Comparative Medicine
Volume 60, Number 3, June 2010
OVERVIEW
Lynch et al. Animals Models of Substance Abuse and Addiction: Implications for
Science, Animal Welfare, and Society, pp. 177-188
Domain 1: Management of spontaneous and experimentally induced disease
SUMMARY: The article describes the background, importance, and concerns in the use
of animals for drug addiction studies. It provides an introduction to the terminology,
administration paradigms, behavioral effects, and welfare issues of these types of
studies. Most drug addiction models involve a number of special considerations in
comparison to other disease models in that they require chronic administration,
instrumentation, may involve food/water restrictions, and the effects of drug
withdrawal. In paradigms involving self-administration of drugs, chance of overdose
must be prevented by limiting either the duration of time the animal has access or the
total volume. There is an issue of public perception regarding these studies, where drug
abuse is believed to be a self-inflicted condition of humans not worthy of the use of
animals. Investigators should collaborate with veterinary staff in ensuring appropriate
animal care, considering any alternatives, and continuing to improve the models.
QUESTIONS:
1. What is one of the compelling reasons for the use of a drug of abuse in a nonhuman primate over a rodent?
2. What is a reinforcer?
3. In what type of experiment is an animal treated with a drug and placed in an
environment with a visual or tactile cue over a series of days, followed by a test
where the animal is allowed to freely roam between two environments and spend the
greater amount of time in the environment in which they were treated?
4. What are the signs of opioid withdrawal in rodents?
ANSWERS:
1. The pharmacokinetics may not allow extrapolation of data from rodents to humans.
2. A stimulus event that strengthens the behavior that follows it.
3. Conditioned place preference.
4. Diarrhea, rhinorrhea, teeth chattering, “wet dog shakes”, anorexia, weight loss.
ORIGINAL RESEARCH
Mouse Models
Paik et al. Effects of Murine Norovirus Infection on a Mouse Model of Diet-Induced
Obesity and Insulin Resistance, pp. 189-195
Domain 3: Research
Primary Species: Mouse
SUMMARY: The objective of this study was to determine if Murine Norovirus (MNV)
infection in C57BL/6 mice (a widely used model of diet-induced obesity) altered obesity
and insulin resistance phenotypes. MNV is prevalent in SPF mouse facilities in the
United States, and we currently lack sufficient data to determine whether it should be
eliminated. It is generally accepted that the virus does not cause clinical symptoms in
immunocompetent mice. However, it was previously reported that MNV infection alters
the phenotype of a mouse model of bacteria-induced inflammatory bowel disease in part
through its effects on dendritic cells. The tropism of MNV towards macrophages and
dendritic cells makes MNV a potential intercurrent variable in murine models of
macrophages-driven inflammatory diseases, such as obesity, insulin resistance, and
artherosclerosis. It was found that MNV infection status did not alter weight gain, food
intake, and glucose metabolism due to high-fat feeding in this model. MNV infection
induced subtle changes in lymphoid tissue. Further studies using other models of
metabolic diseases are needed to provide additional information on the potential role this
‘subclinical’ virus might have on disease progression in mouse models of inflammatory
diseases.
QUESTIONS:
1. T or F: Although MNV does not cause any overt illness in immunocompetent mice,
significant inflammation and mortality can be induced in mice with abnormal innate
immunity.
2. T or F: MNV is known to preferentially infect macrophages and dendritic cells
3. T or F: this study shows that MNV infection did alter glucose homeostasis and lipid
metabolism
4. T or F: In this study, MNV infection was not associated with minimal changes in
lymph nodes
ANSWERS:
1. T
2. T
3. F
4. F
Mordica et al. Male CD81 Knockout Genotype Disrupts Mendelian Distribution of
Offspring, pp. 196-199
Domain 1. K10. Genetics with emphasis on control and treatment of naturally occurring
and experimentally induced disease, predisposition to disease, and modes of inheritance
Primary Species: Mouse (Mus musculus)
SUMMARY: CD81, an adaptor protein in the tetraspanin superfamily, is found on
chromosome 7 of the mouse along with other maternally imprinted genes and is a
required receptor for hepatitis C virus infection. It has been shown to also play a role in
embryo/fetal development. Homozygous null females have been shown to produce
fertilized embryos 40% less frequently than CD81 expressing females. While CD81
expression is important for fertility in female mice, male CD81 knockouts have been
reported as fertile, though it is unknown if genotype distribution was examined by the
investigators. Records from a CD81 knockout breeder colony over 5 years showed
infertility in homozygous null females and skewed genotypes in litters sired by
homozygous null male mice. Successful birth of homozygous null mice in litters sired by
homozygous null males was found to be about 50% of the expected frequency. The 2:1
ratio of heterozygous: homozygous-null births in mice sired by homozygous null males
was observed in two separate strain lines, one backcrossed to the BALB/c line for only 7
generations, while the other had been backcrossed for 20 generations. This suggests
that interactions from the mixed BALB/c and 129 backgrounds likely did not play a role in
the process. Homozygous null mice that were born were found to develop similarly to
their CD81-expressing littermates, suggesting that CD81 deficiency either does not
affect vital functions or can be complemented by other tetraspanins once the mouse is
born. It is currently unknown how the CD81 genotype of the sire affects the genotypic
distribution of the offspring.
QUESTIONS:
1. What chromosome is CD81 found on?
2. What percentage of the expected births of homozygous null offspring from
homozygous null sires were successful?
ANSWERS:
1. Chromosome 7
2. 50%
Lawson. Etiopathogenesis of Mandibulofacial and Maxillofacial Abscesses in
Mice, pp. 200-204
Domain 1: Prevent, Diagnose, Control, and Treat Disease Primary Species - Mouse
SUMMARY: There are very few cases of either mandibulofacial or maxillofacial
abscesses in mice reported in the literature. The reports available discuss the bacteria
that were involved (S. aureus) and the location of the abscesses but only speculate as to
the route of entry. The purpose of this retrospective study was to examine a number of
cases of either maxillary or mandibular abscesses in the rodent colony at UC-Los
Angeles and propose an etiopathogenesis.
The study involved 39 mice of various strains with white, agouti, and black coat colors
identified during routine health checks. The abscesses were cultured immediately after
euthanasia. Complete necropsies were performed followed by formalin fixing of the
heads. The heads were scanned with microcomputed tomography and the molars
examined microscopically.
Two of the mice with mandibular abscesses were
radiographed. All of the skulls were sectioned transversely and prepared for histological
examination.
A review of the records showed that facial abscesses consisted of 0.3% of the health
cases in that particular rodent colony during the time period examined. Maxillofacial
abscesses were more common than mandibulofacial but precise numbers were
unavailable. All of the abscesses examined were associated with hair. The two types of
hair identified histologically were consistent with vibrissiae and pelage hair. They
appeared to penetrate and expand the gingival sulci of the associated molars.
Abscesses were composed of well-defined, multilobulated, botryoid lobules separated by
thick bands of dense fibrous connective tissue. Each lobule contained large numbers of
neutrophils, clusters of coccoid bacteria, and central Splendore-Hoeppli material which is
consistent with S. aureus infection. Osteolysis was common to both abscess locations
but was more severe in those in the mandible and often involved exostosis as well.
Severe cases involved migration into the retroorbital space and inflammation of the
Harderian gland. All cultures were positive for coagulase-positive S. aureus.
The proposed etiopathogenesis is as follows: excessive barbering or grooming activities
leads to mastication and fragmentation of hair, which then becomes entrapped and
impacted in the gingival sulcus of the molar teeth. The hair acts as a foreign body,
resulting in ulceration and inflammation of the periodontium, and carries with it S. aureus
into the submucosa. Abscesses originating at a maxillary gingival sulcus results in an
infection of the maxillary molar tooth roots and rupture through the skin inferior to the
eye. Originating in a mandibular gingival sulcus results in infection of the mandibular
molar tooth roots, with or without osteomyelitis, leading to drainage through the skin of
the ventral mandible.
QUESTIONS:
1. Which bacterial species is the most commonly cultured from a mouse facial abscess?
a. Streptococcus oralis
b. Lactobacillus murinus
c. Staphylococcus aureus
d. Actinomyces bovis
2. What is being proposed as the most common route for infection to be established in a
mouse facial abscess?
a. Fractures of the teeth
b. Penetration by hair into the mucosa of the gingival sulcus
c. Food impaction in the gingival sulcus
d. Hematogenous infection of the tooth pulp
3. T/F. The most common location for facial abscesses to form in the mouse is reported
as the maxilla.
4. T/F. Maxillary abscesses usually involve more severe osteomyelitis than mandibular
abscesses.
ANSWERS:
1. C
2. B
3. T
4. F
Rat Models
Kwiecien. Cellular Compensatory Mechanisms in the CNS of Dysmyelinated Rats,
pp. 205-217
SUMMARY
Introduction: This study was intended to analyze the progressive neuropathology in 2
dysmyelinated mutant rat strains, the Long Evans Shaker (LES) and the Long Evans
Bouncer (LE-bo).
Materials and Methods: They evaluated the CNS tissue of control Long Evans, LES and
LE-bo rats at 1, 2, 4, 8, 12, 16, 20, 24, 28, 32, 36 and 40 wks of age. The natural
longevity of the mutant rats in this study was 93 wks for the LES and 45 wks for the LEbo. The tissues were evaluated using light and electron microscopy (LOTS of pictures).
Results: PCR confirmed that both mutants are affected by the same mutation in the
myelin basic protein gene and both lack normal, compacted myelin sheaths throughout
their lives. In this study, the oligodendrocytes showed degenerative changes
characterized by loss of Golgi structures and accumulation of membranous material.
The membranous material appeared as vesicles limited by a pentolamellar membrane (3
dense lines separated by 2 lucent lines). As the rats aged, the vesicles were replaced
by larger, honeycomb-like stacks. The honeycombs have a periodicity of 5 nm as
opposed to myelin sheaths which have a periodicity of 15 nm. The morphology was the
same in both the LES and the LE-bo rats. Despite an increase in immature
oligodendrocytes in these animals, which occurred well into adulthood, there was only a
very rare occurrence of thin myelin sheaths.
Discussion: Hallmarks of neuropathology in dysmyelinated mutant rats:
 Mitotic glial cells and differentiation of new cells into oligodendrocytes
 Degenerative oligodendrocytes
 Ineffective myelinating activity
 Abundant axonal sprouting
Both mutant strains are prone to seizures which can result in hyperflexion and severe
injury to the midthoracic region of the spinal cord. This can lead to acute hind limb
paralysis which was removal criteria for the study. This can affect up to 50% of animals,
though if they live to be older than 15 wks they are considered resistant to this
syndrome.
The LES and LE-bo rats show differences in:
 The kinetics of the glial cell proliferation
 The age at which ineffective myelinating activity declines and the age at which
oligodendrocyte pathology progresses
The differences indicate that although they have the same functional mutation, they do
have separate phenotypes.
QUESTIONS:
1. What is the mutation present in both the LES and LE-bo mutant rat strains?
2. These mutant rat strains can serve as models for what conditions in humans?
3. What are other rodent dysmyelinated mutant models?
ANSWERS:
1. A mutation of the myelin basic protein gene
2. Multiple Sclerosis, Spinal Cord Injury
3. Jimpy (jp) mice, myelin-deficient (md) rats, shiverer (shi) mice, and old taiep rats
Cox and Kalns.
Development and Characterization of a Rat Model of
Nonpenetrating Liver Trauma, pp. 218-224
SUMMARY: Blunt (nonpenetrating) liver trauma is an important source of morbidity and
mortality in the United States due to multiple causes such as the following: automobile
accidents, falls, wartime operations, etc. Diagnosing and properly treating patients with
hepatic injury may be complicated. Although 71% to 89% of all blunt liver injuries can be
assessed nonsurgically with current diagnostic practices, patients who are
hemodynamically unstable or have peritonitis must undergo either diagnostic peritoneal
lavage or emergency laparotomy before the full extent of hepatic injury can be
diagnosed. Historically, swine have been the animal model of blunt liver trauma of
choice (this trauma has been produced in swine using a variety of mechanical methods
(e.g. striking with a blunt object). Prior to this study, a rodent model for the study of blunt
liver trauma has never been developed and characterized.
In this study, the authors established a reliable and reproducible method for inducing
blunt force liver trauma in the rat. Liver trauma was created by dropping a steel cylinder
through a plastic tube onto the abdomen of supine, anesthetized rats. The experimental
groups were as follows: 1) sham (anesthesia) control; 2) hemorrhage control (internal
hemorrhage in the absence of liver trauma was simulated by instilling fresh blood into
the peritoneum); 3) liver trauma induced by dropping the punch from 50 cm; and 4) liver
trauma induced by dropping from 100 cm. Rats were euthanized and evaluated at 2 and
24 h after treatment (control or trauma). The severity of the injury was indeed found to
be dependent on the distance that the projectile is dropped. Through investigation of
cytokine and chemokine production, gene and protein expression, and
immunohistochemical analysis, the authors demonstrated the usefulness of this model in
detection of physiologic and inflammatory hallmarks associated with traumatic liver
injury. This rat model of liver trauma may be useful in future applications for both
improving clinical diagnosis and treatment of traumatic liver injury and is also noteworthy
in that it may be used to replace the use of higher sentient species (notably swine) in
liver trauma research.
The specific findings were as follows: Multiple cytokines were examined and it was
discovered that Gro-KC appears promising as a potential biomarker for liver trauma.
Gro-KC is a rat homolog to human Groα. Gro proteins are secreted by Kupffer cells, the
interstitial macrophages of the liver that act as its first line of defense. Groα is believed to
be responsible for the chemotaxis of neutrophils in response to injury. Trauma due to
dropping the projectile from 100 cm caused Gro-KC levels to increase 9-fold compared
with those of the anesthesia control. However, injection of blood into the abdominal
cavity increased Gro-KC expression more than 5-fold compared with the anesthesia
control, suggesting that blood in the peritoneal cavity is an important signal leading to
this response.
Levels of platelets in blood were elevated at 2 h after liver trauma but returned to
baseline levels by 24 h. In contrast, platelet count decreased after injection of blood into
the peritoneum, as expected. The observed increase in platelet levels after trauma may
be attributable to an increased expression of PAI1 in liver after trauma but not injection
of blood into the peritoneal cavity. We observed significant increases (77- and 7-fold)
increase in expression in PAI1 at 2 and 24 h, respectively, after trauma. PAI1 is a
glycoprotein that inhibits both plasminogen activator and urokinase activator and
contributes to decreased production of plasmin, which in turn causes inhibition of
fibrinolysis.
Results reveal increased expression of HIF1α 24 h after liver injury. HIF1α is responsible
for cell migration and differentiation, matrix metabolism, angiogenesis, and tissue repair.
Increased
HIF1α expression has been shown to have a direct relation to severity to fibrogenesis
within the liver.
Transcript levels for HSP70 were elevated at 2 h after trauma but returned to baseline by
24 h.
HSP70 transcription is induced when cells respond to increases in calcium generated by
trauma or shock. HSP70 has been shown to protect hepatocytes against oxidative stress
and apoptosis.
Hepatocytes and Kupffer cells release a number of cytokines and chemokines in
response to injury and inflammation in the liver. In turn, SOCS3 and iNOS are released
as regulators to this influx of activity. SOCS3 gene expression was significantly
increased at 2 h and decreased by 24 h. iNOS is responsible for the synthesis of nitric
oxide, a free radical, which protects hepatocytes in acute inflammation and injury.
QUESTIONS:
1. Prior to the establishment of this rat model, which animal model has historically been
used a model of blunt force liver trauma?
2. Multiple cytokines were serologically assessed. Which one was found to be a
promising biomarker for liver trauma?
3. Gro-KC is a rat homolog to which human cytokine?
4. Platelet levels were found to be increased in the blood after the trauma episode as
this appears to be attributable to an increase in PAI1 (a plasminogen activator
inhibitor), which, in turn, results an inhibition of fibrinolysis. T/F
ANSWERS:
1. Pig
2. Gro-KC
3. Groα
4. True
Chinchilla Model
Grieves et al. Mapping the Anatomy of Respiratory Syncytial Virus Infection of the
Upper Airways in Chinchillas (Chinchilla lanigera), pp. 225-233
Domain 3 - Research
Task 3 - Design and conduct research
Tertiary Species – Chinchilla
SUMMARY: Human respiratory syncytial virus (HRSV) is an enveloped, negative strand,
nonsegmented RNA virus of the family Paramyxoviridae. It is the single greatest
causative agent of acute respiratory tract infections in infants and children worldwide.
Upper respiratory tract (URT) infection by HRSV alone does not constitute a serious
problem for healthy adults; its association with the development of bacterial otitis media
in children and exacerbation of asthma in all age groups make it an important health
concern. In addition, strains remain in circulation over several seasons or reappear
many years after they were first detected. HRSV may circulate among seropositive
persons and it has been suggested that persistently infected persons may harbor the
virus between seasonal outbreaks. Therefore, basic scientific questions regarding HRSV
circulation and mechanisms of viral immunoevasion need to be explored. This process
has been slowed by the lack of a suitable animal model. A model of the URT infection
seen in humans is needed and currently two species seem to fulfill these criteria, the
cotton rat (Sigmidon hispidus) and the chinchilla (Chinchilla lanigera).
For this study the group characterized HRSV infection of the uppermost airway in
chinchillas challenged in a manner designed to restrict viral delivery to the nasal cavity
without dose loss to either the GI tract or the lungs. The HRSV spread from the site of
inoculation to the pharyngeal orifice of the Eustachian tube by 48 h after the challenge
but 5 days were required before the virus could be detected in the distal most aspect of
the Eustachian tube. Immunohistochemistry was performed to determine the
microanatomy of the infection and the cells involved. Viral clearance was measured by
plaque assay and confocal microscopy and was complete by 5d post-inoculation;
however, immunopositive epithelial cells were still present in sections taken from
nasoturbinates and ethmoid turbinates on day 14. This suggest that low-level upper
airway infection continues beyond the observation period.
QUESTIONS:
1. Human respiratory syncytial virus is a RNA virus of the family
____________________.
2. URT infection by HRSV does not constitute a serious problem for adults; however, it
does have an association with the development of what in children?
3. What other rodent species besides chinchilla show a susceptibility to URT by HRSV?
ANSWERS:
1. Paramyxoviridae
2. Bacterial otitis media
3. Cotton rats
Nonhuman Primate Model
Hobbs et al. Comparison of Lactate, Base Excess, Bicarbonate, and pH as
Predictors of Mortality after Severe Trauma in Rhesus Macaques (Macaca
mulatta), pp. 233-239
Domain 1, T4: Treat disease or condition as appropriate
Domain 3, Task 3: Design and conduct research
Primary Species: Macaques
One-line synopsis: In this article the authors suggest that clinicians should give more
attention to serum lactate levels as a predictor of 6-d survival in rhesus macaques
recovering from severe trauma and shock.
SUMMARY: The purpose of this retrospective study was to evaluate venous blood
lactate, base excess, bicarbonate, and pH values, obtained before and after treatment of
shock, as predictors of 6-d mortality after trauma in rhesus macaques. All 4 variables
measured prior to shock therapy were associated with survival and were strongly
correlated to each other. However, none of them were highly sensitive or specific preresuscitation.
The data suggested that a guarded or poor prognosis should be given for multiple
trauma cases with 2 or more indicators of acidosis at admission; while a good prognosis
given for multiple trauma cases with values in the normal range at admission. Postresuscitation, the only indicator of acidosis that was associated with survival was serum
lactate.
As outlined by the authors, this work would help in triage decisions, in determining when
humane euthanasia is appropriate and in suggesting a valid measure that can be used
to compare various treatments of traumatic shock in macaques.
QUESTIONS:
1. Which one of the following statements regarding the article is FALSE?
A. A consequence of social housing of macaques is intraspecific aggression
B. The major lesions seen after this aggression are crush injuries from multiple bite
wounds to limbs only.
C. The wounds received can lead to vasodilatory shock which in turn results in a
significant oxygen debt to tissues.
D. The current predominantly used test to assess cellular oxygen utilization is the
measurement of systemic markers of metabolic acidosis.
2. Which one of the following statements regarding the 4 variables used in this study is
FALSE?
A. Lactate is an end-product of anaerobic metabolism
B. pH is a direct measure of acidemia
C. Base excess is determined by using carbon dioxide, pH and lactate values
D. Base excess is viewed as a surrogate marker for lactic acidosis
ANSWERS:
1. B. Among many other lesions, bite wounds are also seen on the face.
2. C. Base excess is determined by using carbon dioxide, pH and, and serum
bicarbonate values.