Search

Background An early-life anxious temperament (AT) is a risk factor for the development of anxiety, depression, and comorbid substance abuse. We validated a nonhuman primate model of early-life AT and identified the dorsal amygdala as a core component of AT's neural circuit. Here, we combine RNA sequencing, viral-vector gene manipulation, functional brain imaging, and behavioral phenotyping to uncover AT's molecular substrates. Methods In response to potential threat, AT and brain metabolism were assessed in 46 young rhesus monkeys. We identified AT-related transcripts using RNA-sequencing data from dorsal amygdala tissue (including central nucleus of the amygdala [Ce] and dorsal regions of the basal nucleus). Based on the results, we overexpressed the neurotrophin-3 gene, NTF3, in the dorsal amygdala using intraoperative magnetic resonance imaging–guided surgery (n = 5 per group). Results This discovery-based approach identified AT-related alterations in the expression of well-established and novel genes, including an inverse association between NTRK3 expression and AT. NTRK3 is an interesting target because it is a relatively unexplored neurotrophic factor that modulates intracellular neuroplasticity pathways. Overexpression of the transcript for NTRK3's endogenous ligand, NTF3, in the dorsal amygdala resulted in reduced AT and altered function in AT's neural circuit. Conclusions Together, these data implicate neurotrophin-3/NTRK3 signaling in the dorsal amygdala in mediating primate anxiety. More generally, this approach provides an important step toward understanding the molecular underpinnings of early-life AT and will be useful in guiding the development of treatments to prevent the development of stress-related psychopathology.

INTRODUCTION The spatial and temporal resolution limits of functional MRI (fMRI) brain mapping provide primarily correlative information on brain connectivity. Determining how one region causally modulates and mediates activity in other regions remains difficult with fMRI. We demonstrate a simple means to add causality in resting state functional connectivity MRI (rs-fcMRI) using techniques developed first for convection-enhanced delivery (CED) of therapeutics. Here we use CED to guide and monitor pharmacologic alteration of a local brain region in anaesthetized Rhesus monkeys while monitoring rs-fcMRI signal changes. METHODS Pre-surgical MRI was used to determine skull locations for craniotomies for installation of NavigusTMbrain ports in two untrained monkeys slated for euthanasia. The ports were aligned in real–time to provide trajectories aimed at the central nucleus of the amygdala (CeA). Fused silica catheters were then inserted into the CeA where 24 mg of muscimol (inhibitory agent) was infused in 24 ml of buffered solution under pressure over 12 minutes, first on the right side and then on the left side. rs-fcMRI studies were done for 45 minutes before and after the unilateral infusion and then after the bilateral infusion. RESULTS Catheters were successfully aligned and inserted into the CeA targets with sub-mm accuracy. T2-weighted imaging detected the enhanced T2 from the infusion’s buffer. Pre-infusion rs-fcMRI provides results consistent with prior studies, which have shown that the CeA is most strongly connected to the contralateral CeA. This connectivity was significantly reduced following both unilateral and bilateral injections of muscimol into the CeA, demonstrating the effectiveness of the muscimol infusions. Conditional Grainger Causality (CGC) analysis shows unexpected new connectivity after the unilateral infusion. Upon the bilateral infusion, global effective connectivity in the region is reduced. CONCLUSION Expected and unexpected changes in resting state functional connectivity resulted from unilateral and bilateral infusions of inhibitory agents.

Background Nonhuman primate models are critical for understanding mechanisms underlying human psychopathology. We established a nonhuman primate model of anxious temperament (AT) for studying the early-life risk to develop anxiety and depression. Studies have identified the central nucleus of the amygdala (Ce) as an essential component of AT's neural substrates. Corticotropin-releasing factor (CRF) is expressed in the Ce, has a role in stress, and is linked to psychopathology. Here, in young rhesus monkeys, we combined viral vector technology with assessments of anxiety and multimodal neuroimaging to understand the consequences of chronically increased CRF in the Ce region. Methods Using real-time intraoperative magnetic resonance imaging-guided convection-enhanced delivery, five monkeys received bilateral dorsal amygdala Ce-region infusions of adeno-associated virus serotype 2 containing the CRF construct. Their cagemates served as unoperated control subjects. AT, regional brain metabolism, resting functional magnetic resonance imaging, and diffusion tensor imaging were assessed before and 2 months after viral infusions. Results Dorsal amygdala CRF overexpression significantly increased AT and metabolism within the dorsal amygdala. Additionally, we observed changes in metabolism in other AT-related regions, as well as in measures of functional and structural connectivity. Conclusions This study provides a translational roadmap that is important for understanding human psychopathology by combining molecular manipulations used in rodents with behavioral phenotyping and multimodal neuroimaging measures used in humans. The results indicate that chronic CRF overexpression in primates not only increases AT but also affects metabolism and connectivity within components of AT's neural circuitry.

Induced pluripotent stem cell (iPSC)-derived neurons represent an opportunity for cell replacement strategies for neurodegenerative disorders such as Parkinson's disease (PD). Improvement in cell graft targeting, distribution, and density can be key for disease modification. We have previously developed a trajectory guide system for real-time intraoperative magnetic resonance imaging (RT-IMRI) delivery of infusates, such as viral vector suspensions for gene therapy strategies. Intracerebral delivery of iPSC-derived neurons presents different challenges than viral vectors, including limited cell survival if cells are kept at room temperature for prolonged periods of time, precipitation and aggregation of cells in the cannula, and obstruction during injection, which must be solved for successful application of this delivery approach. To develop procedures suitable for RT-IMRI cell delivery, we first performed in vitro studies to tailor the delivery hardware (e.g., cannula) and defined a range of parameters to be applied (e.g., maximal time span allowable between cell loading in the system and intracerebral injection) to ensure cell survival. Then we performed an in vivo study to evaluate the feasibility of applying the system to nonhuman primates. Our results demonstrate that the RT-IMRI delivery system provides valuable guidance, monitoring, and visualization during intracerebral cell delivery that are compatible with cell survival.