Chemogenetic dissection of the primate prefronto-subcortical pathways for working memory and decision-making

「何を買うんだっけ」と「どれにしよう」を処理する2つの脳回路を明らかに --霊長類の生体脳で神経経路を可視化・操作する技術で解明、高次脳機能の理解へ大きく前進--. 京都大学プレスリリース. 2021-06-24.The primate prefrontal cortex (PFC) is situated at the core of higher brain functions via neural circuits such as those linking the caudate nucleus and mediodorsal thalamus. However, the distinctive roles of these prefronto-subcortical pathways remain elusive. Combining in vivo neuronal projection mapping with chemogenetic synaptic silencing, we reversibly dissected key pathways from dorsolateral part of the PFC (dlPFC) to the dorsal caudate (dCD) and lateral mediodorsal thalamus (MDl) individually in single monkeys. We found that silencing the bilateral dlPFC-MDl projections, but not the dlPFC-dCD projections, impaired performance in a spatial working memory task. Conversely, silencing the unilateral dlPFC-dCD projection, but not the unilateral dlPFC-MDl projection, altered preference in a decision-making task. These results revealed dissociable roles of the prefronto-subcortical pathways in working memory and decision-making, representing the technical advantage of imaging-guided pathway-selective chemogenetic manipulation for dissecting neural circuits underlying cognitive functions in primates

Characteristics of unique endocytosis induced by weak current for cytoplasmic drug delivery

We previously reported that 20 a weak current (WC, 0.3-0.5mA/cm2) applied to cells can induce endocytosis to promote cytoplasmic delivery of hydrophilic macromolecules (MW: < 70,000), such as dextran and siRNA, which leak from WC-induced endosomes into the cytoplasm (Hasan et al., 2016). In this study, we evaluated the characteristics of WC-mediated endocytosis for application of the technology to cytoplasmic delivery of macromolecular medicines. WC induced significantly higher cellular uptake of exogenous DNA fragments compared to untreated cells; the amount increased in a time-dependent manner, indicating that endocytosis was induced after WC. Moreover, following WC treatment of cells in the presence of an antibody (MW: 150,000) with the lysosomotropic agent chloroquine, the antibody was able to bind to its intracellular target. Thus, high molecular weight protein medicines delivered by WC-mediated endocytosis were functional in the cytoplasm. Transmission electron microscopy of cells treated by WC in the presence of gold nanoparticles covered with polyethylene glycol showed that the WC-induced endosomes exhibited an elliptical shape. In the WC-induced endosomes, ceramide, which makes pore structures in the membrane, was localized. Together, these results suggest that WC can induce unique endocytosis and that macromolecular medicines leak from endosomes through a ceramide pore

Chemogenetic attenuation of cortical seizures in nonhuman primates

「てんかん」の発生を時間的・空間的にピンポイントで抑える画期的な治療法を開発 --世界で初めてサルでの有効性を実証、臨床応用に向け大きく前進--. 京都大学プレスリリース. 2023-03-01.Epilepsy is a disorder in which abnormal neuronal hyperexcitation causes several types of seizures. Because pharmacological and surgical treatments occasionally interfere with normal brain function, a more focused and on-demand approach is desirable. Here we examined the efficacy of a chemogenetic tool—designer receptors exclusively activated by designer drugs (DREADDs)—for treating focal seizure in a nonhuman primate model. Acute infusion of the GABAA receptor antagonist bicuculline into the forelimb region of unilateral primary motor cortex caused paroxysmal discharges with twitching and stiffening of the contralateral arm, followed by recurrent cortical discharges with hemi- and whole-body clonic seizures in two male macaque monkeys. Expression of an inhibitory DREADD (hM4Di) throughout the seizure focus, and subsequent on-demand administration of a DREADD-selective agonist, rapidly suppressed the wide-spread seizures. These results demonstrate the efficacy of DREADDs for attenuating cortical seizure in a nonhuman primate model

Abnormal axon guidance signals and reduced interhemispheric connection via anterior commissure in neonates of marmoset ASD model

In autism spectrum disorder (ASD), disrupted functional and structural connectivity in the social brain has been suggested as the core biological mechanism underlying the social recognition deficits of this neurodevelopmental disorder. In this study, we aimed to identify genetic and neurostructural abnormalities at birth in a non-human primate model of ASD, the common marmoset with maternal exposure to valproic acid (VPA), which has been reported to display social recognition deficit in adulthood. Using a comprehensive gene expression analysis, we found that 20 genes were significantly downregulated in VPA-exposed neonates. Of these, Frizzled3 (FZD3) and PIK3CA were identified in an axon guidance signaling pathway. FZD3 is essential for the normal development of the anterior commissure (AC) and corpus callosum (CC); hence, we performed diffusion tensor magnetic resonance imaging with a 7-Tesla scanner to measure the midsagittal sizes of these structures. We found that the AC size in VPA-exposed neonates was significantly smaller than that in age-matched controls, while the CC size did not differ. These results suggest that downregulation of the genes related to axon guidance and decreased AC size in neonatal primates may be linked to social brain dysfunctions that can happen later in life

Disturbed Vocal Communication in Common Marmoset Family with an Autism-Model Child

Introduction: Autism spectrum disorder (ASD) is one of the most common developmental disorders. Most children with ASD demonstrate delays in language development. Parents of these children may also feel frustration and stress because they cannot communicate with their children (Hastings, 2002). In rodents, exposure embryos to valproic acid (VPA), known as the risk factor of ASD in human (Christensen, 2013), are widely used as an animal model (Crawley, 2012). Here we show that common marmoset (Callithrix jacchus) families with a juvenile with fatal exposure to VPA show vocalization abnormality.Methods: The experiment was conducted using 16 children of marmosets (9 VPA exposed, and 7 unexposed (UE)) consisting of one or two juveniles (ranging from 1 to 5 months of age) and their respective parents. In the families with VPA exposed children, the dams received seven oral administrations of sodium valproate at 200mg/kg/day from day 60 to 66 after conception. To evaluate the kinship social communication, family (one juvenile and its parents) vocalizations were recorded for 30 minutes in their home cage carried at shield room. Vocal spectrogram was used to define and count 9 call types. All procedures were approved by the Animal Research Committee at the National Institute of Neuroscience in Tokyo, Japan.Results & Discussion: There were significant differences of call-uses between VPA and UE families. In VPA families, social isolation call “phee” frequency and agitation call “twitter” was increased, on the other hand, “trill” call frequency, supposed ‘felling affinity’, was decreased. These two groups were significantly discriminated by outlier analysis based on Mahalanobis’ distance. It is suggested that quantitative analysis of verbal communication in family with ASD children could be useful and noninvasive criterion of patient himself and his family.24th International "Stress and Behavior" Neuroscience and Biopsychiatry Conferenc

Social context segmentation analysis in a primate model of Autism Spectrum Disorder

社会的状況における個体間相互作用を理解する事を目指し、(1)自由運動中の身体動作の計測系開発、(2)行動時系列の分節構造解析の開発を行い、(3)脳機能制御を行なったマーモセットの社会行動の評価を進めている。本発表では、(1)について実装レベルに達した結果を報告し、(3)化学遺伝学手法を用いた脳機能制御について現段階での成果をまとめた。次世代脳シンポジウム+新学術班会

Unsupervised segmentation of the contextual structure inherent in goal-directed behavior in non-human primates

Conventional behavioral ethograms rely on experts’ estimation and labeling of specific time-series semantic structures inherent in human and animal behavior. Computational approach is expected to significantly improve the process of behavioral analysis and provide objective measure. In this study we segmented a continuous behavioral sequence in feeding behavior of common marmosets into repetitively observed units using a state-space model that incorporates unsupervised machine learning classification and nonlinear regression. We focused on the behavior because it supposed to have a specific semantic structure in the flow of actions to achieve the goal. A total of 1000-sec time series of feeding behavior of four marmosets was clearly divided into a sequence of 11 different segments. Characteristic segments appeared before and after the feeding timing, and the transition probabilities of these segments allowed us to classify three different feeding patterns. These results suggest that the proposed computational method has the potential to classify and quantitatively compare behavioral ethograms without subjective observation. It has opened the path to understand complex and ambiguous behavioral repertories in social communication and interaction, which are difficult to analyze using conventional methods, as statistical generative models.The 81st Annual meeting of Japanese society for animal psycholog

Familiarity perception call elicited under restricted sensory cues in peer-social interactions of the domestic chick.

Social cognitive mechanisms are central to understanding developmental abnormalities, such as autistic spectrum disorder. Peer relations besides parent-infant or pair-bonding interactions are pivotal social relationships that are especially well developed in humans. Cognition of familiarity forms the basis of peer socialization. Domestic chick (Gallus gallus) studies have contributed to our understanding of the developmental process in sensory-motor cognition but many processes remain unknown. In this report, we used chicks, as they are precocial birds, and we could therefore focus on peer interaction without having to consider parenting. The subject chick behavior towards familiar and unfamiliar reference peers was video-recorded, where the subject and the reference were separated by either an opaque or transparent wall. Spectrogram and behavior correlation analyses based on principal component analysis, revealed that chicks elicited an intermediate contact call and a morphologically different distress call, more frequently towards familiar versus unfamiliar chicks in acoustic only conditions. When both visual and acoustic cues were present, subject chicks exhibited approaching and floor pecking behavior, while eliciting joyful (pleasant) calls, irrespective of whether reference peers were familiar or unfamiliar. Our result showed that chicks recognized familiarity using acoustic cues and expressed cognition through modified distress calls. These finding suggests that peer affiliation may be established by acoustic recognition, independent of visual face recognition, and that eventually, both forms of recognition are integrated, with modulation of acoustic recognition