Rui M. Costa, PhD - US grants

Chronic stress biases behavioral strategies during decision-making: structural and physiological correlates The stress response is vital to maintain homeostasis. However, chronic stress can trigger maladaptive response and predispose to conditions ranging from neuropsychiatric disorders to everyday lapses of attention. Although previous reports have implicated chronic stress in executive function impairment, effects on decision-making processes remain to be clarified. Competing corticostriatal circuits are thought to control heterogeneous decision strategies. While the prelimbic (PL) cortex and the dorsomedial striatum (DMS, or associative striatum) have been implicated in goal-directed actions, the dorsolateral striatum (DLS, or sensorimotor striatum) has been implicated in automatic or habitual choices. We uncovered that in rats and mice chronic stress impairs the decision-making process, predisposing to habitual behavior in detriment to goal-directed strategies. We tested for action-outcome behavior in a lever pressing task and found that responses from rodents submitted to chronic stress became insensitive to both outcome devaluation and contingency degradation. Furthermore, we found that chronic stress causes opposing structural changes in associative and sensorimotor corticostriatal cricuits. Whereas chronic stress resulted in selective atrophy of pyramidal neurons in layer II/III of the PL and infralimbic (IL) sub-regions of medial prefrontal cortex (mPFC) and in medium spiny neurons (MSNs) of the DMS, it triggered an opposite effect in MSNs of the DLS. To determine if this structural reorganization of frontostriatal circuits has functional consequences, we recorded the simultaneous activity of neuronal ensembles in mPFC, DMS and DLS of control and stressed mice during behavioral training and testing. This approach will allow us to investigate if the changes in wiring observed in the associative and sensorimotor circuits underlies changes in neural activity in these circuits that could explain the bias from goal-directed towards habitual behavior observed in stressed subjects. Dynamic reorganization of striatal circuits via region and pathway-specific plasticity during the acquisition and consolidation of a skill Learning to execute and automatize certain actions is essential for survival. The learning of novel skills by trial and error, like riding a bicycle or playing a piano, is characterized by an initial stage of rapid improvement in performance, followed by a phase of more gradual improvements as the skills are consolidated and performance asymptotes (Kargo and Nitz, 2004;Karni et al., 1998;Miyachi et al., 2002;Miyachi et al., 1997). The different phases of skill learning have distinct behavioral and physiological hallmarks (Karni et al., 1998;Kleim et al., 2004;Muellbacher et al., 2002;Shiffrin and Schneider, 1977). For example, the early fast phase is susceptible to interference, while the later, more automatic phase is more resistant to interference (Shiffrin and Schneider, 1977). After the initial acquisitin phase, the memory of how to do things is gradually consolidated and for well-learned skills it can last a lifetime. Previous studies have shown changes in neural activity in the striatum, the major input nucleus of the basal ganglia, during motor and procedural learning (Barnes et al., 2005;Brasted and Wise, 2004;Carelli et al., 1997;Doyon et al., 1996;Jenkins et al., 1994;Ungerleider et al., 2002) . Some studies also suggested that the striatal circuits and processes engaged during the early and late phases of skill learning may differ (Costa et al., 2004;Miyachi et al., 2002;Miyachi et al., 1997). For example, the DMS, which receives input primarily from association cortices such as the prefrontal cortex (McGeorge and Faull, 1989;Voorn et al., 2004), seems to be preferentially involved in the initial stages of visuomotor learning annnd during the rapid acquisition of action-outcome contingencies (Miyachi et al., 2002;Miyachi et al., 1997;Yin et al., 2005). On the other hand, the DLS, which receives inputs from sensorimotor cortex (McGeorge and Faull, 1989;Voorn et al., 2004), is critical for the more gradual acquisition of habitual and automatic behavior (Miyachi et al., 2002;Miyachi et al., 1997;Yin et al., 2004). We have recently recorded neuronal activity in the DMS and DLS regions during the different stages of skill learning in vivo, and found that the task-related activity in these striatal regions differed during the acquisition and consolidation of a novel skill, with the DMS being engaged during the early phase, and the DLS during the late phase. We confirmed the differential involvement of these striatal regions in the different stages of skill learning using selective excitotoxic lesions of the dorsal striatum. We also investigated whether the changes in striatal neural activity observed during skill learning could be mediated by synaptic plasticity or excitability changes in medium spiny projection neurons in the dorsal striatum by using an ex vivo approach, and found that learning was accompanied by long-lasting changes in glutamatergic transmission. These changes evolved dynamically during the different phases of skill learning: changes in the DMS were predominant early in training, while changes in the DLS evolved only after extensive training. In summary, our previous studies indicated that during the automatization or consolidation of a skill, there is extensive potentiation of glutamatergic transmission in MSNs from the DLS, and that this region is necessary for the performance of automatized actions and skills. Neurons from the DLS do not all project to the same downstream basal ganglia structures. Medium spiny neurons projecting preferentially to the substantia nigra (striatonigral or direct pathway), and MSNs projecting to the external globus pallidus (striatopallidal or indirect pathway) have different dopamine receptor expression, different physiological properties, and different plasticity mechanisms (Gerfen et al., 1990;Kreitzer and Malenka, 2007;Shen et al., 2008;Shen et al., 2007). Until recently, LTP induction in the striatum was thought to always depend on D1-receptor activation (Kerr and Wickens, 2001), suggesting that it occurs preferentially in D1-expressing striatonigral neurons, and less in D2-expressing striatopalidal neurons. However, recent studies have shown that LTP can occur in both MSN types via different mechanisms (Shen et al., 2008). Thus, to understand the mechanisms underlying the consolidation and automatization of skills, it is crucial to determine whether the long-lasting potentiation observed in DLS after extended training occurs in striatonigral or striatopalidal MSNs, or in both. To this end we recorded from MSNs in the DLS of D2-EGFP mice and D1-EGFP mice that are naive or extensively trained on the rotarod. These mice allow the visualization of MSNs that express D1 receptors, which are almost exclusively striatonigral, and MSNs that express D2 receptors, which are almost exclusively striatopalidal, respectively. Using the D2-EGFP mice we obtained preliminary data indicating that extensive rotarod training resulted only in a slight potentiation in the non-D2 expressing neurons (putative striatonigral MSNs, direct pathway), but in a much greater potentiation in the D2-expressing MSNs (striatopallidal, indirect pathway) in the DLS of extensively trained animals. Concomitantly, the performance of the skill became less dependent on the activation of D1 receptors. These findings demonstrate that, as a skill becomes automatized, region- and pathway-specific plasticity sculpt the circuits involved in skill performance, and could elucidate why in Parkinsons disease voluntary movements are more affected than automatized movements.

Abstract (Administrative Core) The Administrative Core will oversee coordination and integration of all U19 Program functions; guide and facilitate interactions between the Project and Core leaders, Principal Investigators, and research staff; provide rigorous and regular fiscal oversight of the research projects and cores; ensure that the U19 TeamBCP maximizes the utilization of existing and established resources; and facilitate data, technology and information sharing and collaboration with key BRAIN Initiative stakeholders and the greater neuroscience research community. The core will be lead by our Team Directors, Rui Costa and Tom Jessell, and have an Internal Advisory Committee, a Data Science management sub-committee, an External Advisory Committee and a dedicated staff Program Manager. The core will a) provide overall coordination and support for the TeamBCP U19 research activities, b) foster growth of the Motor Control research within and beyond our research team, c) oversee and ensure resource dissemination and outreach, and d) fiscal and administrative management of cores and research projects. The Internal Advisory Committee will meet monthly to discuss all administrative issues and to provide scientific direction for the program. Drs. Costa and Jessell will co- chair these meetings. The needs, usage and effectiveness of the Cores will be assessed at these meetings and any obstacles to progress will be identified. Each Core, including the Administrative Core, will present a status update on any outstanding scientific, administrative or budgetary issues that require immediate attention. These regular meetings will ensure that the program can address problems in a timely manner. They will also serve to make decisions about the resources, priorities, efforts and data access for the whole program, and to resolve potential conflicts. The Internal Data Science Subcommittee will meet at least once a year, with more frequent ad hoc meetings with individual advisory members to review Data Science Core specific issues and report back to the IAC.

Understanding the mechanisms that the nervous system uses to control movement is critical for understanding brain and behavior, and one of the fundamental questions in neuroscience. The control of movement emerges from the activity of different motor control centers, that converge onto output systems, mostly located in the spinal cord. While the spinal circuits that underlie different aspects of motor control have been relatively well characterized, the way by which these circuits are coordinated by supraspinal motor control centers remains elusive. In this project, we aim to understand the functional and computational logic of connectivity between a motor control centers, the motor cortex, and the spinal cord and muscle. We will anatomically and functionally characterize the role of projection-specific populations of corticospinal neurons during particular modes of motor control. Because even the simplest motor program requires the activation of many neuronal populations across multiple brain areas, we will also investigate the contribution of other cortical and subcortical areas to the output of the brain to the spinal cord, and to muscle activity. This understanding requires It also requires extracting the information that is carried between brain areas and neuronal cell types, and understanding the computations that are operated in the circuits in order to achieve specific patterns of muscle activation. We will extract computational principles governing the relation between brain activity and muscle activity that are conserved between rodents and , and will construct predictive models of . In order to achieve a mechanistic understanding of the brain circuits underlying motor control, we will dissect the contributions of activity in specific neural populations using closed-loop optogenetic manipulations. The level of understanding that we are seeking requires a dynamic back and forth between anatomical and functional mapping experiments, computational and conceptual models, and causal testing of predictions. We put together a a multidisciplinary team of PIs working in a tight network, sharing the latest technologies to measure and manipulate the brain through an Advanced Imaging and Instrumentation core, creating and refining circuit models based on data that generate testable predictions, and establishing real-time knowledge exchange between team members through a Data Science Core. Our U19BCP Motor Control team proposes a comprehensive and ambitious project to establish the computational and circuit mechanisms underlying classical modes of motor control based on cell-type specific connectivity between brain and spinal cord, novel technology to measure and manipulate functionally and genetically-defined neural populations, and state-of-the-art computational tools. primates multi-area dynamics during motor control

Abstract The ability to understand neural circuit mechanisms underlying behavior and motor control is critically dependent on our ability to modulate the activity of functional neural circuits reversibly, and with single cell precision. This project will causally test the predictions from Projects1-4 and in turn inform the models, data analyses and data collection in other Projects 2-4. For a clear understanding of the circuit mechanisms underlying motor control, cell-specific cortical and subcortical areas will be optogenetic manipulated through the use of closed-loop paradigms to directly assess the contributions of particular neural populations, or particular types of activity dynamics, on motor control. We will use 3 types of closed-loop operant paradigms: i) closed-loop paradigms where we optogenetically manipulate the activity of specific neuronal populations based on the behavioral state, ii) closed-loop paradigms where we optogenetically manipulate the activity of specific neuronal populations based on their activity, and iii) closed-loop paradigms where the neural activity will produce a specific behavioral outcome. We will test the contribution of cortical and subcortical areas to the activity of specific CSN populations and motor behavior by triggering the optogenetic manipulations on particular epochs of the behavior. We will also characterize the role of functionally identified neurons by using holographic optogenetic manipulations triggered by the neural activity patterns. Finally, we will use optical closed-loop operant brain-machine paradigms to investigate how upstream neural populations contribute to activity of corticospinal neurons versus non-corticospinal neurons in motor cortex. These experiments will test with multiple approaches the same hypotheses, and will permit us to causally establish causal connections between neural activity in different areas, and neural activity and behavior. They will provide unvaluable knowledge about the role of particular corticospinal neurons with specific projection patterns or with specific activity patterns, and also inform us about the identity and the contributions of cortical and subcortical neural populations to corticospinal activity, and to movement.

Abstract Adaptive control of behavior is critical for survival. Even a simple movement, like extending the arm, requires the activation of many neuronal populations across the nervous system. Our lab has used a combination of anatomical, genetic, optical and behavioral approaches to unravel how animals move, and learn to control movement. However, adaptive responses are not effected only through muscles, but also through other organs. For example, planning to pick an apple will trigger not only muscle activity but also the expectation of food, and the conditioned release of insulin. Hence adaptive behavior requires the coordination of an organism's actions with its physiological internal states. We propose to leverage our expertise to dissect the neural circuits and principles governing the learning and adaptive ?motor? control of internal organ function. We will spearhead this new research direction by investigating conditioned insulin release and conditioned immunosuppression, mediated by the innervation of the pancreas and spleen, respectively. We will leverage state of the art viral and RNA-seq approaches to map with high-resolution the first, second and third-order innervation of spleen and pancreas. Our preliminary anatomical mapping of the innervation of these organs revealed that different populations of celiac-mesenteric ganglia sympathetic neurons innervate pancreas versus spleen. Remarkably, most innervation of the thoracic preganglionic spinal cord targeting these organs emerges from the cortex: motor cortex, but also sensory and prefrontal. We therefore hypothesize that learning to select the appropriate responses in internal organs after conditioning is mediated by higher-order brain circuits, and follows principles similar to those used for motor responses. We propose to use both targeted and unbiased approaches to identify and manipulate the activity of descending neural populations responsible for the learned control of spleen and pancreatic function. This new line of research is innovative but trackable with our expertise, and the Pioneer award support will help us attack this novel research area. Importantly, the proposed research has the potential to conceptually position the nervous system as a ?smart? regulator of organism homeostasis, and hence impact health in unexpected ways - mental disorders like anxiety and depression, or neurological problems like stroke, are associated with abnormal physiological states likely emerging from these brain-internal organ interactions.