Aging and neurodegeneration are often accompanied by a functionally impaired ubiquitin–proteasome system (UPS). In tauopathies and polyglutamine diseases, a mutant form of ubiquitin B (UBB+1) accumulates in disease-specific aggregates. UBB+1 mRNA is generated at low levels in vivo during transcription from the ubiquitin B locus by molecular misreading. The resulting mutant protein has been shown to inhibit proteasome function. To elucidate causative effects and neuropathological consequences of UBB+1 accumulation, we used a UBB+1 expressing transgenic mouse line that models UPS inhibition in neurons and exhibits behavioral phenotypes reminiscent of Alzheimer’s disease (AD). In order to reveal affected organs and functions, young and aged UBB+1 transgenic mice were comprehensively phenotyped for more than 240 parameters. This revealed unexpected changes in spontaneous breathing patterns and an altered response to hypoxic conditions. Our findings point to a central dysfunction of respiratory regulation in transgenic mice in comparison to wild-type littermate mice. Accordingly, UBB+1 was strongly expressed in brainstem regions of transgenic mice controlling respiration. These regions included, e.g., the medial part of the nucleus of the tractus solitarius and the lateral subdivisions of the parabrachial nucleus. In addition, UBB+1 was also strongly expressed in these anatomical structures of AD patients (Braak stage #6) and was not expressed in non-demented controls. We conclude that long-term UPS inhibition due to UBB+1 expression causes central breathing dysfunction in a transgenic mouse model of AD. The UBB+1 expression pattern in humans is consistent with the contribution of bronchopneumonia as a cause of death in AD patients.

Neurobeachin (Nbea) regulates neuronal membrane protein trafficking and is required for the development and functioning of central and neuromuscular synapses. In homozygous knockout (KO) mice, Nbea deficiency causes perinatal death. Here, we report that heterozygous KO mice haploinsufficient for Nbea have higher body weight due to increased adipose tissue mass. In several feeding paradigms, heterozygous KO mice consumed more food than wild-type (WT) controls, and this consumption was primarily driven by calories rather than palatability. Expression analysis of feeding-related genes in the hypothalamus and brainstem with real-time PCR showed differential expression of a subset of neuropeptide or neuropeptide receptor mRNAs between WT and Nbea+/- mice in the sated state and in response to food deprivation, but not to feeding reward. In humans, we identified two intronic NBEA single-nucleotide polymorphisms (SNPs) that are significantly associated with body-mass index (BMI) in adult and juvenile cohorts. Overall, data obtained in mice and humans suggest that variation of Nbea abundance or activity critically affects body weight, presumably by influencing the activity of feeding-related neural circuits. Our study emphasizes the importance of neural mechanisms in body weight control and points out NBEA as a potential risk gene in human obesity.

Despite extensive investigations of Cbl-interacting protein of 85 kDa (CIN85) in receptor trafficking and cytoskeletal dynamics, little is known about its functions in vivo. Here, we report the study of a mouse deficient of the two CIN85 isoforms expressed in the central nervous system, exposing a function of CIN85 in dopamine receptor endocytosis. Mice lacking CIN85 exon 2 (CIN85(Deltaex2)) show hyperactivity phenotypes, characterized by increased physical activity and exploratory behaviour. Interestingly, CIN85(Deltaex2) animals display abnormally high levels of dopamine and D2 dopamine receptors (D2DRs) in the striatum, an important centre for the coordination of animal behaviour. Importantly, CIN85 localizes to the post-synaptic compartment of striatal neurons in which it co-clusters with D2DRs. Moreover, it interacts with endocytic regulators such as dynamin and endophilins in the striatum. Absence of striatal CIN85 causes insufficient complex formation of endophilins with D2DRs in the striatum and ultimately decreased D2DR endocytosis in striatal neurons in response to dopamine stimulation. These findings indicate an important function of CIN85 in the regulation of dopamine receptor functions and provide a molecular explanation for the hyperactive behaviour of CIN85(Deltaex2) mice.

Background: In a variety of organisms, including mammals, caloric restriction improves metabolic status and lowers the incidence of chronic-degenerative diseases, ultimately leading to increased lifespan.Methodology/Principal Findings: Here we show that knockout mice for Eps8, a regulator of actin dynamics, display reduced body weight, partial resistance to age-or diet-induced obesity, and overall improved metabolic status. Alteration in the liver gene expression profile, in behavior and metabolism point to a calorie restriction-like phenotype in Eps8 knockout mice. Additionally, and consistent with a calorie restricted metabolism, Eps8 knockout mice show increased lifespan. The metabolic alterations in Eps8 knockout mice correlated with a significant reduction in intestinal fat absorption presumably caused by a 25% reduction in intestinal microvilli length.Conclusions/Significance: Our findings implicate actin dynamics as a novel variable in the determination of longevity. Additionally, our observations suggest that subtle differences in energy balance can, over time, significantly affect bodyweight and metabolic status in mice.

With the completion of the mouse genome sequence an essential task for biomedical sciences in the twenty-first century will be the generation and functional analysis of mouse models for every gene in the mammalian genome. More than 30,000 mutations in ES cells will be engineered and thousands of mouse disease models will become available over the coming years by the collaborative effort of the International Mouse Knockout Consortium. In order to realize the full value of the mouse models proper characterization, archiving and dissemination of mouse disease models to the research community have to be performed. Phenotyping centers (mouse clinics) provide the necessary capacity, broad expertise, equipment, and infrastructure to carry out large-scale systemic first-line phenotyping. Using the example of the German Mouse Clinic (GMC) we will introduce the reader to the different aspects of the organization of a mouse clinic and present selected methods used in first-line phenotyping.

It has been proposed that two amino acid substitutions in the transcription factor FOXP2 have been positively selected during human evolution due to effects on aspects of speech and language. Here, we introduce these substitutions into the endogenous Foxp2 gene of mice. Although these mice are generally healthy, they have qualitatively different ultrasonic vocalizations, decreased exploratory behavior and decreased dopamine concentrations in the brain suggesting that the humanized Foxp2 allele affects basal ganglia. In the striatum, a part of the basal ganglia affected in humans with a speech deficit due to a nonfunctional FOXP2 allele, we find that medium spiny neurons have increased dendrite lengths and increased synaptic plasticity. Since mice carrying one nonfunctional Foxp2 allele show opposite effects, this suggests that alterations in cortico-basal ganglia circuits might have been important for the evolution of speech and language in humans.For a video summary of this article, see the PaperFlick file available with the online Supplemental Data.

The German Mouse Clinic (GMC) is a large scale phenotyping center where mouse mutant lines are analyzed in a standardized and comprehensive way. The result is an almost complete picture of the phenotype of a mouse mutant line--a systemic view. At the GMC, expert scientists from various fields of mouse research work in close cooperation with clinicians side by side at one location. The phenotype screens comprise the following areas: allergy, behavior, clinical chemistry, cardiovascular analyses, dysmorphology, bone and cartilage, energy metabolism, eye and vision, host-pathogen interactions, immunology, lung function, molecular phenotyping, neurology, nociception, steroid metabolism, and pathology. The German Mouse Clinic is an open access platform that offers a collaboration-based phenotyping to the scientific community (www.mouseclinic.de). More than 80 mutant lines have been analyzed in a primary screen for 320 parameters, and for 95% of the mutant lines we have found new or additional phenotypes that were not associated with the mouse line before. Our data contributed to the association of mutant mouse lines to the corresponding human disease. In addition, the systemic phenotype analysis accounts for pleiotropic gene functions and refines previous phenotypic characterizations. This is an important basis for the analysis of underlying disease mechanisms. We are currently setting up a platform that will include environmental challenge tests to decipher genome-environmental interactions in the areas nutrition, exercise, air, stress and infection with different standardized experiments. This will help us to identify genetic predispositions as susceptibility factors for environmental influences.

Much of our understanding of physiology and metabolism is derived from investigating mouse mutants and transgenic mice, and open-access platforms for standardized mouse phenotyping such as the German Mouse Clinic (GMC) are currently viewed as one powerful tool for identifying novel gene-function relationships. Phenotyping or phenotypic screening involves the comparison of wild-type control mice with their mutant or transgenic littermates. In our study, we explored the extent to which standardized phenotyping will succeed in detecting biologically relevant phenotypic differences in mice generated and provided by different collaborators. We analyzed quantitative metabolic data (body mass, energy intake, and energy metabolized) collected at the GMC under the current workflow, and used them for statistical power considerations. Our results demonstrate that there is substantial variability in these parameters among lines of wild-type C57BL/6 (B6) mice from different sources. Given this variable background noise in mice that serve as controls, subtle phenotypes in mutant or transgenic littermates may be overlooked. Furthermore, a phenotype observed in one cohort of a mutant line may not be reproducible (to the same extent) in mice coming from a different environment or supplier. In the light of these constraints, we encourage researchers to incorporate information on intrastrain variability into future study planning, or to perform advanced hierarchical analyses. Both will ultimately improve the detectability of novel phenotypes by phenotypic screening.

Housing conditions are known to influence laboratory animal behavior. However, it is not known whether housing mice in individually ventilated cages (IVCs) to maintain optimal hygienic conditions alters behavioral baselines established in conventional housing. This issue is important with regard to comparability and reproducibility of data. Therefore, we investigated the impact of IVC housing on emotionality and fear learning in male C3HeB/FeJ (C3H) and C57BL/6J (B6J) mice housed singly either in conventional type II cages with wire bar lids (Conventional), or in IVCs of the same size, but with smooth, untextured lids (IVC classic), thus acoustically attenuated from external stimuli and with limited climbing facilities compared to Conventional. To evaluate the role of climbing, additional mice were kept in IVCs with lids having wire bars ("grid") added to the inner surface (IVC grid). Spontaneous behavior, sensorimotor behavior, and fear learning were measured. IVC housing reduced activity and enhanced anxiety-related behavior in both strains, whereas grooming latency was reduced in B6J only. IVC housing increased Acoustic Startle Response in C3H but not in B6J mice. The "grid" did not compensate for these IVC housing effects. In contrast, B6J mice in IVC grid performed best in fear potentiated startle while B6J mice in IVC classic performed the worst, suggesting that climbing facilities combined with IVC housing facilitate FPS performance in singly-housed B6J males. Our data show that IVC housing can affect behavioral performance and can modulate behavioral parameters in a general and a strain-specific manner, thus having an impact on mouse functional genomics.

Effects of a Protein Optimized Diet Combined with Moderate Resistance Training on the Postoperative Course in Older Patients with Hip Fracture

Housing conditions are known to influence laboratory animal behaviour. However, it is not known whether housing mice in individually ventilated cages (IVCs) to maintain optimal hygienic conditions alters behavioural baselines established in conventional housing. This issue is important regarding comparability and reproducibility of data. Therefore, we investigated the impact of IVC housing on emotionality and fear learning in male C3HeB/FeJ (C3H) and C57BL/6J (B6J) mice housed singly either in conventional type II cages with wire bar lids (Conventional), or in IVCs of the same size, but with smooth, untextured lids (IVC classic), thus acoustically attenuated from external stimuli and with limited climbing facilities as compared to Conventional. To evaluate the role of climbing, additional mice were kept in IVCs with lids having wire bars (“grid”) added to the surface (IVC grid). Spontaneous behaviour, sensorimotor behaviour and fear learning were measured. IVC housing reduced activity and enhanced anxiety-related behaviour in both strains, whereas grooming latency was reduced in B6J only. IVC housing increased Acoustic Startle response in C3H, not in B6J mice. The “grid” did not compensate for these IVC housing effects. In contrast, B6J mice in IVC grid performed best in Fear Potentiated Startle, B6J mice in IVC classic performed worst, suggesting climbing facilities combined with IVC housing facilitate FPS performance in singly housed B6J males. Our data show that IVC housing can affect behavioural performance, and can modulate behavioural parameters in a general as well as a strainspecific manner, both relevant for mouse functional genomics.

The German Mouse Clinic was established as a phenotyping center to provide the scientific community with systematic standardized phenotyping of mouse models from various genetic backgrounds. We found metabolic phenotypes in nine out of 20 mutant lines screened in a primary screen. Based on these findings, the mutants were analyzed in secondary and tertiary screens. Mice of a sample mutant line, isolated from the ENU-screen at the National Research Center for Environment and Health in Munich, were found to have lower body weight, consume less food, but have higher ratios of metabolized energy per unit body weight compared with their wild-type littermates. Basal metabolic rate and heat production were simultaneously increased by 16-18%, whereas body fat content was reduced by 11-16%. The combination of various parameters of energy consumption, expenditure and energy storage illustrate the metabolic demands of the sample mutant mouse line and demonstrate the utility of the powerful phenotyping tool used at the German Mouse Clinic.

The golden spiny mouse (Acomys russatus) is an omnivorous desert rodent that does not store food, but can store large amounts of body fat. Thus, it provides a good animal model to study physiological and behavioural adaptations to changes in food availability. The aim of this study was to investigate the time course of metabolic and behavioural responses to prolonged food restriction. Spiny mice were kept at an ambient temperature of 27°C and for 3 weeks their food was reduced individually to 30% of their previous ad libitum food intake. When fed ad libitum, their average metabolic rate was 82.77±3.72 ml O2 h−1 during the photophase and 111.19±4.30 ml O2 h−1 during the scotophase. During food restriction they displayed episodes of daily torpor when the minimal metabolic rate gradually decreased to 16.07±1.07 ml O2 h−1, i.e. a metabolic rate depression of approximately 83%. During the hypometabolic bouts the minimum average body temperature Tb, decreased gradually from 32.6±0.1°C to 29.0±0.4°C, with increasing duration of consecutive bouts. In parallel, the animals increased their activity during the remaining daytime. Torpor as well as hyperactivity was suppressed immediately by refeeding. Thus golden spiny mice used two simultaneous strategies to adapt to shortened food supply, namely energysaving torpor during their resting period and an increase in locomotor activity pattern during their activity period.