Society for Neuroscience
Doctor of Philosophy, Stanford University, NEURS-PHD (2014)
Bachelor of Arts, Harvard University, Neurobiology (2007)
In vivo experimental evidence indicates that acute neuronal activation increases Aβ release from presynaptic terminals, whereas long-term effects of chronic synaptic activation on Aβ pathology remain unclear. To address this issue, we adopted optogenetics and transduced stabilized step-function opsin, a channelrhodopsin engineered to elicit a long-lasting neuronal hyperexcitability, into the hippocampal perforant pathway of APP transgenic mice. In vivo microdialysis revealed a ∼24% increase in the hippocampal interstitial fluid Aβ42 levels immediately after acute light activation. Five months of chronic optogenetic stimulation increased Aβ burden specifically in the projection area of the perforant pathway (i.e., outer molecular layer of the dentate gyrus) of the stimulated side by ∼2.5-fold compared with that in the contralateral side. Epileptic seizures were observed during the course of chronic stimulation, which might have partly contributed to the Aβ pathology. These findings implicate functional abnormalities of specific neuronal circuitry in Aβ pathology and Alzheimer disease.
View details for DOI 10.1016/j.celrep.2015.04.017
View details for Web of Science ID 000354406900002
View details for PubMedID 25937280
Linking the activity of defined neural populations with behavior is a key goal of neuroscience. In the context of controlling behavior, electrical stimulation affords researchers precision in the temporal domain with gross regional specificity, whereas pharmacology allows for more specific manipulation of cell types, but in the absence of temporal precision. The use of microbial opsins-light activated, genetically encoded ion channels and pumps-to control mammalian neurons now allows researchers to "sensitize" genetically and/or topologically defined populations of neurons to light to induce either depolarization or hyperpolarization in both a cell-type-specific and temporally precise manner not achievable with previous techniques. Here, we describe the use of transgenic mice expressing the blue-light gated cation channel Channelrhodopsin-2 (ChR2) under control of the Thy1 promoter for the purpose of linking neuronal activity to behavior through restricted delivery of light to an anatomic region of interest. The surgical procedure for implanting a fiber-optic light delivery guide into the mouse brain, the process of optically stimulating the brain in a behaving animal, and post hoc evaluation are given, along with necessary reagents and discussion of common technical problems and their solutions.
View details for DOI 10.1101/pdb.prot075598
View details for PubMedID 26034299
Clinical and research efforts have focused on promoting functional recovery after stroke. Brain stimulation strategies are particularly promising because they allow direct manipulation of the target area's excitability. However, elucidating the cell type and mechanisms mediating recovery has been difficult because existing stimulation techniques nonspecifically target all cell types near the stimulated site. To circumvent these barriers, we used optogenetics to selectively activate neurons that express channelrhodopsin 2 and demonstrated that selective neuronal stimulations in the ipsilesional primary motor cortex (iM1) can promote functional recovery. Stroke mice that received repeated neuronal stimulations exhibited significant improvement in cerebral blood flow and the neurovascular coupling response, as well as increased expression of activity-dependent neurotrophins in the contralesional cortex, including brain-derived neurotrophic factor, nerve growth factor, and neurotrophin 3. Western analysis also indicated that stimulated mice exhibited a significant increase in the expression of a plasticity marker growth-associated protein 43. Moreover, iM1 neuronal stimulations promoted functional recovery, as stimulated stroke mice showed faster weight gain and performed significantly better in sensory-motor behavior tests. Interestingly, stimulations in normal nonstroke mice did not alter motor behavior or neurotrophin expression, suggesting that the prorecovery effect of selective neuronal stimulations is dependent on the poststroke environment. These results demonstrate that stimulation of neurons in the stroke hemisphere is sufficient to promote recovery.
View details for DOI 10.1073/pnas.1404109111
View details for PubMedID 25136109
Precisely defining the roles of specific cell types is an intriguing frontier in the study of intact biological systems and has stimulated the rapid development of genetically encoded tools for observation and control. However, targeting these tools with adequate specificity remains challenging: most cell types are best defined by the intersection of two or more features such as active promoter elements, location and connectivity. Here we have combined engineered introns with specific recombinases to achieve expression of genetically encoded tools that is conditional upon multiple cell-type features, using Boolean logical operations all governed by a single versatile vector. We used this approach to target intersectionally specified populations of inhibitory interneurons in mammalian hippocampus and neurons of the ventral tegmental area defined by both genetic and wiring properties. This flexible and modular approach may expand the application of genetically encoded interventional and observational tools for intact-systems biology.
View details for DOI 10.1038/nmeth.2996
View details for PubMedID 24908100
Social interaction is a complex behavior essential for many species and is impaired in major neuropsychiatric disorders. Pharmacological studies have implicated certain neurotransmitter systems in social behavior, but circuit-level understanding of endogenous neural activity during social interaction is lacking. We therefore developed and applied a new methodology, termed fiber photometry, to optically record natural neural activity in genetically and connectivity-defined projections to elucidate the real-time role of specified pathways in mammalian behavior. Fiber photometry revealed that activity dynamics of a ventral tegmental area (VTA)-to-nucleus accumbens (NAc) projection could encode and predict key features of social, but not novel object, interaction. Consistent with this observation, optogenetic control of cells specifically contributing to this projection was sufficient to modulate social behavior, which was mediated by type 1 dopamine receptor signaling downstream in the NAc. Direct observation of deep projection-specific activity in this way captures a fundamental and previously inaccessible dimension of mammalian circuit dynamics.
View details for DOI 10.1016/j.cell.2014.05.017
View details for Web of Science ID 000340941900010
View details for Web of Science ID 000310541600336
Diverse optogenetic tools have allowed versatile control over neural activity. Many depolarizing and hyperpolarizing tools have now been developed in multiple laboratories and tested across different preparations, presenting opportunities but also making it difficult to draw direct comparisons. This challenge has been compounded by the dependence of performance on parameters such as vector, promoter, expression time, illumination, cell type and many other variables. As a result, it has become increasingly complicated for end users to select the optimal reagents for their experimental needs. For a rapidly growing field, critical figures of merit should be formalized both to establish a framework for further development and so that end users can readily understand how these standardized parameters translate into performance. Here we systematically compared microbial opsins under matched experimental conditions to extract essential principles and identify key parameters for the conduct, design and interpretation of experiments involving optogenetic techniques.
View details for DOI 10.1038/nmeth.1808
View details for PubMedID 22179551
The capture and utilization of light is an exquisitely evolved process. The single-component microbial opsins, although more limited than multicomponent cascades in processing, display unparalleled compactness and speed. Recent advances in understanding microbial opsins have been driven by molecular engineering for optogenetics and by comparative genomics. Here we provide a Primer on these light-activated ion channels and pumps, describe a group of opsins bridging prior categories, and explore the convergence of molecular engineering and genomic discovery for the utilization and understanding of these remarkable molecular machines.
View details for DOI 10.1016/j.cell.2011.12.004
View details for Web of Science ID 000298403400011
View details for PubMedID 22196724
Parkinson's disease (PD) is an incurable age-related neurodegenerative disorder affecting both the central and peripheral nervous systems. Although common, the etiology of PD remains poorly understood. Genetic studies infer that the disease results from a complex interaction between genetics and environment and there is growing evidence that PD may represent a constellation of diseases with overlapping yet distinct underlying mechanisms. Novel clinical approaches will require a better understanding of the mechanisms at work within an individual as well as methods to identify the specific array of mechanisms that have contributed to the disease. Induced pluripotent stem cell (iPSC) strategies provide an opportunity to directly study the affected neuronal subtypes in a given patient. Here we report the generation of iPSC-derived midbrain dopaminergic neurons from a patient with a triplication in the ?-synuclein gene (SNCA). We observed that the iPSCs readily differentiated into functional neurons. Importantly, the PD-affected line exhibited disease-related phenotypes in culture: accumulation of ?-synuclein, inherent overexpression of markers of oxidative stress, and sensitivity to peroxide induced oxidative stress. These findings show that the dominantly-acting PD mutation is intrinsically capable of perturbing normal cell function in culture and confirm that these features reflect, at least in part, a cell autonomous disease process that is independent of exposure to the entire complexity of the diseased brain.
View details for DOI 10.1371/journal.pone.0026159
View details for Web of Science ID 000297555400007
View details for PubMedID 22110584
Cortico-thalamo-cortical circuits mediate sensation and generate neural network oscillations associated with slow-wave sleep and various epilepsies. Cortical input to sensory thalamus is thought to mainly evoke feed-forward synaptic inhibition of thalamocortical (TC) cells via reticular thalamic nucleus (nRT) neurons, especially during oscillations. This relies on a stronger synaptic strength in the cortico-nRT pathway than in the cortico-TC pathway, allowing the feed-forward inhibition of TC cells to overcome direct cortico-TC excitation. We found a systemic and specific reduction in strength in GluA4-deficient (Gria4(-/-)) mice of one excitatory synapse of the rhythmogenic cortico-thalamo-cortical system, the cortico-nRT projection, and observed that the oscillations could still be initiated by cortical inputs via the cortico-TC-nRT-TC pathway. These results reveal a previously unknown mode of cortico-thalamo-cortical transmission, bypassing direct cortico-nRT excitation, and describe a mechanism for pathological oscillation generation. This mode could be active under other circumstances, representing a previously unknown channel of cortico-thalamo-cortical information processing.
View details for DOI 10.1038/nn.2896
View details for Web of Science ID 000294284900017
View details for PubMedID 21857658
Both observational and perturbational technologies are essential for advancing the understanding of brain function and dysfunction. But while observational techniques have greatly advanced in the last century, techniques for perturbation that are matched to the speed and heterogeneity of neural systems have lagged behind. The technology of optogenetics represents a step toward addressing this disparity. Reliable and targetable single-component tools (which encompass both light sensation and effector function within a single protein) have enabled versatile new classes of investigation in the study of neural systems. Here we provide a primer on the application of optogenetics in neuroscience, focusing on the single-component tools and highlighting important problems, challenges, and technical considerations.
View details for DOI 10.1016/j.neuron.2011.06.004
View details for Web of Science ID 000292806200004
View details for PubMedID 21745635
Anxiety--a sustained state of heightened apprehension in the absence of immediate threat--becomes severely debilitating in disease states. Anxiety disorders represent the most common of psychiatric diseases (28% lifetime prevalence) and contribute to the aetiology of major depression and substance abuse. Although it has been proposed that the amygdala, a brain region important for emotional processing, has a role in anxiety, the neural mechanisms that control anxiety remain unclear. Here we explore the neural circuits underlying anxiety-related behaviours by using optogenetics with two-photon microscopy, anxiety assays in freely moving mice, and electrophysiology. With the capability of optogenetics to control not only cell types but also specific connections between cells, we observed that temporally precise optogenetic stimulation of basolateral amygdala (BLA) terminals in the central nucleus of the amygdala (CeA)--achieved by viral transduction of the BLA with a codon-optimized channelrhodopsin followed by restricted illumination in the downstream CeA--exerted an acute, reversible anxiolytic effect. Conversely, selective optogenetic inhibition of the same projection with a third-generation halorhodopsin (eNpHR3.0) increased anxiety-related behaviours. Importantly, these effects were not observed with direct optogenetic control of BLA somata, possibly owing to recruitment of antagonistic downstream structures. Together, these results implicate specific BLA-CeA projections as critical circuit elements for acute anxiety control in the mammalian brain, and demonstrate the importance of optogenetically targeting defined projections, beyond simply targeting cell types, in the study of circuit function relevant to neuropsychiatric disease.
View details for DOI 10.1038/nature09820
View details for Web of Science ID 000288444000041
View details for PubMedID 21389985
Genetically encoded, single-component optogenetic tools have made a significant impact on neuroscience, enabling specific modulation of selected cells within complex neural tissues. As the optogenetic toolbox contents grow and diversify, the opportunities for neuroscience continue to grow. In this review, we outline the development of currently available single-component optogenetic tools and summarize the application of various optogenetic tools in diverse model organisms.
View details for DOI 10.1146/annurev-neuro-061010-113817
View details for Web of Science ID 000293772100017
View details for PubMedID 21692661
Despite a rapidly-growing scientific and clinical brain imaging literature based on functional magnetic resonance imaging (fMRI) using blood oxygenation level-dependent (BOLD) signals, it remains controversial whether BOLD signals in a particular region can be caused by activation of local excitatory neurons. This difficult question is central to the interpretation and utility of BOLD, with major significance for fMRI studies in basic research and clinical applications. Using a novel integrated technology unifying optogenetic control of inputs with high-field fMRI signal readouts, we show here that specific stimulation of local CaMKIIalpha-expressing excitatory neurons, either in the neocortex or thalamus, elicits positive BOLD signals at the stimulus location with classical kinetics. We also show that optogenetic fMRI (of MRI) allows visualization of the causal effects of specific cell types defined not only by genetic identity and cell body location, but also by axonal projection target. Finally, we show that of MRI within the living and intact mammalian brain reveals BOLD signals in downstream targets distant from the stimulus, indicating that this approach can be used to map the global effects of controlling a local cell population. In this respect, unlike both conventional fMRI studies based on correlations and fMRI with electrical stimulation that will also directly drive afferent and nearby axons, this of MRI approach provides causal information about the global circuits recruited by defined local neuronal activity patterns. Together these findings provide an empirical foundation for the widely-used fMRI BOLD signal, and the features of of MRI define a potent tool that may be suitable for functional circuit analysis as well as global phenotyping of dysfunctional circuitry.
View details for DOI 10.1038/nature09108
View details for Web of Science ID 000278551800047
View details for PubMedID 20473285
In the study of complex mammalian behaviours, technological limitations have prevented spatiotemporally precise control over intracellular signalling processes. Here we report the development of a versatile family of genetically encoded optical tools ('optoXRs') that leverage common structure-function relationships among G-protein-coupled receptors (GPCRs) to recruit and control, with high spatiotemporal precision, receptor-initiated biochemical signalling pathways. In particular, we have developed and characterized two optoXRs that selectively recruit distinct, targeted signalling pathways in response to light. The two optoXRs exerted opposing effects on spike firing in nucleus accumbens in vivo, and precisely timed optoXR photostimulation in nucleus accumbens by itself sufficed to drive conditioned place preference in freely moving mice. The optoXR approach allows testing of hypotheses regarding the causal impact of biochemical signalling in behaving mammals, in a targetable and temporally precise manner.
View details for DOI 10.1038/nature07926
View details for Web of Science ID 000265412900042
View details for PubMedID 19295515
Human embryonic stem cells possess the unique ability to differentiate into any adult cell type. Recent advances in the understanding of stem cell biology make new applications possible for stem cell based technology. Of note, it is now possible to reprogram terminally differentiated human somatic cells into pluripotent cells that are functionally equivalent to embryonic stem cells. These induced pluripotent cells may become the substrate for future disease models and cell-based therapies. In addition, novel techniques for genetic manipulation have increased the ease with which genes can be modified into stem cells. In this review, we describe these novel technologies as well as developments in the understanding of basic biology of stem cell pluripotency and differentiation.
View details for DOI 10.1016/j.gde.2008.06.004
View details for Web of Science ID 000260700500007
View details for PubMedID 18625311