A Practical Guide to the Histology of the Mouse

Cheryl L. Scudamore

• Publisher: Wiley-Blackwell
• Year: 2013
• ISBN:
  9781118789476
• Edition: 1
• Topic: Medicine
• Subtopic: Medical Theory, Practice & Reference

A Practical Guide to the Histology of the Mouse provides a full-colour atlas of mouse histology. Mouse models of disease are used extensively in biomedical research with many hundreds of new models being generated each year. Complete phenotypic analysis of all of these models can benefit from histologic review of the tissues.

This book is aimed at veterinary and medical pathologists who are unfamiliar with mouse tissues and scientists who wish to evaluate their own mouse models. It provides practical guidance on the collection, sampling and analysis of mouse tissue samples in order to maximize the information that can be gained from these tissues. As well as illustrating the normal microscopic anatomy of the mouse, the book also describes and explains the common anatomic variations, artefacts associated with tissue collection and background lesions to help the scientist to distinguish these changes from experimentally- induced lesions.

This will be an essential bench-side companion for researchers and practitioners looking for an accessible and well-illustrated guide to mouse pathology.

• Written by experienced pathologists and specifically tailored to the needs of scientists and histologists
• Full colour throughout
• Provides advice on sampling tissues, necropsy and recording data
• Includes common anatomic variations, background lesions and artefacts which will help non-experts understand whether histologic variations seen are part of the normal background or related to their experimental manipulation

Chapter 1

Necropsy of the mouse

Lorna Rasmussen and Elizabeth McInnes

Cerberus Sciences, Thebarton, Australia

Necropsies on mice are a fundamental part of the research process (Fiete and Slaoui 2011) and it is vital that, in every laboratory where animal research is conducted, prosectors (persons who perform necropsies or post mortem examinations) are trained to perform a complete mouse necropsy.

Necropsies on mice are performed for a number of reasons including harvesting of tissues for research, health surveillance and investigation of disease. The process may involve collection of tissues for pathology (e.g. for phenotyping or analysis of research models), but also collection of appropriate samples for microbiological and parasitological examinations for disease identification. Autolysis after death begins immediately after the onset of hypoxia as a result of cessation of blood flow (Slauson and Cooper 2000). Autolysis of the small intestine will commence within 10 minutes of the death of the animal, resulting in the swelling of villus tips and epithelial denudation of the villi (Pearson and Logan 1978a,b). Bone marrow and adrenals are also susceptible to rapid autolysis. Storage of mouse carcases in a refrigerator (2–4 °C) is recommended to avoid rapid autolysis, which may occur in the warm atmosphere of an animal house.

A systematic approach to the mouse necropsy, which allows the examination of all the tissues in the animal in the most expedient manner, is recommended (Slaoui and Fiette 2011). In this chapter, the authors describe a recommended protocol, but variations on this method may exist, depending on the target organ or disease model. It is important to conduct a complete necropsy and to avoid the temptation of just looking at the organs of interest or selection of organs (Seymour et al. 2004). Necropsy results should always be viewed in conjunction with ante mortem clinical signs and haematology and biochemistry results. It is advisable to prepare all instruments, sample collection materials, camera and forms before beginning the necropsy (Knoblaugh et al. 2011). Necropsy personnel should always have access to the experimental study plan so that they can collect particular tissues that are pertinent to the study (Fiette and Slaoui 2011).

Mouse necropsies carry risks of zoonoses, allergen exposure and exposure to hazardous materials such as formalin (Fiette and Slaoui 2011). Appropriate equipment is thus necessary to conduct the mouse necropsy procedure. This includes equipment to conduct appropriate and ethical methods of euthanasia, if the mouse is still alive. Different methods of euthanasia of laboratory mice include carbon dioxide asphyxiation, barbiturate overdose and cervical dislocation (Seymour et al. 2004). Carbon dioxide asphyxiation is a rapid and humane form of euthanasia for mice over the age of seven days (Seymour et al. 2004), but can result in significant agonal haemorrhage, which can complicate microscopic examination of the lungs. Only one mouse should be placed in the perspex container at a time and carbon dioxide gas slowly added to the chamber. A flow rate of 20% V/min CO₂ as a gradual fill or slow filling method for the chamber results in least evidence of stress in mice (Valentine et al. 2012). Barbiturate overdose is an effective and efficient form of euthanasia and requires the use of pentobarbital sodium (Seymour et al. 2004). Decapitation of adult mice should be avoided because the method has welfare concerns and may not be accepted by the ethics committee of an institute or the Home Office (United Kingdom) (Seymour et al. 2004). Cervical dislocation was first approved for mouse euthanasia in 1972 by the AVMA Panel on Euthanasia (Carbone et al. 2012). The disadvantages of cervical dislocation are that although it may be a quick and efficient method of euthanasia, it causes damage to the tissues in the cervical area and may cause the release of large amounts of blood into the body cavities (Seymour et al. 2004). In addition, Carbone and co-workers (2012) examined spinal dislocation and noted that of the 81 mice that underwent cervical dislocation, 17 (21%) continued to breathe and euthanasia was scored as unsuccessful.

Further equipment for mouse necropsy includes a controlled air-flow cabinet or down draft table and plastic containers of formalin and syringes. The controlled air-flow cabinet is not always available in all facilities and is not essential, however it does reduce the risk of noxious substance inhalation (e.g. formalin), or allergen exposure and it is also essential to control the spread of known pathogens or zoonotic agents from the mouse carcase. Cover slips and glass slides may also be required for the preparation of cytology and parasitology specimens. It is important to have a flat and contained area in which to perform the necropsy. A flat board made of rubber or plastic is advisable so that it can be decontaminated or autoclaved if necessary. A metal tray to hold the mouse carcase may also be useful. Furthermore, two pairs of forceps, scalpel blades and scalpel blade holder, one pair of sharp-edged scissors, disinfectant spray and racks for Eppendorf containers and other plastic containers as necessary depending on the sampling protocol will be required. In addition, paper towels are necessary throughout the necropsy procedure (Knoblaugh et al. 2011). Some prosectors will prefer to pin the mouse carcase to a flat cork board while others may prefer to move the mouse during the necropsy procedure. A metric ruler is important for measuring organs and lesions (Knoblaugh et al. 2011). Plastic containers of 10% neutral buffered formalin (NBF) or 4% paraformaldehyde as well as containers of other fixatives (such as modified Davidson's for the fixation of testes and eyes) and containers for microbiology and molecular biology samples should be present at the start of the necropsy process and should all be labelled with the correct mouse identification number. A syringe and plastic cannula or needle (22G) for perfusing the lungs with formalin should also be present at the start (Braber et al. 2010). Mouse adrenals, pituitary gland and lymph nodes are notoriously small and difficult to handle and the use of cassettes with foam pads or biopsy bags to store them in so that they are not mislaid is highly recommended (Knoblaugh et al. 2011).

1.1 Recording of findings

During the necropsy the prosector must record, in some form, all the observations made during the necropsy examination. This will provide a valuable aid at the end of the necropsy and after the histopathological examination of the slides has been conducted, to form conclusions about what abnormalities were observed and may form part of the data set for the experimental group (Chapter 2). The observations may be written down on a specific form designed for that purpose or they may be entered into a computer data-collection program. It is a good idea to develop a checklist for the mouse necropsy procedure, which is referred to each time and on which organ systems may be ticked off as they are inspected and collected.

The correct identification of the animal is very important. The researcher must examine the information on the cage lid, the ear tag of the animal or the ear notches (Figure 1.1) or scan the microchip in order to confirm the exact identification of the animal to be necropsied. The prosector may have to consult a specific key indicating what number each ear notch represents as ear notch keys are usually specific to particular research institutions. It is also important to label all samples generated from the necropsy with the same mouse identification number. The age, sex, strain, genotype, reason for submission and study number (if appropriate) should also be recorded if they are known (Seymour et al. 2012) the body weight, in grams, at the time of necropsy should be recorded. Retaining the identification (ear tag, ears or chip) with the fixed org...

Table of contents

• Cover
• Title Page
• Copyright
• List of contributors
• Foreword
• Preface
• About the companion website
• Chapter 1: Necropsy of the mouse
• Chapter 2: Practical approaches to reviewing and recording pathology data
• Chapter 3: Gastrointestinal system
• Chapter 4: Cardiovascular system
• Chapter 5: Urinary system
• Chapter 6: Reproductive system
• Chapter 7: Endocrine system
• Chapter 8: Nervous system
• Chapter 9: Lymphoid and haematopoietic system
• Chapter 10: Integument and adipose tissue
• Chapter 11: The respiratory system
• Chapter 12: Special senses
• Chapter 13: Musculoskeletal system
• Index