Administration of thimerosal-containing vaccines to infant rhesus macaques does not result in autism-like behavior or neuropathology.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder. Some anecdotal reports suggest that ASD is related to exposure to ethyl mercury, in the form of the vaccine preservative, thimerosal, and/or receiving the measles, mumps, rubella (MMR) vaccine. Using infant rhesus macaques receiving thimerosal-containing vaccines (TCVs) following the recommended pediatric vaccine schedules from the 1990s and 2008, we examined behavior, and neuropathology in three brain regions found to exhibit neuropathology in postmortem ASD brains. No neuronal cellular or protein changes in the cerebellum, hippocampus, or amygdala were observed in animals following the 1990s or 2008 vaccine schedules. Analysis of social behavior in juvenile animals indicated that there were no significant differences in negative behaviors between animals in the control and experimental groups. These data indicate that administration of TCVs and/or the MMR vaccine to rhesus macaques does not result in neuropathological abnormalities, or aberrant behaviors, like those observed in ASD.

Examination of the safety of pediatric vaccine schedules in a non-human primate model: assessments of neurodevelopment, learning, and social behavior.

BACKGROUND: In the 1990s, the mercury-based preservative thimerosal was used in most pediatric vaccines. Although there are currently only two thimerosal-containing vaccines (TCVs) recommended for pediatric use, parental perceptions that vaccines pose safety concerns are affecting vaccination rates, particularly in light of the much expanded and more complex schedule in place today. OBJECTIVES: The objective of this study was to examine the safety of pediatric vaccine schedules in a non-human primate model. METHODS: We administered vaccines to six groups of infant male rhesus macaques (n = 12-16/group) using a standardized thimerosal dose where appropriate. Study groups included the recommended 1990s Pediatric vaccine schedule, an accelerated 1990s Primate schedule with or without the measles-mumps-rubella (MMR) vaccine, the MMR vaccine only, and the expanded 2008 schedule. We administered saline injections to age-matched control animals (n = 16). Infant development was assessed from birth to 12 months of age by examining the acquisition of neonatal reflexes, the development of object concept permanence (OCP), computerized tests of discrimination learning, and infant social behavior. Data were analyzed using analysis of variance, multilevel modeling, and survival analyses, where appropriate. RESULTS: We observed no group differences in the acquisition of OCP. During discrimination learning, animals receiving TCVs had improved performance on reversal testing, although some of these same animals showed poorer performance in subsequent learning-set testing. Analysis of social and nonsocial behaviors identified few instances of negative behaviors across the entire infancy period. Although some group differences in specific behaviors were reported at 2 months of age, by 12 months all infants, irrespective of vaccination status, had developed the typical repertoire of macaque behaviors. CONCLUSIONS: This comprehensive 5-year case-control study, which closely examined the effects of pediatric vaccines on early primate development, provided no consistent evidence of neurodevelopmental deficits or aberrant behavior in vaccinated animals.

Four decades of leading-edge research in the reproductive and developmental sciences: the Infant Primate Research Laboratory at the University of Washington National Primate Research Center.

The Infant Primate Research Laboratory (IPRL) was established in 1970 at the University of Washington as a visionary project of Dr. Gene (Jim) P. Sackett. Supported by a collaboration between the Washington National Primate Research Center and the Center on Human Development and Disability, the IPRL operates under the principle that learning more about the causes of abnormal development in macaque monkeys will provide important insights into the origins and treatment of childhood neurodevelopmental disabilities. Over the past 40 years, a broad range of research projects have been conducted at the IPRL. Some have described the expression of normative behaviors in nursery-reared macaques while others have focused on important biomedical themes in child health and development. This article details the unique scientific history of the IPRL and the contributions produced by research conducted in the laboratory. Past and present investigations have explored the topics of early rearing effects, low-birth-weight, prematurity, birth injury, epilepsy, prenatal neurotoxicant exposure, viral infection (pediatric HIV), diarrheal disease, vaccine safety, and assisted reproductive technologies. Data from these studies have helped advance our understanding of both risk and resiliency in primate development. New directions of research at the IPRL include the production of transgenic primate models using our embryonic stem cell-based technology to better understand and treat heritable forms of human intellectual disabilities such as fragile X.

Post-drug consequences of chronic atypical antipsychotic drug administration on the ability to adjust behavior based on feedback in young monkeys.

RATIONALE: Atypical antipsychotic drugs are characterized by their affinity for serotonin and dopamine receptors. The dopaminergic system undergoes developmental changes during childhood, making it vulnerable to external influences such as drug administration. OBJECTIVE: The purpose of this study was to assess the long-term effects of administering risperidone and quetiapine to 12-24-month-old macaque monkeys on cognitive development, a maturational equivalent to 4-8-year-old children. METHODS: Forty male pigtailed macaques were used in the study (n = 20 placebo, n = 10 risperidone, n = 10 quetiapine). Following a 4-month pre-drug period, animals were orally administered 2 mg/kg of quetiapine and .025 mg/kg of risperidone daily for 4 months, then the dosage was doubled for another 4 months. They were followed up for 4 months after cessation of the drug. Animals were assessed through all phases of the study on two-object discrimination and learning set. RESULTS: Cognitive development was not negatively affected while the animals were being administered the drug. However, the risperidone group had significant decrements in performance on the learning set task after cessation of the drug (p = 0.006, η (p) (2) = 0.59). Analysis of errors showed that the risperidone group had a significant increase in perseverative responding during the post-drug phase (p = 0.002, η (p) (2) = 0.67). CONCLUSION: As with human studies, neither risperidone nor quetiapine had a negative impact on cognitive development during the drug phases. However, the results show that the risperidone group had behavioral impairment post-drug, suggesting that the drug may have impacted the development of the dopaminergic system.

Some effects of risperidone and quetiapine on growth parameters and hormone levels in young pigtail macaques.

OBJECTIVE: Atypical antipsychotic drugs are prescribed to young children for a number of symptoms, some with no Food and Drug Administration approval for children. Effects on growth of children have received little experimental study. We assessed the effects of two atypicals, risperidone and quetiapine, on growth, prolactin, and thyroid hormones of young pigtail macaques (macaca nemestrina), modeling potential effects on 4-8-year-old children. METHODS: Subjects were studied blindly after random assignment to risperidone (N = 10), quetiapine (N = 10), or placebo (N = 20). Four phases were studied: (1) predrug, 9-12 months of age; (2) low dose (risperidone 0.025 mg/kg, quetiapine 2 mg/kg), 13-16 months; (3) high dose (risperidone 0.05 mg/kg, quetiapine 4 mg/kg), 17-20 months; (4) postdrug, 21-24 months. Body weight was measured daily, skeletal dimension monthly, and bone mineralization and hormones bimonthly. RESULTS: Our primary result showed that, compared with placebo, neither drug had detrimental effects on body weight, skeletal dimensions, or thyroid hormones. However, in a transient effect, bone density was lower following low-dose risperidone than either quetiapine or placebo. In both drug phases, risperidone prolactin was higher than the other groups, which did not differ. The higher prolactin of the risperidone group may partially explain the bone density effect. CONCLUSION: This 16-month study of young, developing pigtail macaques given risperidone at doses from 0.025 to 0.05 mg/kg or quetiapine at doses from 2 to 4 mg/kg suggests that these drugs are safe for normal body weight and skeletal growth in young pigtail macaques given an adequate diet, although these drugs are known to cause significant weight gain and other metabolic changes in some children, adolescents, and adult humans. In addition, the results, although transient in our study, also suggest that research in children on bone mineralization effects of risperidone, and possibly other antipsychotic drugs, may be warranted.

Delayed acquisition of neonatal reflexes in newborn primates receiving a thimerosal-containing hepatitis B vaccine: influence of gestational age and birth weight.

This study examined whether acquisition of neonatal reflexes in newborn rhesus macaques was influenced by receipt of a single neonatal dose of hepatitis B vaccine containing the preservative thimerosal (Th). Hepatitis B vaccine containing a weight-adjusted Th dose was administered to male macaques within 24 h of birth (n = 13). Unexposed animals received saline placebo (n = 4) or no injection (n = 3). Infants were tested daily for acquisition of nine survival, motor, and sensorimotor reflexes. In exposed animals there was a significant delay in the acquisition of root, snout, and suck reflexes, compared with unexposed animals. No neonatal responses were significantly delayed in unexposed animals. Gestational age (GA) and birth weight (BW) were not significantly correlated. Cox regression models were used to evaluate main effects and interactions of exposure with BW and GA as independent predictors and time-invariant covariates. Significant main effects remained for exposure on root and suck when controlling for GA and BW, such that exposed animals were relatively delayed in time-to-criterion. Interaction models indicated there were various interactions between exposure, GA, and BW and that inclusion of the relevant interaction terms significantly improved model fit. This, in turn, indicated that lower BW and/or lower GA exacerbated the adverse effects following vaccine exposure. This primate model provides a possible means of assessing adverse neurodevelopmental outcomes from neonatal Th-containing hepatitis B vaccine exposure, particularly in infants of lower GA or BW. The mechanisms underlying these effects and the requirements for Th requires further study.

WITHDRAWN: Delayed acquisition of neonatal reflexes in newborn primates receiving a thimerosal-containing Hepatitis B vaccine: Influence of gestational age and birth weight.

This article has been withdrawn at the request of the editor. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at http://www.elsevier.com/locate/withdrawalpolicy.

Comparability of developmental cognitive assessments between standard and computer testing methods.

Substantial questions have been raised about the validity of using computer-based testing to assess cognitive development with young children. However, little work has been done to assess the comparability of performance elicited using computerized methods with performance garnered using standard testing methods. The purpose of this study was to establish whether computerized testing resulted in performance that was different than established performance norms for infant monkeys (Macaca nemestrina) tested on four highly used cognitive tasks. Infants performed comparably on simple discrimination, reversal learning, and delayed nonmatch to sample rule learning. However, the infants tested in a computer testing-environment appeared to have difficulty on a task that required them to form response strategies. The results of the study reveal some apparent limitations of computer-based testing with infants, but do show that performance on several common cognitive tasks is comparable between the environments.

A computer touch screen system and training procedure for use with primate infants: Results from pigtail monkeys (Macaca nemestrina).

Computerized cognitive and perceptual testing has resulted in many advances towards understanding adult brain-behavior relations across a variety of abilities and species. However, there has been little migration of this technology to the assessment of very young primate subjects. We describe a training procedure and software that was developed to teach infant monkeys to interact with a touch screen computer. Eighteen infant pigtail macaques began training at 90-postnatal days and five began at 180-postnatal days. All animals were trained to reliably touch a stimulus presented on a computer screen and no significant differences were found between the two age groups. The results demonstrate the feasibility of using computers to assess cognitive and perceptual abilities early in development.

The effect of experience on the development of tactual-visual transfer in pigtailed macaque monkeys.

The study described here is the first to experimentally demonstrate the effects of experience on the development of tactual-visual transfer. Infant pigtailed macaque monkeys (Macaca nemestrina) were reared from birth to 2 months of age in special cages that allowed the separation of tactual and visual experience. When assessed on a battery of measures at the end of the 2-month period, animals reared without the opportunity to integrate information across the two sensory modalities performed at chance levels on a paired-comparison measure of tactual-visual transfer and performed worse than controls in a visually guided reaching task. After living in the standard laboratory environment for 2 additional months, they were reassessed. While their visually guided reaching now no longer differed from that of controls, they continued to perform at chance on the tactual-visual transfer assessment and their performance on this task was significantly worse than the control groups. Performance on visual acuity and visual recognition memory measures did not differ between groups at either age, suggesting that the deficit was limited to tactual-visual functioning. The results are discussed in terms of a possible sensitive period during which specific environmental input is required for the development of normal tactual-visual cross-modal processing.

Beta-endorphin levels in longtailed and pigtailed macaques vary by abnormal behavior rating and sex.

Frequent or severe abnormal behavior may be associated with the release of endorphins that positively reinforce the behavior with an opiate euphoria or analgesia. One line of research exploring this association involves the superhormone, proopiomelanocortin (POMC). The products of POMC appear to be dysregulated in some human subjects who exhibit self-injurious behavior (SIB). Macaque monkeys have POMC very similar to humans, and some laboratory macaques display SIB or frequent stereotypies. We investigated associations between plasma levels of three immunoreactive POMC fragments with possible opioid action and abnormal behavior ratings in macaques. In 58 adult male and female macaques (24 Macaca fascicularis and 34 Macaca nemestrina), plasma levels of intact beta-endorphin (betaE) and the N-terminal fragment (BEN) were significantly higher in animals with higher levels of abnormal behavior. The C-terminal fragment (BEC) was significantly higher in males but unrelated to ratings of abnormal behavior. Levels of ACTH, cortisol, and (betaE-ACTH)/betaE dysregulation index were unrelated to abnormal behavior. None of the POMC products differed significantly by subjects' species, age, or weight. The finding that intact beta-endorphin is positively related to abnormal behavior in two species of macaque is consistent with some previous research on human subjects and nonprimates. The positive relation of the N-terminal fragment of betaE to abnormal behavior is a new finding.

Statistical methods for describing developmental milestones with censored data: effects of birth weight status and sex in neonatal pigtailed macaques.

Neurobehavioral tests are used to assess early neonatal behavioral functioning and detect effects of prenatal and perinatal events. However, common measurement and data collection methods create specific data features requiring thoughtful statistical analysis. Assessment response measurements are often ordinal scaled, not interval scaled; the magnitude of the physical response may not directly correlate with the underlying state of developmental maturity; and a subject's assessment record may be censored. Censoring occurs when the milestone is exhibited at the first test (left censoring), when the milestone is not exhibited before the end of the study (right censoring), or when the exact age of attaining the milestone is uncertain due to irregularly spaced test sessions or missing data (interval censoring). Such milestone data is best analyzed using survival analysis methods. Two methods are contrasted: the non-parametric Kaplan-Meier estimator and the fully parametric interval censored regression. The methods represent the spectrum of survival analyses in terms of parametric assumptions, ability to handle simultaneous testing of multiple predictors, and accommodation of different types of censoring. Both methods were used to assess birth weight status and sex effects on 14 separate test items from assessments on 255 healthy pigtailed macaques. The methods gave almost identical results. Compared to the normal birth weight group, the low birth weight group had significantly delayed development on all but one test item. Within the low birth weight group, males had significantly delayed development for some responses relative to females.

Neonatal behavior and infant cognitive development in rhesus macaques produced by assisted reproductive technologies.

Assisted reproductive technologies (ART) used in fertility clinics include in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI), followed by embryo transfer into the biological or a surrogate mother. Over 1,000,000 liveborn offspring--an estimated 1 in 150 United States newborns--have been produced worldwide by ART since 1978. IVF appears to produce healthy children in singleton pregnancies, though concerns remain regarding preterm deliveries, multiple pregnancies, as well as the longer-term consequences of all ART procedures. Clinical studies remain difficult to interpret and subject to confounding variables, as developmental problems may be due to a parent's reproductive conditions rather than, or in addition to, an ART procedure. Also, because of expense and time commitments, the United States ART clinical population is not fully representative of society diversities. This socio-economic skewing might compensate for negative effects, masking small, or modest developmental deficits. Embryo splitting (ES), an ART procedure used only with animals, can produce genetically identical offspring. ES involves dividing four- to eight-cell embryos into separate blastomeres and implanting them into empty zona pellucida, followed by embryo transfer. Although these ART techniques have produced nonhuman primate offspring, there has been no research on behavioral safety. Here, we report the first study of behavioral development by rhesus macaques infants produced through ES, ICSI, and IVF. We assessed neonatal reflexes, self-feeding ability, recognition memory, object concept attainment, simple discrimination learning and reversal, and learning set (LS) acquisition. Although the sample sizes are small, we found no overall ART group delayed development. Surprisingly, the ES and ICSI monkeys appeared to be accelerated in attaining age milestones involving sensory-motor behaviors and a difficult Well Hiding object concept task. We conclude that macaque monkeys may provide an excellent model for the study of early human development by offspring of parents with conditions requiring ART pregnancies, as well as a model for the behavioral study of genetic-environment interactions using identical twins produced by ES.

Trisomy 16 in a pigtailed macaque (M. nemestrina) with multiple anomalies and developmental delays.

A female pigtailed macaque (Macaca nemestrina) with unusual physical characteristics, deficits in learning and cognitive tasks, abnormal social behavior, and abnormal reflexes and motor control was followed from birth until 3 years of age and found to have trisomy 16, which is homologous to trisomy 13 in humans. The animal described here showed similar features to cases of trisomy 16 and 18 (human trisomy 13 and 18, respectively) reported previously in nonhuman primates. However, both significant differences and similarities were found when compared with the homologous human trisomy. Evaluation of the genetic components of these disorders as well as systematic developmental evaluation can lead to new insights into the genetic basis of speciation, development, and the underlying differences between humans and their closest living relatives.

Markovian models for the developmental study of social behavior.

Behavioral development involves changes in the probabilities of both social and nonsocial activities and the sequential pattern of activities over time. A number of methods have been offered for the analysis of these patterns of behavioral sequences. However, there continue to be problematic issues, including the analysis of nonstationary data; accommodation of changes in patterns within an observation period, or over repeated observations or age; and identification of differences in pattern changes between individuals or groups, and the factors responsible for these differences. In this work, we analyze data from 15 young monkeys (Macaca nemestrina) using classification and Markovian methods, including a new approach to nonstationary data called the double-chain Markov model (DCCM). These methods allowed us to identify differences in behavior patterns that differentiate between normal subjects and those presenting developmental anomalies.

Survival, growth, health, and reproduction following nursery rearing compared with mother rearing in pigtailed monkeys (Macaca nemestrina).

Nursery-reared primates do not experience psychological "maternal bonding" or immunological benefits of breast milk, so they are expected to be inferior to mother-raised monkeys in growth, health, survival, reproduction, and maternal abilities. Studies of nursery-reared monkeys support aspects of this prediction for infants deprived of social contact or raised in pairs. We present colony record data on 1,187 mother and 506 nursery-raised monkeys, 2-10 yr of age, living in mixed groups. We found no group differences in survival, growth, clinical treatments for disease or bite wounds, or pregnancy outcome and neonatal deaths. Nursery males given breeding opportunities produced an average of 24 offspring. In addition to 24-hr personnel present on every day of the year, we believe that three of our procedures account for differences between our results and other reports. Our infants received 1) intensive human handling, 2) daily social interaction in a playroom, and 3) success and failure experience during learning and cognitive testing. We do not advocate rearing primates without mothers, but we conclude that these procedures are sufficient for producing physical health and adaptive juvenile and adult social skills in nursery-raised monkeys.

Characteristics of early rhythmic behaviors in children at risk for developmental disorders.

OBJECTIVE: Abnormal stereotyped behaviors are a significant problem for many individuals with mental retardation or mental illness. To increase understanding of the development of abnormal stereotyped behaviors, the authors investigated the early rhythmic behaviors of children at increased risk for developmental delays. METHOD: Rhythmic behaviors in 13-month-old children born prematurely and in children born at term were coded from laboratory videotapes of structured interaction segments. RESULTS: While few differences in early rhythmic behaviors were found between children born prematurely and control children, some relationships with cognitive outcomes at 2 years were identified after controlling for family socioeconomic status. However, the direction of the association was dependent on the specific coded situation. No differences in the durations of rhythmic behavior bouts were found between the groups. CONCLUSIONS: The findings indicate that even topographically similar behaviors have different functional significance in different settings. In at least some settings, rhythmic behaviors in infancy and early childhood seem to facilitate development. The fact that specific forms of rhythmic behaviors did not differ on the basis of prematurity status suggests that abnormal stereotyped behaviors have an onset later than 13 months.

Behavioral responses of longtailed macaques to different cage sizes and common laboratory experiences.

The authors tested the effects of varying cage size on the behavior of 10 female and 10 male Macaca fascicularis by singly caging them for 2 weeks in each of 5 cage sizes, ranging from approximately 20% to 148% of regulation size. Behavior in the regulation cage size, a size 23% smaller, and a size 48% larger did not differ in any analysis. Locomotion was significantly less in the 2 smallest cage sizes. Abnormal behavior occurred only 5% of the time, did not increase as cage size decreased, and did not change significantly over nearly 3 years. Disruption of the normal activity budget in the laboratory environment proved to be a useful indicator of psychological well-being. Moving to a new room and, to a lesser extent, moving into a new, clean cage, regardless of size, was associated with disrupted sleep the 1st night and suppressed activity, especially self-grooming, the next day.

Ultrasound measurement of fetal growth in Macaca nemestrina.

Longitudinal ultrasound data were collected for 18 structures in 37 pigtailed macaque (Macaca nemestrina) fetuses to 1) generate standards of normal fetal growth, 2) measure operator reliability, 3) assess the accuracy of linear and nonlinear regression models to estimate gestational age and dates of delivery, and 4) evaluate the portability of equations and absolute values derived from data at one facility (Seattle) to describe independently collected data on the same species at another facility (Medical Lake Breeding Colony). Femur length, biparietal diameter, and head area were found to be the best predictors of gestational age, as judged by maximum "explained" variance (R2) with minimum error estimates. Days to delivery could not be adequately predicted by any single fetal growth parameter or by the best combination of parameters. Operator reliability was very good: error was ⩽5% over all parameters studied. Although the form of growth curves from each facility was generally the same, variability was much greater for most parameters at Medical Lake, and both femur and humerus lengths were overestimated relative to the Seattle data. The same result was obtained for femur length when Seattle data on pigtailed macaques were compared with published data on rhesus macaques. We believe that these differences in facilities and populations may be due to the practices of the ultrasound operators. We suggest that portability of regression coefficients and absolute values for structures at a given gestational age can be accomplished only if operators are trained to use the same standard methods and receive periodic reliability checks. © 1995 Wiley-Liss, Inc.