Abnormality of anxious behaviors and functional connectivity between the amygdala and the frontal lobe in maternally deprived monkeys.

OBJECTIVE: Anxious behaviors often occur in individuals who have experienced early adversity. Anxious behaviors can bring many hazards, such as social withdrawal, eating disorders, negative self-efficacy, self-injurious thoughts and behaviors, anxiety disorders, and even depression. Abnormal behavior are is closely related to changes in corresponding circuit functions in the brain. This study investigated the relationship between brain circuits and anxious behaviors in maternal-deprived rhesus monkey animal model, which mimic early adversity in human. METHODS: Twenty-five rhesus monkeys (Macaca mulatta) were grouped by two different rearing conditions: 11 normal control and mother-reared (MR) monkeys and 14 maternally deprived and peer-reared (MD) monkeys. After obtaining images of the brain areas with significant differences in maternal separation and normal control macaque function, the relationship between functional junction intensity and stereotypical behaviors was determined by correlation analysis. RESULTS: The correlation analysis revealed that stereotypical behaviors were negatively correlated with the coupling between the left lateral amygdala subregion and the left inferior frontal gyrus in both MD and MR macaques. CONCLUSION: This study suggests that early adversity-induced anxious behaviors are associated with changes in the strength of the amygdala-prefrontal connection. The normalization of the regions involved in the functional connection might reverse the behavioral abnormality. It provides a solid foundation for effective intervention in human early adversity. SIGNIFICANCE STATEMENT: This study suggests that early adversity-induced anxious behaviors are associated with changes in the strength of the amygdala-prefrontal connection. The higher the amygdala-prefrontal connection strength, the less stereotyped behaviors exhibited by monkeys experiencing early adversity. Thus, in the future, changing the strength of the amygdala-prefrontal connection may reverse the behavioral abnormalities of individuals who experience early adversity. This study provides a solid foundation for effective intervention in humans' early adversity.

Transient neurogenesis in ischemic cortex from Sox2+ astrocytes.

The adult cortex has long been regarded as non-neurogenic. Whether injury can induce neurogenesis in the adult cortex is still controversial. Here, we report that focal ischemia stimulates a transient wave of local neurogenesis. Using 5'-bromo-2'-deoxyuridine labeling, we demonstrated a rapid generation of doublecortin-positive neuroblasts that died quickly in mouse cerebral cortex following ischemia. Nestin-CreER-based cell ablation and fate mapping showed a small contribution of neuroblasts by subventricular zone neural stem cells. Using a mini-photothrombotic ischemia mouse model and retrovirus expressing green fluorescent protein labeling, we observed maturation of locally generated new neurons. Furthermore, fate tracing analyses using PDGFRα-, GFAP-, and Sox2-CreER mice showed a transient wave of neuroblast generation in mild ischemic cortex and identified that Sox2-positive astrocytes were the major neurogenic cells in adult cortex. In addition, a similar upregulation of Sox2 and appearance of neuroblasts were observed in the focal ischemic cortex of Macaca mulatta. Our findings demonstrated a transient neurogenic response of Sox2-positive astrocytes in ischemic cortex, which suggests the possibility of inducing neuronal regeneration by amplifying this intrinsic response in the future.

Bimetal heterointerfaces towards enhanced electro-activation of O2 under room condition.

Efficient catalysts for oxygen (O2) activation under room condition are required for effective wet air oxidation (WAO) technology. Here, we report a novel manganese-cobalt-based composite (MnO-CoO@Co) fabricated on a graphite felt (GF) support for catalyzing the electro-activation of O2 under room condition. Abundant Co-MnO and CoO-MnO heterointerfaces are formed in the composite. In comparison to the single-metal counterparts, i.e. CoO@Co/GF (16.99 wt% Co) and MnO/GF (26.83 wt% Mn), the bimetal MnO-CoO@Co/GF (5.29 wt% Co and 8.79 wt% Mn) displays an improved oxygen storage capacity and provides more active sites to accommodate surface adsorbed oxygen species. Notably, the strong synergy derived from bimetal heterointerfaces enhances the electron transfer and oxygen mobilization during the electro-activation of O2, thereby significantly reducing the reaction barrier. MnO-CoO@Co/GF exhibits excellent efficiency and stability in electrocatalytic WAO (ECWAO) towards the removal of pharmaceuticals and personal care products (PPCPs) over a wide pH range from 4.0 to 10.0. A model pollutant sulfamethoxazole (SMX) acquires mineralization efficiency of 78.4 ± 2.1% and mineralization current efficiency of 157.89% at +1.0 V of electrode potential. The toxicity of PPCPs can be totally eliminated after the ECWAO treatment. This work highlights the synergy derived from bimetal heterointerfaces in O2 electrocatalysis, and provides a promising approach for advanced WAO catalysts in PPCPs pollution control.

Biological implications and limitations of a cynomolgus monkey with naturally occurring Parkinson's disease.

We recently identified a cynomolgus monkey with naturally occurring Parkinson's disease (PD), indicating that PD may not be a uniquely human disease (Li et al, 2020). In our previous study, four lines of evidence, including typical PD clinical symptoms, pharmacological responses, pathological hallmarks, and genetic mutations, strongly supported the identification of a monkey with spontaneous PD (Figure 1). To the best of our knowledge, this is the first reported case of naturally developed PD in animals. This suggests that PD is not a disease restricted to humans, with its existence in a non-human primate providing a novel evolutionary angle for understanding PD. As a close relative to humans (Buffalo et al, 2019; Phillips et al, 2014; Yan et al, 2011), this rare case of PD in another primate species provides solid evidence that monkeys are ideal candidates for the development of a genuine "animal version of PD", with conserved etiology and pathogenesis (Li et al, 2020). Furthermore, it allows us to compare similarities and differences in PD development between species and to understand PD pathogenesis from an evolutionary point of view.

Induction of core symptoms of autism spectrum disorder by in vivo CRISPR/Cas9-based gene editing in the brain of adolescent rhesus monkeys.

Although CRISPR/Cas9-mediated gene editing is widely applied to mimic human disorders, whether acute manipulation of disease-causing genes in the brain leads to behavioral abnormalities in non-human primates remains to be determined. Here we induced genetic mutations in MECP2, a critical gene linked to Rett syndrome (RTT) and autism spectrum disorders (ASD), in the hippocampus (DG and CA1-4) of adolescent rhesus monkeys (Macaca mulatta) in vivo via adeno-associated virus (AAV)-delivered Staphylococcus aureus Cas9 with small guide RNAs (sgRNAs) targeting MECP2. In comparison to monkeys injected with AAV-SaCas9 alone (n = 4), numerous autistic-like behavioral abnormalities were identified in the AAV-SaCas9-sgMECP2-injected monkeys (n = 7), including social interaction deficits, abnormal sleep patterns, insensitivity to aversive stimuli, abnormal hand motions, and defective social reward behaviors. Furthermore, some aspects of ASD and RTT, such as stereotypic behaviors, did not appear in the MECP2 gene-edited monkeys, suggesting that different brain areas likely contribute to distinct ASD symptoms. This study showed that acute manipulation of disease-causing genes via in vivo gene editing directly led to behavioral changes in adolescent primates, paving the way for the rapid generation of genetically engineered non-human primate models for neurobiological studies and therapeutic development.

In vivo Neuroregeneration to Treat Ischemic Stroke Through NeuroD1 AAV-Based Gene Therapy in Adult Non-human Primates.

Stroke may cause severe death and disability but many clinical trials have failed in the past, partially because the lack of an effective method to regenerate new neurons after stroke. In this study, we report an in vivo neural regeneration approach through AAV NeuroD1-based gene therapy to repair damaged brains after ischemic stroke in adult non-human primates (NHPs). We demonstrate that ectopic expression of a neural transcription factor NeuroD1 in the reactive astrocytes after monkey cortical stroke can convert 90% of the infected astrocytes into neurons. Interestingly, astrocytes are not depleted in the NeuroD1-converted areas, consistent with the proliferative capability of astrocytes. Following ischemic stroke in monkey cortex, the NeuroD1-mediated astrocyte-to-neuron (AtN) conversion significantly increased local neuronal density, reduced microglia and macrophage, and surprisingly protected parvalbumin interneurons in the converted areas. Furthermore, the NeuroD1 gene therapy showed a broad time window in AtN conversion, from 10 to 30 days following ischemic stroke. The cortical astrocyte-converted neurons showed Tbr1+ cortical neuron identity, similar to our earlier findings in rodent animal models. Unexpectedly, NeuroD1 expression in converted neurons showed a significant decrease after 6 months of viral infection, indicating a downregulation of NeuroD1 after neuronal maturation in adult NHPs. These results suggest that in vivo cell conversion through NeuroD1-based gene therapy may be an effective approach to regenerate new neurons for tissue repair in adult primate brains.

CircGRIA1 shows an age-related increase in male macaque brain and regulates synaptic plasticity and synaptogenesis.

Circular RNAs (circRNAs) are abundant in mammalian brain and some show age-dependent expression patterns. Here, we report that circGRIA1, a conserved circRNA isoform derived from the genomic loci of α-mino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor subunit Gria1, shows an age-related and male-specific increase in expression in the rhesus macaque prefrontal cortex and hippocampus. We show circGRIA1 is predominantly localized to the nucleus, and find an age-related increase in its association with the promoter region of Gria1 gene, suggesting it has a regulatory role in Gria1 transcription. In vitro and in vivo manipulation of circGRIA1 negatively regulates Gria1 mRNA and protein levels. Knockdown of circGRIA1 results in an age-related improvement of synaptogenesis, and GluR1 activity-dependent synaptic plasticity in the hippocampal neurons in males. Our findings underscore the importance of circRNA regulation and offer an insight into the biology of brain aging.

Age-associated changes in amyloid-ß and formaldehyde concentrations in cerebrospinal fluid of rhesus monkeys.

Rhesus monkeys ( Macaca mulatta) are valuable experimental animals for studies on neurodegenerative diseases due to their evolutionarily close relationship to humans (Zhang et al., 2014). Rhesus monkeys also display similar hallmarks of aging and neurodegeneration as humans, including formation of senile plaques in the brain (Beckman et al., 2019; Paspalas et al., 2018). However, changes in formaldehyde (FA) levels in the cerebrospinal fluid (CSF) of rhesus monkeys with aging have not been reported. Additionally, whether changes in CSF FA are correlated with changes in amyloid-ß (Aß) concentrations have not yet been explored. Here, the CSF levels of Aß 40, Aß 42, and FA were measured in 56 rhesus monkeys of different ages, ranging from 4 to 26 years old. Results revealed significant declines in Aß 40 and Aß 42, and an increase in FA with age. Interestingly, the increase in FA levels was negatively correlated with Aß 40 and Aß 42 concentrations in aged rhesus monkeys but not in young and middle-aged monkeys. These results appear to parallel changes seen within human aging, i.e., decreased levels of CSF Aß and increased levels of FA in normal aged adults and Alzheimer's disease (AD) patients. These findings further indicate that rhesus monkeys are a reliable model for studying age-related neurological disorders such as AD and suggest that FA is an important factor in AD development and may be used as a diagnostic indicator of such disease.

Potential use of actigraphy to measure sleep in monkeys: comparison with behavioral analysis from videography.

Sleep is indispensable for human health, with sleep disorders initiating a cascade of negative consequences. As our closest phylogenetic relatives, non-human primates (NHPs) are invaluable for comparative sleep studies and exhibit tremendous potential for improving our understanding of human sleep and related disorders. Previous work on measuring sleep in NHPs has mostly used electroencephalography or videography. In this study, simultaneous videography and actigraphy were applied to observe sleep patterns in 10 cynomolgus monkeys ( Macaca fascicularis) over seven nights (12 h per night). The durations of wake, transitional sleep, and relaxed sleep were scored by analysis of animal behaviors from videography and actigraphy data, using the same behavioral criteria for each state, with findings then compared. Here, results indicated that actigraphy constituted a reliable approach for scoring the state of sleep in monkeys and showed a significant correlation with that scored by videography. Epoch-by-epoch analysis further indicated that actigraphy was more suitable for scoring the state of relaxed sleep, correctly identifying 97.57% of relaxed sleep in comparison with video analysis. Only 34 epochs (0.13%) and 611 epochs (2.30%) were differently interpreted as wake and transitional sleep compared with videography analysis. The present study validated the behavioral criteria and actigraphy methodology for scoring sleep, which can be considered as a useful and a complementary technique to electroencephalography and/or videography analysis for sleep studies in NHPs.

Author Correction: 547 transcriptomes from 44 brain areas reveal features of the aging brain in non-human primates.

Following publication of the original paper [1], the authors reported an error in the affiliation of Xin-Tian Hu, who is also affiliated with "Kunming Primate Research Center, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan, China".

547 transcriptomes from 44 brain areas reveal features of the aging brain in non-human primates.

BACKGROUND: Brain aging is a complex process that depends on the precise regulation of multiple brain regions; however, the underlying molecular mechanisms behind this process remain to be clarified in non-human primates. RESULTS: Here, we explore non-human primate brain aging using 547 transcriptomes originating from 44 brain areas in rhesus macaques (Macaca mulatta). We show that expression connectivity between pairs of cerebral cortex areas as well as expression symmetry between the left and right hemispheres both decrease after aging. Although the aging mechanisms across different brain areas are largely convergent, changes in gene expression and alternative splicing vary at diverse genes, reinforcing the complex multifactorial basis of aging. Through gene co-expression network analysis, we identify nine modules that exhibit gain of connectivity in the aged brain and uncovered a hub gene, PGLS, underlying brain aging. We further confirm the functional significance of PGLS in mice at the gene transcription, molecular, and behavioral levels. CONCLUSIONS: Taken together, our study provides comprehensive transcriptomes on multiple brain regions in non-human primates and provides novel insights into the molecular mechanism of healthy brain aging.

Direct sunlight exposure reduces hair cortisol levels in rhesus monkeys (Macaca mulatta).

Major depressive disorder (MDD), commonly known as depression, is a mental disease characterized by a core symptom of low mood. It lasts at least two weeks (Badamasi et al., 2019; Wang et al., 2019) and is frequently accompanied by low self-esteem, loss of interest in routinely enjoyable activities, low energy, and unexplained pain (Huey et al., 2018; Park et al., 2012; Post & Warden, 2018; Rice et al., 2019; Xiao et al., 2018). Approximately 2%-8% of adults with MDD commit suicide (Richards & O'Hara, 2014; Strakowski & Nelson, 2015), and around half of suicidal individuals suffer depression or other mood disorders (Bachmann, 2018).

Comparative study of the transfection efficiency of commonly used viral vectors in rhesus monkey (Macaca mulatta) brains.

Viral vector transfection systems are among the simplest of biological agents with the ability to transfer genes into the central nervous system. In brain research, a series of powerful and novel gene editing technologies are based on these systems. Although many viral vectors are used in rodents, their full application has been limited in non-human primates. To identify viral vectors that can stably and effectively express exogenous genes within non-human primates, eleven commonly used recombinant adeno-associated viral and lentiviral vectors, each carrying a gene to express green or red fluorescence, were injected into the parietal cortex of four rhesus monkeys. The expression of fluorescent cells was used to quantify transfection efficiency. Histological results revealed that recombinant adeno-associated viral vectors, especially the serotype 2/9 coupled with the cytomegalovirus, human synapsin I, or Ca2+/calmodulin-dependent protein kinase II promoters, and lentiviral vector coupled with the human ubiquitin C promoter, induced higher expression of fluorescent cells, representing high transfection efficiency. This is the first comparison of transfection efficiencies of different viral vectors carrying different promoters and serotypes in non-human primates (NHPs). These results can be used as an aid to select optimal vectors to transfer exogenous genes into the central nervous system of non-human primates.

The RNA editome of Macaca mulatta and functional characterization of RNA editing in mitochondria.

RNA editing was first discovered in mitochondrial RNA molecular. However, whether adenosine-to-inosine (A-to-I) RNA editing has functions in nuclear genes involved in mitochondria remains elusive. Here, we retrieved 707,246 A-to-I RNA editing sites in Macaca mulatta leveraging massive transcriptomes of 30 different tissues and genomes of nine tissues, together with the reported data, and found that A-to-I RNA editing occurred frequently in nuclear genes that have functions in mitochondria. The mitochondrial structure, the level of ATP production, and the expression of some key genes involved in mitochondrial function were dysregulated after knocking down the expression of ADAR1 and ADAR2, the key genes encoding the enzyme responsible for RNA editing. When investigating dynamic changes of RNA editing during brain development, an amino-acid-changing RNA editing site (I234/V) in MFN1, a mediator of mitochondrial fusion, was identified to be significantly correlated with age, and could influence the function of MFN1. When studying transcriptomes of brain disorder, we found that dysregulated RNA editing sites in autism were also enriched within genes having mitochondrial functions. These data indicated that RNA editing had a significant function in mitochondria via their influence on nuclear genes.

Iron oxide nanoclusters for T 1 magnetic resonance imaging of non-human primates.

Iron-oxide-based contrast agents for magnetic resonance imaging (MRI) had been clinically approved in the United States and Europe, yet most of these nanoparticle products were discontinued owing to failures to meet rigorous clinical requirements. Significant advances have been made in the synthesis of magnetic nanoparticles and their biomedical applications, but several major challenges remain for their clinical translation, in particular large-scale and reproducible synthesis, systematic toxicity assessment, and their preclinical evaluation in MRI of large animals. Here, we report the results of a toxicity study of iron oxide nanoclusters of uniform size in large animal models, including beagle dogs and the more clinically relevant macaques. We also show that iron oxide nanoclusters can be used as T 1 MRI contrast agents for high-resolution magnetic resonance angiography in beagle dogs and macaques, and that dynamic MRI enables the detection of cerebral ischaemia in these large animals. Iron oxide nanoclusters show clinical potential as next-generation MRI contrast agents.

Neurons Differentiated from Transplanted Stem Cells Respond Functionally to Acoustic Stimuli in the Awake Monkey Brain.

Here, we examine whether neurons differentiated from transplanted stem cells can integrate into the host neural network and function in awake animals, a goal of transplanted stem cell therapy in the brain. We have developed a technique in which a small "hole" is created in the inferior colliculus (IC) of rhesus monkeys, then stem cells are transplanted in situ to allow for investigation of their integration into the auditory neural network. We found that some transplanted cells differentiated into mature neurons and formed synaptic input/output connections with the host neurons. In addition, c-Fos expression increased significantly in the cells after acoustic stimulation, and multichannel recordings indicated IC specific tuning activities in response to auditory stimulation. These results suggest that the transplanted cells have the potential to functionally integrate into the host neural network.