Non-Human Primate Models of Neonatal Brain Injury
Terrie Inder, MD, PhD,* Jeffrey Neil, MD, PhD,† Bradley Yoder, MD,‡
and Sandra Rees, PhD§
Nonhuman primate species have been selectively used in the scientific investigation of
adult and newborn neurological diseases. The rhesus monkey has been utilized in models
of term asphyxial insults, accurately reflecting the mechanisms and neuropathology demonstrated in the newborn human infant. More recently, a premature baboon model developed for evaluation of bronchopulmonary dysplasia has been applied to the investigation of
cerebral development and injury, revealing high similarity in neuropathology to the premature human infant. Given the differences in the outcomes of neuroprotective therapies
between lower order species, such as the rat, and human trials in disorders such as stroke,
nonhuman primate models may provide an invaluable resource for safety and efficacy
testing before trials in human newborns. This article summarizes both models of brain
injury. The histologic findings from the models are compared with neuropathological
studies in human infants.
Semin Perinatol 28:396-404 © 2004 Elsevier Inc. All rights reserved.
T
he animal species most suitable for modeling human
neurological disease remains controversial. The use of
nonhuman primates for the study of neurological diseases is
clearly not appropriate for all disorders because of the cost in
time and resources necessary to develop and use these models combined with ethical considerations. Further, many important fundamental pathophysiological mechanisms are being successfully addressed in other species. Rats are the most
commonly used species for the investigation of neurological
disease in adults and newborns (Table 1; see later chapter
“Experimental models of perinatal brain injury”). Although
there are undeniable differences between rats and primates
with respect to the complexity of cortical motor organization,
there is an increasing recognition that there are marked functional similarities1,2 including the cortical and basal-ganglia
circuits controlling the development, maintenance and selection of adaptive motor response.3 Thus, many aspects of neurological disease and its functional correlates can be tested in
lower order species. However, nonhuman primate models,
*Neonatal Neurology, Royal Women’s and Royal Children’s Hospital, Murdoch Children’s Research Institute, Parkville, Victoria, Australia.
†Departments of Neurology, Pediatrics, and Radiology, Washington University School of Medicine, St. Louis, MO.
‡Southwest Foundation for Biomedical Research, and Department of Pathology, University of Texas, San Antonio, TX.
§Department of Anatomy and Cell Biology, University of Melbourne,
Parkville.
Address reprint requests to: Dr. Terrie E. Inder, Department of Neurology,
Royal Children’s Hospital, 6th Floor, Flemington Road, Parkville, VIC
3052, Australia. E-mail: terrie.inder@rch.org.au
396
0146-0005/04/$-see front matter © 2004 Elsevier Inc. All rights reserved.
doi:10.1053/j.semperi.2004.10.002
such as the baboon model described below, provide unprecedented fidelity to the human condition. Consequently, they
may be appropriate for testing agents and interventions
proven successful in lower order species before their evaluation in human studies. Thus, lower order and nonhuman
primate models can serve complementary roles in the evaluation of neurologic disorders.
Primate Taxonomy
The order primate includes humans, apes, monkeys, prosimians and related animals. All of the 190 or more species of
primates are classified into one taxonomic order because they
are mammals with close evolutionary roots. Nonhuman primates are limited in their natural habitats to the tropical and
subtropical areas of the New World (Americas) and Old
World (Africa and Asia). There are striking morphometric
similarities in primates, including the retention of pentadactylism and erectness in posture, while intellect and lively
success in complex environmental situations are recognizable higher cognitive skills. The primate family is illustrated
in Fig. 1. The hominoidea superfamily, which represents the
highest order in the primate family, includes gibbons and
orangutans as well as gorillas, chimpanzees and humans
(Fig. 1). The majority of nonhuman primate research is performed in a limited number of species: the squirrel monkey,
the rhesus monkey, the macaque monkey and the baboon,
with such research being undertaken predominantly for vaccine work, pharmacological and toxological studies, and im-
Primate models of neonatal brain injury
397
Table 1 Animal Models Used in Common Neonatal and Adult Neurological Diseases
Human Disorder
Term infant with
hypoxic—
ischemic injury
Premature infant
with white
matter injury
Parkinsons disease
Ischemic stroke
Traumatic brain
injury
Species Used
Models
Rhesus monkey
Rat PN 10-12 days
Sheep 119-133 gestational
days
Rabbit PN 1 day (33 days
gestation)
Dog > 2 weeks postnatal age
Piglet Term gestation
Premature baboon
Rat PN 3-7 days
Sheep 93-96 days (65% term
equivalent)
Rabbit 20-28 days gestation
(70-85% term equivalent)
Non-human primates, dog,
cat, minipig, mouse, guinea
pig, rat
Non-human primates, dog,
cat, rabbit, gerbil, guinea
pig, rat, mouse
Non-human primates, sheep,
pig, dog, rabbit, rat, mouse
Hypoxic–ischemia
with vascular
occlusion ⴙ
hypoxia
1. Paucity of functional correlative
studies
2. Models often severe
3. High utility in mechanism of injury
1. In-utero or perinatal
infection/endotoxin
2. Excititoxic injury
(ibotenate)
3. Chronic hypoxia
4. Hypoxic-ischemic
injury vascular
occlusion
1. Systemic MPTP
2. Intracerebral 6OHDA (rat)
1. Vascular occlusion
by ligation,
electrocautery
2. Injection of clotforming agents
3. Intracerebral
endothelium-1
1. Fluid-percussion
injury
2. Controlled impact
3. Inertial impact
(primates)
1. Paucity of functional correlative
studies.
2. Models often produce cystic PVL
only.
3. Focus on single mechanism allows
focused pathophysiology
munology. Less than 10% of studies in nonhuman primates
occur for neuroscience.4
Animal Models for Neurological
Injury in the Newborn
Before investigating the role of any animal species in the
study of neurological diseases in the newborn brain, it is
important to answer several fundamental questions:
Primates
Prosimii
Lemurs
Anthropoidea
Tarsioidea
Ayes Ayes
Tarsiers
Old World
Monkeys
New World
Monkeys
Callithricidae
Marmosets
Tamarins
Cebidae
Squirrel monkeys
Spider monkeys
Cercopithecinae
Baboons
Macaques
Figure 1 The taxonomy of the primate family.
Colobinae
Langurs
Probiscis
Comments
Homonidae
Apes
Humans
High similarity in clinical features, initial
drug trials in rats, pig, cat and then
non-human primate
1. Similarity in lesion and functional
outcome in species
2. Marked discrepancy in results from
neuroprotective therapies between
species
1. Variation in species in mechanism
sensitivity and functional outcomes
1. What is the normal ontogeny of this species, and is the
model developmentally appropriate for the neonatal
brain injury research paradigm?
2. Are there similarities in biochemical response, physiology and neuropathology that accurately mimic the human condition?
3. Are there functional correlates in chronic outcomes,
relevant to human premature and term born infants,
that support the relevance of the model and the utility
of therapeutic neuroprotective interventions?
Ontogeny of the
Nonhuman Primate Brain
Fetal studies of the macaque and baboon have demonstrated
that they share the same multivillous placental blood flow
pattern shown by the human and are highly relevant to studies of altered placental function.5,6 There are, however, differences between the species in the patterns of amniotic fluid
composition and in fetal breathing movements with the baboon most closely resembling the human fetus.
We have recently assessed the ontogeny of cerebral development over the last third of gestation in the baboon species
to establish a background for studies of the effects of prema-
T. Inder et al.
398
A
B
C
Figure 2 The gross anatomical appearances of the premature baboon
brain on gestational controls at 160 days gestation (A), 140 days
gestation (B) and 125 days gestation (C). Note the increase in gyral
formation occurring maximally in the occipital regions and progressing to the frontal regions by 160 days gestation.
ture delivery in the baboon on brain structure. We defined
the normal ontogeny of cortical development, including gyral
formation and white matter development. The premature
baboons (Papio sp) studied were delivered by hysterotomy
under general anesthesia at 125 (n ⫽ 3), 140 (n ⫽ 5) and 160
(n ⫽ 2) days gestation (dg). They were euthanased immediately with sodium pentobarbitone (130 mg/kg IV) and the
brains removed and immersion fixed. Magnetic resonance
imaging and histopathological analysis was done on the brain
tissue. Term delivery is considered 184 days gestation in this
model.
During the developmental period from 125 to 160 dg, the
fetal baboon brain undergoes a period of considerable
growth, with a 42% increase in whole brain weight between
125 and 140 dg (125 dg: 38.8 ⫾ 1.3 g; 149 dg: 55.3 ⫾ 2.4 g)
followed by a further 16% increase in weight between 140
and 160 days (160 dg: 64.1 ⫾ 3.5 g). At 125 dg, the main
sulci are present, including the lateral, central, lunatus, calcarine, superior, temporal and frontal (Fig. 2C); the inferior
temporal and inferior frontal sulci are present but rudimentary. Between 125 and 140 dg (Fig. 2B) these primary indentations deepen, gyri become more prominent, and secondary
gyri are seen in the orbital, frontal, insular and occipital regions. Gyral formation further progresses by 160 dg, with
tertiary gyri now evident (Fig. 2A). There is no obvious asymmetry between hemispheres in gyral development.
At 125 dg, all six cortical layers are present and there is a
significant volume of deep white matter. Cortical thickness
does not increase significantly (P ⫽ 0.43) between 125 dg
(1.49 ⫾ 0.03 mm) and 140 dg (1.54 ⫾ 0.03 mm), but does
increase by 20% (P ⬍ 0.001) between 140 and 160 dg (1.84
⫾ 0.04 mm). There is a dramatic increase of 73% in gyral
formation between 125 and 140 dg, followed by a slower
increase of 30% from 140 to 160 dg. Myelination in the
baboon forebrain was examined at the level of the central
sulcus and 5 mm rostral and caudal to this point. Myelination
was evident in the thalamus and internal capsule at 125 dg.
By 140 dg, myelination it had increased in these regions and
was also present in the alveus of the hippocampus, posterior
limb of the internal capsule and the white matter of the preand postcentral gyri. At 160 dg, myelination has increased in
the internal capsule, alveus, and lateral geniculate nucleus.
Subcortical myelination further proceeds in a dorsomedial to
lateral direction, with staining apparent in the gyri flanking
the lateral fissure, including the superior temporal gyrus.
The extent of hemispheric gyral folding, cortical growth,
astrocyte development and myelination can be used to establish the gestational ages at which equivalent brain development occurs in the baboon and human.7-9 At 125 dg in the
baboon, all layers are present in the cerebral cortex, the main
sulci and gyri have developed, and myelination has commenced in the internal capsule and diencephalon. In humans, the six cortical layers are established by at least 28
weeks10 and the primary gyri by 26 to 28 weeks.11 Taken
together, this suggests that development of the baboon brain
at 0.7 of gestation (125 dg) equates to human development at
26 to 28 weeks (182-196 dg). In baboons, there is a brain
growth spurt from 125 to 140 dg, followed by a slower rate of
growth from 140 to 160 dg. However, cortical thickness increases more significantly during the latter time period. By
140 dg, myelin is present in the posterior limb of the internal
capsule and extends into the dorsal gyri of the frontal and
parietal lobes in the region around the central sulcus. Fetal
human brains display a growth spurt around 28 weeks, and
there is prominent growth in cortical gray matter between 29
and 40 weeks.12 Myelinogenesis begins in the periventricular
white matter at around 30 weeks in the human13 although
mature myelin is not evident in the forebrain until about 44
to 52 weeks postconception,14,15 appearing first in the posterior limb of the internal capsule at about 44 weeks, in the
temporal gyrus at about 48 weeks and the optic radiation at
52 weeks. Taken together, this implies that the baboon brain
at 140 dg is equivalent to 32 to 34 weeks, and 160 dg to term,
in humans. Overall, there is a marked similarity in the sequence of development, albeit with a somewhat more advanced pattern of myelination in the baboon. Thus, as has
been previously noted, the “term” nonhuman primate baboon (184 dg) is much more advanced in cerebral structural
development and may differ in neuropathology and physiology in comparison to the “term” human infant. The baboon
would be more appropriately studied at 0.8 to 0.85 of full
term, or 160 dg, for comparison with term human infants. An
accurate understanding of the appropriate development neurobiology of the period of cerebral development is clearly
crucial to selecting the correct model for investigation, but is
often sadly lacking in full definition in many animal species
studied for neonatal neuropathology. Of note, many aspects
of cerebral development have been studied in rhesus and
macaque monkeys.7-9
Are There Similarities
in Nonhuman Primate
Models to the Human
Infant with Term Perinatal
Hypoxic–Ischemic Insult?
The major cause of neurological morbidity in the term-born
human infant is hypoxic–ischemic brain injury. The se-
Primate models of neonatal brain injury
quence of pathophysiological events leading to neonatal hypoxic–ischemic brain injury is ultimately dominated by the
occurrence of global cerebral ischemia. The neuropathological features of human neonatal hypoxic–ischemic encephalopathy are determined by factors such as the severity, pattern and type of insult, as well as the gestational age and
metabolic status, including temperature, of the human infant.
The most common variety of neuronal injury resulting
from hypoxic-ischemic injury in the term infant is selective
neuronal necrosis. Neurons demonstrate the greatest sensitivity to hypoxic-ischemic injury in the term infant. Among
term born human infants, neurons of the CA1 region of the
hippocampus (Sommer’s sector), deeper layers of the cerebral cortex, and cerebellar Purkinje cells are injured most
frequently by hypoxic–ischemic insult.16-18 Neuronal necrosis is most prominent in the watershed regions of the cerebral
cortex and in the depth of the sulci, reflecting the greater
effect of ischemia in these regions. Parasagittal cerebral injury
is the principal ischemic lesion of the full term infant with
injury to the cerebral cortex and subcortical white matter in
the parasagittal and superomedial aspects of the cerebral convexities. The injury is bilateral and usually symmetrical. Status mamoratus describes a pattern of injury involving the
basal ganglia and thalamus, characterized initially by neuronal loss, gliosis and hypermyelination. Other patterns of hypoxic–ischemic injury consist of focal and multifocal ischemic brain necrosis, including infarction, porencephaly,
hydranencephaly and multicystic encephalomalacia.17
Nonhuman Primate Studies of
Term Hypoxic–Ischemic Injury
Studies defining the perinatal patterns of brain injury occurring during experimental asphyxia in the nonhuman primate
with associated cardiovascular and blood gas alterations have
been conducted exclusively by Myers and Brann.19-23 Four
patterns of perinatal brain damage resembling that of asphyxial insults to the human term infant have been described.21
The first pattern of injury described in the rhesus monkey
by Myers is that of “total asphyxia”—a pattern of brainstem
injury noted when respiratory gas exchange of the fetus is
completely stopped. The brain structure that is most vulnerable is the inferior colliculus, with damage to the superior
olives, Purkinje cells, vestibular nuclei, posterior and lateral
ventral nuclei of the thalamus, and gracile/cuneate nuclei
appearing sequentially with increasing duration of asphyxia.
Of interest in this model is that the duration of asphyxia
insult necessary to produce microscopic brain injury increased dramatically with decreasing gestational age, ie, 12
minutes at term compared with 30 minutes at midgestation.
The second pattern of injury is that of “partial asphyxia
with acidosis,” for which the monkey fetus is subjected to
partial asphyxia by impairment of placental gas exchange in a
chronic fashion. In eight Rhesus monkeys, following prolonged partial insult, there is clinical behavior similar to the
term asphyxiated newborn infant, with a prolonged time to
399
first gasp and clinical seizure activity.20 The average time of
first seizure is 23 hours of age. Neuropathology reveals pale
or hemorrhagic necrosis involving the entire cortex of both
hemispheres or restricted to a parasagittal posterior parietal
distribution. Animals subjected to partial asphyxia of shorter
duration tend to survive and later show focal cortical injury
with predominance in the posterior parietal parasagittal regions. Rarely these animals survive beyond the newborn period. With later survival there is symmetrical bilateral porencephaly or cerebral necrosis with cystic degeneration of gray
and white matter, centered in the middle paracentral region.
The third pattern of injury described is that of “partial
asphyxia without acidosis,” which is associated with selective
white-matter injury. This pattern of injury appears when respiratory gas exchange of the fetus is diminished gradually
and maintained for long periods of time. In contrast to the
low oxygen levels, pH and pCO2 levels were virtually unaffected in this model. The neuropathological findings are similar to those noted in juvenile monkeys exposed to cyanide or
carbon monoxide. Both processes are known to impair cerebral oxidative metabolism without altering CO2 or pH.
The final pattern described is that of “partial asphyxia in
association with total asphyxia.” In this model, partial asphyxia in association with total asphyxia leads to selective
basal ganglia injury. Necrosis and later gliosis affect the caudate, putamen, and globus pallidus, with less conspicuous
lesions of the neocortex.21 These pathological findings of status marmoratus bear a striking similarity to human neuropathological lesions in neonatal hypoxic–ischemic encephalopathy. The relative degrees of partial and total asphyxia are
reflected in varying degrees of involvement of cerebral cortex.
Animals subjected to severe partial asphyxia and relatively
short episodes of total asphyxia show lesions of both basal
ganglia and cerebral cortex, whereas those subjected to more
severe episodes of total asphyxia show more striking brainstem abnormalities.
Correlations with
the Human Model
The parasagittal distribution of lesions described by Brann
and Myers20 with partial asphyxia correlate with the parasagittal distribution of cerebral injury recognized in the human
infant by both technetium brain scan24 and positron– omission tomography.25 There is some variation in the functional
clinical correlate to the parasagittal lesions between the human and nonhuman primate models. In the human infant,
the clinical correlation is of a greater degree of weakness of
the proximal limbs, upper greater than lower extremities. In
contrast, long-term evaluation of the monkey revealed spastic paraparesis. Hypotonia was also noted in the monkeys, as
recognized in human infants who had manifest prolonged
periods of abnormal fetal heart rate patterns.26 The white
matter lesions with minimal gliosis are well described in premature human neonates and correlate with the midgestational rhesus model of oligoacidotic hypoxemia.27 Human
neonates with apparent acute total asphyxia and brain stem
T. Inder et al.
400
injury have also been reported. Thus, the clinical and pathological findings described by Myers and coworkers are remarkably similar to those that have been noted in the human
infant, rendering this a model of potential high utility, particularly for neuroprotective drug trials for birth asphyxia
before human trial.
Are There Similarities
in Nonhuman Primate
Models to the Human Infant
with Preterm Cerebral Injury?
Very low birth weight (VLBW) infants now represent approximately 1.5% of all live-born infants, increasing slightly as a
proportion of total births over the last 3 decades.28,29 More
notably, the survival of VLBW infants has increased to close
to 90% with recent improvements in obstetric and neonatal
management.29-31 This increased survival has been greatest in
the extremely low birth weight (ELBW) cohort of infants who
weigh less than 1000 g and are at greatest risk of later neurological deficits.30-32 With this improved survival, there has
been no reduction in the documented rates of neurological
disability occurring in premature infants, with a recognized
increase in the absolute numbers of infants with long term
neurological sequelae.33 For all VLBW survivors, 5 to 15%
later exhibit major spastic motor deficits, while an additional
25-50% exhibit less prominent developmental disabilities,
involving not only motility but also cognition and behavior,
with school disturbance a nearly uniform result.33-38 Improvement in the detection, diagnosis and management of
neurological injuries and, ultimately, formulation of strategies to prevent the accompanying morbidity require an understanding of the pathogenesis of glial and neuronal injury
in the brain of the premature infant.
The principal neuropathological substrates for the neurological disturbances in the human premature infant appear to
involve the cerebral white matter. These lesions include
periventricular hemorrhagic infarction and periventricular
leukomalacia (PVL). The most common neuropathology
found at autopsy in the premature infant is PVL. PVL refers to
necrosis of white matter in a characteristic distribution, ie,
dorsal and lateral to the external angles of the lateral ventricles, involving particularly the centrum semiovale (frontal
horn and body), optic radiation (trigone and occipital horn),
and acoustic radiation (temporal horn).17 The incidence of
the disorder at autopsy varies considerably from one medical
center to another, ie, from approximately 25% to 75%. Despite this variation, several facts remain clear: the lesion is
observed particularly often in infants who (1) are prematurely born (⬍30 weeks gestation), (2) survive more than a
few days, (3) have intraventricular hemorrhage (IVH), (4)
have evidence of cardiorespiratory disturbance, and (5) have
evidence of antenatal/placental/fetal infection.17,39-51 The
clinical consequences of PVL are a significant increase in the
risk of cerebral palsy. In a meta-analysis, it was found that
58% of 272 infants with cystic PVL developed cerebral palsy,
compared with 2.6% of 655 infants with normal head ultra-
sound scans.52 The etiology of white matter injury is not
known, but clinical data suggest that ischemia–reperfusion
and/or infection–inflammation are important factors.53
Other major forms of neuropathology in the human premature infant, less common than PVL, include IVH, selective
neuronal injury, and focal cerebral ischemic lesions. IVH, in
addition to being associated with an increased risk of
PVL,17,54-56 is also intimately associated with the occurrence
of periventricular hemorrhagic infarction. Apart from these
major forms of neuropathology, it is also likely that the extremely immature infant is at risk for more subtle alterations
in cerebral structure and development. Our research has described alterations in cortical gray matter development in
premature infants with PVL57,58 and in premature infants
receiving dexamethasone.59 These more subtle alterations
may include a reduction in cortical gyral development, impaired or delayed cerebral myelination, impairment in cortical laminar organization, and/or a reduction in axonal and
dendritic growth and synaptogenesis. Such subtle abnormalities have not been extensively examined on previous human
neuropathology studies, but MR studies of premature infants
are increasingly suggesting that such alterations in cerebral
structure are common, particularly in the extremely immature infant.60-63
Nonhuman Primate
Studies of Cerebral
Injury in the Premature Infant
We have evaluated the premature nonhuman primate as a
model of cerebral injury in the immature brain. Employing a
model of premature delivery of the baboon, we have attempted to determine the nature and severity of the cerebral
injury resulting from premature delivery followed by intensive care similar to that received by the human premature
infant. In this regard, the experimental conditions for this
baboon model are nearly identical to those experienced by
premature human infants. The respiratory and cardiovascular development of premature baboons64,65 suggests that
125 dg corresponds to 26 to 28 weeks in humans. Our pilot
work on the ontogeny of cerebral development also confirm
that 125 dg is equivalent to 26 to 28 weeks human gestation
(vide supra). Thus, the experimental cohort of baboons we
have studied was delivered at 125 dg and subsequently
nursed for two weeks in an intensive care facility with euthanasia at 139 to 140 dg. The neuropathological examination
included assessment of white matter damage, neuronal necrosis, presence of reactive astrogliosis or activated microglia,
ventricular size, and the presence of subarachnoid, germinal
matrix or intraventricular hemorrhage. The pattern of cerebral injury correlates with that seen in the preterm human
infant, validating the prematurely born baboon as a highly
appropriate model for the study of preterm human birth.
Preterm Baboon Model
Infants are delivered by elective hysterotomy under general anesthesia at 125 ⫾ 2 days gestation. The infants were weighed,
Primate models of neonatal brain injury
A
401
B
Figure 3 The presence of astrogliosis in the periventricular white
matter in the premature baboon delivered at 125 days and receiving
neonatal intensive care for 14 days demonstrated by glial fibrillary
acidic staining (A, B). Note also the presence of micro-hemorrhage
in the white matter (B).
anesthetized with intramuscular ketamine (5-10 mg/kg) and
intravenous valium (0.1-0.5 mg/kg), and intubated with a 2.0 to
2.5 mm diameter endotracheal tube. All animals received surfactant replacement therapy (Survanta or Curosurf) and were
ventilated on a standard, time-cycled, pressure-regulated infant
ventilator (Infant Star) with humidifier maintained at 36 to
37°C. Hypotension was managed with volume replacement,
dobutamine, dopamine and/or epinephrine infusions as
needed.65 For all animals, brains were removed at postmortem after euthanasia at 139 dg and placed immediately
into 10% formalin. Sections were stained for hematoxylin
and eosin (H&E), glial fibrillary acidic protein (GFAP)
immunohistochemistry, and lectin histochemistry.
Histological Findings in the
Premature Baboon Model
Histological examination revealed various neuropathologies,
the two most common of which were white matter injury and
hemorrhage.
Evidence of white matter injury was found in 50% of the
prematurely delivered animals. The injury ranged from small
patches of reactive astrocytosis (Fig. 3) to more extensive
damage with activated microglia, small cystic lesions with
dense cellular borders, and endothelial hypertrophy. The extent of white matter injury ranged from approximately 0.52.5% of total white matter. White matter injury occurred
most frequently in the parietal and occipital lobes. There was
often ventriculomegaly present in those animals with significant white matter injury. One quarter of the prematurely
delivered neonates displayed enlarged lateral ventricles
(Fig. 4) in relation to the surrounding brain tissue when
compared with gestational control brains. In particular, the
posterior and inferior horns were markedly enlarged.
Subarachnoid hemorrhages were observed in 40% of the
prematurely delivered neonates. Hemorrhages also occurred
in the lateral ventricles (5%), germinal matrix (9%), white
matter (28%) and cerebellum (9%). Distinct regions of cell
loss were seen in the CA2/3 region of the pyramidal cell layer
in the hippocampus in some animals, and reactive astrogliosis was observed in association with this cell loss.
Correlations with
the Human Model
Neonatal white matter injury, the most common form of
damage in the baboon, is also the most common form of
injury in human infants. In baboon, this damage is usually
the diffuse form, with cystic infarction occurring rarely. Diffuse white matter injury is associated with the attraction of
infiltrating cells, although the injury is not as extensive as that
reported in humans, which may be due to the absence of
potentiating or priming factors such as perinatal infection or
hypoxia.
The frequency of intraventricular and subarachnoid hemorrhage also closely correlates with human data.17,30 Hippocampal cell loss occurs in about 20% of the premature
baboons and may be directly relevant to the recent observations of hippocampal atrophy in older, prematurely born
children.66
Overall, there are marked similarities in the pattern of
cerebral injury between prematurely delivered baboons and
premature human infants. Most notably, the premature baboon displays a marked predominance of cerebral white matter injury in the absence of an experimental interventional
insult. There is also striking similarity in the distribution and
frequency of cerebral injury to that seen in the prematurely
born human infant (Table 2). Thus, this model has the potential to shed light on the nature of cerebral injury in the
prematurely delivered human. Clearly there are important
differences between this model and the human setting, such
as the presence of potentiating or priming factors precipitating preterm labor, including perinatal fetal infection, although these may be possible to replicate in the model in the
future. The functional correlates of the neuropathologies in
the prematurely born baboon are unknown, but clearly worthy of study.
A
B
Figure 4 Ventricular size was enlarged in 25% of experimental animals as compared here in two experimental animals without (A) and
with (B) ventriculomegaly. One can also note the relative white
matter atrophy in the animal displaying significant ventriculomegaly.
T. Inder et al.
402
Table 2 The Neuropathologic Findings in the Premature Baboon (125 days surviving with neonatal intensive care 14 days) in
Comparison to Those Found in the Human Premature Infant
Commonest Form of Injury
Regional of Cerebral
Injury
Periventricular leukomalacia
(white matter injury)
Hemorrhage
Hippocampal injury
Cortical and deep nuclear
gray matter injury
Premature Human Infant
Cystic PVL (5%)
Diffuse PVL (20%)
Moderate ventriculomegaly (40-50%)
Intraventricular hemorrhage 20%
(Vermont–Oxford)
Subarachnoid haemorrhage, isolated
30%
Recent MR studies highlight
significant hippocampal atrophy in
older prematurely born children
and the subiculum of the
hippocampus vulnerable to injury
in premature infants
Injury to the thalamus, globus
pallidus. Volumetric MR studies
reduction in cortical gray matter
volumes in preterm infants by
term and later
Disadvantages of the
Nonhuman Primate Models
It is important to point out the disadvantages in the use of
these models, which must be balanced against their advantages. They include:
1. The requirement for a facility specifically staffed and
equipped for nonhuman primates
2. The significant cost of the animals and their care in full
neonatal intensive care setting
3. Ethical considerations in the use of a higher order primate species
4. The need for professional individuals with high surgical, medical, and laboratory skills with nonhuman
primates.
One means of reducing cost is to perform studies of multiple organ systems in a single experimental setting. For example, the baboon data described above were derived from
brains removed from animals that underwent various ventilation protocols for optimization of mechanical ventilation
strategies. Thus, the same model can be used to evaluate
brain injury and ventilation schemes. Another approach is to
apply these models in a selective fashion. For example, therapies and interventions could be screened in lower species.
Those that prove successful would be evaluated in nonhuman primate models before their application to human studies.
Conclusion
Nonhuman primate models of term and preterm brain injury
have a remarkably high similarity to human brain injury in
Premature Baboon
(125 dg treated for 14 days)
Cystic PVL (5%)
Diffuse PVL (50%)
Moderate ventriculomegaly (25%)
Intraventricular hemorrhage (6%)
Subarachnoid hemorrhage, isolated (40%)
Hippocampal cellular injury and reactive
astrocytosis (20%)
Gray matter injury in gray matter (25%)
and basal ganglia (6%)
Further subtle cortical laminar study
under investigation
etiological mechanism, neuropathological lesion and clinical
outcome. Though their implementation is resource intensive,
their selective use for evaluation of brain injury and protective strategies is well justified.
References
1. Iwaniuk AN, Whishaw IQ: On the origin of skilled forelimb movements. Trends Neurosci 23:372-376, 2000
2. Cenci MA, Whishaw IQ, Schallert T: Animal models of neurological
deficit: How relevant is the rat? Nature Neurosci 3:574-579, 2002
3. Redgrave P, Prescott TJ, Gurney K: The basal ganglia: A vertebrate
solution to the selection problem? Neuroscience 89:1009-1023, 1999
4. Meldrum BS: Future problems and possibilities. Adv Neurol 10:349356, 1975
5. Martin CB, Murata U: Perspective of the fetus. The laboratory primate
as a model for fetal physiology and disease, in Brans YW, Kuehl TJ
(eds): Non-Human Primates in Perinatal Research. New York, NY,
Wiley & Sons, 1978
6. Painter MJ: Animal models of perinatal asphyxia: Contributions, contradictions, clinical relevance. Semin Ped Neurol 2:37-56, 1995
7. Kostovic I, Rakic P: Developmental history of the transient subplate
zone in the visual and somatosensory cortex of the macaque monkey
and human brain. J Comp Neurol 297:441-470, 1990
8. Eckenhoff MF, Rakic P: Radial organization of the hippocampal dentate
gyrus: A Golgi, ultrastructural, and immunocytochemical analysis in
the developing rhesus monkey. J Comp Neurol 223:1-21, 1984
9. Levitt P, Rakic P: Immunoperoxidase localization of glial fibrillary
acidic protein in radial glial cells and astrocytes of the developing rhesus monkey brain. J Comp Neurol 193:815-840, 1980
10. Marin-Padilla M: Prenatal and early postnatal ontogenesis of the human
motor cortex: A golgi study. I. The sequential development of the
cortical layers. Brain Res 23:167-183, 1970
11. Chi JG, Dooling EC, Gilles FH: Gyral development of the human brain.
Ann Neurol 1:86-93, 1977
12. Huppi PS, Warfield S, Kikinis R, et al: Quantitative magnetic resonance
imaging of brain development in premature and mature newborns.
Ann Neurol 43:224-235, 1998
13. Back SA, Luo NL, Borenstein NS, et al: Arrested oligodendrocyte lin-
Primate models of neonatal brain injury
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
33.
34.
35.
36.
37.
38.
eage progression during human cerebral white matter development:
dissociation between the timing of progenitor differentiation and
myelinogenesis. J Neuropathol Exp Neurol 61:197-211, 2002
Brody BA, Kinney HC, Kloman AS, et al: Sequence of central nervous
system myelination in human infancy. I. An autopsy study of myelination. J Neuropathol Exp Neurol 46:283-301, 1987
Kinney HC, Brody BA, Kloman AS, et al: Sequence of central nervous
system myelination in human infancy. II. Patterns of myelination in
autopsied infants. J Neuropathol Exp Neurol 47:217-234, 1988
Larroche JC: Developmental Pathology of the Neonate. New York, NY,
Excerpta Medica, 1977
Volpe JJ: Neurology of the Newborn (ed 4). Philadelphia, PA, W.B.
Saunders, 2001
Rivkin MJ: Hypoxic-ischemic brain injury in the term newborn. Neuropathology, clinical aspects, and neuroimaging. Clin Perinatol 24:
607-625, 1997
Myers RA: Atrophic cortical sclerosis associated with status mamoratus
in a perinatally damaged monkey. Neurology 19:1177-1188, 1969
Brann AW, Myers RE: Central nervous system findings in newborn
monkey following severe in utero partial asphyxia. Neurology 25:327338, 1975
Myers RE: Experimental models of perinatal brain damage: Relevance
to human pathology, in Gluck L (ed): Intrauterine Asphyxia and the
Developing Fetal Brain. Chicago, IL, Year Book, 1977, pp 37-97
Myers RE: Two patterns of perinatal brain damage and their conditions
of occurrence. Am J Obstet Gynecol 112:246-262, 1971
Adamsons K, Mueller-Heubach E, Myers RE: Production of fetal asphyxia in the rhesus monkey by administration of catecholamines to
the mother. Am J Obstet Gynecol 109:248-262, 1971
Volpe JJ, Pasternak JF: Parasagittal cerebral injury in neonatal hypoxicischemic encephalopathy: Clinical and neuroradiologic features. J Pediatr 91:472-476, 1977
Volpe JJ, Herscovitch P, Perlman JM, et al: Positron emission tomography in the asphyxiated term newborn: Parasagittal impairment of cerebral blood flow. Ann Neurol 17:287-296, 1985
Painter MJ, Dyrp R, O’Donoghue P: Fetal heart rate patterns and development in the first year of life. Am J Obstet Gynecol 132:271, 1978
Barkovich AJ, Truvit C: Brain damage from perinatal asphyxia: Revelation of MR findings with gestational age. AJNR Am J Neuroradiol 11:
1087-1096, 1990
Volpe JJ: Brain injury in the premature infant. Neuropathology, clinical
aspects, pathogenesis, and prevention. Clin Perinatol 24:567-587,
1997
Hack M, Friedman H, Fanaroff AA: Outcomes of extremely low birth
weight infants. Pediatrics 98:931-937, 1996
Horbar JD: Vermont-Oxford Network 1997 Database Summary. Burlington, VT, Vermont–Oxford Network, 1997
Network, A.A.N.Z.N., Australian and New Zealand Neonatal Network
Series number 2. Australian Institute of Health and Welfare Perinatal
Statistics Unit: Sydney, 1995
Improved outcome into the 1990s for infants weighing 500-999 g at
birth. The Victorian Infant Collaborative Study Group. Arch Dis Child
Fetal Neonatal Ed 77:F91-F94, 1997
Pharoah PO, Platt MJ, Cooke T: The changing epidemiology of cerebral
palsy. Arch Dis Child Fetal Neonatal Ed 75:F169-F173, 1996
Pharoah PO, Cooke T, Cooke RW, et al: Birthweight specific trends in
cerebral palsy. Arch Dis Child 65:602-606, 1990
Piecuch RE, Leonard CH, Cooper BA, et al: Outcome of extremely low
birth weight infants (500 to 999 grams) over a 12-year period. Pediatrics 100:633-639, 1997
Powls A, Botting N, Cooke RW, et al: Visual impairment in very low
birthweight children. Arch Dis Child Fetal Neonatal Ed 76:82-87, 1997
Skranes JS, Vik T, Nilsen G, et al: Cerebral magnetic resonance imaging
and mental and motor function of very low birth weight children at six
years of age. Neuropediatrics 28:149-154, 1997
Singer L, Yamashita T, Lilien L, et al: A longitudinal study of developmental outcome of infants with bronchopulmonary dysplasia and very
low birth weight. Pediatrics 100:987-993, 1997
403
39. Banker BQ, Larroche JC: Periventricular leukomalacia of infancy. Arch
Neurol 7:386-410, 1962
40. Back SA, Volpe JJ: Cellular and molecular pathogenesis of periventricular white matter injury. MRDD Res Rev 3:96-107, 1997
41. Barth PG, Stam FC, Oosterkamp RF, et al: On the relationship between
germinal layer haemorrhage and telencephalic leucoencephalopathy in
the preterm infant. Neuropediatrie 11:17-26, 1980
42. Carson SC, Hertzberg BS, Bowie JD, et al: Value of sonography in the
diagnosis of intracranial hemorrhage and periventricular leukomalacia:
A postmortem study of 35 cases. Am J Neuroradiol 155:595-601, 1990
43. Dammann O, Leviton A: Maternal intrauterine infection, cytokines,
and brain damage in the preterm newborn. Pediatr Res 42:1-8, 1997
44. de Vries LS, Eken P, Dubowitz LMS: The spectrum of leukomalacia
using cranial ultrasound. Behav Brain Res 49:1-6, 1992
45. Iida KS, Takashima S, Ueda K: Immunohistochemical study of myelination and oligodendrocyte in infants with periventricular leukomalacia. Pediatr Neurol 13:296-304, 1995
46. Leviton A, Gilles FH: Acquired perinatal leukoencephalopathy. Ann
Neurol 16:1-10, 1984
47. Paneth N, Rudelli R, Monte W, et al: White matter necrosis in very low
birth weight infants: Neuropathologic and ultrasonographic findings in
infants surviving six days or longer. J Pediatr 116:975-984, 1990
48. Perlman JM, Risser R, Broyles RS: Bilateral cystic periventricular leukomalacia in the premature infant: Associated risk factors. Pediatrics
97:822-827, 1996
49. Yoon BH, Romero R, Yang SH, et al: Interleukin-6 concentrations in
umbilical cord plasma are elevated in neonates with white matter lesions associated with periventricular leukomalacia. Am J Obstet Gynecol 174:1433-1440, 1996
50. Yoon BH, Romero R, Kim CJ, et al: High expression of tumor necrosis
factor-alpha and interleukin-6 in periventricular leukomalacia. Am J
Obstet Gynecol 177:406-411, 1997
51. Zupan V, Gonzalez P, Lacaze-Masmonteil T, et al: Periventricular leukomalacia: Risk factors revisited. Dev Med Child Neurol 38:10611067, 1996
52. Holling EE, Leviton A: Characteristics of cranial ultrasound white matter echolucencies that predict disability: A review. Dev Med Child
Neurol 41:136-139, 1998
53. Inder T, Volpe JJ: Mechanisms of perinatal brain injury. Semin Neonatol 5:3-16, 2000
54. Armstrong DL, Sauls CD, Goddard-Finegold J: Neuropathologic findings in short-term survivors of intraventricular hemorrhage. Am J Dis
Child 141:617-621, 1987
55. Takashima S, Mito T, Houdou S, et al: Relationship between periventricular hemorrhage, leukomalacia and brainstem lesions in prematurely born infants. Brain Dev 11:121-124, 1989
56. Leviton A, Gilles F: Ventriculomegaly, delayed myelination, white matter hypoplasia, and “periventricular” leukomalacia. How are they related? Pediatr Neurol 15:127-136, 1996
57. Inder TE, Huppi PS, Warfield S, et al: Periventricular white matter
injury in the premature infant is associated with a reduction in cerebral
cortical gray matter volume at term. Ann Neurol 46:755-760, 1999
58. Inder TE, Warfield SK, Wang HX, et al: Abnormal cerebral structure at
term in premature infants. Pediatrics 2004 (in press)
59. Murphy BP, Inder TE, Huppi PS, et al: Impaired cerebral cortical gray
matter growth after treatment with dexamethasone for neonatal
chronic lung disease. Pediatrics 107:217-221, 2001
60. Maalouf EF, Duggan PJ, Rutherford MA, et al: Magnetic resonance
imaging of the brain in a cohort of extremely preterm infants. J Pediatr
135:351-357, 1999
61. Childs AM, Cornette L, Ramenghi LA, et al: Magnetic resonance and
cranial ultrasound characteristics of periventricular white matter abnormalities in newborn infants. Clin Radiol 56:647-655, 2001
62. Valkama AM, Vainionpaa LK, Lanning FP, et al: Magnetic resonance
imaging at term and neuromotor outcome in preterm infants. Acta
Paediatr 89:348-355, 2000
63. Inder TE, Wells SJ, Mogridge NB, et al: Defining the nature of
404
the cerebral abnormalities in the premature infant: A qualitative
magnetic resonance imaging study. J Pediatr 143:171-179, 2003
64. Coalson JJ, Winter VT, Siler-Khodr T, et al: Neonatal chronic lung
disease in extremely immature baboons. Am J Respir Crit Care Med
160:1333-1346, 1999
T. Inder et al.
65. Yoder B, Martin H, McCurnin DC, et al: Impaired urinary cortisol
excretion and early cardiopulmonary dysfunction in immature baboons. Pediatr Res 51:426-432, 2002
66. Isaacs E, Lucas A, Chong WK, et al: Hippocampal volume and everyday
memory in children of low birthweight. Ped Res 47:713-720, 2000