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.

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.

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.

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.

Sex differences in compatibility of pair-housed adult longtailed macaques.

This research was designed to evaluate the effects of same-sex pair housing on the psychological well-being of adult wild-born longtailed macaques (Macaca fascicularis). We studied behavioral compatibility and stress as measured by urinary cortisol excretion in 15 pairs of each sex. Before they were housed together, the pairs were categorized by noncontact pairedpreference testing as preferred, nonpreferred, or randomly assigned partners. Every aspect of data analysis indicated that the success of pairing was strongly related to gender. Whereas 100% of female pairs were compatible, only eight of the 15 male pairs were still together after two weeks, and only five (33%) showed a degree of compatibility resembling that of females. The psychological well-being of virtually all females seemed to be improved during the physical contact paired-housing conditions; they spent more than one-third of the day engaged in social grooming. Paired adult males had much lower interaction rates than adult females. On average, males were initially somewhat stressed by the introduction to a cagemate as indicated by increased urinary cortisol excretion. The noncontact preference testing procedure was no more predictive of pair success than random assignment. For males, the presence of fighting combined with the absence of grooming during the first 90 min opportunity for physical contact ("introduction") was associated with pair incompatibility, but not to a statistically significant extent. For research protocols permitting social grouping of this species, the social contact requirement of the USDA Animal Welfare Rules usually can be met for adult females by pair housing. For males, pairing with other adult males often is unsuccessful; by our estimates, at least 20% of males cannot be pair-housed with other males. These sex differences in response to same-sex adults are consistent with the known socioecology of macaques. Further research is necessary to determine whether adult males have a lower need for social contact than females, or whether their needs are better met by other types of social contact. © 1994 Wiley-Liss, Inc.

Urinary cortisol responses of longtailed macaques to five cage sizes, tethering, sedation, and room change.

Urinary free cortisol responses to five cage sizes, cage level, room change, tethering adaptation, chronic catheterization, and ketamine sedation were measured in 14 female and 14 male wild-born adult Macaca fascicularis. Urinary free cortisol, a physiological measure of psychological well-being that can be collected unobtrusively, provided a measure of the animals' general adrenocortical response to various conditions over a time course of hours. Urinary free cortisol values in response to stimulation with adrenocorticotropic hormone (ACTH) validated the measure as a reflection of blood values. Cortisol values were expressed as a ratio to creatinine, which normalized for differences in urinary output and body weight (muscle mass). Cage size (20-140% of regulation floor area) and housing level (upper vs. lower cage) had no effect on stress, as measured by cortisol excretion. Room change elicited a slight increase in cortisol excretion for the first day, but not to a level suggesting stress. Sedation, surgery, some aspects of tethering adaptation, and chronic catheterization produced urinary cortisol evidence of stress. Even so, animals varied in their responses and all showed adaptation. Males and females did not differ in normal mean values but females tended to have higher cortisol levels in response to potential stressors investigated in this study. Cortisol levels continued to decline gradually throughout the study. © 1993 Wiley-Liss, Inc.

Growth of nursery-raised Macaca nemestrina infants: Effects of feeding schedules, sex, and birth weight.

Ponderal growth, caloric intake, growth velocity, and growth efficiency were studied from birth through 90 days in 523 nursery-raised pigtailed macaque infants. The male and female subjects ranged in birth weight from the 1st through 100th centiles of the colony distribution and were all judged to be clinically healthy. One group of infants was fed on an ad lib schedule, with formula continuously available from dispensers which were replenished every 8 h. A second group was fed on a distributed regimen, with formula available on a 4-h-on/4-h-off schedule. The latter schedule was instituted because of bacterial growth in the milk after 4 h. Infants fed the distributed diet had 20-25% lower caloric intake than those fed ad lib, but equaled or exceeded ad lib infants in body weight, growth velocity, and grams gained per 100 kilocalories (growth efficiency). The poorer utilization of calories by the ad lib group was attributed to a combination of bacteria in the milk, which caused diarrhea, and inability to metabolize the excess nutrients gained from overconsumption of fluids. The effects of the distributed diet occurred for both males and females, and across the full range of birth weights. Peak velocity and growth efficiency for both males and females occurred at 2 weeks after birth for high-birth-weight (HBW) infants and at 3 weeks for low-birth-weight (LBW) infants. However, the growth efficiency of LBW infants was equal to, or greater than, that of HBW infants after postnatal day 2. Descriptive statistics and regression equations based on this large sample are presented for quantitative comparisons of growth parameters with those of other primate species reared in laboratory or field settings. © 1992 Wiley-Liss, Inc.

Rearing infant monkeys (Macaca nemestrina) in pairs produces deficient social development compared with rearing in single cages.

Many scientists and colony managers assume that social housing is a beneficial living condition for all captive primates. Several older studies of primate development question the generality of this assumption. We recently tested this assumption by comparing the social development of pigtailed macaque infants raised in pairs and those that were raised in individual cages. All animals received 30 min of daily socialization in a playroom. Infants paired from postnatal week 3 through month 4 developed a playroom behavioral repertoire consisting largely of mutual clinging, fear, and social withdrawal. This was especially true of females. Unlike the singly caged infants, pair-reared monkeys did not successfully adapt to living in a large social group at 8-10 months of age. In this situation, pair-reared infants were subordinate and spent almost all of their time huddling on the pen floor. It was concluded that rearing macaque infants in pairs produces a behavioral repertoire that is maladaptive with respect to social development.

Effects of management methods, social organization, and physical space on primate behavior and health.

The incidence of clinical treatment in a large colony of pigtailed macaques (Macaca, nemestrina) was studied retrospectively to assess the effectiveness of three sequential housing and management protocols. In the initial standard protocol, groups were housed in single rooms, with each room occupied by a different group after daily cleaning. In the experimental protocol, animals were housed in two-room suites dedicated exclusively to the resident group. This protocol was meant to reduce enteritis due to communication of Shigella flexneri across groups. Isolation of groups was coupled with the doubling of available space in an attempt to reduce stress. Under a final compromise protocol, a two-room suite was dedicated to each group, but the group occupied only one room at a time. The compromise protocol maximized isolation and reduced the available space to the level under the standard protocol. Thus, the study involved two main factors, the degree of group isolation and the amount and configuration of physical space available to each group. Under the experimental conditions enteritis increased, other communicable diseases did not decrease, and trauma from fighting increased by over 45% even though more space was available. Under the compromise conditions, trauma, enteritis, respiratory disorders, and inflammation all decreased markedly. Behavioral studies identified beneficial social behavior patterns associated with spatial reduction as the basis of the decrease in aggression, trauma, and disease under the compromise conditions.

Sires influence fetal death in pigtailed macaques (Macaca nemestrina).

Computerized colony records were used to identify dams and sires whose reproductive histories while breeding in harem groups indicated high or low risk for producing fetal deaths. The monkeys were then singly caged and bred within and between risk conditions during 24- to 72-hour matings. Although the low- and high-risk sires did not differ in ability to achieve conceptions, the results revealed that reproductive history predicted subsequent fetal death rates, and sires made a substantial contribution. Overall, high-risk sires had a 4.86 times higher relative risk of producing fetal death than low-risk sires. Compared with this figure, high-risk dams had only a 2.64 times higher risk for fetal death than low-risk dams. The male effect was most evident in comparisons of between- and within-risk matings. Females with excellent reproductive histories experienced a threefold increase in fetal deaths when mated with high-risk males, whereas females with poor reproductive histories experienced almost a 50% reduction in fetal deaths when mated with low-risk males. The results show that male factors beyond fertility need to be considered when examining the causes of reproductive failure in captive primates.

Measurement of excreted steroids in Macaca nemestrina.

A practical method for the quantitative measurement of the estrogenic steroid estradiol-17ß in the feces of pigtailed macaques (Macaca nemestrina) is described. The method, which includes homogenization, filtration, ether extraction, and sephadex purification, produces an 85.3% recovery of 3H-estradiol. Comparable recovery of 3H-progesterone (64.1%) and 3H-testoster-one (67.9%) is also obtained. Estradiol, measured by radioimmunoassay methodology, has inter- and intra-assay variations of 7.1% and 4.3%, respectively. Linearity of estradiol was checked by varying the amount of feces analyzed. Results show a correlation of 0.9945, Y = 32.526x + 0.205. Accuracy of estradiol, determined by adding radioinert estradiol to the feces before homogenization, was 98.7% with a correlation of 0.9950, Y = 1.027x - 0.543. The menstrual cycles of several animals were analyzed and corresponded well with serum levels of estradiol-17ß. Similar correspondence was obtained in matched fecal and serum samples from pregnant animals. These methods provide a practical and accurate solution to the problem of collecting hormone data in field studies without the potential complications of capture in laboratory research.