Morphological Mouse Phenotyping: Anatomy, Histology and Imaging

By Jesus Ruberte, Ana Carretero and Marc Navarro

Morphological Mouse Phenotyping: Anatomy, Histology and Imaging is an atlas of explanatory diagrams and text that guides the reader through normal mouse anatomy, histology, and imaging. The book is targeted for mouse researchers and veterinarian and human pathologists, and presents a complete, integrative description of normal mouse morphology.

Disease animal models are fundamental in research to improve human health. The success of using genetically engineered mice to evaluate molecular disease hypotheses has encouraged the development of massive global projects, making the mouse the most used animal disease model. Laboratory mouse populations are straining the housing capacity of pharmaceutical and biotechnology companies, as well as public research institutions. However, the scientific community lacks sufficient expertise in morphological phenotyping to effectively characterize and validate these animal models. The mouse displays fundamental morphological similarities to humans; however, a mouse is not a man.

• Features more than 2,200 original images showing the anatomy, histology, and cellular structure of mouse organs
• Includes images specifically produced for this book in the Mouse Imaging Platform (Center for Animal Biotechnology and Gene Therapy, Universitat Autònoma de Barcelona)
• Offers an integrative vision of mouse morphology using correlative X-ray, computed tomography, magnetic resonance, and ultrasound images
• Employs classical anatomical techniques such as conventional dissection, skeletal preparations, vascular injections, and histological, immunohistochemical, and electron microscopy techniques to characterize mouse morphology

• Language: English
• Publisher: Academic Press
• Release date: Jan 27, 2017
• ISBN: 9780128128770

Jesus Ruberte

Jesus Ruberte is Head of the Mouse Imaging Platform in the Center for Animal Biotechnology and Gene Therapy, Department of Animal Health and Anatomy, Veterinary School, Universitat Autonoma de Barcelona, Spain.

Book preview

9780128128770_FC

Morphological Mouse Phenotyping

Anatomy, Histology and Imaging

Prof. Dr. Jesús Ruberte, (DVM, PhD)

Head of the Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain

Associate Prof. Dr. Ana Carretero, (DVM, PhD)

Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain

Associate Prof. Dr. Marc Navarro, (DVM, PhD)

Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain

Table of Contents

Cover image

Title page

Copyright

Authors

Dedication

Foreword

Preface

Acknowledgments

List of abbreviations

1: Introduction

Origin of the Laboratory Mouse

Anatomical Nomenclature

2: Osteology

Bones of the Skull and Mandible

Vertebral Column and Thorax

Bones of the Thoracic Limbs

Bones of the Pelvic Limbs

3: Arthrology

Joints without Articular Cavity

Joints with Articular Cavity

4: Myology

Form and Structure of Muscles

The Skeletal Muscle Fiber

Comparative Anatomy of Mouse Musculature

5: Digestive tract

Oral Cavity

Pharynx and Esophagus

Stomach

Intestine

Liver

6: Respiratory apparatus

Nasal Cavity

Larynx

Trachea

Lungs

7: Urinary organs

Kidneys

Ureter

Urinary Bladder

Urethra

8: Male genital organs

Testicles and Epididymis

Accessory Genital Glands

Penis

9: Female genital organs

Ovaries

Uterine Tubes

Uterus

Vagina

Vulva and Clitoris

10: Anatomy of development

Ontogenic Development

Placenta

11: Circulatory System

Heart

Arteries

Veins

Lymphatic System

Blood and Lymph

12: The endocrine glands

Pancreas

Adrenal Gland

Thyroid Glands

Parathyroid Gland

Hypophysis

13: Nervous System

Central Nervous System

Peripheral Nervous System

14: Eye and related structures

Eyelids and Lacrimal System

Eyeball

Orbit

15: Vestibulocochlear organ

External Ear

Middle Ear

Internal Ear

16: Common integument

Skin

Modified Skin Structures

Bibliography

Index

Copyright

The original English edition of Morphological Mouse Phenotyping: Anatomy, Histology and Imaging

(ISBN: 9788479035006) is published by:

Editorial Medica Panamericana, S.A., Madrid, Spain

Copyright © 2017. All rights reserved.

This reprinted edition is published and distributed by Elsevier, Inc. in cooperation with Editorial Medica Panamericana, S.A.

No part of this publication may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher. Details on how to seek permission, further information about the Publisher’s permissions policies and our arrangements with organizations such as the Copyright Clearance Center and the Copyright Licensing Agency, can be found at our website: www.elsevier.com/permissions.

This book and the individual contributions contained in it are protected under copyright by the Publisher (other than as may be noted herein).

Notices

Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary. Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility.

To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume any liability for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas contained in the material herein.

Library of Congress Cataloging-in-Publication Data

A catalog record for this book is available from the Library of Congress

Library Cataloguing-in-Publication Data

A catalogue record for this book is available from the British Library

ISBN: 978-0-12-812972-2

For Information on all Elsevier publications visit our website at https://www.elsevier.com

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Authors

Prof. Dr. Jesús Ruberte (DVM PhD),     Head of the Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Associate Prof. Dr. Ana Carretero (DVM PhD),     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Associate Prof. Dr. Marc Navarro (DVM PhD),     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Associate Prof. Dr. Víctor Nacher (PhD),     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Auxiliar Prof. Dr. Luisa Mendes-Jorge (DVM PhD),     Centre for Interdisciplinary Research in Animal Health. Veterinary School. Universidade de Lisboa. Alto da Ajuda – Polo Universitario 1300-477 Portugal.

Dr. David Ramos (DVM PhD),     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain. Centre for Interdisciplinary Research in Animal Health. Veterinary School. Universidade de Lisboa. Alto da Ajuda – Polo Universitario 1300-477 Portugal.

Dr. Mariana López-Luppo (DVM PhD),     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Department of Animal Health and Anatomy. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Dr. Pedro Otaegui (DVM PhD, DiplECLAM),     Director of Servei d’Estabulari. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Dr. Anna Pujol (PhD),     Head of the Transgenic Animal Unit. Center for Animal Biotechnology and Gene Therapy. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Associate Prof. Dr. Yvonne Espada (DVM PhD),     Head of the Diagnostic Imaging Service. Fundació Hospital Clinic Veterinari. Department of Animal Medicine and Surgery. Veterinary School. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Dr. Silvia Lope (PhD),     Nuclear Magnetic Resonance Facility. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Dr. Elisabet Domínguez (DVM PhD, DiplECVDI),     Diagnostic Imaging. Centre for Small Animal Studies. Animal Health Trust. Lanwades Park Kentford, Newmarket CB8 7UU. United Kingdom.

Em. Prof. Dr. Horst Erich König (DVM PhD),     Institut für Anatomie, Histologie und Embryologie. Veterinärmedizinische Universität Wien. Veterinärplatz 1, A-1210 Wien, Austria.

Prof. Dr. Luis Puelles (MD PhD),     Department Human Anatomy. Faculty of Medicine. Universidad de Murcia. Campus de Espinardo 30100, Spain.

TECHNICIANS

Verónica Melgarejo,     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Lorena Noya,     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain.

Ángel Vazquez,     Mouse Imaging Platform. Center for Animal Biotechnology and Gene Therapy. Universitat Autònoma de Barcelona. Cerdanyola del Vallès 08193, Spain

ILLUSTRATIONS

Dr. Eva Polsterer (Diplomtierärztin Magister),     A-2431 Enzersdorf/Fischa, Austria.

TRANSLATION

Dr. Christopher John Mann (PhD),     08002 Barcelona, Spain

Dedication

Dedicated to our children Carmen, Andrea and Aleix

Foreword

Professor Steve Brown Director MRC Harwell, Chair IMPC Steering Committee

We still remain profoundly ignorant of the function of the majority of genes in the human and mouse genomes, and importantly, their role in disease. One of the grand challenges for human and mouse genetics over the coming decades is to identify and dissect mammalian gene function and to uncover the contributions of the gene landscape to the proper functioning of body systems. Importantly, this challenge needs to be extended to defining the context in which gene variation contributes to disease with respect to environment and lifestyle, and how that genetic variation impinges upon the efficacy of therapeutic strategies. Thus, projects such as the Precision Medicine Initiative in the US, and the 100,000 genomes project in the UK aim to begin to develop a global genome view of the contributions of genetic variation alongside behavioral and environmental factors to disease outcome, and how this intersects with the possibilities of disease treatment at an individual level. The expectation is that advances in the developments in preventative medicine, the effective targeting of treatments, and the discovery of novel therapeutic strategies will all emerge from these endeavors.

However, it is also clear that a fundamental underpinning for these advances is a more comprehensive and profound understanding of mammalian gene function in the development, physiology and biochemistry of all body systems. In addition, we need to understand the underlying mechanisms that govern mammalian development and physiology. Though the last few decades have been a triumph in terms of the discovery of genetic mechanisms of mammalian biology, in many areas we have only scratched the surface and much remains to be uncovered. Given the unrivalled ability to intervene genetically in the mouse and our understanding of its development, anatomy and histology, nowhere better exemplified than by the comprehensive analysis conveyed in this remarkable volume, we are in privileged position in mouse genetics to begin to develop a comprehensive catalogue of mammalian gene function.

The International Mouse Phenotype Consortium (IMPC) has embarked upon a major programme to generate and phenotype mouse mutants for every gene in the mouse genome. This project is already illuminating many of the dark places in the mammalian genome, shedding light as it proceeds on the thousands of genes for which we have no clue as to function or role in disease processes. Each mutant that is generated by IMPC is subject to a broad based phenotyping pipeline that aims to uncover comprehensive information about the impact of individual genes on development, anatomy, histology and physiology. It is vital for such efforts that we have an extraordinarily fine, accurate and comprehensive description of the anatomy and histology of the mouse – one that also utilizes the ongoing developments in imaging - all of which contributes to the capture and analysis of morphological variation and how it relates to underlying genetics. This volume provides the foundation for our studies of morphological variation, developing a comprehensive atlas of mouse anatomy and histology.

Morphology and anatomy are the bedrocks on which biology and medicine have been based. This vital area of knowledge is now more important than ever as we begin to develop a multidimensional view of body systems incorporating genetic factors and the environment. The efforts of Jesús Ruberte, Ana Carretero and Marc Navarro to assemble an extraordinary atlas of mouse anatomy is one of the fundamental pillars on which developments in mammalian gene function will be based in the coming years.

Preface

Jesús Ruberte; Ana Carretero; Marc Navarro

The mouse displays fundamental similarities in anatomy, physiology and genome organization to humans. This makes the mouse model one of the most important tools to understand gene function and their possible role in human disease. Specific advantages of the mouse as a model system include: 1) the mouse genome is > 95% identical to the human genome, 2) there are powerful toolkits available for the manipulation of the mouse genome, 3) mice breed rapidly, with about ten weeks per generation, and are relatively inexpensive to feed and house, and 4) the mouse is a model used for drug discovery and to test therapies, including cell and gene therapy. All these reasons, in addition to the start up of large scale global mouse functional genomics projects, such as the International Mouse Phenotyping Consortium (IMPC), which aims to create more than 20,000 strains with one specific deactivated gene, mean that mice literally dominate biomedical laboratories worldwide.

The bottleneck in the process of establishing suitable mouse models for human diseases is quite often appropriate mouse phenotyping. Whilst novel gene editing methodologies like CRISPR/Cas9 represent an easy and fast way to produce mouse models, the ability of mouse researchers to make morphological phenotyping remains slower and more complicated. In addition, there is a growing appreciation of imaging technologies for mouse morphological phenotyping. The reason for this is that if we aim to develop mouse models for human diseases, we should have the same diagnostic imaging capabilities for mice and humans.

In this scenario, unfortunately few books, atlases or internet resources explain in detail mouse morphology. Furthermore, to date a complete comprehensive set of images that describe mouse morphology with an integrative vision has not been generated. The different levels of mouse morphology, cellular, tissular, organic, or systemic have no boundaries. In fact, they form a continuum that researchers must go over to accomplish an accurate morphological phenotyping of mouse models.

Here we present a complete and integrative description of mouse normal morphology. More than 2,200 original images have been specifically produced for this atlas in the Mouse Imaging Platform (Center for Animal Biotechnology and Gene Therapy, Universitat Autònoma de Barcelona). These images show the anatomy, histology and cellular structure of mouse organs. In addition, correlative X-ray, Computed Tomography, Magnetic Resonance and Ultrasound images complete this integrative vision of the mouse morphology. Classical anatomical techniques such as conventional dissection, skeletal preparations, vascular injections, as well as histological, immunohistochemical and electron microscopy techniques have been employed to prepare this atlas.

This book is the culmination of twelve years of work. The interaction established during this time with European (EUMORPHIA, EUMODIC, INFRAFRONTIER) and global (IMPC) mouse consortia has allowed us to learn firsthand the evolution of mouse phenotyping and the specific morphological knowledge demands from mouse researchers. This book, basically an atlas, but also containing explanatory diagrams and text, guides the reader through normal mouse anatomy, histology and imaging, and is aimed at the mouse researcher as well as the veterinary and human pathologists.

Barcelona, Spring 2016

Acknowledgments

Many people and institutions have supported this editorial project over the years. We especially want to thank Prof. Dr. Fátima Bosch, director of Center for Animal Biotechnology and Gene Therapy, for believing in the project and providing all the necessary resources for its realization. We are also indebted to outstanding members of the international community of mouse functional genomics for their advice and support; especially we want to thank Prof. Dr. Steve Brown (Medical Research Center, Harwell), Prof. Dr. Martin Hrabé de Angelis (Helmholtz Zentrum Muenchen Deutsches Forschungszentrum fuer Gesundheit und Umwelt GMBH), Dr. Yann Herault (Centre Européen de Recherche en Biologie et en Medicine – Institut Clinique de la Souris, Strasbourg), Dr. Colin McKerlie (The Centre for Phenogenomics Inc, Toronto) and Dr. Karen Svenson (The Jackson Laboratory, Bar Harbor). The preparation of this book has been partially financed by grants from the Ministerio de Ciencia e Innovación (SAF2008-04581-E/) and the Instituto de Salud Carlos III (PI030496, PI061837, PS09/01152 and PI12/00605) of Spain. We want to thank also the team of Editorial Médica Panamericana for its commitment, effort and professionalism devoted over the years to the publication of a book with these characteristics. Finally, thank you very much to Prof. Dr. Alfonso Rodriguez Baeza, Prof. Dr. Bernardo Castellano, Dr. Clara Armengol, Dr. Cristina Llombart, Dr. Jaume Martorell, Dr. Josep Maria Pons, Myriam Torralba, Diego Rubén Mediano, Joana Catita, Andreia Valença, Fernando Hernández and all those who have somehow helped in this project.

List of abbreviations

ATPase Adenosine triphosphatase.

DAB 3,3′-Diaminobenzidine.

DAPI 4′,6-Diamidino-2-phenylindole.

EGFP Enhanced green fluorescent protein.

FITC Fluorescein isothiocyanate.

GABA Gamma-aminobutyric acid.

GFAP Glial fibrillary acidic protein.

GPDH Glycerol-3-Phosphate Dehydrogenase.

GS Glutamine synthetase.

HRP Horseradish peroxidase.

Iba1 Ionized calcium binding adaptor molecule 1.

Ki67 Antigen KI-67.

m muscle.

NADH Nicotinamide adenine dinucleotide.

NG2 Neural/glial antigen 2.

PAS Periodic acid–Schiff.

PECAM-1 Platelet endothelial cell adhesion molecule 1.

PGP Protein gene product 9.5.

PKCα Protein kinase C alpha.

RIP Rip-antigen (2′,3′-cyclic nucleotide 3′-phosphodiesterase).

SDH Succinate dehydrogenase.

VEGF Vascular endothelial growth factor.

α-SMA Alpha-smooth muscle actin.

1

Introduction

A. Carretero; J. Ruberte; M. Navarro; P. Otaegui

By definition, any study of the mouse anatomy should take into consideration the structure, disposition and shape of the organs that constitute the body of the mouse. Etymologically, the word «anatomy» comes from the Greek, meaning «to cut up». Currently, the dissection procedure is just one of the many that morphologists employ. From its origins, as gross and descriptive anatomy, the concept of anatomy has been enriched with other meanings and approaches, such as functional, comparative, microscopic or applied. In addition, thanks to technological advances in the methods of imaging, we can also talk about other specializations of anatomy, such as radiological anatomy, ultrasound anatomy, etc.

In accordance with the modern definition of anatomy, this book attempts to provide an overview of the different levels of morphology of the mouse. It ranges from gross anatomy and topographical anatomy, to explain the relative position of the organs and structures of a particular body region, down to the microscopic anatomy. In addition, we have also introduced the latest imaging technologies applicable to the study of mouse anatomy, including computed tomography (CT), magnetic resonance imaging (MRI), ultrasonography and angiography.

Origin of the Laboratory Mouse

The mouse belongs to the order of rodents and, within this, to the murine family. More specifically, the mouse belongs to the genus Mus, which comprises at least a dozen species quite homogeneous in appearance. The origin of the genus Mus goes back five million years to Southeast Asia. In fact, the word mouse comes from the Sanskrit word «mush». Later, in Roman times with improved communications and changes in agricultural practices, the mouse spread across Europe and Africa. Within the genus Mus, the laboratory mouse belongs to the Mus musculus species. Within this species there are four subspecies or populations: Mus musculus musculus, which was the first to be described and occupies most of northern Europe and Asia; Mus musculus domesticus, which occupies Western Europe, North Africa, the Arabian Peninsula and the Middle East; Mus musculus castaneus, which is located in Southeast Asia; and Mus musculus bactrianus, which predominates in Iran, Pakistan and India. The subspecies domesticus was transported by man to America, Australia and South Pacific countries. There is some controversy about the existence of a fifth subspecies, Mus musculus molossinus, which are proposed to inhabit the islands of Japan, although they are generally considered to be a hybrid between musculus and castaneus.

Mice commonly used in laboratories are mainly derived from the domesticus subspecies and to a lesser extent from the musculus and castaneus subspecies. These lines were created from albino mice, which were used as pets in Egypt. It has been shown that all inbred albino lines carry the same mutation, which occurs very rarely in nature, and they thus represent in a certain way a totally artificial mouse population.

One of the known origins for the use of the mouse as a laboratory animal took place in 1664, when English scientist Robert Hooke used them in his studies of the properties of air. However, it was not until 1902 when their use becomes widespread in genetic studies. It was in this year that Lucien Cuénot first crossed mice carrying recessive mutations related to coat color and showed that Mendel’s laws could be applied to animals. At the same time, William Castle began his studies on inheritance in mice, thanks to the generosity of Mrs. Abbie Lathrop, a retired teacher, who kept as an entertainment and small business a mouse breeding farm in Massachusetts. Many of the current laboratory mouse lines were originally derived in her farm. In 1909 C. C. Little and William Castle created the first inbred line for use in cancer research, which today is known as the DBA (Dilute, Brown and non-Agouti) line. It was Little, who went on to found the «Jackson Laboratory» in 1929, who first published crucial data linking genetic inheritance with resistance to growth of certain tumors using these mice. In 1913, J. Hasley Bagg used albino mice for behavioral studies, whereas in 1920, Leonell C. Strong crossed the DBA line with Bagg’s albino mice to form hybrids. From these hybrids he developed a series of inbred lines that includes the C3H line. In 1921, Strong further crossed Bagg’s albino mice with Little’s mice to produce the line A, characterized by a high incidence of breast and lung tumors. In the same year, and operating from Bussey Institute for Research in Applied Biology at Harvard, Little crossed female 57 with male 52 from the farm of Mrs. Lathrop. The mice obtained were segregated into two populations, one black and one brown, thereby creating the respective C57BR and C57BL lines. Subsequently, E. Carleton MacDowell received from C. C. Little the descendants of Lathrop’s mice and created the line C58. MacDowell also raised Bagg’s albinos and sent some of them to George D. Snell in 1932. Snell used the letter «c» to indicate that the animals were white and henceforward this albino strain was called BALB/c and is still commonly used today. Swiss albino mice were maintained in colonies that were not inbred (cosanguinous) and are derived from mice that were originally acquired in 1926 by Clara J. Lynch at the Rockefeller Institute from A. Coulon of Lausanne in Switzerland. Descendants of these mice were spread across different laboratories and other lines were created including the SWR/J and SJL/J lines during the 50s. Also in the 50s, to assess the risk of the use of nuclear energy on genetic inheritance, millions of mice were irradiated at the Oak Ridge Centre (Tennessee) and at Harwell (Great Britain). Here they found hundreds of new mutations, which permitted a very precise study of the mouse genome.

Most of these mouse lines were developed for use in cancer research, in order to prove or refute the existence of genetic factors involved in this disease. By using inbred crossings, mouse lines were obtained that had a higher frequency of occurrence, whereas others were resistant to the development of tumors. In parallel, it was found that in some of these inbred lines there appeared a high incidence of other diseases, such as anemia, eye and ear defects, neuromuscular disorders, digestive problems, obesity, bone malformations or urinary tract diseases, etc. It was by this manner that many diseases were first linked to genetic factors.

In 1966, A. K. Tarkowski and B. Mintz produced in vitro mouse chimeras from four different embryos, and them in 1969 R. Gardner obtained chimeric mice by injecting cells into the embryonic blastocyst cavity of a recipient mouse. In 1976, R. Jaenisch produced the first transgenic mice by infecting embryos with a retrovirus. Later, in 1980 and 1981, five laboratories (Gordon et al., 1980; Brinster et al., 1981; Costantini and Lacy, 1981; Harbers et al., 1981; Wagner et al., 1981) more or less simultaneously produced transgenic mice by injecting a DNA fragment into one of the pronuclei of a fertilized oocyte. The next breakthrough, which launched a wide range of applications, was the possibility of replacing genes in the mouse. Initially total «knockouts» were created by deactivating a gene in the whole body, which subsequently became more tissue specific in the so-called conditional «knockouts». Perhaps the last big step towards the current state of the art occurred in December 5, 2002, when the full sequence of the mouse genome was published in Nature (Waterston et al., 2002). Subsequent studies have shown that 99.5% of mouse genes have a homologous gene in human, making the mouse one of the most important models for studying human diseases.

Anatomical Nomenclature

The mouse body is divided externally into the head, neck, trunk, tail and the thoracic (fore) and pelvic (hind) limbs. To situate in a precise position the different parts of the body, it is necessary to know a number of descriptive terms and directional planes (Fig. 1-1). Holding the mouse in its anatomical position, that is, with all four limbs resting on the ground, the term dorsal is used in the trunk to refer to structures located or projecting towards the top of the trunk. The term ventral is used for the structures located or directed towards the belly. By extension, the same terminology is used in the head and tail. The term cranial is used for structures that are positioned or projected towards the head (or cranium), whereas caudal is used for structures located or directed towards the tail. Within the head itself, structures located toward the nose are considered rostral. The mouse body is divided in two halves, the right and left, by the median plane. The structures that lie closest to this plane are said to be medial, in contrast to the lateral structures that are located towards the exterior of the mouse. Planes parallel to the median plane, but that do not pass exactly through the centre of the animal, are called sagittal planes. Furthermore, dorsal planes are horizontal planes parallel to the back, and the transversal planes are perpendicular to the longitudinal axis of the body. A specific nomenclature is also used for the limbs. Structures located near the junction with the trunk are called proximal, whereas the more distant structures are said to be distal. In the forearm and forepaw (Fig. 1-1), the structures located more cranially are called dorsal, while those in the caudal face are called palmar in the forearm and plantar in the forepaw. Further terms are applied to the fingers, where axial and abaxial refer to structures located closer or farther from the longitudinal axis of the third finger.

f01-01-9780128129722

Figure 1-1 Sectional planes. A) Median plane. B) Horizontal plane. C and D) Transverse planes.

1: Cranial; 2: Caudal; 3: Dorsal; 4: Ventral; 5: Lateral; 6: Medial; 7: Rostral; 8: Palmar; 9: Plantar; 10: Proximal; 11: Distal; 12: Brain; 13: Spinal cord; 14: Heart; 15: Lung; 16: Liver; 17: Stomach; 18: Kidney; 19: Coxal bone; 20: Forepaw; 21: Hindpaw; 22: Thoracic limb; 23: Pelvic limb; 24: Radius; 25: Ulna; 26: Aorta; 27: Hepatic portal vein; 28: Thoracic vertebrae; 29: Ribs.

Superficially, the body of the mouse is divided into regions (Figs. 1-2 to 1-5). Firstly there are the regions of the head, which are divided into the skull and face. In the trunk are located the neck regions (dorsal, lateral and ventral), the pectoral regions situated in the thoracic wall, the abdominal regions located at the surface of the abdomen, and the pelvic regions. Finally, the thoracic limb regions and the pelvic limb regions are named in according with bones located in deep.

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Figure 1-2 Body regions. Lateral view.

1: Parotid region; 2: Thoracic vertebral region; 3: Scapular region; 4: Shoulder joint region; 5: Brachial region; 6: Ulnar region (region of elbow joint); 7: Antebrachial region; 8: Forepaw; 9: Costal region; 10: Lumbar region; 11: Caudal abdominal region; 12: Sacral region; 13: Femoral region (region of the thigh); 14: Stifle region; 15: Leg region; 16: Tarsal region; 17: Hindpaw region.

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Figure 1-3 Regions of the head . Lateral view.

1: Frontal region; 2: Orbital region; 3: Infraorbital region; 4: Nasal region; 5: Nostril region; 6: Buccal region; 7: Oral region; 8: Zygomatic arch; 9: Masseteric region; 10: Auricular region.

f01-04-9780128129722

Figure 1-4 Body regions. A) Dorsal view. B) Ventral view.

1: Dorsal midline; 2: Dorsal neck region; 3: Interscapular region; 4: Thoracic vertebrae region; 5: Lumbar region; 6: Sacral region; 7: Region of the root of the tail; 8: Ventral midline; 9: Sternal region; 10: Costal region; 11: Costal arch; 12: Cranial abdominal region; 13: Middle abdominal region; 14: Caudal abdominal region; 15: Pubic region; 16: Preputial region; 17: Scrotal region.

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Figure 1-5 Regions of the forepaw and hindpaw. A) Forepaw dorsal view. B) Forepaw palmar view. C) Hindpaw dorsal view. D) Hindpaw plantar view. The roman numerals indicate the number of the digits.

1: Carpal region; 2: Metacarpal region; 3: Metacarpophalangeal region; 4: Proximal phalangeal region: 5: Middle phalangeal region; 6: Distal phalangeal region; 7: Interdigital space; 8: Tarsal region; 9: Metatarsal region; 10: Calcaneal region; 11: Abaxial surface; 12: Axial surface; 13: Longitudinal axis.

The anatomical terms that have been used here to describe the morphology of the mouse mainly correspond to the «Nomina Anatomica Veterinaria (NAV)» (4th edition, 1992). However, there are anatomical structures that exist in the mouse, the clavicle for example, which are very similar to humans, but different from domestic animals. In these situations, we have used the human anatomical nomenclature published in the «Anatomisches Bildwörterbuch der interantionalen Nomenklatur» (3rd edition, 1993), by H. Feneis; and the «International Anatomical Terminology», by The International Federation of Association of Anatomist (IFAA) and the Federative Committee of Anatomical Terminology (FCAT) (1st edition). For the description of the mouse nervous system, we have also followed the terminology of the book «The Mouse Nervous System» (2012), edited by C. Watson, G. Paxinos and L. Puelles. The histological and cellular description used the terms proposed in the «Nomina Histologica» (2nd edition, 1992). Finally, we have also taken into account the mouse anatomical and pathological ontologies contained in Pathbase (http://eulep.pdn.cam.ac.uk), which have been designed specifically to morphologically phenotype genetically modified mice.

2

Osteology

A. Carretero; J. Ruberte; M. Navarro; V. Nacher; L. Mendes-Jorge

As in other mammals, most of the mouse bones are derived from embryonic mesoderm with the exception of the cranial bones which originate from the cephalic ectomesenchyme (Fig. 2-1). Bone cells are differentiated by the processes of chondrogenesis and osteogenesis, first forming chondroblasts and osteoblasts and later chondrocytes and osteocytes (Figs. 2-3 to 2-7). Bones can be formed from direct ossification processes mediated by osteoblasts, such as occurs in the clavicle and some bones of the skull, or they may be formed by indirect ossification processes. The latter are called endochondral bones and pass through an intermediate cartilage stage. During development, the cartilaginous skeleton of the embryo is progressively replaced by bone tissue via ossification centers. The growth of long bones continues during the early stages of life from the growth plates (Figs. 2-2, 2-7 and 2-8) which are located between the shaft and the epiphysis. On the surface of the bone is found the compact bone tissue, whereas the interior is the so-called trabecular bone (Figs. 2-2 and 2-3), which is arranged in alignment with the mechanical forces which support the bone.

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Figure 2-1 Mouse skeleton. A) Lateral view. B) Dorsal view.

1: Cervical vertebrae; 2:Thoracic vertebrae; 3: Lumbar vertebrae; 4: Sacral vertebrae; 5: Caudal vertebrae; 6: Skull; 7: Orbit; 8: Zygomatic arch; 9: Incisor tooth. 10: Mandible; 11: Tympanic bulla; 12: Scapula; 13: Floating ribs; 14: Asternal ribs; 15: Costal arch; 16: Humerus; 17: Radius; 18: Ulna; 19: Interosseous space of forearm; 20: Forepaw; 21: Coxal bone; 22: Obturator foramen; 23: Femur; 24: Tibia; 25: Fibula; 26: Tarsal bones; 27: Metatarsals and phalanges. 28: Hindpaw; 29: Interparietal bone; 30: Parietal bone; 31: Frontal bone; 32: Nasal bone; 33: Atlanto-occipital space; 34: Atlantoaxial space; 35: Lumbosacral space.

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Figure 2-2 Structure of a long bone (femur). A) Isolated bone. B) Radiography. C) Histological longitudinal section. Hematoxylin-eosin stain.

1: Head of femur; 2: Greater trochanter; 3: Lesser trochanter; 4: Third trochanter; 5: Body of femur; 6: Lateral condyle; 7: Medial condyle; 8: Facets for sesamoid bones of m. gastrocnemius; 9: Sesamoid bones of m. gastrocnemius; 10: Compact bone (Substantia compacta); 11: Bone marrow; 12: Vascular sinus; 13: Growth plate.

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Figure 2-3 Structure of a long bone (femur). A) Diamond saw longitudinal section. B) Histological section. Hematoxylineosin stain (1,000X). C) Histological section of the distal epiphysis. Hematoxylin-eosin stain (100X). D) Histological section of the distal epiphysis. Masson trichrome stain (100X). E) Histological section of the distal epiphysis. Alcian blue stain (100X).

1: Compact bone; 2: Trabeculae; 3: Bone marrow; 4: Endosteum (osteoblasts and osteoclasts); 5: Osteocyte; 6: Megacariocyte; 7: Growth plate; 8: Articular cartilage; 9: Vascular sinus.

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Figure 2-4 Structure of a long bone (femur). A) Transverse histological section. Hematoxylin-eosin stain (100X). B) Endosteum (1,000X). C) Periosteum (1,000X).

1: Third trochanter; 2: Medullary cavity; 3: Bone marrow; 4: Compact bone; 5: Endosteum; 6: Periosteum; 7: Osteocyte.

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Figure 2-5 Structure of a long bone (femur) . A and C) Histological section. Thionin stain (1,000X). B) Histological section. Hematoxylin-eosin stain (1,000X).

1: Osteocyte; 2: Haversian canal; 3: Capillary; 4: Compact bone; 5: Medullary cavity.

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Figure 2-6 Structure of the femoral head. A) Scanning electron microscopy image. B) Histological section. Masson’s trichrome stain (40X and 400X). C) Histological section. Thionine stain (40X and 400X). D) Histological section. PAS and Alcian blue stain (40X and 400X).

1: Head of femur; 2: Greater trochanter; 3: Lesser trochanter; 4: Trochanteric fossa; 5: Bone marrow; 6: Trabeculae; 7: Calcified cartilage; 8: Articular cartilage.

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Figure 2-7 Structure of the tibial growth plate. A) Hematoxylin-eosin stain (200X). B) Masson’s trichrome stain (200X). C) Confocal laser microscopy image. Griffonia simplicifolia (red), nuclei counterstained with Sytox Green (200X).

1: Zone of proliferation; 2: Mature zone; 3: Zone of hypertrophied chondrocytes; 4: Zone of destruction; 5: Zone of ossification; 6: Capillary.

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Figure 2-8 Zone of destruction and calcification in the humeral growth plate . A) Hematoxylin-eosin stain (400X). B) Masson’s trichrome stain (400X). C) Acid phosphatase stain (400X).

1: Osteoclast; 2: Osteoblast; 3: Osteocyte; 4: Bone marrow.

The skeleton of the mouse can be divided into:

• Axial skeleton (skull, mandible, vertebral column and thoracic bones).

• Appendicular skeleton (thoracic and pelvic limbs).

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Figure 2-9 Skeletal ossification. Alizarin red stain. A, C and E) Forelimb and hindlimb from a eighteen days of gestation fetus (E18). B, D and F) Forelimb and hindlimb from a one week old mouse (P7).

1: Bones of cranium; 2: Body of mandible; 3: Ramus of mandible; 4: Zygomatic arch; 5: Clavicle; 6: Sternebrae; 7: Xiphoid process; 8: Ribs; 9: Lumbar vertebrae; 10: Costal cartilage; 11: Scapula; 12: Humerus; 13: Radius; 14: Ulna; 15: Metacarpal bones; 16: Digital skeleton (forepaw); 17: Carpal bones; 18: Ilium; 19: Ischium; 20: Pubis; 21: Femur; 22: Tibia; 23: Fibula; 24: Calcaneus; 25: Talus; 26: Metatarsal bones; 27: Digital skeleton (hindpaw) 28: Tarsal bones.

Bones of the Skull and Mandible

The skull (Figs. 2-10 to 2-18) is formed by the union of several bones usually pairs, which define a series of cavities that house the brain, the sensory organs of sight, hearing, smell and taste, as well as the entrance to the respiratory and digestive tracks.

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Figure 2-10 Skull. A) Diagram. B) Lateral view.

1: Occipital bone; 2: Occipital condyle; 3: Paracondylar process; 4: Interparietal bone; 5: Parietal bone; 6: Temporal bone. Squamous part; 7: Temporal bone. Tympanic part (tympanic bulla); 8: External acoustic pore; 9: Frontal bone; 10: Maxilla; 11: Zygomatic process (maxilla); 12: Infraorbital foramen; 13: Body of maxilla; 14: Basisphenoid bone; 15: Incisive bone; 16: Nasal bone; 17: Incisor teeth; 18: Molar teeth; 19: Pterygoid bone; 20: Lacrimal bone; 21: Zygomatic process (temporal bone); 22: Petrosquamous fissure.

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Figure 2-11 Occipital and temporal bones. A) Skull, lateral view. B) Squamous part of the temporal bone. Lateral view. C) Occipital bone. Lateral view. D) Tympanic part of the temporal bone. Latero-cranial view. E) Petrous part of the temporal bone (the tympanic part was removed). Lateral view.

1: Temporal bone; 2: Occipital bone; 3: Zygomatic process; 4: Parietal border; 5: Frontal border; 6: Sphenoidal margin; 7: Petrosquamous fissure; 8: Occipital process; 9: Retrotympanic process; 10: Parietal margin; 11: Mastoid border; 12: Occipital condyle; 13: Basilar part; 14: Pharyngeal tubercle; 15: Tympanic bulla; 16: External acoustic meatus; 17: Muscular process; 18: Malleus; 19: Mastoid process; 20: Groove for stapedial artery; 21: Groove for occipital artery; 22: Facial canal; 23: Vestibular (oval) window; 24: Cochlear (round) window; 25: Mastoid foramen; 26: Promontory.

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Figure 2-12 Cranium and facial bones. A) Medial view. B) Laterolateral radiography.

1: Occipital bone; 2: Occipital condyle; 3: Hypoglossal canal; 4: Interparietal bone; 5: Parietal bone; 6: Temporal bone; 7: Tympanic bulla; 8: Petrosquamous fissure; 9: Anterior semicircular canal; 10: Internal acoustic meatus; 11: Tentorium cerebelli osseum; 12: Frontal bone; 13: Maxilla; 14: Incisive bone; 15: Nasal bone; 16: Perpendicular plate (ethmoid bone); 17: Incisor tooth; 18: Pterygoid bone; 19: Vomer; 20: Dorsal nasal concha; 21: Paracondylar process; 22: Ethmoidal labyrinth.

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Figure 2-13 Ethmoid bone. A) Ethmoidal labyrinth. B) Ethmoid bone inside the skull. Caudal view. C) Isolated ethmoid bone. Lateral, dorsal, caudal and rostral views, respectively.

1: Cribriform plate; 2: Ectoturbinates; 3: Endoturbinates (I-IV); 4: Ethmoidal meatus; 5: Sphenopalatine foramen; 6: Nasopharyngeal meatus; 7: Incisive bone; 8: Nasal bone; 9: Vomeronasal bone; 10: Crista galli; 11: Wing (presphenoid bone); 12: Optic canal; 13: Frontal bone; 14: Basisphenoid bone; 15: Ethmoidal labyrinth (lateral mass); 16: Ethmoidal cells; 17: Ethmoidal infundibulum; 18: Basal plate; 19: Ethmoidal bulla; 20: Perpendicular plate; 21: Ala of crista galli; 22: Uncinate process.

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Figure 2-14 Cranium and facial bones. Ventral view. A) Diagram. B) Isolated skull.

1: Incisor tooth; 2: Incisive bone; 3: Palatine process (incisive bone); 4: Incisive canal; 5: Maxilla; 6: Zygomatic process (maxilla); 7: Palatine fissure; 8: Palatine process (maxilla); 9: Molar teeth; 10: Palatine bone; 11: Basisphenoid bone; 12: Pterygoid bone; 13: Temporal bone; 14: Zygomatic process (temporal bone); 15: Tympanic bulla (tympanic part, temporal bone); 16: Basioccipital bone; 17: Paracondylar process; 18: Hypoglossal canal; 19: Foramen magnum; 20: Foramen ovale; 21: Major palatine groove and major palatine foramen; 22: Jugular foramen; 23: Sphenotympanic fissure; 24: Occipital condyle; 25: Vomeronasal bone.

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Figure 2-15 Cranium and facial bones. Ventral view. A) Dorsoventral radiography. B) Fetus of 18 days of gestation (E18). Alizarin red stain.

1: Incisor tooth; 2: Palatine process (incisine bone); 3: Incisive bone; 4: Maxilla; 5: Zygomatic process (maxilla); 6: Palatine process (maxilla); 7: Infraorbital foramen; 8: Molar teeth; 9: Zygomatic bone; 10: Vomeronasal bone; 11: Palatine bone; 12: Pterygoid bone; 13: Zygomatic process (temporal bone); 14: Alar canal; 15: Tympanic bulla; 16: Osseous labyrinth; 17: Sphenotympanic fissura; 18: Petrooccipital canal; 19: Petrooccipital fissure; 20: Jugular foramen; 21: Stylomastoid foramen; 22: Hypoglossal canal; 23: Basisphenoid bone 24: Basioccipital bone.

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Figure 2-16 Base of the cranium. Dorsal view of isolated bones.

1: Presphenoid bone; 2: Basisphenoid bone; 3: Occipital bone; 4: Body (presphenoid bone); 5: Jugum sphenoidale; 6: Wing (presphenoid bone); 7: Optic canal; 8: Sphenoidal rostrum; 9: Rostral clinoid process; 10: Body (basisphenoid bone); 11: Sella turcica; 12: Hypophysial fossa; 13: Wing (basisphenoid bone); 14: Infratemporal crest; 15: Carotid sulcus; 16: Groove for trigeminal nerve; 17: Groove for middle meningeal artery; 18: Foramen rotundum; 19: Foramen ovale; 20: Foramen magnum; 21: Basioccipital bone; 22: Pontine impression; 23: Medulla impression; 24: Squamous part of occipital bone; 25: Hypoglossal canal.

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Figure 2-17 Cranium and facial bones. Dorsal view. A) Diagram. B) Skull. C) Isolated nasal, parietal and interparietal bones. D) Nasal bone. Lateral view.

1: Nasal bone; 2: Incisive bone; 3: Maxilla; 4: Frontal bone; 5: Temporal bone; 6: Parietal bone; 7: Interparietal bone; 8: Occipital bone; 9: Infraorbital foramen; 10: Zygomatic process (maxilla); 11: Zygomatic process (temporal bone); 12: Nasal process; 13: External surface (nasal bone); 14: Internasal suture; 15: Frontonasal suture; 16: External surface (frontal bone); 17: Frontal suture; 18: Frontoparietal suture (coronal suture); 19: Temporal line; 20: Frontal crest; 21: Frontal angle; 22: Sagittal suture; 23: Interparietal border; 24: Parietal plane; 25: Parietotemporal suture (squamous suture); 26: Squamous border; 27: Parietal border; 28: Occipital border; 29: Septal process; 30: Ethmoidal groove.

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Figure 2-18 Cranium. A) Diagram. Caudal view.