermoeffector organs-brown adipose tissue and skeletal muscle-in developing deer mice native to low and high altitude. We demonstrate that the developmental delay in thermogenesis is associated with adaptive shifts in the expression of genes involved in nervous system development, fuel/O2 supply, and oxidative metabolism pathways. Our results demonstrate that selection has modified the developmental trajectory of the thermoregulatory system at high altitude and has done so by acting on the regulatory systems that control the maturation of thermoeffector tissues. We suggest that the cold and hypoxic conditions of high altitude force a resource allocation tradeoff, whereby limited energy is allocated to developmental processes such as growth, versus active thermogenesis, during early development.

Differential Expression in Testis and Liver Transcriptomes from Four Species of Peromyscus (Rodentia: Cricetidae).

The genus Peromyscus represents a rapidly diverged clade of Cricetid rodents that contains multiple cryptic species and has a propensity for morphologic conservation across its members. The unresolved relationships in previously proposed phylogenies reflect a suspected rapid adaptive radiation. To identify functional groups of genes that may be important in reproductive isolation in a reoccurring fashion across the Peromyscus phylogeny, liver and testis transcriptomes from four species (P. attwateri, P. boylii, P. leucopus, and P. maniculatus) were generated and differential expression (DE) tests were conducted. Taxa were selected to represent members diverged from a common ancestor: P. attwateri + P. boylii (clade A), and P. leucopus + P. maniculatus (clade B). Comparison of clades (A vs. B) suggested that 252 transcripts had significant DE in the liver data set, whereas significant DE was identified for 657 transcripts in the testis data set. Further, 45 genes had DE isoforms in the 657 testis transcripts and most of these functioned in major reproductive roles such as acrosome assembly, spermatogenesis, and cell cycle processes (meiosis). DE transcripts in the liver mapped to more broad gene ontology terms (metabolic processes, catabolic processes, response to chemical, and regulatory processes), and DE transcripts in the testis mapped to gene ontology terms associated with reproductive processes, such as meiosis, sperm motility, acrosome assembly, and sperm-egg fusion. These results suggest that a suite of genes that conduct similar functions in the testes may be responsible for the adaptive radiation events and potential reoccurring speciation of Peromyscus in terms of reproduction through varying expression levels.

Long-Term Effects of Fatherhood on Morphology, Energetics, and Exercise Performance in California Mice (Peromyscus californicus).

In male mammals that provide care for their offspring, fatherhood can lead to changes in behavioral, morphological, and physiological traits, some of which might constitute trade-offs. However, relatively little is known about these changes, especially across multiple reproductive bouts, which are expected to magnify differences between fathers and nonreproductive males. We evaluated consequences of fatherhood in the monogamous, biparental California mouse (Peromsycus californicus) across seven consecutive reproductive bouts. We compared breeding adult males (housed with sham-ovariectomized females) with two control groups: nonbreeding males (housed with ovariectomized females treated with estrogen and progesterone to induce estrous behavior) and virgin males (housed with untreated ovariectomized females). At five time points (before pairing, early postpartum of the first litter, late postpartum of the second litter, early postpartum of the sixth litter, and late postpartum of the seventh litter or comparable time points for nonbreeding and virgin males), we measured males' body composition, hematocrit, predatory aggression, resting metabolic rate, maximal oxygen consumption (V˙O2 max⁡), grip strength, and sprint speed. We also weighed organs at the final time point. We predicted that fathers would have lower relative body fat and lower performance abilities compared with control groups and that these effects would become more pronounced with increasing parity. Contrary to predictions, breeding and control males differed in surprisingly few measures, and the number and magnitude of differences did not increase with parity. Thus, our expectations regarding trade-offs were not met. As reported in studies of single reproductive events, these results suggest that fatherhood has few costs in this species when housed under standard laboratory conditions, even across multiple reproductive bouts.

Testosterone does not mediate variation in basal metabolic rate and activity in relation to reproductive condition and photoperiod in white-footed mice (Peromyscus leucopus).

The photoperiodic response of many temperate zone rodents, including white-footed mice (Peromyscus leucopus), is a heritable life-history trait with underlying physiological variation. Previous studies of intact male P. leucopus utilized two wild-derived bidirectional selection lines, a short photoperiod responsive (R) line selected for reproductive suppression in short-day conditions (SD) and a nonresponsive (NR) line selected for reproductive maturity in SD. NR mice in SD had greater food intake, but also higher levels of locomotor activity, and basal metabolic rate (BMR) than R mice. We hypothesized that testosterone may be a key mediator of this metabolic difference, as it is likely to be significantly reduced in R SD mice. Male P. leucopus from either line in SD were castrated and given either an implant containing testosterone (T) or a sham control (C). They were then tested for variation in metabolic rate and activity in SD, thermoneutral conditions. T mice had significantly higher levels of food intake, testosterone, and seminal vesicle dry weight than C mice. Seminal vesicle dry weight was significantly and positively correlated with average testosterone level, indicating an effect of the T implants. There was no statistically significant difference among treatment groups in BMR and average daily metabolic rate, suggesting that differences in testosterone alone are not the cause of differences in metabolic rate between selection lines.

Regulation of catecholamine release from the adrenal medulla is altered in deer mice (Peromyscus maniculatus) native to high altitudes.

High-altitude natives have evolved to overcome environmental hypoxia and provide a compelling system to understand physiological function during reductions in oxygen availability. The sympathoadrenal system plays a key role in responses to acute hypoxia, but prolonged activation of this system in chronic hypoxia may be maladaptive. Here, we examined how chronic hypoxia exposure alters adrenal catecholamine secretion and how adrenal function is altered further in high-altitude natives. Populations of deer mice (Peromyscus maniculatus) native to low and high altitudes were each born and raised in captivity at sea level, and adults from each population were exposed to normoxia or hypobaric hypoxia for 5 mo. Using carbon fiber amperometry on adrenal slices, catecholamine secretion evoked by low doses of nicotine (10 µM) or acute hypoxia (Po2 ∼15-20 mmHg) was reduced in lowlanders exposed to hypobaric hypoxia, which was attributable mainly to a decrease in quantal charge rather than event frequency. However, secretion evoked by high doses of nicotine (50 µM) was unaffected. Hypobaric hypoxia also reduced plasma epinephrine and protein expression of 3,4-dihydroxyphenylalanine (DOPA) decarboxylase in the adrenal medulla of lowlanders. In contrast, highlanders were unresponsive to hypobaric hypoxia, exhibiting typically low adrenal catecholamine secretion, plasma epinephrine, and DOPA decarboxylase. Highlanders also had consistently lower catecholamine secretion evoked by high nicotine, smaller adrenal medullae with fewer chromaffin cells, and a larger adrenal cortex compared with lowlanders across both acclimation environments. Our results suggest that plastic responses to chronic hypoxia along with evolved changes in adrenal function attenuate catecholamine release in deer mice at high altitude.

Social environment affects central distribution of estrogen receptor-α in Peromyscus californicus.

Social environment has well-established effects on an animal's social behavior and associated neuroendocrine responses. The presence of estrogen receptor alpha (ERα) in limbic system brain regions is related to the expression of a variety of social, reproductive and aggressive behaviors. We hypothesized that alterations to the social environment, specifically social isolation, would cause changes in ERα throughout the limbic system. The number of ERα immunoreactive (ERα-ir) cells within specific limbic system brain regions was quantified in male and female California mice (Peromyscus californicus), isolated or same sex pair-housed for 4 or 24 days. Peromyscus californicus is a highly social rodent species (monogamous and bi-parental) and therefore, may be particularly sensitive to manipulations of its social environment. Isolated males had a significantly greater number of ERα-ir cells in the ventromedial nucleus of the hypothalamus (VMH) and similar patterns within the bed nucleus of the stria terminalis (BST) and medial preoptic area (MPOA). Males housed for 24 days had a significantly greater number of ERα-ir cells in the BST, VMH, MPOA when compared with males housed for 4 days. Females housed for 24 days had significantly greater ERα-ir in the dentate gyrus of the hippocampus (DG) when compared with females housed for 4 days. No differences were found in the medial amygdala (MeA). These data demonstrate that social environment has region and sex specific effects on ERα-ir cells in this species. These results add to the comparative evidence regarding ERα, demonstrating a consistent role for ERα in species specific responsiveness to changes in the social environment.

Peromyscus maniculatus bairdii as a naturalistic mammalian model of obsessive-compulsive disorder: current status and future challenges.

Obsessive-compulsive disorder (OCD) is a prevalent and debilitating condition, characterized by intrusive thoughts and repetitive behavior. Animal models of OCD arguably have the potential to contribute to our understanding of the condition. Deer mice (Permomyscus maniculatus bairdii) are characterized by stereotypic behavior which is reminiscent of OCD symptomology, and which may serve as a naturalistic animal model of this disorder. Moreover, a range of deer mouse repetitive behaviors may be representative of different compulsive-like phenotypes. This paper will review work on deer mouse behavior, and evaluate the extent to which this serves as a valid and useful model of OCD. We argue that findings over the past decade indicate that the deer mouse model has face, construct and predictive validity.

Alteration of the α1ß2/α2ß1 subunit interface contributes to the increased hemoglobin-oxygen affinity of high-altitude deer mice.

BACKGROUND: Deer mice (Peromyscus maniculatus) that are native to high altitudes in the Rocky Mountains have evolved hemoglobins with an increased oxygen-binding affinity relative to those of lowland conspecifics. To elucidate the molecular mechanisms responsible for the evolved increase in hemoglobin-oxygen affinity, the crystal structure of the highland hemoglobin variant was solved and compared with the previously reported structure for the lowland variant. RESULTS: Highland hemoglobin yielded at least two crystal types, in which the longest axes were 507 and 230 Å. Using the smaller unit cell crystal, the structure was solved at 2.2 Å resolution. The asymmetric unit contained two tetrameric hemoglobin molecules. CONCLUSIONS: The analyses revealed that αPro50 in the highland hemoglobin variant promoted a stable interaction between αHis45 and heme that was not seen in the αHis50 lowland variant. The αPro50 mutation also altered the nature of atomic contacts at the α1ß2/α2ß1 intersubunit interfaces. These results demonstrate how affinity-altering changes in intersubunit interactions can be produced by mutations at structurally remote sites.

Effects of repeated pup exposure on behavioral, neural, and adrenocortical responses to pups in male California mice (Peromyscus californicus).

In biparental mammals, the factors facilitating the onset of male parental behavior are not well understood. While hormonal changes in fathers may play a role, prior experience with pups has also been implicated. We evaluated effects of prior exposure to pups on paternal responsiveness in the biparental California mouse (Peromyscus californicus). We analyzed behavioral, neural, and corticosterone responses to pups in adult virgin males that were interacting with a pup for the first time, adult virgin males that had been exposed to pups 3 times for 20min each in the previous week, and new fathers. Control groups of virgins were similarly tested with a novel object (marble). Previous exposure to pups decreased virgins' latency to approach pups and initiate paternal care, and increased time spent in paternal care. Responses to pups did not differ between virgins with repeated exposure to pups and new fathers. In contrast, repeated exposure to a marble had no effects. Neither basal corticosterone levels nor corticosterone levels following acute pup or marble exposure differed among groups. Finally, Fos expression in the medial preoptic area, ventral and dorsal bed nucleus of the stria terminalis was higher following exposure to a pup than to a marble. Fos expression was not, however, affected by previous exposure to these stimuli. These results suggest that previous experience with pups can facilitate the onset of parental behavior in male California mice, similar to findings in female rodents, and that this effect is not associated with a general reduction in neophobia.

Cytokine mRNA expression in Peromyscus yucatanicus (Rodentia: Cricetidae) infected by Leishmania (Leishmania) mexicana.

Peromyscus yucatanicus, the main reservoir of Leishmania (Leishmania) mexicana in the Yucatan peninsula of Mexico, reproduces clinical and histological pictures of LCL in human as well as subclinical infection. Thus, we used this rodent as a novel experimental model. In this work, we analyzed cytokine mRNA expression in P. yucatanicus infected with L. (L.) mexicana. Animals were inoculated with either 2.5×10(6) or 1×10(2) promastigotes and cytokine expressions were analyzed by real-time RT-PCR in skin at 4 and 12weeks post-infection (wpi). Independently of the parasite inoculum none of the infected rodents had clinical signs of LCL at 4wpi and all expressed high IFN-γ mRNA. All P. yucatanicus inoculated with 2.5×10(6) promastigotes developed signs of LCL at 12wpi while the mice inoculated with 1×10(2) remained subclinical. At that time, both IFN-γ and IL-10 were expressed in P. yucatanicus with clinical and subclinical infections. Expressions of TNF-α and IL-4 were significantly higher in clinical animals (2.5×10(6)) compared with subclinical ones (1×10(2)). High TGF-ß expression was observed in P. yucatanicus with clinical signs when compared with healthy animals. Results suggested that the clinical course of L. (L.) mexicana infection in P. yucatanicus was associated with a specific local pattern of cytokine production at 12wpi.

Estrogen Receptor Alpha Distribution and Expression in the Social Neural Network of Monogamous and Polygynous Peromyscus.

In microtine and dwarf hamsters low levels of estrogen receptor alpha (ERα) in the bed nucleus of the stria terminalis (BST) and medial amygdala (MeA) play a critical role in the expression of social monogamy in males, which is characterized by high levels of affiliation and low levels of aggression. In contrast, monogamous Peromyscus males display high levels of aggression and affiliative behavior with high levels of testosterone and aromatase activity. Suggesting the hypothesis that in Peromyscus ERα expression will be positively correlated with high levels of male prosocial behavior and aggression. ERα expression was compared within the social neural network, including the posterior medial BST, MeA posterodorsal, medial preoptic area (MPOA), ventromedial hypothalamus (VMH), and arcuate nucleus in two monogamous species, P. californicus and P. polionotus, and two polygynous species, P. leucopus and P. maniculatus. The results supported the prediction, with male P. polionotus and P. californicus expressing higher levels of ERα in the BST than their polygynous counter parts, and ERα expression was sexually dimorphic in the polygynous species, with females expressing significantly more than males in the BST in both polygynous species and in the MeA in P. leucopus. Peromyscus ERα expression also differed from rats, mice and microtines as in neither the MPOA nor the VMH was ERα sexually dimorphic. The results supported the hypothesis that higher levels of ERα are associated with monogamy in Peromyscus and that differential expression of ERα occurs in the same regions of the brains regardless of whether high or low expression is associated with social monogamy. Also discussed are possible mechanisms regulating this differential relationship.

Genetic variation in total number and locations of GnRH neurons identified using in situ hybridization in a wild-source population.

The evolution of brain function in the regulation of physiology may depend in part upon the numbers and locations of neurons. Wild populations of rodents contain natural genetic variation in the inhibition of reproduction by winter-like short photoperiod, and it has been hypothesized that this functional variation might be due in part to heritable variation in the numbers or location of gonadotropin releasing hormone (GnRH) neurons. A naturally variable wild-source population of white-footed mice was used to develop lines artificially selected for or against mature gonads in short, winter-like photoperiods. We compared a selection line that is reproductively inhibited in short photoperiod (Responsive) to a line that is weakly inhibited by short photoperiod (Nonresponsive) for differences in counts of neurons identified using in situ hybridization for GnRH mRNA. There was no effect of photoperiod, but there were 60% more GnRH neurons in total in the Nonresponsive selection line than the Responsive selection line. The lines differed specifically in numbers of GnRH neurons in more anterior regions, whereas numbers of GnRH neurons in posterior areas were not statistically different between lines. We compare these results to those of an earlier study that used immunohistochemical labeling for GnRH neurons. The results are consistent with the hypothesis that the selection lines and natural source population contain significant genetic variation in the number and location of GnRH neurons. The variation in GnRH neurons may contribute to functional variation in fertility that occurs in short photoperiods in the laboratory and in the wild source population in winter.

Paracellular nutrient absorption is higher in bats than rodents: integrating from intact animals to the molecular level.

Flying vertebrates have been hypothesized to rely heavily on paracellular absorption of nutrients to compensate for having smaller intestines than non-flyers. We tested this hypothesis in an insectivorous bat (Myotis lucifugus) and two insect-eating rodents (Onychomys leucogaster and Peromyscus leucopus). In intact animals, the fractional absorption of orally dosed l-arabinose (Mr 150) was 82% in M. lucifugus, which was more than twice that of the rodents. Absorption of creatinine (Mr 113) was greater than 50% for all species and did not differ between M. lucifugus and the rodents. We also conducted intestinal luminal perfusions on anesthetized animals. Absorption of l-arabinose per nominal surface area in M. lucifugus was nearly double that of the rodents, while absorption of creatinine was not different among species. Using an everted sleeve preparation, we demonstrated that high concentrations of l-arabinose and creatinine did not inhibit their own uptake, validating their use as passive, paracellular probes. Histological measurements indicated that M. lucifugus has more cells, and presumably more tight junctions, per nominal surface area than P. leucopus. This seems unlikely to explain entirely the higher absorption of l-arabinose in M. lucifugus during perfusions, because l-arabinose absorption normalized to the number of enterocytes was still double that of P. leucopus. As an alternative, we investigated tight junction gene expression. M. lucifugus had higher expression of claudin-1 and claudin-15, and lower expression of claudin-2 relative to P. leucopus. Expression of claudin-7 and occludin did not differ among species. Taken together, our results support the hypothesis that bats have evolved higher paracellular nutrient absorption than non-flying animals, and that this phenomenon might be driven by both histological characteristics and differences in tight junction gene expression.

The effects of exogenous melatonin and melatonin receptor blockade on aggression and estrogen-dependent gene expression in male California mice (Peromyscus californicus).

Photoperiodic regulation of aggression has been well established in several vertebrate species, with rodents demonstrating increased aggression in short day photoperiods as compared to long day photoperiods. Previous work suggests that estrogens regulate aggression via rapid nongenomic pathways in short days and act more slowly in long days, most likely via genomic pathways. The current study therefore examines the role of melatonin in mediating aggression and estrogen-dependent gene transcription. In Experiment 1, male California mice were housed under long day photoperiods and were treated with either 0.3 µg/g of melatonin, 40 mg/kg of the melatonin receptor antagonist luzindole, or vehicle for 10 days. We found that melatonin administration significantly increased aggression as compared to mice receiving vehicle, but this phenotype was not completely ameliorated by luzindole. In Experiment 2, male California mice were injected with either 1mg/kg of the aromatase inhibitor letrozole or vehicle, and oxytocin receptor (OTR), estrogen receptor alpha (ERα), and c-fos gene expression was examined in the bed nucleus of the stria terminalis (BNST) and medial preoptic area (MPOA). In the BNST, but not MPOA, OTR mRNA was significantly downregulated following letrozole administration, indicating that OTR is an estrogen-dependent gene in the BNST. In contrast, ERα was not estrogen dependent in either brain region. In the MPOA, OTR mRNA was inhibited by melatonin, and luzindole suppressed this effect. C-fos and ERα did not differ between treatments in any brain region examined. These results suggest that it is unlikely that melatonin facilitates aggression via broad spectrum regulation of estrogen-dependent gene expression. Instead, melatonin may act via regulation of other transcription factors such as extracellular signal regulated kinase.

Functional genomics of adaptation to hypoxic cold-stress in high-altitude deer mice: transcriptomic plasticity and thermogenic performance.

In species that are distributed across steep environmental gradients, adaptive variation in physiological performance may be attributable to transcriptional plasticity in underlying regulatory networks. Here we report the results of common-garden experiments that were designed to elucidate the role of regulatory plasticity in evolutionary adaptation to hypoxic cold-stress in deer mice (Peromyscus maniculatus). We integrated genomic transcriptional profiles with measures of metabolic enzyme activities and whole-animal thermogenic performance under hypoxia in highland (4350 m) and lowland (430 m) mice from three experimental groups: (1) wild-caught mice that were sampled at their native elevations; (2) wild-caught/lab-reared mice that were deacclimated to low-elevation conditions in a common-garden lab environment; and (3) the F(1) progeny of deacclimated mice that were maintained under the same low-elevation common-garden conditions. In each experimental group, highland mice exhibited greater thermogenic capacities than lowland mice, and this enhanced performance was associated with upregulation of transcriptional modules that influence several hierarchical steps in the O(2) cascade, including tissue O(2) diffusion (angiogenesis) and tissue O(2) utilization (metabolic fuel use and cellular oxidative capacity). Most of these performance-related transcriptomic changes occurred over physiological and developmental timescales, suggesting that regulatory plasticity makes important contributions to fitness-related physiological performance in highland deer mice.