Exposure of the Pig CNS for Histological Analysis: A Manual for Decapitation, Skull Opening, and Brain Removal

Summary

The goal of this paper and instructional video is to describe how to expose and remove the postmortem pig brain and pituitary gland in an intact state, suitable for subsequent macroscopic and histological analysis.

Abstract

Pigs have become increasingly popular in large-animal translational neuroscience research as an economically and ethically feasible substitute to non-human primates. The large brain size of the pig allows the use of conventional clinical brain imagers and the direct use and testing of neurosurgical procedures and equipment from the human clinic. Further macroscopic and histological analysis, however, requires postmortem exposure of the pig central nervous system (CNS) and subsequent brain removal. This is not an easy task, as the pig CNS is encapsulated by a thick, bony skull and spinal column. The goal of this paper and instructional video is to describe how to expose and remove the postmortem pig brain and the pituitary gland in an intact state, suitable for subsequent macroscopic and histological analysis.

Introduction

Translational neuroscience studies in pigs have become increasingly popular during the last two decades. The large size of the pig brain enables the use of conventional clinical brain imagers and the direct use and testing of neurosurgical procedures and equipment from the human clinic^{1,2,3,4,5,6,7,8}. In the last 20 years, pigs, especially minipigs (e.g., Göttingen minipig), have been used ....

Protocol

Animal anesthesia and euthanesia was performed in accordance with "Principles of laboratory animal care" (NIH publication No. 86-23, revised 1985) and approved by the Danish Council for Animal Research Ethics.

1. Instruments

1. Collect the instruments presented in the video and listed in the Table of Materials.

2. Decapitation

NOTE: Anesthesia was induced by an intramuscular injection of 5 mL of midazolam (5 mg/mL) and 5 mL of ketamine (25 mg/mL). 5-10 min later, when the animal was deeply sedated, an ear vein was cannulated and a lethal overdose (100 mg/....

Results

To prevent the tissue material from drying out, it is recommended to store the removed brain and pituitary in a jar filled with fixative or isotonic saline immediately after macroscopic analysis has been performed. The tissue material may be stored in the fixative for years, whereas storage in isotonic saline, even in a refrigerator, will lead to tissue decay with time.

The removed pituitary may also be directly frozen by immers.......

Discussion

Most experimental neuroscience studies are performed in small animal species, such as mice and rats, where access to the CNS is facilitated by a thin skull- and dural-thickness. However, in larger experimental animals like pigs^1,4,8, sheep³², and non-human primates, the considerable thickness of these structures necessitates the use of robust instruments (table of materials) and proper entry points for sk.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Heavy Scalpel Handle #4	FST (Fine Science Tools)	10008-13	Good for skin incision and soft tissue removal
Non-Sterile Scalpel Blades #23	FST	10023-00
Scalpel Handle #7	FST	10007-12	Optimal for dural incision and precision work
Non-Sterile Scalpel Blades #11	FST	10011-00
Surgical Forceps	FST	11024-18	The tip of the surgical forceps ensure a firm grip
Kerrison Bone Punch	Aesculap Neurosurgery	FF713R	Must be robust, bite size 3-5 mm
Bone Rongeur	Aesculap Neurosurgery	MD615	Must be robust, bite size 15 x 5 mm
Bone Rongeur	Aesculap Neurosurgery	FO551R	Must be robust, bite size 25 x 15 mm
Bone Chisel	Lawton	67-0335	The size of the chisel head should not exceed 20 mm
Mallet (Hammer)	Millarco	5624108	Weigth 300 g, length 30 cm, head hit area size 2 x 2 cm
Micro-Scissor	FST	14002-14
Dissector	Aesculap Neurosurgery	OL165R
Göttingen minipigs	Ellegaard Göttingen Minipigs A/S, Denmark
Euthanimal			pentobarbital
Ketamine	Pfizer
Midazolam	Hameln Pharmaceuticals