Anatomical Phenotypes Research Articles

Gene × Environment Effects: Stress and Memory Dysfunctions Caused by Stress and Gonadal Factor Irregularities during Puberty in Control And TGF-α Hypomorphic Mice

The maturation of many neural functions occurs during puberty. An abnormal development of these processes, in the context of genetic vulnerability, may result in sex- and age-dependent penetrance of neuropsychiatric disorders. Reduced transforming growth factors-alpha (TGF-alpha) expression in Waved-1 (Wa-1) mice impairs the stress response and fear memory in adult males, but are absent or far less prominent in adult females and in pubertal males. Gonadectomy around the onset of puberty, when the mutant anatomical and behavioral phenotypes are undetectable, results in significant gene x environment effects. Adult control males show reduced physiological stress response as a result of gonadectomy, but not adult Wa-1 males. In females, pubertal gonadectomy elevates specific anxiety parameters only in adult control mice. There also are general sex-specific effects of pubertal gonadectomy on adult stress and fear memory. Surgical stress alone also induces sex- and genotype-dependent effects, albeit in different behavioral parameters than those affected by gonadectomy. We conclude that normal development of stress and memory processes is reliant on the levels of stress and gonadal factors during puberty, the effects of which are modulated by genetic factors and sex.

Magnetic resonance imaging as a tool for in vivo and ex vivo anatomical phenotyping in experimental genetic models.

This article describes a suite of computational approaches suitable for deriving various quantitative phenotypes from structural magnetic resonance (MR) images obtained in rodents and used subsequently in genetic studies of complex traits. We begin by introducing the basic principles of genetic studies of complex traits in experimental models. We then illustrate the use of MR-based computational anatomy in vivo and ex vivo, and in combination with histology. This work was carried out in two inbred strains of rats, namely spontaneously hypertensive rats and Brown Norway rats; these are parental strains of the only existing panel of recombinant inbred strains of rats. The rats were scanned in vivo at two time points (at 8 and 12 weeks of age) and ex vivo (at 12 weeks of age). We describe between-strain differences and across-time changes in brain and kidney volumes, as well as regional variations in brain structure using surface- and deformation-based approaches. We conclude by discussing the power of the population-based computational analysis of MR images, and their fusion with histology, in studies of complex traits.

The Genetics of Brain Wiring: From Molecule to Mind

What makes some people neurotic or schizophrenic or right-handed or fearless? The challenge in answering this is to map from genotype to anatomical and physiological phenotypes and beyond to behavior and cognition.

Anatomical and Functional Phenotyping of Mice Models of Alzheimer's Disease by MR Microscopy

The wide variety of transgenic mouse models of Alzheimer's disease (AD) reflects the search for specific genes that influence AD pathology and the drive to create a clinically relevant animal model. An ideal AD mouse model must display hallmark AD pathology such as amyloid plaques, neurofibrillary tangles, reactive gliosis, dystrophic neurites, neuron and synapse loss, and brain atrophy and in parallel behaviorally mimic the cognitive decline observed in humans. Magnetic resonance (MR) microscopy (MRM) can detect amyloid plaque load, development of brain atrophy, and acute neurodegeneration. MRM examples of AD pathology will be presented and discussed. What has lagged behind in preclinical research using transgenic AD mouse models is functional phenotyping of the brain; in other words, the ability to correlate a specific genotype with potential aberrant brain activation patterns. This lack of information is caused by the technical challenges involved in performing functional MRI (fMRI) in mice including the effects of anesthetic agents and the lack of relevant "cognitive" paradigms. An alternative approach to classical fMRI using external stimuli as triggers of brain activation in rodents is to electrically or pharmacologically stimulate regions directly while simultaneously locally tracking the activated interconnected regions of rodents using, for example, the manganese-enhanced MRI (MEMRI) technique. Finally, transgenic mouse models, MRM, and future AD research would be strengthened by the ability to screen for AD-like pathology in other non-AD transgenic mouse models. For example, molecular biologists may focus on cardiac or pulmonary pathologies in transgenic mice models and as an incidental finding discover behavioral AD phenotypes. We will present MRM data of brain and cardiac phenotyping in transgenic mouse models with behavioral deficits.

Growth in stature and head circumference in high-functioning autism and Asperger disorder during the first 3 years of life

Little effort has been made to characterize the developmental anatomic phenotype of autism; although there is evidence of an increased head circumference and brain size, few other physical characteristics have been studied. The head circumference, body length/height, and weight measurements of infants, who were later diagnosed with high-functioning autism (HFA, n = 16) and Asperger disorder (AsD, n = 12), were extracted from health records over the first 3 years of life and compared to the measurements of a matched normal control group (n = 19). Using linear mixed-effects models, no differences were found in the average growth rate for head circumference, stature, or weight between the children with HFA and AsD. However, a significantly higher growth rate in body length/height and weight was found for the combined group of children with HFA and AsD compared to the normal control group. A trend toward higher growth rate in head circumference was also found among the former group. The results indicate that growth dysregulation in autism is not specific to the brain but also involves growth in stature.

Anatomical phenotyping in the brain and skull of a mutant mouse by magnetic resonance imaging and computed tomography

Since genetically modified mice have become more common in biomedical research as models of human disease, a need has also grown for efficient and quantitative methods to assess mouse phenotype. One powerful means of phenotyping is characterization of anatomy in mutant vs. normal populations. Anatomical phenotyping requires visualization of structures in situ, quantification of complex shape differences between mouse populations, and detection of subtle or diffuse abnormalities during high-throughput survey work. These aims can be achieved with imaging techniques adapted from clinical radiology, such as magnetic resonance imaging and computed tomography. These imaging technologies provide an excellent nondestructive method for visualization of anatomy in live individuals or specimens. The computer-based analysis of these images then allows thorough anatomical characterizations. We present an automated method for analyzing multiple-image data sets. This method uses image registration to identify corresponding anatomy between control and mutant groups. Within- and between-group shape differences are used to map regions of significantly differing anatomy. These regions are highlighted and represented quantitatively by displacements and volume changes. This methodology is demonstrated for a partially characterized mouse mutation generated by N-ethyl-N-nitrosourea mutagenesis that is a putative model of the human syndrome oculodentodigital dysplasia, caused by point mutations in the gene encoding connexin 43.

Progression of Inner Ear Pathology in Ames Waltzer Mice and the Role of Protocadherin 15 in Hair Cell Development

The Ames waltzer (av) mouse mutant exhibits auditory and vestibular abnormalities resulting from mutation of protocadherin 15 (Pcdh15). Ames waltzer has been identified as an animal model for inner ear pathology associated with Usher syndrome type 1F. Studies correlating anatomical phenotype with severity of genetic defect in various av alleles are providing better understanding of the role played by Pcdh15 in inner ear development and of sensorineural abnormalities associated with alterations in Pcdh15 protein structure as a result of gene mutation. In this work we present new findings on inner ear pathology in four alleles of av mice with differing mutations of Pcdh15 as well as varying alterations in inner ear morphology. Two alleles with in-frame deletion mutations (Pcdh15 (av-J) and Pcdh15 (av-2J)) and two presumptive functional null alleles (Pcdh15 (av-3J) and Pcdh15 (av-Tg)) were studied. Light and electron microscopic observations demonstrated that the severity of cochlear and vestibular pathology in these animals correlates positively with the extent of mutation in Pcdh15 from embryonic day 18 (E18) up to 12 months. Electron microscopic analysis of immature ears indicated early abnormalities in the arrangement of stereocilia and the inner and outer hair cell cuticular plates, stereocilia rootlets, and the actin meshwork within the cuticular plate. In severe cases, displacement of the kinocilium and alterations in the shape of the cuticular plate was also observed. Mice harboring in-frame deletion mutations showed less disorganization of stereocilia and cuticular plates in the organ of Corti than the presumptive functional null alleles at P0-P10. A slower progression of pathology was also seen via light microscopy in older animals with in-frame deletions, compared to the presumptive functional null mutations. In summary, our results demonstrate that mutation in Pcdh15 affects the initial formation of stereocilia bundles with associated changes in the actin meshwork within the cuticular plate; these effects are more pronounced in the presumed null mutation compared to mutations that only affect the extracellular domain. The positive correlation of severity of effects with extent of mutation can be seen well into adulthood.

Dendritic cells heterogeneity and its role in cancer immunity

Dendritic cells are the most potent 'professional' antigen-presenting cells, with high ability of primary immune response initiation. Dendritic cells originate from bone marrow progenitors, which circulate in peripheral blood and subsequently give rise to immature dendritic cells, which reside in peripheral tissues. When dendritic cells encounter danger signals, they undergo differentiation and maturation; thereafter they migrate to lymphatic tissues, where they synapse with T-cells and initiate primary immune response. The immune response efficiency is determined by Th1/Th2 balance. Although dendritic cells represent a rare group of leukocytes, their functional and phenotypical heterogeneity confer a great challenge to immunologists. In this review, the myeloid and lymphoid development pathways of dendritic cells, are discussed. The heterogeneity of dendritic cells will be reviewed, based on their anatomical locations, phenotypes and functions. This section focuses on blood and lymphoid tissue dendritic cells. Subsequently, the roles of dendritic cells in the immunity of cancer and how cancer bypasses the dendritic cell-mediated immune responses, are discussed.

Neuroanatomical Phenotyping in the Mouse: The Dopaminergic System

Voluntary movement in animals is modulated by a number of subcortical systems. One of these resides in the basal nuclei and their associated projections and utilizes dopamine as a neurotransmitter. Apart from regulating movement, the dopaminergic axis is also involved in the control of goal-oriented behavior, cognition, and mood. Disorders of this system result in common human neurologic disorders such as Parkinson's and Huntington's diseases, as well contributing to a host of behavioral conditions, such as schizophrenia, attention deficit hyperactivity disorder, and addiction. Many individual mouse models of human dopaminergic dysfunction have been described in varying degrees of detail. However, when evaluating this region of the brain, the veterinary pathologist is confronted by a paucity of information summarizing the comparative aspects of the anatomy, physiology, and pathology of the central dopaminergic system. In this review, a systematic approach to anatomic phenotyping of the central dopaminergic system in the mouse is described and illustrated using tyrosine hydroxylase immunohistochemistry. Differences between murine neuroanatomy and comparable regions of the nonhuman primate brain are highlighted. Although the mouse is the focus of this review, conditions in domestic animals characterized by lesions within the basal nuclei and its projections are also briefly described. Murine behavioral and motor tests that accompany abnormalities of specific anatomic regions of the dopaminergic axis are summarized. Finally, we review mouse models of Parkinson's and Huntington's diseases, as well as those genetically altered mice that elucidate aspects of dopamine metabolism and receptor function.

Molecular Control of Physiological and Pathological T-Cell Recruitment after Mouse Spinal Cord Injury

The intraspinal cues that orchestrate T-cell migration and activation after spinal contusion injury were characterized using B10.PL (wild-type) and transgenic (Tg) mice with a T-cell repertoire biased toward recognition of myelin basic protein (MBP). Previously, we showed that these strains exhibit distinct anatomical and behavioral phenotypes. In Tg mice, MBP-reactive T-cells are activated by spinal cord injury (SCI), causing more severe axonal injury, demyelination, and functional impairment than is found in non-Tg wild-type mice (B10.PL). Conversely, despite a robust SCI-induced T-cell response in B10.PL mice, no overt T-cell-mediated pathology was evident. Here, we show that chronic intraspinal T-cell accumulation in B10.PL and Tg mice is associated with a dramatic and sustained increase in CXCL10/IP-10 and CCL5/RANTES mRNA expression. However, in Tg mice, chemokine mRNA were enhanced 2- to 17-fold higher than in B10.PL mice and were associated with accelerated intraspinal T-cell influx and enhanced CNS macrophage activation throughout the spinal cord. These data suggest common molecular pathways for initiating T-cell responses after SCI in mice; however, if T-cell reactions are biased against MBP, molecular and cellular determinants of neuroinflammation are magnified in parallel with exacerbation of neuropathology and functional impairment.

A GFP-based genetic screen reveals mutations that disrupt the architecture of the zebrafish retinotectal projection

The retinotectal projection is a premier model system for the investigation of molecular mechanisms that underlie axon pathfinding and map formation. Other important features, such as the laminar targeting of retinal axons, the control of axon fasciculation and the intrinsic organization of the tectal neuropil, have been less accessible to investigation. In order to visualize these processes in vivo, we generated a transgenic zebrafish line expressing membrane-targeted GFP under control of the brn3c promoter/enhancer. The GFP reporter labels a distinct subset of retinal ganglion cells (RGCs), which project mainly into one of the four retinorecipient layers of the tectum and into a small subset of the extratectal arborization fields. In this transgenic line, we carried out an ENU-mutagenesis screen by scoring live zebrafish larvae for anatomical phenotypes. Thirteen recessive mutations in 12 genes were discovered. In one mutant, ddl, the majority of RGCs fail to differentiate. Three of the mutations, vrt, late and tard, delay the orderly ingrowth of retinal axons into the tectum. Two alleles of drg disrupt the layer-specific targeting of retinal axons. Three genes, fuzz, beyo and brek, are required for confinement of the tectal neuropil. Fasciculation within the optic tract and adhesion within the tectal neuropil are regulated by vrt, coma, bluk, clew and blin. The mutated genes are predicted to encode molecules essential for building the intricate neural architecture of the visual system.

Identification of a cluster of X-linked imprinted genes in mice

Complete or partial monosomy with respect to the X chromosome is the genetic basis of Turner syndrome in human females. Individuals with Turner syndrome have a spectrum of anatomical, physiological and behavioral phenotypes with expressivity dependent on the extent of monosomy and the parental origin of the single X. Parent-of-origin influences on social cognition in Turner syndrome might be due to the presence of imprinted genes on the X. Imprinting of X-linked genes has also been implicated in the male prevalence of autistic spectrum disorders, in male sexual orientation and in the developmental delay of XO mouse embryos. The only molecular evidence for X-chromosome imprinting, however, concerns X-chromosome inactivation in specific circumstances and does not account for these phenotypes. Using a mouse model for Turner syndrome, we searched for locus-specific imprinting of X-linked genes in developing brain. We identified a cluster of X-linked genes containing at least three genes that show transcriptional repression of paternal alleles. Imprinting of these three genes, Xlr3b, Xlr4b and Xlr4c, is independent of X-chromosome inactivation and has a dynamic and complex pattern of tissue and stage specificity.

Paraxial protocadherin coordinates cell polarity during convergent extension via Rho A and JNK.

Convergent extension movements occur ubiquitously in animal development. This special type of cell movement is controlled by the Wnt/planar cell polarity (PCP) pathway. Here we show that Xenopus paraxial protocadherin (XPAPC) functionally interacts with the Wnt/PCP pathway in the control of convergence and extension (CE) movements in Xenopus laevis. XPAPC functions as a signalling molecule that coordinates cell polarity of the involuting mesoderm in mediolateral orientation and thus selectively promotes convergence in CE movements. XPAPC signals through the small GTPases Rho A and Rac 1 and c-jun N-terminal kinase (JNK). Loss of XPAPC function blocks Rho A-mediated JNK activation. Despite common downstream components, XPAPC and Wnt/PCP signalling are not redundant, and the activity of both, XPAPC and PCP signalling, is required to coordinate CE movements.

Three-dimensional anatomical characterization of the developing mouse brain by diffusion tensor microimaging

Investigation of three-dimensional (3D) morphometry of developing brains has been hindered by a lack of imaging modalities that can monitor the 3D evolution of various anatomical structures without sectioning and staining processes. In this study, we combined magnetic resonance microimaging and diffusion tensor imaging techniques to accomplish such visualization. The application of this approach to developing mouse embryos revealed that it could clearly delineate early critical structures such as neuroepithelium, cortical plate, and various axonal structures, and follow their developmental evolution. The technique was applied to the study of the Netrin-1 mutant, allowing verification of its anatomical phenotype.

Relation between the anatomical genital phenotype and cystic fibrosis transmembrane conductance regulator gene mutations in the absence of the vas deferens

Objective: To study the correlation between genital phenotype and cystic fibrosis genotype in men lacking at least one vas deferens. Design: Prospective study. Setting: Institut Rhônalpin pour la Reproduction Humaine, Lyon-Bron, France. Patient(s): Forty-seven infertile men lacking at least one vas deferens. Intervention(s): All patients were screened for the 13 most common CFTR gene mutations and for the 5-thymidine variant of intron 8. Renal, scrotal, and transrectal ultrasonography were systematically performed. Main Outcome Measure(s): Epididymal and seminal vesicular abnormalities and testicular volume were compared among men with two, one, or no CFTR gene mutation, with or without the 5T allele. Results: Seminal vesicles and the symmetry of epididymal and vesicular abnormalities did not differ between patients with and those without the CFTR gene mutation. Epididymal abnormalities were more frequent in men without the mutation. Testicular volumes were significantly lower in men without the mutation and those with the 5T allele only. Conclusion: Men with the CFTR mutation, the 5T allele only, and those without CFTR mutation have few differences in genital phenotype. Low testicular volume is observed in men without the CFTR mutation and those with the 5T allele only.

Motor deficits in fibroblast growth factor receptor-3 null mutant mice.

Fibroblast growth factor receptor-3 (FGFR-3) regulates aspects of bone development. Mutations in the FGFR-3 gene (Fgfr3) in humans and mice produce vertebral abnormalities and bone deformities. The present study evaluated the behavioural concomitants of the Fgfr3-/- mutation. Fgfr3-/- null mutant mice displayed severe impairments of motor abilities as detected on the rotarod, wire hang and open field tests. Absence of prepulse inhibition of acoustic startle was seen at prepulse levels from 74 to 86 dB. The motor deficits appear to be a direct and predicted consequence of the skeletal kyphosis, scoliosis and long bone overgrowth previously reported in Fgfr3 null mutant mice. The behavioural phenotype displayed by these mutant mice complements their anatomical, physiological and biochemical phenotypes, to complete the characterization of the functional outcome of a single gene mutation. Simple, robust behavioural symptoms, such as poor rotorod performance in Fgfr3 knockout mice, can provide useful surrogate markers to evaluate pharmacological treatments and gene therapies for human genetic diseases.

Non-invasive, quantitative assessment of the anatomical phenotype of corticotropin-releasing factor-overexpressing mice by MRI.

High resolution magnetic resonance imaging (MRI) was applied to quantify alterations in thymus and adrenal volumes, as well as body fat in genetically engineered corticotropin-releasing factor (CRF)-overexpressing mice. When compared to the organs in age-matched wild-type animals, the adrenals in CRF-overexpressing male mice were significantly enlarged and the thymus volume in females was significantly smaller. The fat content was significantly larger in CRF-overexpressing mice. The anatomical alterations observed in the MRI studies were in perfect line with post-mortem data (weights of organs). Furthermore, the observed interstrain differences are in agreement with recently published data on (i) the effect of continuous, intraventricular infusion of CRF in rats and (ii) the presence of atrophic adrenals in CRF-knockout mice. The present studies demonstrate that MRI can provide reliable measures of relatively small structures such as the adrenal glands and the thymus in mice. This makes MRI an attractive, non-terminal tool to monitor in laboratory animals, including transgenic mice, the consequence of continuous stress on relevant organs.

Anatomical origin of dendritic cells determines their life span in peripheral lymph nodes.

Dendritic cells (DCs) exhibit considerable heterogeneity in their anatomical location, surface phenotype, and functional properties. In this study, we demonstrate that peripheral lymph nodes contain at least four major, functionally separable, and independently derived, DC subsets, which can be clearly demarcated by their CD11c, CD40, and CD8 expression pattern. Surprisingly, all DCs derived directly from the bone marrow, the myeloid- and the lymphoid-related subsets, turned over fast with t(1/2) of a couple of days. In contrast, DCs exported from the skin, both dermal and epidermal, accumulated 3- to 4-fold slower, turnover that is dramatically increased by cutaneous inflammation.

Magnetic Resonance Microscopy of the C57BL Mouse Brain

With the rapid progression in gene technologies, transgenic, targeted, and chemically induced mutations in mice are continually created. The major goal of these studies is to understand and characterize the effects of genotype on anatomy, physiology, and behavior and ultimately the role of genotype in development of disease. The demand for imaging techniques with high spatial resolution potential is rising because such imaging tools would expedite anatomical phenotyping in the genetically altered mice. Magnetic resonance microscopy (MRM) is a noninvasive, inherently three-dimensional (3D) imaging technique capable of visualizing several anatomical structures in the small mouse. The 3D nature of MRM also allows for interpretation of complex spatial relationships between substructures, which is important when phenotyping anatomically. The goal of this paper is to systematically describe three major brain regions in the C57BL/6J mouse at microanatomical spatial resolution ranges using in vitro MRM. We explore different MR contrast parameters, voxel sizes, and signal-to-noise ratios to best characterize C57BL/6J mouse brain microstructure by MRM. Further, we compare all MRM images with Nissl-stained brain sections. Major findings were as follows: T2* MR images visualized several gross anatomical regions in the mouse brain but not, for example, subregions within the hippocampus. Diffusion proton stains on the other hand were superior to T2* MR images and delineated many subregions within the hippocampus proper. Finally, contrast enhancement facilitated visualization of hippocampal anatomy on the T2* MR images. The results of this study are part of an ongoing initiative at our Center focused on creating a complete C57BL/6J mouse anatomical 3D image database by MRM.

CTL priming by CD8+ and CD8 dendritic cells in vivo

Two distinct developmental pathways are driving the formation of myeloid- and lymphoid-related dendritic cells (DC) which differ in anatomical localization and phenotype. In terms of function, it has been hypothesized that only the myeloid-related CD8– DC are able to initiate immune responses, whereas the lymphoid-related CD8+ DC have been suggested to induce tolerance. Here we show that both subsets activate CD8+ T cells in vitro and induce protective anti-viral CTL responses in vivo. Thus, vaccine strategies using peptide-pulsed DC do not have to take into account DC subsets for priming.