Diverse Mouse Strains Research Articles

Further Evidence That the Soluble Urokinase Plasminogen Activator Receptor Does Not Directly Injure Mice or Human Podocytes.

The role of the soluble urokinase plasminogen activator receptor (suPAR) in focal segmental glomerulosclerosis (FSGS) as the circulating factor or as a predictor of recurrence after transplantation remains controversial. Previously published studies in mice and isolated podocytes produced conflicting results on the effect of suPAR on podocyte injury, effacement of foot processes, and proteinuria. These discordant results were in part due to diverse experimental designs and different strains of mice. The aim of our study was to determine the reasons for the inconsistencies of the previous studies results with suPAR by using uniform methods and studies in different strains of mice. We utilized a primary culture of human podocytes and 2 mouse models, the wild type (WT) and the urokinase plasminogen activator receptor (uPAR) KO (uPAR), in an attempt to resolve the reported conflicting results. In both WT and uPAR mouse models, injection of recombinant uPAR, even at a high dose (100 μg), did not induce proteinuria, effacement of podocytes, or disruption of the cytoskeleton. Injection of suPAR resulted in its deposition exclusively in the glomerular endothelial cells and not in the podocytes of WT mice and was not detected at the uPAR KO mice. Kidneys from patients with recurrent FSGS had negative immunostaining for uPAR. We also evaluated the effect of recombinant uPAR on primary culture of human podocytes. uPAR did not result in podocytes damage. suPAR by itself is not the cause for direct podocyte injury, in vitro or in vivo. These findings suggest a more complex and still poorly understood role of suPAR in FSGS.

DNA Methylation Changes More Slowly Than Physiological States in Response to Weight Loss in Genetically Diverse Mouse Strains.

Responses to a high fat, high sucrose (HFHS) diet vary greatly among inbred strains of mice. We sought to examine the epigenetic (DNA methylation) changes underlying these differences as well as variation in weight loss when switched to a low-fat chow diet. We surveyed DNA methylation from livers of 45 inbred mouse strains fed a HFHS diet for 8 weeks using reduced-representation bisulfite sequencing (RRBS). We observed a total of 1,045,665 CpGs of which 83 candidate sites were significantly associated with HFHS diet. Many of these CpGs correlated strongly with gene expression or clinical traits such as body fat percentage and plasma glucose. Five inbred strains were then studied in the context of weight loss to test for evidence of epigenetic “memory.” The mice were first fed a HFHS diet for 6 weeks followed by a low-fat chow diet for 4 weeks. Four of the five strains returned to initial levels of body fat while one strain, A/J, retained almost 50% of the fat gained. A total of 36 of the HFHS diet responsive CpGs exhibited evidence of persistent epigenetic modifications following weight normalization, including CpGs near the genes Scd1 and Cdk1. Our study identifies DNA methylation changes in response to a HFHS diet challenge that revert more slowly than overall body fat percentage in weight loss and provides evidence for epigenetic mediated “memory.”

Isoproterenol-Induced Cardiac Diastolic Dysfunction in Mice: A Systems Genetics Analysis

We examined an isoproterenol heart failure model across a panel of diverse inbred strains of mice, the Hybrid Mouse Diversity Panel (HMDP), using left atrial (LA) and lung weights as well as echocardiogram parameters as surrogates for cardiac diastolic function. We identified gene transcripts that significantly correlated with diastolic function. In addition, we mapped echocardiographic parameters associated with diastolic function. We identified a locus near Tns3-Hus1 to be associated with baseline E/A ratio in mice (p = 1.65E-06), the syntenic region of which was recently associated with E/A ratio in a genome-wide association study (GWAS) meta-analysis of the EchoGen consortium in humans. We also identified a locus near Cdkn2a-Cdkn2b, which is a region syntenic to the human 9p21 locus, to be associated with week 3 A/E ratio (p = 2.15E-06). Our study is the first study to map diastolic dysfunction in mice, in which a locus was found to be shared with a recent human GWAS on diastolic function. Moreover, our cardiac transcriptome correlation and eQTL analysis generated hypotheses for future basic investigations. These results showed that, although technical and physiological challenges limit diastolic function assessment in mice and humans, future investigations examining the genetic architecture of diastolic function among a diverse mouse population, such as the HMDP, in controlled experimental settings, offer distinct advantages in understanding the genetic determinants of diastolic function.

Abstract 2745: Tumor microenvironment and host genetics impact glioma progression in a Collaborative Cross-based orthotopic allograft model

Abstract Gliomas are diffusely invasive brain tumors with fatal outcomes and few effective treatments. Precision medicine focuses on targeting the genetics of individual tumors, but not host genetics, despite studies that have linked germline polymorphisms with glioma risk. Accordingly, glioma survival studies in mice utilize genetically variable tumors on identical host genetic backgrounds, which fails to differentiate between cancer cell-autonomous (CCA) and tumor microenvironment (TME) effects on glioma progression and host survival. The Collaborative Cross (CC) is a panel of genetically diverse mouse strains derived from both wild- and traditional inbred laboratory strains that facilitates high-resolution genetic mapping in models of complex disease. Here, we implement a novel platform to discover genetic modifiers of both CCA and TME phenotypes using genetically defined orthotopic murine allograft gliomas and CC hosts. We stereotactically injected Nf1;Trp53-/-oligodendrocyte progenitor-derived mouse tumor cells into syngeneic C57BL/6 control mice and 14 different CC strains. Seven strains survived significantly longer than controls (P<0.05), suggesting slower tumor growth (Gs, growth slow). The remaining 7 strains survived similarly to controls, suggesting fast growth (Gf, growth fast). Variable tumor growth in CC mice suggests that genetic background influences molecular processes in the TME that inhibit or potentiate tumor growth, respectively. To identify candidate genes, we performed RNA sequencing on 36 tumors from 3 Gf strains, 4 Gs strains, and controls. 134 genes were differentially expressed among Gf, Gs, and control tumors (P<0.05). Hierarchical clustering on these genes revealed that Gs strains clustered separately from Gf and controls. Gene ontology analysis using GOrilla showed 30 enriched processes, (FDR q<0.001), all of which were involved in immune responses or extracellular matrix biology. These results suggest that Gs strains activate immune and TME processes that slow tumor growth. Quantitative trait locus (QTL) analyses of host genetics and tumor data are pending and will facilitate identification of genetic variants that influence TME effects on tumor progression. Citation Format: Kasey Skinner, Martin Ferris, Ryan Bash, Abigail Shelton, Erin Smithberger, Steve Angus, Brian Golitz, Noah Sciaky, Jeremy Simon, Jason Stein, Glenn Matsushima, Quinn Ostrom, Lindsay Stetson, Jill Barnholtz-Sloan, Harshil Dhruv, Michael Berens, Fernando Pardo Manuel de Villena, C. Ryan Miller. Tumor microenvironment and host genetics impact glioma progression in a Collaborative Cross-based orthotopic allograft model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2745.

Comparison and Functional Genetic Analysis of Striatal Protein Expression Among Diverse Inbred Mouse Strains.

C57BL/6J (B6) and DBA/2J (D2) inbred mouse strains are highly variable genetically and differ in a large number of behavioral traits related to striatal function, including depression, anxiety, stress response, and response to drugs of abuse. The genetic basis of these phenotypic differences are, however, unknown. Here, we present a comparison of the striatal proteome between B6 and D2 and relate differences at the protein level to strain differences at the mRNA level. We also leverage a recombinant inbred BXD population derived from B6 and D2 strains to investigate the role of genetic variation on the regulation of mRNA and protein levels. Finally, we test the hypothesis that differential protein expression contributes to differential behavioral responses between the B6 and D2 strain. We detected the expression of over 2,500 proteins in membrane-enriched protein fractions from B6 and D2 striatum. Of these, 160 proteins demonstrated significant differential expression between B6 and D2 strains at a 10% false discovery level, including COMT, GABRA2, and cannabinoid receptor 1 (CNR1)—key proteins involved in synaptic transmission and behavioral response. Similar to previous reports, there was little overlap between protein and transcript levels (25%). However, the overlap was greater (51%) for proteins demonstrating genetic regulation of cognate gene expression. We also found that striatal proteins with significantly higher or lower relative expression in B6 and D2 were enriched for dopaminergic and glutamatergic synapses and processes involved in synaptic plasticity [e.g., long-term potentiation (LTP) and long-term depression (LTD)]. Finally, we validated higher expression of CNR1 in B6 striatum and demonstrated greater sensitivity of this strain to the locomotor inhibiting effects of the CNR1 agonist, Δ9-tetrahydrocannabinol (THC). Our study is the first comparison of differences in striatal proteins between the B6 and D2 strains and suggests that alterations in the striatal proteome may underlie strain differences in related behaviors, such as drug response. Furthermore, we propose that genetic variants that impact transcript levels are more likely to also exhibit differential expression at the protein level.

Variable outcomes of human heart attack recapitulated in genetically diverse mice

Clinical variation in patient responses to myocardial infarction (MI) has been difficult to model in laboratory animals. To assess the genetic basis of variation in outcomes after heart attack, we characterized responses to acute MI in the Collaborative Cross (CC), a multi-parental panel of genetically diverse mouse strains. Striking differences in post-MI functional, morphological, and myocardial scar features were detected across 32 CC founder and recombinant inbred strains. Transcriptomic analyses revealed a plausible link between increased intrinsic cardiac oxidative phosphorylation levels and MI-induced heart failure. The emergence of significant quantitative trait loci for several post-MI traits indicates that utilizing CC strains is a valid approach for gene network discovery in cardiovascular disease, enabling more accurate clinical risk assessment and prediction.

Genetic control of the mouse HDL proteome defines HDL traits, function, and heterogeneity

HDLs are nanoparticles with more than 80 associated proteins, phospholipids, cholesterol, and cholesteryl esters. The potential inverse relation of HDL to coronary artery disease (CAD) and the effects of HDL on myriad other inflammatory conditions warrant a better understanding of the genetic basis of the HDL proteome. We conducted a comprehensive genetic analysis of the regulation of the proteome of HDL isolated from a panel of 100 diverse inbred strains of mice (the hybrid mouse diversity panel) and examined protein composition and efflux capacity to identify novel factors that affect the HDL proteome. Genetic analysis revealed widely varied HDL protein levels across the strains. Some of this variation was explained by local cis-acting regulation, termed cis-protein quantitative trait loci (QTLs). Variations in apoA-II and apoC-3 affected the abundance of multiple HDL proteins, indicating a coordinated regulation. We identified modules of covarying proteins and defined a protein-protein interaction network that describes the protein composition of the naturally occurring subspecies of HDL in mice. Sterol efflux capacity varied up to 3-fold across the strains, and HDL proteins displayed distinct correlation patterns with macrophage and ABCA1-specific cholesterol efflux capacity and cholesterol exchange, suggesting that subspecies of HDL participate in discrete functions. The baseline and stimulated sterol efflux capacity phenotypes were associated with distinct QTLs with smaller effect size, suggesting a multigenetic regulation. Our results highlight the complexity of HDL particles by revealing the high degree of heterogeneity and intercorrelation, some of which is associated with functional variation, and support the concept that HDL-cholesterol alone is not an accurate measure of HDL’s properties, such as protection against CAD.

Common Heritable Immunological Variations Revealed in Genetically Diverse Inbred Mouse Strains of the Collaborative Cross

Variations in the proportion and number of specific immune cell types among healthy individuals are influenced by both heritable and nonheritable factors. Mouse models, subjected to fewer nonheritable factors than humans, allow the identification of genetic factors that shape the immune system. We characterized immunological trait variability in the Collaborative Cross (CC), a powerful genetic resource of recombinant inbred mouse strains derived from eight diverse founder strains. Of the 18 immunological traits studied in more than 60 CC strains, eight showed genome-wide significant linkage, revealing new genetic loci linked to specific immune traits. We also found that these traits were highly subject to heritable influences. As for humans, mouse immunological traits varied as a continuum rather than as discrete immunophenotypes. The CC thus represents a useful resource to identify factors that determine immunological variations, as well as defining other immune traits likely to be heritable in humans.

Gene-by-Sex Interactions in Mitochondrial Functions and Cardio-Metabolic Traits

We studied sex differences in over 50 cardio-metabolic traits in a panel of 100 diverse inbred strains of mice. The results clearly showed that the effects of sex on both clinical phenotypes and gene expression depend on the genetic background. In support of this, genetic loci associated with the traits frequently showed sex specificity. For example, Lyplal1, a gene implicated in human obesity, was shown to underlie a sex-specific locus for diet-induced obesity. Global gene expression analyses of tissues across the panel implicated adipose tissue "beiging" and mitochondrial functions in the sex differences. Isolated mitochondria showed gene-by-sex interactions in oxidative functions, such that some strains (C57BL/6J) showed similar function between sexes, whereas others (DBA/2J and A/J) showed increased function in females. Reduced adipose mitochondrial function in males as compared to females was associated with increased susceptibility to obesity and insulin resistance. Gonadectomy studies indicated that gonadal hormones acting in a tissue-specific manner were responsible in part for the sex differences.

Different Outcomes of Experimental Hepatitis E Virus Infection in Diverse Mouse Strains, Wistar Rats, and Rabbits.

Hepatitis E virus (HEV) is the causative agent of acute hepatitis E in humans in developing countries, but autochthonous cases of zoonotic genotype 3 (HEV-3) infection also occur in industrialized countries. In contrast to swine, rats, and rabbits, natural HEV infections in mice have not yet been demonstrated. The pig represents a well-established large animal model for HEV-3 infection, but a suitable small animal model mimicking natural HEV-3 infection is currently missing. Therefore, we experimentally inoculated C57BL/6 mice (wild-type, IFNAR−/−, CD4−/−, CD8−/−) and BALB/c nude (nu/nu) mice, Wistar rats, and European rabbits with a wild boar-derived HEV-3 strain and monitored virus replication and shedding, as well as humoral immune responses. HEV RNA and anti-HEV antibodies were detected in one and two out of eight of the rats and all rabbits inoculated, respectively, but not in any of the mouse strains tested. Remarkably, immunosuppressive dexamethasone treatment of rats did not enhance their susceptibility to HEV infection. In rabbits, immunization with recombinant HEV-3 and ratHEV capsid proteins induced protection against HEV-3 challenge. In conclusion, the rabbit model for HEV-3 infection may serve as a suitable alternative to the non-human primate and swine models, and as an appropriate basis for vaccine evaluation studies.

The E3 ligase MARCH5 is a PPARγ target gene that regulates mitochondria and metabolism in adipocytes.

Mitochondrial dynamics refers to the constant remodeling of mitochondrial populations by multiple cellular pathways that help maintain mitochondrial health and function. Disruptions in mitochondrial dynamics often lead to mitochondrial dysfunction, which is frequently associated with disease in rodents and humans. Consistent with this, obesity is associated with reduced mitochondrial function in white adipose tissue, partly via alterations in mitochondrial dynamics. Several proteins, including the E3 ubiquitin ligase membrane-associated RING-CH-type finger 5 (MARCH5), are known to regulate mitochondrial dynamics; however, the role of these proteins in adipocytes has been poorly studied. Here, we show that MARCH5 is regulated by peroxisome proliferator-activated receptor-γ (PPARγ) during adipogenesis and is correlated with fat mass across a panel of genetically diverse mouse strains, in ob/ob mice, and in humans. Furthermore, manipulation of MARCH5 expression in vitro and in vivo alters mitochondrial function, affects cellular metabolism, and leads to differential regulation of several metabolic genes. Thus our data demonstrate an association between mitochondrial dynamics and metabolism that defines MARCH5 as a critical link between these interconnected pathways.

Hybrid Sterility with Meiotic Metaphase Arrest in Intersubspecific Mouse Crosses.

Although organisms belonging to different species and subspecies sometimes produce fertile offspring, a hallmark of the speciation process is reproductive isolation, characterized by hybrid sterility (HS) due to failure in gametogenesis. In mammals, HS is usually exhibited by males, the heterogametic sex. The phenotypic manifestations of HS are complex. The most frequently observed are abnormalities in both autosomal and sex chromosome interactions that are linked to meiotic prophase arrest or postmeiotic spermiogenesis aberrations and lead to defective or absent gametes. The aim of this study was to determine the HS phenotypes in intersubspecific F1 mice produced by matings between Mus musculus molossinus-derived strains and diverse Mus musculus domesticus-inbred laboratory mouse strains. Most of these crosses produced fertile F1 offspring. However, when female BALB/cJ (domesticus) mice were mated to male JF1/MsJ (molossinus) mice, the (BALBdomxJF1mol)F1 males were sterile, whereas the (JF1molxBALBdom)F1 males produced by the reciprocal crossings were fertile; thus the sterility phenotype was asymmetric. The sterile (BALBdomxJF1mol) F1 males exhibited a high rate of meiotic metaphase arrest with misaligned chromosomes, probably related to a high frequency of XY dissociation. Intriguingly, in the sterile (BALBdomxJF1mol)F1 males we observed aberrant allele-specific expression of several meiotic genes, that play critical roles in important meiotic events including chromosome pairing. Together, these observations of an asymmetrical HS phenotype in intersubspecific F1 males, probably owing to meiotic defects in the meiotic behavior of the XY chromosomes pair and possibly also transcriptional misregulation of meiotic genes, provide new models and directions for understanding speciation mechanisms in mammals.

The Ca2+ transient as a feedback sensor controlling cardiomyocyte ionic conductances in mouse populations.

Conductances of ion channels and transporters controlling cardiac excitation may vary in a population of subjects with different cardiac gene expression patterns. However, the amount of variability and its origin are not quantitatively known. We propose a new conceptual approach to predict this variability that consists of finding combinations of conductances generating a normal intracellular Ca2+ transient without any constraint on the action potential. Furthermore, we validate experimentally its predictions using the Hybrid Mouse Diversity Panel, a model system of genetically diverse mouse strains that allows us to quantify inter-subject versus intra-subject variability. The method predicts that conductances of inward Ca2+ and outward K+ currents compensate each other to generate a normal Ca2+ transient in good quantitative agreement with current measurements in ventricular myocytes from hearts of different isogenic strains. Our results suggest that a feedback mechanism sensing the aggregate Ca2+ transient of the heart suffices to regulate ionic conductances.

Quantifying and Localizing the Mitochondrial Proteome Across Five Tissues in A Mouse Population

We have used SWATH mass spectrometry to quantify 3648 proteins across 76 proteomes collected from genetically diverse BXD mouse strains in two fractions (mitochondria and total cell) from five tissues: liver, quadriceps, heart, brain, and brown adipose (BAT). Across tissues, expression covariation between genes' proteins and transcripts-measured in the same individuals-broadly aligned. Covariation was however far stronger in certain subsets than others: only 8% of transcripts in the lowest expression and variance quintile covaried with their protein, in contrast to 65% of transcripts in the highest quintiles. Key functional differences among the 3648 genes were also observed across tissues, with electron transport chain (ETC) genes particularly investigated. ETC complex proteins covary and form strong gene networks according to tissue, but their equivalent transcripts do not. Certain physiological consequences, such as the depletion of ATP synthase in BAT, are thus obscured in transcript data. Lastly, we compared the quantitative proteomic measurements between the total cell and mitochondrial fractions for the five tissues. The resulting enrichment score highlighted several hundred proteins which were strongly enriched in mitochondria, which included several dozen proteins were not reported in literature to be mitochondrially localized. Four of these candidates were selected for biochemical validation, where we found MTAP, SOAT2, and IMPDH2 to be localized inside the mitochondria, whereas ABCC6 was in the mitochondria-associated membrane. These findings demonstrate the synergies of a multi-omics approach to study complex metabolic processes, and this provides a resource for further discovery and analysis of proteoforms, modified proteins, and protein localization.

The impact of host genetic background in the Pseudomonas aeruginosa respiratory infections.

Understanding the significance of human genetic diversity in modulating host susceptibility to opportunistic infections is an emerging challenge in the field of respiratory illnesses. While it is recognized that diverse bacterial strains account for differential disease manifestations, emerging data indicate that host genetic diversity is an important determinant factor that influences the severity of opportunistic infections. With particular regard to respiratory illnesses mediated by the gram-negative bacterium Pseudomonas aeruginosa, diverse genetic background is also emerging as a key contributor. Human-genome-wide association studies are a common approach for determining the inter-individual genetic variation associated with variability of the pulmonary infections. Historically, diverse murine inbred mouse strains and ex-vivo cellular models were considered complementary to human studies for establishing the contribution of genetic background to P. aeruginosa respiratory infections. More recently, the development of a new mouse model of infection, mirroring human airway diseases, combined with innovative murine resource populations, modelling human genetic variation, provides additional insights into the mechanisms of genetic susceptibility. In this review, we cover the recent state of the art of human and animal studies and we discuss future potential challenges in the field of P. aeruginosa respiratory infections.

Gene Therapy induced Tregs: A treatment for relapsing-remitting MS in mice

Abstract Autoimmune disease is caused by a breakdown of tolerance against endogenous protein(s). New therapies are being developed that utilize the natural abilities of regulatory T (Treg) cells to suppress autoimmune effector cells in an antigen-specific manner. Recently, we demonstrated a clinically relevant in vivo gene immunotherapy approach for restoring Ag-specific tolerance using hepatic adeno-associated virus (AAV) gene transfer. This approach was shown to be remarkably successful at preventing and reversing a monophasic, MOG-induced EAE disease. Additionally, we have shown the dynamic ability of this treatment to prevent and/or reverse disease induced with various MOG epitopes, in genetically diverse mouse strains. However, it is unclear if this approach would be effective in the more complicated, PLP induced relapsing-remitting disease model. Thus, we re-engineered our therapeutic vector to contain the full-coding sequence of PLP. This is critically important as the established RR-EAE (SJL) model is characterized by epitope spreading; a phenomenon that may more closely resemble multiple sclerosis. SJL mice received a single peripheral injection of AAV. PLP vector, or control, 2 weeks prior to PLP 139–151 induced EAE. By day 14, control mice developed significant neurological deficits followed by a relapsing-remitting disease course. In contrast, the treated mice failed to exhibit any neurological symptoms of disease throughout the duration of the experiment, ~75 days. The long-term prevention of PLP induced disease, provides the first evidence of an effective Treg based therapy for combating epitope drift in RR-EAE, and potentially a new paradigm for treating the autoimmune disease multiple sclerosis.

Integrative analysis of sex differences in adipose tissue gene expression

Sex differences in adipose tissue distribution and function are associated with cardiometabolic disease risk. While extensive research efforts have revealed sex differences in cell signaling and physiology, there is a relative dearth of information regarding the contribution of transcriptional regulation to sex differences in adipose function. We investigated sex‐dependent features of transcriptional regulation in subcutaneous adipose tissues using multiple publicly available omic‐scale gene expression data sets from geographically and ethnically diverse healthy human populations (N = 111–404 samples). Gene set enrichment analyses revealed that fatty acid metabolism, glycolysis, oxidative phosphorylation, and reactive oxygen species production pathways were enriched for differentially expressed genes (FDR < 0.1). We further identified a core set of 79 genes that were consistently differentially expressed in subcutaneous adipose tissue, but not other metabolic tissues (e.g., ELOVL3, FADS1, RORB, FDR < 0.05). A preponderance of these sex‐dependent genes exhibited expression correlations with metabolic traits that were measured in parallel with genes expression (e.g., LDL cholesterol, blood pressure, and body mass index). The genes that were positively correlated with deleterious metabolic traits were enriched for upregulation in males. Many of these results from humans were replicated in adipose tissue transcriptomic data from a population of 97 genetically diverse mouse strains. To further interrogate the molecular causes and consequences of sex‐dependent gene expression differences, we employed bioinformatic and network inference analyses. We identified putative upstream regulators of gene expression sex differences including PPARG and CEBPB. We identified subtle differences in gene expression coordination based on network analysis. A related but distinct aspect of sex differences in adipose function is that there are sex differences in the associations between genetic variants and body fat distribution‐related traits. Current efforts are underway to determine whether these sex‐dependent influences of genetics on adipose distribution could be mediated by influences on gene expression in adipose tissue, as suggested in the literature but not previously demonstrated. Our findings and related investigations have the potential to impact precision medicine approaches that account for the influences of sex on the development and perpetuation of cardiometabolic diseases.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Genetic and Environmental Contributions to Variation in the Posterior Communicating Collaterals of the Circle of Willis.

Variation in blood flow mediated by the posterior communicating collateral arteries (PComs) contributes to variation in the severity of tissue injury in obstructive disease. Evidence in animals and humans indicates that differences in the extent of PComs, i.e., their anatomic lumen diameter and whether they are present bilaterally, unilaterally, or absent, are a major factor. These differences arise during development since they are present at birth. However, the causal mechanisms are unknown. We used angiography after maximal dilation to examine involvement of genetic, environmental, and stochastic factors. The extent of PComs varied widely among seven genetically diverse strains of mice. Like pial collaterals in the microcirculation, aging and hypertension reduced PCom diameter, while in contrast, obesity, hyperlipidemia, metabolic syndrome, and diabetes mellitus had no effect. Naturally occurring intrauterine growth restriction had no effect on extent of PCom or pial collaterals in the adult. The number and diameter of PComs evidenced much larger apparent stochastic-dependent variation than pial collaterals. In addition, both PComs underwent flow-mediated outward remodeling after unilateral permanent MCA occlusion that varied with genetic background and was greater on the ipsilesional side. These findings indicate that variation in the number and diameter of PCom collateral arteries arises from stochastic factors and naturally occurring genetic variants that differ from those that cause variation in pial collateral arterioles. Environmental factors also contribute: aging and hypertension reduce PCom diameter. Our resultssuggest possible sources of variation of PComs in humans and provide information relevant when studying mouse models of occlusive cerebrovascular disease.

Differences in Exercise Capacity and Responses to Training in 24 Inbred Mouse Strains.

Changes in cardiorespiratory fitness in response to a standardized exercise training protocol differ substantially between individuals. Results from cross-sectional, twin, and family studies indicate genetics contribute to individual differences in both baseline exercise capacity and the response to training. Exercise capacity and responses to training also vary between inbred strains of mice. However, such studies have utilized a limited number of inbred strains. Therefore, the aim of this study was to characterize exercise-training responses in a larger number of genetically diverse strains of inbred mice and estimate the contribution of genetic background to exercise training responses. Eight-week old male mice from 24 inbred strains (n = 4–10/strain) performed a graded exercise test before and after 4 weeks of exercise training. Before training, exercise capacity was significantly different between strains when expressed as time (range = 21–42 min) and work performed (range = 0.42–3.89 kg·m). The responses to training also were significantly different between strains, ranging from a decrease of 2.2 min in NON/ShiLtJ mice to an increase of 8.7 min in SWR/J mice. Changes in work also varied considerably between the lowest (−0.24 kg·m in NON/ShiLtJ) and highest (+2.30 kg·m in FVB/NJ) performing strains. Heart and skeletal muscle masses also varied significantly between strains. Two broad sense heritability estimates were calculated for each measure of exercise capacity and for responses to training. For change in run time, the intraclass correlation between mice within the same inbred strain (rI) was 0.58 and the coefficient of genetic determination (g2) was 0.41. Heritability estimates were similar for the change in work: rI = 0.54 and g2 = 0.37. In conclusion, these results indicate genetic background significantly influences responses to exercise training.

Genetic control of oromotor phenotypes: A survey of licking and ingestive behaviors in highly diverse strains of mice

In order to examine genetic influences on fluid ingestion, 20-min intake of either water or 0.1M sucrose was measured in a lickometer in 18 isogenic strains of mice, including 15 inbred strains and 3 F1 hybrid crosses. Intake and licking data were examined at a number of levels, including lick rate as defined by mean or median interlick interval, as well as several microstructural parameters (i.e. burst-pause structure). In general, strain variation for ingestive phenotypes were correlated across water and sucrose in all strains, indicating fundamental, rather than stimulus-specific, mechanisms of intake. Strain variation was substantial and robust, with heritabilities for phenotypes ranging from 0.22 to 0.73. For mean interlick interval (MPI; a measure of lick rate) strains varied continuously from 94.3 to 127.0ms, a range consistent with previous studies. Furthermore, variation among strains for microstructural traits such as burst size and number suggested that strains possess different overall ingestive strategies, with some favoring more short bursts, and others favoring fewer, long bursts. Strains also varied in cumulative intake functions, exhibiting both linear and decelerated rates of intake across the session.