Immunological and microbial shifts in the aging rhesus macaque lung during nontuberculous mycobacterial infection.

Nontuberculous mycobacteria (NTM) are environmentally ubiquitous organisms that predominately cause NTM pulmonary disease (NTMPD) in individuals over the age of 65. The incidence of NTMPD has increased in the U.S., exceeding that of Mycobacterium tuberculosis. However, the mechanisms leading to higher susceptibility and severity of NTMPD with aging are poorly defined in part due to the lack of animal models that accurately recapitulate human disease. Here, we compared bacterial load, microbial communities, and host responses longitudinally between three young (two female and one male) and two aged (two female) rhesus macaques inoculated with Mycobacterium avium subsp. hominissuis (MAH) in the right caudal lobe. Unilateral infection resulted in a low bacterial load in both young and aged animals confined to the infected side. Although a robust inflammatory response was only observed in the inoculated lung, immune cell infiltration and antigen-specific T cells were detected in both lungs. Computed tomography, gross pathology, and histopathology revealed increased disease severity and persistence of bacterial DNA in aged animals. Additional analyses showed the translocation of gut and oral-pharyngeal bacterial DNA into the lower respiratory microbiome. Finally, single-cell RNA sequencing revealed a heightened inflammatory response to MAH infection by alveolar macrophages in aged animals. These data are consistent with the model that increased disease severity in the aged is mediated by a dysregulated macrophage response that may be sustained through persistent antigen presence. IMPORTANCE: Nontuberculous mycobacteria (NTM) are emerging as pathogens of high consequence, as cases of NTM pulmonary disease (NTMPD) have exceeded those of Mycobacterium tuberculosis. NTMPD can be debilitating, particularly in patients over 65 years of age, as it causes chronic cough and fatigue requiring prolonged treatments with antibiotics. The underlying mechanisms of this increased disease severity with age are poorly understood, hampering the development of therapeutics and vaccines. Here, we use a rhesus macaque model to investigate the impact of age on host-NTM interactions. This work shows that aging is associated with increased disease severity and bacterial persistence in aged rhesus macaques, thus providing a preclinical model to develop and test novel therapeutics and interventions.

Age-related increase in the expression of 11ß-hydroxysteroid dehydrogenase type 1 in the hippocampus of male rhesus macaques.

Introduction: The hippocampus is especially susceptible to age-associated neuronal pathologies, and there is concern that the age-associated rise in cortisol secretion from the adrenal gland may contribute to their etiology. Furthermore, because 11ß-hydroxysteroid dehydrogenase type 1 (HSD11B1) catalyzes the reduction of cortisone to the active hormone cortisol, it is plausible that an increase in the expression of this enzyme enhances the deleterious impact of cortisol in the hippocampus and contributes to the neuronal pathologies that underlie cognitive decline in the elderly. Methods: Rhesus macaques were used as a translational animal model of human aging, to examine age-related changes in gene and protein expressions of (HSD11B1/HSD11B1) in the hippocampus, a region of the brain that plays a crucial role in learning and memory. Results: Older animals showed significantly (p < 0.01) higher base-line cortisol levels in the circulation. In addition, they showed significantly (p < 0.05) higher hippocampal expression of HSD11B1 but not NR3C1 and NR3C2 (i.e., two receptor-encoding genes through which cortisol exerts its physiological actions). A similar age-related significant (p < 0.05) increase in the expression of the HSD11B1 was revealed at the protein level by western blot analysis. Discussion: The data suggest that an age-related increase in the expression of hippocampal HSD11B1 is likely to raise cortisol concentrations in this cognitive brain area, and thereby contribute to the etiology of neuropathologies that ultimately lead to neuronal loss and dementia. Targeting this enzyme pharmacologically may help to reduce the negative impact of elevated cortisol concentrations within glucocorticoid-sensitive brain areas and thereby afford neuronal protection.

Modulation of neural gene networks by estradiol in old rhesus macaque females.

The postmenopausal decrease in circulating estradiol (E2) levels has been shown to contribute to several adverse physiological and psychiatric effects. To elucidate the molecular effects of E2 on the brain, we examined differential gene expression and DNA methylation (DNAm) patterns in the nonhuman primate brain following ovariectomy (Ov) and subsequent subcutaneous bioidentical E2 chronic treatment. We identified several dysregulated molecular networks, including MAPK signaling and dopaminergic synapse response, that are associated with ovariectomy and shared across two different brain areas, the occipital cortex (OC) and prefrontal cortex (PFC). The finding that hypomethylation (p = 1.6 × 10-51) and upregulation (p = 3.8 × 10-3) of UBE2M across both brain regions provide strong evidence for molecular differences in the brain induced by E2 depletion. Additionally, differential expression (p = 1.9 × 10-4; interaction p = 3.5 × 10-2) of LTBR in the PFC provides further support for the role E2 plays in the brain, by demonstrating that the regulation of some genes that are altered by ovariectomy may also be modulated by Ov followed by hormone replacement therapy (HRT). These results present real opportunities to understand the specific biological mechanisms that are altered with depleted E2. Given E2's potential role in cognitive decline and neuroinflammation, our findings could lead to the discovery of novel therapeutics to slow cognitive decline. Together, this work represents a major step toward understanding molecular changes in the brain that are caused by ovariectomy and how E2 treatment may revert or protect against the negative neuro-related consequences caused by a depletion in estrogen as women approach menopause.

Pathological Markers of Alzheimer's Disease and Related Dementia in the Rhesus Macaque Amygdala.

Rhesus macaques develop amyloid-ß (Aß) plaques during old age, but it is unclear how extensively they express other pathological hallmarks of dementia. Here we used immunohistochemistry to examine expression of phosphorylated tau (pTau) protein and cytoplasmic inclusions of TAR DNA binding protein 43 kDa (TDP-43) within the amygdala of young and old males, and also in old surgically-menopausal females that were maintained on regular or obesogenic diets. Only one animal, a 23-year-old female, showed pTau expression and none showed TDP-43 inclusions. What genetic and/or environmental factors protect macaques from expressing more severe human neuro-pathologies remains an interesting unresolved question.

Brain volumetrics across the lifespan of the rhesus macaque.

The rhesus macaque is a long-lived nonhuman primate (NHP) with a brain structure similar to humans, which may represent a valuable translational animal model in which to study human brain aging. Previous magnetic resonance imaging (MRI) studies of age in rhesus macaque brains have been prone to low statistical power, unbalanced sex ratio and lack of a complete age range. To overcome these problems, the current study surveyed structural T1-weighted magnetic resonance imaging scans of 66 animals, 34 females (aged 6-31 years) and 32 males (aged 5-27 years). Differences observed in older animals, included enlargement of the lateral ventricles and a smaller volume in the frontal cortex, caudate, putamen, hypothalamus, and thalamus. Unexpected, greater volume, were measured in older animals in the hippocampus, amygdala, and globus pallidus. There were also numerous differences between males and females with respect to age in both white and gray matter regions. As an apparent model of normative human aging, the macaque is ideal for studying induction and mitigation of neurodegenerative disease.

A class of anti-inflammatory lipids decrease with aging in the central nervous system.

Lipids contribute to the structure, development, and function of healthy brains. Dysregulated lipid metabolism is linked to aging and diseased brains. However, our understanding of lipid metabolism in aging brains remains limited. Here we examined the brain lipidome of mice across their lifespan using untargeted lipidomics. Co-expression network analysis highlighted a progressive decrease in 3-sulfogalactosyl diacylglycerols (SGDGs) and SGDG pathway members, including the potential degradation products lyso-SGDGs. SGDGs show an age-related decline specifically in the central nervous system and are associated with myelination. We also found that an SGDG dramatically suppresses LPS-induced gene expression and release of pro-inflammatory cytokines from macrophages and microglia by acting on the NF-κB pathway. The detection of SGDGs in human and macaque brains establishes their evolutionary conservation. This work enhances interest in SGDGs regarding their roles in aging and inflammatory diseases and highlights the complexity of the brain lipidome and potential biological functions in aging.

Modulation of neural gene networks by estradiol in old rhesus macaque females.

The postmenopausal decrease in circulating estradiol (E2) levels has been shown to contribute to several adverse physiological and psychiatric effects. To elucidate the molecular effects of E2 on the brain, we examined differential gene expression and DNA methylation (DNAm) patterns in the nonhuman primate brain following ovariectomy (Ov) and subsequent E2 treatment. We identified several dysregulated molecular networks, including MAPK signaling and dopaminergic synapse response, that are associated with ovariectomy and shared across two different brain areas, the occipital cortex (OC) and prefrontal cortex (PFC). The finding that hypomethylation (p=1.6×10-51) and upregulation (p=3.8×10-3) of UBE2M across both brain regions, provide strong evidence for molecular differences in the brain induced by E2 depletion. Additionally, differential expression (p=1.9×10-4; interaction p=3.5×10-2) of LTBR in the PFC, provides further support for the role E2 plays in the brain, by demonstrating that the regulation of some genes that are altered by ovariectomy may also be modulated by Ov followed by hormone replacement therapy (HRT). These results present real opportunities to understand the specific biological mechanisms that are altered with depleted E2. Given E2's potential role in cognitive decline and neuroinflammation, our findings could lead to the discovery of novel therapeutics to slow cognitive decline. Together, this work represents a major step towards understanding molecular changes in the brain that are caused by ovariectomy and how E2 treatment may revert or protect against the negative neuro-related consequences caused by a depletion in estrogen as women approach menopause.

Effect of hormone replacement therapy on amyloid beta (Aß) plaque density in the rhesus macaque amygdala.

Background: Amyloid beta (Aß) plaque density was examined in the amygdala of rhesus macaques, to elucidate the influence of age, diet and hormonal environment. Methods: Luminex technology was used to measure cerebrospinal fluid (CSF) concentrations of Aß40 and Aß42 across three decades, while immunohistochemistry was used to examine Aß plaque density in the amygdala. Results: Aß40 was found to be the predominant isoform of Aß in the CSF, but neither Aß40 or Aß42 concentrations showed an age-related change, and the ratio of Aß42 to Aß40 showed only a marginal increase. Significantly fewer Aß plaques were detected in the amygdala of old ovariectomized animals if they received estradiol HRT (p < 0.001); similar results were obtained regardless of whether they had been maintained on a regular monkey chow for ∼48 months or on a high-fat, high-sugar, Western-style diet for ∼30 months. Conclusion: The results demonstrate that HRT involving estrogen can reduce Aß plaque load in a cognitive brain region of aged non-human primates. The results from this translational animal model may therefore have clinical relevance to the treatment of AD in post-menopausal women, whether used alone, or as a supplement to current pharmacological and monoclonal antibody-based interventions.

A macaque clonal hematopoiesis model demonstrates expansion of TET2-disrupted clones and utility for testing interventions.

Individuals with age-related clonal hematopoiesis (CH) are at greater risk for hematologic malignancies and cardiovascular diseases. However, predictive preclinical animal models to recapitulate the spectrum of human CH are lacking. Through error-corrected sequencing of 56 human CH/myeloid malignancy genes, we identified natural CH driver mutations in aged rhesus macaques matching genes somatically mutated in human CH, with DNMT3A mutations being the most frequent. A CH model in young adult macaques was generated via autologous transplantation of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9-mediated gene-edited hematopoietic stem and progenitor cells (HSPCs), targeting the top human CH genes with loss-of-function (LOF) mutations. Long-term follow-up revealed reproducible and significant expansion of multiple HSPC clones with heterozygous TET2 LOF mutations, compared with minimal expansion of clones bearing other mutations. Although the blood counts of these CH macaques were normal, their bone marrows were hypercellular and myeloid-predominant. TET2-disrupted myeloid colony-forming units isolated from these animals showed a distinct hyperinflammatory gene expression profile compared with wild type. In addition, mature macrophages purified from the CH macaques showed elevated NLRP3 inflammasome activity and increased interleukin-1ß (IL-1ß) and IL-6 production. The model was used to test the impact of IL-6 blockage by tocilizumab, documenting a slowing of TET2-mutated expansion, suggesting that interruption of the IL-6 axis may remove the selective advantage of mutant HSPCs. These findings provide a model for examining the pathophysiology of CH and give insights into potential therapeutic interventions.

Functional, transcriptional, and microbial shifts associated with healthy pulmonary aging in rhesus macaques.

Older individuals are at increased risk of developing severe respiratory infections. However, our understanding of the impact of aging on the respiratory tract remains limited as samples from healthy humans are challenging to obtain and results can be confounded by variables such as smoking and diet. Here, we carry out a comprehensive cross-sectional study (n = 34 adult, n = 49 aged) to define the consequences of aging on the lung using the rhesus macaque model. Pulmonary function testing establishes similar age and sex differences as humans. Additionally, we report increased abundance of alveolar and infiltrating macrophages and a concomitant decrease in T cells were in aged animals. scRNAseq reveals shifts from GRZMB to IFN expressing CD8+ T cells in the lungs. These data provide insight into age-related changes in the lungs' functional, microbial, and immunological landscape that explain increased prevalence and severity of respiratory diseases in the elderly.

Effects of estradiol supplementation on the brain transcriptome of old rhesus macaques maintained on an obesogenic diet.

Obesity, the cessation of ovarian steroids with menopause, and age are risk factors for mood disorders, dementia, and Alzheimer's disease (AD). However, immediate hormone therapy (HT) after menopause may have beneficial effects in different brain regions involved in memory and cognition. To more closely replicate the age, endocrine, and metabolic environment of obese postmenopausal women, either on or off HT, middle-aged female rhesus macaques were ovariectomized/hysterectomized (OvH) and maintained on a high-fat, high-sugar, obesogenic Western-style diet (WSD) for 30 months; half of the animals received HT immediately after OvH and half served as placebo controls. RNAseq of the occipital (OC) and prefrontal cortex (PFC), hippocampus (HIP), and amygdala (AMG) identified 293, 379, 505, and 4993 differentially expressed genes (DEGs), respectively. Pathway enrichment analysis identified an activation of neuroinflammation in OC and HIP, but an inhibition in the AMG with HT. Synaptogenesis, circadian rhythm, mitochondrial dysfunction, mTOR, glutamate, serotonin, GABA, dopamine, epinephrine/norepinephrine, glucocorticoid receptor signaling, neuronal NOS, and amyloid processing were exclusively enriched in AMG. As compared to the placebo control group, most of these signaling pathways are downregulated after HT, suggesting a protective effect of HT in OvH females under a WSD. Overall, our results suggest that a chronic obesogenic diet may induce a wide range of alterations in multiple signaling pathways that are linked to age-associated brain pathology and dementia. In these individuals, HT seems to have a protective effect against neuroinflammation, amyloid beta depositions, and tau tangle formation.

MRI characteristics of Japanese macaque encephalomyelitis: Comparison to human diseases.

BACKGROUND AND PURPOSE: To describe MRI findings in Japanese macaque encephalomyelitis (JME) with emphasis on lesion characteristics, lesion evolution, normal-appearing brain tissue, and similarities to human demyelinating disease. METHODS: MRI data were obtained from 114 Japanese macaques, 30 presenting neurological signs of JME. All animals were screened for presence of T2 -weighted white matter signal hyperintensities; animals with behavioral signs of JME were additionally screened for contrast-enhancing lesions. Whole-brain quantitative T1 maps were collected, and histogram analysis was performed with regression across age to evaluate microstructural changes in normal appearing brain tissue in JME and neurologically normal animals. Quantitative estimates of blood-brain-barrier (BBB) permeability to gadolinium-based-contrast agent (GBCA) were obtained in acute, GBCA-enhancing lesions. Longitudinal imaging data were acquired for 15 JME animals. RESULTS: One hundred and seventy-three focal GBCA-enhancing lesions were identified in 30 animals demonstrating behavioral signs of neurological dysfunction. JME GBCA-enhancing lesions were typically focal and ovoid, demonstrating highest BBB GBCA permeability in the lesion core, similar to acute, focal multiple sclerosis lesions. New GBCA-enhancing lesions arose rapidly from normal-appearing tissue, and BBB permeability remained elevated for weeks. T1 values in normal-appearing tissue were significantly associated with age, but not with sex or disease. CONCLUSIONS: Intense, focal neuroinflammation is a key MRI finding in JME. Several features of JME compare directly to human inflammatory demyelinating diseases. Investigation of JME combined with the development and validation of noninvasive imaging biomarkers offers substantial potential to improve diagnostic specificity and contribute to the understanding of human demyelinating diseases.

Myelin-specific T cells in animals with Japanese macaque encephalomyelitis.

OBJECTIVE: To determine whether animals with Japanese macaque encephalomyelitis (JME), a spontaneous demyelinating disease similar to multiple sclerosis (MS), harbor myelin-specific T cells in their central nervous system (CNS) and periphery. METHODS: Mononuclear cells (MNCs) from CNS lesions, cervical lymph nodes (LNs) and peripheral blood of Japanese macaques (JMs) with JME, and cervical LN and blood MNCs from healthy controls or animals with non-JME conditions were analyzed for the presence of myelin-specific T cells and changes in interleukin 17 (IL-17) and interferon gamma (IFNγ) expression. RESULTS: Demyelinating JME lesions contained CD4+ T cells and CD8+ T cells specific to myelin oligodendrocyte glycoprotein (MOG), myelin basic protein (MBP), and/or proteolipid protein (PLP). CD8+ T-cell responses were absent in JME peripheral blood, and in age- and sex-matched controls. However, CD4+ Th1 and Th17 responses were detected in JME peripheral blood versus controls. Cervical LN MNCs from eight of nine JME animals had CD3+ T cells specific for MOG, MBP, and PLP that were not detected in controls. Mapping myelin epitopes revealed a heterogeneity in responses among JME animals. Comparison of myelin antigen sequences with those of JM rhadinovirus (JMRV), which is found in JME lesions, identified six viral open reading frames (ORFs) with similarities to myelin antigen sequences. Overlapping peptides to these JMRV ORFs did not induce IFNγ responses. INTERPRETATIONS: JME possesses an immune-mediated component that involves both CD4+ and CD8+ T cells specific for myelin antigens. JME may shed new light on inflammatory demyelinating disease pathogenesis linked to gamma-herpesvirus infection.

Lack of effect of short-term DHEA supplementation on the perimenopausal ovary†.

Dehydroepiandrosterone (DHEA) hormonal supplementation can improve oocyte quality in women with diminished ovarian function. However, it is unclear whether DHEA supplementation can also enhance ovarian function during the perimenopause (i.e., when the number of follicles in the ovary has undergone a marked reduction). To address this question, we examined the impact of 2.5-months of daily 5-mg oral DHEA supplementation on the number of ovarian follicles and the concentration of anti-Müllerian hormone (AMH) in perimenopausal rhesus macaques. Like women, these long-lived nonhuman primates have ~ 28-day menstrual cycles and eventually undergo menopause. They also show similar age-related neuroendocrine changes, including a marked decrease in circulating concentrations of DHEA and DHEA sulfate (DHEAS). Our experimental design involved the following three groups of animals (N = 6 per group): Young adult (mean age = 11.6 years), Old control (mean age = 23.1 years), and Old DHEA-treated (mean age = 23.5 years). Histological examination of the ovaries revealed a significant age-related decrease in the mean number of primordial follicles despite DHEA supplementation. Moreover, AMH concentrations within the ovaries and circulation, assessed by Western analysis and ELISA, respectively, showed significant age-related decreases that were not attenuated by DHEA supplementation. Taken together, these results fail to show a clear effect of short-term physiological DHEA supplementation on the perimenopausal ovary. However, they do not exclude the possibility that alternative DHEA supplementation paradigms (e.g., involving an earlier start date, longer duration and using pharmacological doses) may extend reproductive potential during aging.

Longitudinal Effects of Immediate and Delayed Estradiol on Cognitive Performance in a Spatial Maze and Hippocampal Volume in Menopausal Macaques Under an Obesogenic Diet.

The consumption of a diet high in fat and refined sugars has several health risks, including the development of cognitive decline and neurodegeneration. For women, menopause carries additional health risks that may interact with a high-fat diet in negative ways. Some symptoms of menopause, including cognitive impairments, can be modulated by hormone replacement therapy (HRT), but the hormonal formulation and the timing of the treatment relative to the onset of menopause are critical factors determining its efficacy. Little is known about how obesogenic, high-fat, high-sugar diets interact with HRT in menopause to affect cognition and neurodegeneration. Given the high prevalence of the consumption of an obesogenic Western-style diet, understanding how the effects of HRT are modulated by an obesogenic diet is critical for developing optimized therapeutic strategies for peri- and post-menopausal women. In this study, we investigated by magnetic resonance imaging (MRI) the effects of either immediate or delayed estradiol hormone therapy on cognition and neuroanatomy following ovo-hysterectomy (OvH) of aged, female rhesus macaques on an obesogenic diet. The macaques were followed for 2.5 years after ovo-hysterectomy, with four time points at which anatomical MRIs were acquired. Analysis of hippocampal volumes revealed an interaction between time point and treatment; hippocampal volumes in the delayed estrogen group, but not the immediate estrogen group, increased over time compared to those in untreated controls. Performance on a hippocampal-dependent spatial maze task showed improved performance in estrogen treated animals compared to OvH macaques given placebo. These results indicate that HRT may contribute to beneficial cognitive outcomes after menopause under an obesogenic diet.

Regional Molecular Mapping of Primate Synapses during Normal Healthy Aging.

Normal mammalian brain aging is characterized by the selective loss of discrete populations of dendritic spines and synapses, particularly affecting neuroanatomical regions such as the hippocampus. Although previous investigations have quantified this morphologically, the molecular pathways orchestrating preferential synaptic vulnerability remain to be elucidated. Using quantitative proteomics and healthy rhesus macaque and human patient brain regional tissues, we have comprehensively profiled the temporal expression of the synaptic proteome throughout the adult lifespan in differentially vulnerable brain regions. Comparative profiling of hippocampal (age vulnerable) and occipital cortex (age resistant) synapses revealed discrete and dynamic alterations in the synaptic proteome, which appear unequivocally conserved between species. The generation of these unique and important datasets will aid in delineating the molecular mechanisms underpinning primate brain aging, in addition to deciphering the regulatory biochemical cascades governing neurodegenerative disease pathogenesis.

Preconditioning in the Rhesus Macaque Induces a Proteomic Signature Following Cerebral Ischemia that Is Associated with Neuroprotection.

Each year, thousands of patients are at risk of cerebral ischemic injury, due to iatrogenic responses to surgical procedures. Prophylactic treatment of these patients as standard care could minimize potential neurological complications. We have shown that protection of brain tissue, in a non-human primate model of cerebral ischemic injury, is possible through pharmacological preconditioning using the immune activator D192935. We postulate that preconditioning with D192935 results in neuroprotective reprogramming that is evident in the brain following experimentally induced cerebral ischemia. We performed quantitative proteomic analysis of cerebral spinal fluid (CSF) collected post-stroke from our previously published efficacy study to determine whether CSF protein profiles correlated with induced protection. Four groups of animals were examined: naïve animals (no treatment or stroke); animals treated with vehicle prior to stroke; D192935 treated and stroked animals, further delineated into two groups, ones that were protected (small infarcts) and those that were not protected (large infarcts). We found that distinct protein clusters defined the protected and non-protected animal groups, with a 16-member cluster of proteins induced exclusively in D192935 protected animals. Seventy percent of the proteins induced in the protected animals have functions that would enhance neuroprotection and tissue repair, including several members associated with M2 macrophages, a macrophage phenotype shown to contribute to neuroprotection and repair during ischemic injury. These studies highlight the translational importance of CSF biomarkers in defining mechanism and monitoring responses to treatment in development of stroke therapeutics.

Discovery of a CLN7 model of Batten disease in non-human primates.

We have identified a natural Japanese macaque model of the childhood neurodegenerative disorder neuronal ceroid lipofuscinosis, commonly known as Batten Disease, caused by a homozygous frameshift mutation in the CLN7 gene (CLN7-/-). Affected macaques display progressive neurological deficits including visual impairment, tremor, incoordination, ataxia and impaired balance. Imaging, functional and pathological studies revealed that CLN7-/- macaques have reduced retinal thickness and retinal function early in disease, followed by profound cerebral and cerebellar atrophy that progresses over a five to six-year disease course. Histological analyses showed an accumulation of cerebral, cerebellar and cardiac storage material as well as degeneration of neurons, white matter fragmentation and reactive gliosis throughout the brain of affected animals. This novel CLN7-/- macaque model recapitulates key behavioral and neuropathological features of human Batten Disease and provides novel insights into the pathophysiology linked to CLN7 mutations. These animals will be invaluable for evaluating promising therapeutic strategies for this devastating disease.

Gene expression profiling of the SCN in young and old rhesus macaques.

In mammals, the suprachiasmatic nucleus (SCN) is the location of a master circadian pacemaker. It receives photic signals from the environment via the retinal hypothalamic tract, which play a key role in synchronizing the body's endogenously generated circadian rhythms with the 24-h rhythm of the environment. Therefore, it is plausible that age-related changes within the SCN contribute to the etiology of perturbed activity-rest cycles that become prevalent in humans during aging. To test this hypothesis, we used gene arrays and quantitative RT-PCR to profile age-related gene expression changes within the SCN of male rhesus macaques - a pragmatic translational animal model of human aging, which similarly displays an age-related attenuation of daytime activity levels. As expected, the SCN showed high expression of arginine vasopressin, vasoactive intestinal polypeptide, calbindin and nuclear receptor subfamily 1, group D, member 1 (NR1D1) (also known as reverse strand of ERBA (REV-ERBα), both at the mRNA and protein level. However, no obvious difference was detected between the SCNs of young (7-12 years) and old animals (21-26 years), in terms of the expression of core clock genes or genes associated with SCN signaling and neurotransmission. These data demonstrate the resilience of the primate SCN to normal aging, at least at the transcriptional level and, at least in males, suggest that age-related disruption of activity-rest cycles in humans may instead stem from changes within other components of the circadian system, such as desynchronization of subordinate oscillators in other parts of the body.