2009 — 2010 |
Feng, Guoping |
RC1Activity Code Description: NIH Challenge Grants in Health and Science Research |
Optogenetic Mice For Cell Type-Specific Manipulation of Neuronal Activity @ Massachusetts Institute of Technology
DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (06) Enabling Technologies and specific Challenge Topic, 06-MH-102: Technologies to study neuronal signaling, plasticity, and neurodevelopment. The complex and diverse functions of the brain depend on the unique properties of neural circuits formed by various subtypes of neurons with distinct molecular and/or electrical properties. Furthermore, many neurological disorders are often due to the dysfunction of specific subsets of neurons or neural circuits. Thus, elucidating the unique roles of each subtype of neurons in shaping circuitry function is critical for our understanding of both normal and abnormal brain functions. Genetic tools incorporating spatial and temporal control over neural activity in neuronal subsets would greatly enhance our capability to precisely map circuitry function and dysfunction in the brain. Manipulating activity in this way requires a tool that can be genetically targeted to specific populations of neurons and that allows simple and rapid control of neuronal firing. This has been made possible by the recent development of the genetically encoded light-activated cation channel channelrhodopsin-2 (ChR2) for photoactivation and the light-driven chloride pump halorhodopsin (NpHR) for photoinhibition. Recent studies from several laboratories have highlighted the tremendous potentials of using ChR2 and NpHR in mapping neuronal connectivity and manipulating circuitry function. The goal of this research proposal is to fully realize the potentials of these optogenetic tools by generating a series of transgenic mice that express improved ChR2 and NpHR selectively in molecularly defined subtypes of neurons in the brain, thus providing a set of powerful optogenetic mice for interrogating brain circuitry function and dysfunction using high-speed photostimulation and photoinhibition in brain slices and in vivo. PUBLIC HEALTH RELEVANCE: Abnormal neuronal connectivity and neuronal activity in the brain contribute to many neurological and neuropsychiatric disorders such as epilepsy and autism. The goal of this research proposal is to develop new genetic tools that will allow neuroscientists to manipulate neuronal activity in mouse models of human diseases, thus helping to elucidate the pathogenic mechanisms of neurological and neuropsychiatric disorders.
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0.921 |
2012 — 2013 |
Feng, Guoping |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Cell Type-Specific Halorhodopsin Mice For Neuronal Silencin @ Massachusetts Institute of Technology
DESCRIPTION (provided by applicant): The complex and diverse functions of the brain depend on the unique properties of neural circuits formed by various subtypes of neurons with distinct molecular and/or electrical properties. Furthermore, many neurological and psychiatric disorders are often due to the dysfunction of specific subsets of neurons or neural circuits. Thus, elucidating the unique roles of each subtype of neurons in shaping circuitry function is critical t our understanding of both normal and abnormal brain functions. Genetic tools that incorporating spatial and temporal control over neural activity in neuronal subsets would greatly enhance our capability to precisely map circuitry function and dysfunction in the brain. Manipulating activity n this way requires a tool that can be genetically targeted to specific populations of neurons and that allows simple and rapid control of neuronal firing. This has been made possible by the recent development of the genetically encoded light-activated cation channel channelrhodopsin-2 (ChR2) for photoactivation and the light-driven chloride pump halorhodopsin (NpHR) for photoinhibition. Recent studies from several laboratories have highlighted the tremendous potentials of using ChR2 and NpHR in mapping neuronal connectivity and manipulating circuitry function. The goal of this research proposal is to generate a series of transgenic mice that express improved NpHR selectively in molecularly defined subtypes of neurons in the brain. Together with our recently generated cell type-specific ChR2 transgenic mice, it will provide a set of powerful genetic tools for interrogating brain circuitry function and dysfunction using high speed photostimulation and photoinhibition in brain slices and in vivo.
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0.921 |
2012 — 2016 |
Feng, Guoping |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Shank3 in Synaptic Function and Autism @ Massachusetts Institute of Technology
DESCRIPTION (provided by applicant): Recent genetic and genomic studies have identified a large number of candidate genes for autism spectrum disorders (ASDs), many of which encode synaptic proteins, suggesting synaptic dysfunction may play a critical role in ASDs. One of the most promising ASD candidate genes is Shank3. Shank family proteins (Shank1-3) directly bind SAPAP to form the PSD95/SAPAP/Shank complex. This core of proteins is thought to function as a scaffold, orchestrating the assembly of the macromolecular postsynaptic signaling complex. They have been proposed to play important roles in trafficking and anchoring of postsynaptic ionotropic glutamate receptors and the development of glutamatergic synapses. Shank3 is the only member of the Shank family highly expressed in the striatum, a brain region strongly implicated in ASDs. To investigate the in vivo function of Shank3 at synapses and to elucidate how a disruption of Shank3 may lead to ASDs, we generated Shank3 mutant mice. We found that disruption of Shank3 resulted in both structural and functional changes in cortico-striatal synapses. Furthermore, Shank3 mutant mice exhibit compulsive/repetitive behavior and impaired social interaction, which resemble two of the cardinal features of ASDs. Together, our studies demonstrate a critical role for Shank3 in cortico-striatal synaptic structure and function n vivo and establish causality between a disruption in the Shank3 gene and the genesis of autistic like-behaviors in mice. Thus, the Shank3 mutant mice provide us with an excellent opportunity to dissect the neural circuitry mechanisms underlying the abnormal behaviors relevant to human ASD. We propose to combine genetic, optogenetic, electrophysiological and behavioral approaches to achieve the following goals: (1) To investigate the intrastriatal microcircuitry dysfunction in Shank3 mutant mice. (2) To determine the relative contributions of the direct and indirect pathway of the basal ganglia in repetitive behavior. (3) To dissect neural circuits involved in social interaction deficits in Shank3 mutant mice. Together, these studies may significantly enhance our understanding of neural circuitry mechanisms of autistic-like behaviors and may help to develop novel strategies for more effective treatment.
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0.921 |
2016 — 2020 |
Feng, Guoping Halassa, Michael M (co-PI) [⬀] |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Dissecting the Role of Thalamic Inhibition in Neurodevelopmental Diseases @ Massachusetts Institute of Technology
PROJECT SUMMARY/ABSTRACT Sensory abnormalities characterize a wide range of neurodevelopmental disorders. In autism spectrum disorder (ASD), for example, sensory overload is one of the most frequently reported symptoms. Abnormal regulation of sensory information flow (sensory gating) is also observed in schizophrenia and ADHD, and is thought to contribute to overall cognitive dysfunction across all these conditions. Despite its central importance, little is known about the neurobiology of sensory gating, and even less is known about its failure in disease. This proposal aims to address this critical gap. The neocortex is requires for higher level sensory processing, but early processing and transmission of sensory information is performed by the thalamus. We and others have found that thalamic sensory input is controlled by the thalamic reticular nucleus (TRN), a shell of GABAergic neurons surrounding thalamic relay nuclei. The TRN is composed of individual subnetworks, each controlling thalamic flow in a modality-specific manner. Recent clinical data have shown thalamic and TRN dysfunction in neurodevelopmental disorders. Given the critical role for TRN in sensory processing, we expect perturbations in its circuits to pathologically augment cortical sensory input, explaining several clinical symptoms. In sleep, TRN dysfunction may result in increased sensory-related arousals, while in attention irrelevant inputs may become much more distracting. As such, a `leaky thalamus' may have profound consequences on behavior and cognition across disorders. In this proposal, we will test the leaky thalamus framework by manipulating thalamic inhibition in mice while monitoring the impact on sensory function and related behaviors. In addition, we will investigate the therapeutic potential of reversing thalamic inhibition deficits in models of human neurodevelopmental disorders.
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0.921 |
2017 — 2021 |
Feng, Guoping |
U01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
A Molecular and Cellular Atlas of the Marmoset Brain @ Massachusetts Institute of Technology
PROJECT SUMMARY/ABSTRACT The complexity of the mammalian brain is unparalleled by any other organ, and understanding its cellular composition is essential to understand how it gives rise to cognition and behavior. It is clear that brain contains many more cell types than have been described to date. Many cell types can now be distinguished by their patterns of gene expression, and knowledge of these patterns can provide genetic access to specific populations of neurons. The ability to manipulate and measure activity in genetically defined cell types and circuits will allow us to move from a static anatomical description to a dynamic understanding of brain function. Although genetic tools have dramatically advanced our understanding of brain function, they have largely been confined to mice. While mice are essential models for many areas of neuroscience, there are also many aspects of higher brain function that cannot be adequately modeled in rodents. Similarly, many brain disorders affect higher cognitive functions that have no clear parallels in rodents. There is thus an urgent need for new genetic models that are phylogenetically closer to humans. A promising emerging primate model is the common marmoset, a small new world primate that has many advantages for neuroscience and genetic research. In the past three years, we have established a large marmoset colony and a genetic engineering platform at MIT to generate marmoset genetic models for various brain disorders. We have successfully demonstrated efficient gene knockout and knockin techniques in marmoset embryos. We have also generated and assembled high quality marmoset genome sequence. In addition, we are developing hardware and software for automated behavioral analysis as well as electrophysiological recording and multi-photon imaging approaches. Our goal is to make this a national technology and resource center for using marmosets to study brain function and dysfunction. Here propose to add another important dataset to this potentially transformative model?using single cell RNAseq to systematically define cell types, their location and morphology. These data will be critical for generating cell type-specific genetic tools as well as for monitoring and manipulating circuit activity in a cell type-specific manner, key approaches to understand brain function and dysfunction.
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0.921 |
2018 — 2021 |
Feng, Guoping |
U24Activity Code Description: To support research projects contributing to improvement of the capability of resources to serve biomedical research. |
Knockin Marmoset Reporters For Non-Invasive Measuring of Genome-Editing Efficiency @ Massachusetts Institute of Technology
Project Summary The past decade has seen unprecedented advances in genome editing technologies and the approval of several targeted gene therapies by the Food & Drug Administration. As such, targeted endonuclease-based approaches to gene therapy now hold realistic promise for the widespread treatment and reversal of a large number of human diseases. Although early versions of CRISPR/Cas systems suffered from problems with efficiency and specificity, recent advances achieved through directed-evolution, targeted protein engineering, and the identification of editing enhancers have put widespread use of gene therapy within reach. However, one major hurdle between the basic science and the implementation of safe and effective clinical interventions is a lack of suitable large animal models for pre-clinical testing of new technologies and therapeutic strategies. Here, we propose a strategy to generate two reporter lines in the common marmoset (Callithrix jacchus) that will be used for monitoring of both on- and off-target editing by Cas9 and adenine base editors at single-cell resolution. Large animal reporters must be highly efficient and capable of testing multiple aspects of gene editing if they are to be broadly useful and capable of overcoming the slow sexual maturation and gestational periods of primates. Taking this into consideration, we will utilize well-established fluorescent reporters in conjunction with a newly-developed variant of luciferase called Akaluc that can be used for non-invasive bioluminescent imaging in marmosets with single-cell sensitivity. We will generate an initial set of two marmoset reporters capable of testing knock-in and adenine base editing in a non-invasive manner through on-target editing-mediated activation of Venus-Akaluc expression. Following initial screening for on-target editing via whole-body imaging with an electron-multiplying charge-couple device (EMCCD) camera, tissue can be collected from these reporter animals and used for single-cell analyses of both on- and off-target editing through fluorescence activated cell sorting (FACS) and single-cell sequencing. Our approach establishes a versatile and efficient platform for testing editing technologies in a non-human primate species and can be easily and rapidly adapted to evolving technologies, making it a valuable system for accelerating the development of safe and effective gene therapies.
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0.921 |
2018 — 2021 |
Feng, Guoping |
P50Activity Code Description: To support any part of the full range of research and development from very basic to clinical; may involve ancillary supportive activities such as protracted patient care necessary to the primary research or R&D effort. The spectrum of activities comprises a multidisciplinary attack on a specific disease entity or biomedical problem area. These grants differ from program project grants in that they are usually developed in response to an announcement of the programmatic needs of an Institute or Division and subsequently receive continuous attention from its staff. Centers may also serve as regional or national resources for special research purposes. |
Res Support Core
Abstract The work to be done in the laboratory of Dr. Guoping Feng at MIT covers the following activities: 1. Provide Shank3 ASD mouse models and assistance for investigating PV neuron dysfunction. In addition to the Shank3 knockout mice we originally shared with Takao Hensch, we have recently developed two new Shank3 models: a conditional Shank3 knockin mouse that allows us to restore Shank3 expression at any developmental stages with Cre-ER, and a conditional Shank3 knockout mouse that allows us to delete Shank3 at any developmental stages with Cre-ER. We will provide these mice for a variety of structural and functional analysis. 2. Develop and provide genetic tools and resources for studying PV neuron function in marmosets. As a Core Facility, we will provide marmoset brain tissues of various developmental stages for studying PV neuron development and function in a primate brain. We will develop GABAergic neuron- specific AAV system for gene manipulations and calcium imaging. Most importantly, we will provide PV neuron-specific knockin marmoset (on-going effort) for viral-mediated, Cre-dependent manipulation of gene function and neuronal imaging in PV neurons. 3. Develop and characterize marmoset models for studying PV neuron dysfunction in ASD. A recent collaborative work between the laboratories of Hensch and Feng using mouse models suggested that PV neuron defects may be a common neuropathology in ASD. Here we propose to further this study by generating and characterizing Shank3 knockout marmosets. These models will allow us to study PV neuron dysfunction in a well-developed primate prefrontal cortex but also dissect cellular and circuitry basis of higher brain function that are disrupted in severe ASD such as eye-gazing, social interaction and cognitive impairment. 4. Provide human stem cells with Shank3 mutations for studying human PV neurons in organoids. In collaboration with Kevin Eggan?s group at Harvard and Lindy Barret at Stanley Center for Psychiatric Research at Broad Institute, we have developed human ES cell lines with Shank3 mutations using CRISPR/Cas9 system. We will provide these cells to Paola Arlotta?s group for developing organoids.
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0.909 |
2019 — 2021 |
Feng, Guoping Halassa, Michael M (co-PI) [⬀] |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Functional Dissection of Thalamocortical Interactions Through Genetically-Defined Trn Subnetworks @ Massachusetts Institute of Technology
PROJECT SUMMARY The thalamic reticular nucleus (TRN), the major source of thalamic inhibition, plays essential roles in sensory processing, arousal and cognition. Receiving inputs from cortical and subcortical regions, this structure is strategically positioned to influence thalamo-cortical interactions. During quiescence, the TRN participate in sleep rhythm generation, sleep stability and memory consolidation, while in active states, TRN neurons contribute to sensory filtering underlying attention. Perturbed TRN function may underlie behavioral deficits in disorders ranging from schizophrenia and autism to ADHD. Despite its importance, however, several key challenges have limited our ability to determine exactly how TRN circuitry contributes to various brain functions, a prerequisite for determining how it malfunctions in diseases and how its circuitry can be leveraged for diagnostic and therapeutic purposes. This proposal aims to address this critical gap in knowledge by capitalizing on a novel set of findings and tools that we generated. The TRN is a thin shell of GABAergic neurons surrounding thalamic projection nuclei. Within the TRN, neurons that have distinct structural and functional properties can be partially intermingled. This anatomical feature has been a major impediment for functional studies, since selective targeting of TRN neurons that share structural and functional properties with traditional methods is challenging. Using single cell RNAseq, we have recently discovered that TRN neurons can be dissociated into two major subtypes with distinct transcriptomic profiles, anatomical localizations, electrophysiological properties and thalamic connectivity. One group, located in the ?core? region of the TRN and can be marked by the expression of the Spp1 gene, targets first-order sensory thalamic nuclei, and the other, located in the ?shell? region of the TRN and marked by the expression of Ecel1 gene, targets higher- order ones. We have generated transgenic mice expressing Cre recombinase in each of these two populations individually. Here, we propose to use these new knowledge and genetic tools to answer fundamental questions about TRN structure-function organization as well as the contribution of this brain region to sensory processing, arousal and cognition.
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0.921 |
2021 |
Feng, Guoping |
U01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
A Genetic Engineering Toolbox For Marmosets (Getmarm): Development and Optimization of Genome Editing and Assisted Reproduction Techniques For Marmoset Models @ Massachusetts Institute of Technology
PROJECT SUMMARY While mice are essential models for many areas of neuroscience, there are also many aspects of higher brain function and dysfunction that cannot be adequately modeled in rodents. Thus, there is a need for new genetic models that have brain structure and function closer to humans. For these reasons, non-human primates (NHP) provide an attractive model to study higher brain function and brain disorders. A promising emerging NHP model is the common marmoset, a small New World primate that has many advantages as a genetic model. Although the adaptation of bacterial CRISPR/Cas systems for targeted genome engineering and model creation has revolutionized modern biology, editing of the marmoset genome is still in its infancy. Given the significant time and money required for marmoset genome editing, new methods to increase editing efficiency, decrease mosaicism, and identify correctly-edited embryos prior to transfer to recipient females are critical. Additionally, new methods for controlling the zygosity of founder animals are necessary to enable analysis of homozygous F0 animals and to avoid homozygous editing when targeting essential genes that cause embryonic lethality upon biallelic disruption. To these ends, we propose a research program that will significantly enhance our ability to introduce multiple types of edits into the marmoset genome, reduce mosaicism, control the zygosity of edits, and identify successfully edited embryos through prenatal genetic testing. We will disseminate these technologies and models through direct resource sharing, in-person trainings, deposition to NIH-supported Marmoset Coordination Center. Together, the proposed advances will significantly reduce the time, effort, costs and animal numbers necessary to marmoset genetic models and will unlock the true potential of marmosets for basic and translational neuroscience research.
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0.921 |
2021 |
Feng, Guoping Mitra, Partha Pratim |
UG3Activity Code Description: As part of a bi-phasic approach to funding exploratory and/or developmental research, the UG3 provides support for the first phase of the award. This activity code is used in lieu of the UH2 activity code when larger budgets and/or project periods are required to establish feasibility for the project. |
Developing Cell Type-Specific Enhancers and Connectivity Mapping Pipelines For Marmosets @ Massachusetts Institute of Technology
PROJECT SUMMARY Although genetic tools have dramatically advanced our understanding of brain function, they have largely been confined to mice. While mice are essential models for many areas of neuroscience, there are also many aspects of higher brain function that cannot be adequately modeled in rodents. Similarly, many brain disorders affect higher cognitive functions that have no clear parallels in rodents. Furthermore, recent large- scale single cell transcriptomic analyses have revealed many neuron types, connections and gene expression patterns that are unique to primates. Thus, there is an urgent need for new genetic models that have brain structure and function closer to humans. Non-human primates (NHP) are much more closely related to humans than are rodents, and this is reflected in their brain development, structure and physiology. Hence, it is increasingly recognized that they provide an attractive model to study higher brain function and brain disorders. A promising emerging NHP model is the common marmoset, a small new world primate that has many advantages for neuroscience and genetic research. However, lack of tools with cell type specificity has been a major obstacle in advancing structural and functional studies in NHP. With the combined single cell RNA-seq and single cell ATAC-seq, it is now possible to nominate short cell type-specific enhancer sequences. If validated, these enhancers will provide an effective tool to map connectivity and interrogate function using virus mediated expression. The difficulty lies in the identification of functional enhancers from the hundreds or thousands of nominated potential enhancer sequences in NHP. Here we propose (1) to use a novel high throughput in vivo approach to identify functional enhancers, and (2) to establish a whole-brain circuit mapping pipeline for use striatal circuitry to validate our approach for cell type-specific connectivity mapping in marmosets. When completed, these studies will provide much needed essential tools, methods and computational pipelines for cell type-specific mapping and functional interrogation of the marmoset brain in healthy and disease models.
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0.921 |