A neural network for abstracting nociception into the affective dimension of pain
Gregory Corder, Biafra Ahanonu, Benjamin Grewe, Mark Schnitzer, Gregory Scherrer

Stanford University

In Parkinson’s Disease (PD), midbrain dopamine neurons are lost. Dopamine replacement therapy with levodopa is effective in treating symptoms, but is commonly complicated by levodopa-induced dyskinesia (LID). One hypothesis, supported by immediate early gene expression, is that LID is caused by aberrant activity in the striatum, the input nucleus of the basal ganglia. To test this hypothesis, we combined optogenetics and a novel method to capture activated neurons, Targeted Recombination of Activated Populations (TRAP), to establish a direct link between striatal activity and dyskinesia. We observed that LID-associated TRAPed cells are a stable subset of striatal neurons, primarily those of the direct pathway, and that reactivation of TRAPed striatal neurons caused dyskinesia in the absence of levodopa. Inhibition of TRAPed cells, but not a nonspecific subset of direct pathway neurons, ameliorated LID. These results support the idea that a distinct and stable subset of striatal neurons participates in LID.

A subpopulation of striatal neurons mediates levodopa-induced dyskinesia
Allison E Girasole, Matthew Y Lum, Diane Nathaniel, Chloe J. Bair-Marshall, Casey J. Guenthner, Liqun Luo, Anatol C. Kreitzer, Alexandra B. Nelson

UCSF

In Parkinson’s Disease (PD), midbrain dopamine neurons are lost. Dopamine replacement therapy with levodopa is effective in treating symptoms, but is commonly complicated by levodopa-induced dyskinesia (LID). One hypothesis, supported by immediate early gene expression, is that LID is caused by aberrant activity in the striatum, the input nucleus of the basal ganglia. To test this hypothesis, we combined optogenetics and a novel method to capture activated neurons, Targeted Recombination of Activated Populations (TRAP), to establish a direct link between striatal activity and dyskinesia. We observed that LID-associated TRAPed cells are a stable subset of striatal neurons, primarily those of the direct pathway, and that reactivation of TRAPed striatal neurons caused dyskinesia in the absence of levodopa. Inhibition of TRAPed cells, but not a nonspecific subset of direct pathway neurons, ameliorated LID. These results support the idea that a distinct and stable subset of striatal neurons participates in LID.

Acute effects of locomotion on visual plasticity
Yujiao Jennifer Sun, Michael Stryker

UCSF

Locomotion increases visual responses to exposed stimulus in adult mouse visual cortex, while responses to other stimuli were unaffected. (Kaneko et Stryker, 2017, Journal of Neuroscience). Is it a result of the high-gain state of visual cortex induced by locomotion, and how long does this effect last? Here we presented two type of visual stimulus under two states: one during the locomotion and the other in the stationary state, with a 250ms transitional period between the two states. Before and after the visual exposure, we used wide-field imaging and two-photon imaging to trace populational and single-neuron changes in the awake and anesthetized animals to dissect the top-down and bottom-up circuit involved in the adult visual plasticity.

An amygdala to brainstem connection gates defensive locomotion
Thomas K Roseberry, Anatol Kreitzer

UCSF

An animal under threat must rapidly choose between running away from danger or freezing to avoid detection. The circuitry that mediates this choice is unknown. Here we describe a novel projection from the central amygdala (CeA) to the mesencephalic locomotor region (MLR) which gates locomotion in response to aversive stimuli. We find that CeA terminal stimulation rapidly stops locomotion while many neurons in the MLR are inhibited at putatively monosynaptic latencies. Terminal inhibition increases MLR firing and locomotor initiations in response to air puff. Using a cued active avoidance task we show that glutamatergic neurons in the MLR are required for learned locomotor avoidance responses while increased activity in the CeA to MLR projection can stop these responses. Finally, inhibiting CeA terminals in the MLR increases locomotor avoidance responses. Together these data indicate that the CeA to MLR projection gates defensive locomotor responses to both learned and innately painful stimuli.

An ARL-8 GEF in regulating axonal transport of synaptic vesicle precursors
Shinsuke Niwa, Li Tao, Sharon Y. Lu, Gerald M. Liew, Maxence V. Nachury, Kang Shen

Stanford University

Intracellular transport is essential to a cell’s function and survival. In particular, neurons rely heavily on directional transport to send various cargos between the cell body and the distal processes. Previous studies have identified the KIF1A/UNC-104 kinesin as the critical motor for synaptic vesicle precursor (SV) transport. However, it is not clear how UNC-104 activity is regulated in vivo. Our lab has shown that a highly conserved vesicle-bound small G protein, ARL-8, directly binds to and activates UNC-104 by suppressing its autoinhibition. What activates ARL-8 and localizes it to SV remains unknown. In this study, we identified a conserved, SV-localized protein, SAM-4, that acts as a GEF of ARL-8. We used both genetic and biochemical experiments to demonstrate that SAM-4 is essential in regulating the activity of ARL-8 for axonal transport in vivo.

Computational Illumination for Phase Microscopy
Michael Chen, Zachary F. Phillips, Lei Tian and Laura Waller

UC Berkeley

We demonstrate a phase microscopy with an LED array microscope, which measures phase of transparent sample using spatially partially coherent light. By collecting intensity images with coded illumination, optical path difference of the sample can be recovered with a proposed algorithm. In addition, resolution is two times compared to coherent phase imaging techniques. To maximize the throughput, one can adopt wavelength multiplexing. A color camera was used to capture intensity images as we illuminated the sample with RGB LEDs. In this case, three independent measurements can be acquired within single exposure, resulting in camera limited frame rate. Finally, the proposed technique can be easily extended to 3D phase imaging. By taking through focus image stacks with coded illumination, the refractive indices and 3D geometry of the sample can be reconstructed.

DiffuserCam
Grace Kuo, Nick Antipa, Ren Ng, Laura Waller

UC Berkeley

Traditional cameras require a lens which optically forms an image on the sensor. In this work, we replace the lens with a thin piece of bumpy plastic (a diffuser) and we use computational algorithms to recover an image.

Three-dimensional spatiotemporal focusing of holographic patterns
O. Hernandez, E. Papagiakoumou, D. Tanese, K. Fidelin, C. Wyart and V. Emiliani

Stanford University

Genetically encoded light-sensitive channels and reporters enable both neuronal activity optical control and read-out. Full exploitation of these optogenetic tools requires single-cell scale methods to pattern light into neural tissue.
Computer Generated Holography (CGH) can powerfully enhance optogenetic stimulation by efficiently shaping light onto multiple cellular targets. However, a linear proportionality between lateral shape area and axial extent degrades axial precision for cases demanding extended lateral patterning i.e., to cover entire soma of multiple cells. To address this limitation, we previously combined CGH with temporal focusing (TF) to stretch laser pulses outside of the focal plane, which combined with two-photon’s nonlinear fluorescence dependence, axially confines fluorescence regardless of lateral extent. However, this configuration restricts nonlinear excitation to a single spatiotemporal focal plane, precluding simultaneous confinement of axially separated light patterns. Here we combine CGH with TF using a novel scheme enabling arbitrary spatiotemporally focused pattern generation in three dimensions. We demonstrate simultaneous photoconversion of tens of zebrafish larvae spinal cord neurons occupying separate axial planes.

Input-specific, dopaminergic modulation of gain at long-range inputs to medial prefrontal cortex
Kenneth J. Burke & Kevin J. Bender

UCSF

The prefrontal cortex (PFC) supports working memory and decision-making tasks by integrating excitatory and neuromodulatory inputs from several brain regions. Performance on these tasks is dependent on the neuromodulator dopamine (Brozoski et al, 1979). The dopamine D1 receptor has been previously shown to suppress glutamatergic transmission in PFC (Urban et al, 2002), however it has not been established whether this suppression is ubiquitous or specific to a subset of PFC inputs. Here, we show that activation of prefrontal D1 receptors preferentially suppresses a subset of excitatory long-range inputs by modulating calcium influx in glutamatergic terminals. Interestingly, this presynaptic modulation lowers release probability without affecting short-term synaptic plasticity, thus allowing for input-specific modulation of synaptic gain. These findings suggest that D1 receptors can bias information transfer through suppression of specific inputs into PFC.

Habenular cell recruitment measures the magnitude of challenge to orchestrate brain-wide and behavioral responses
Aaron Andalman, Vanessa Burns, Matthew Lovett-Barron, Michael Broxton, Ben Poole, Samuel Yang, Logan Grosenick, Talia Lerner, Philippe Mourrain, Marc Levoy, and Karl Deisseroth

Stanford University

The ability of prior experiences to affect future action is central to the generation of adaptive behavior. Behavioral challenge assays use inescapable stressors to probe how the repeated failure of escape actions cause a transition to passive coping strategy. Here, we develop an assay that induces passive coping in the larval zebrafish and use brain-wide imaging to perform an unbiased search for where the negative value of ongoing experience is encoded. We show that during behavioral challenge habenular cells are continuously recruited into an activated ensemble. This causes a ramping of neural activity that is confined to the habenula and can be bi-directionally modulated, while other regions display plateauing changes in activity. We use optogenetic stimulation combined with brain-wide imaging to demonstrate that lateral habenula activation is sufficient to reduce mobility as well as selectively reduce activity in the raphe nuclei. Together, our results identify the temporal recruitment of habenular neurons as a mechanism by which the negative value of ongoing experience is encoded to regulate behavioral strategy.

High-throughput automated time-lapse imaging of neuron degeneration within a live animal using robotic microscopy
Jeremy Linsley, Elliot Mount, David Kokel, Steve Finkbeiner

Gladstone Institute/UCSF

Elucidating the cellular changes that destine a neuron to live or to die is a fundamental challenge to understanding neurodegenerative disease. To quantitatively relate intermediate changes within a neuron to its fate within the degenerating brain requires a robust imaging platform that can relate changes of individual neurons within a live brain over time. Combining robotic microscopy, which facilitates longitudinal single cell analysis of neurons, with the ability to perform time-lapse imaging on immobilized zebrafish larvae, we have created a high-throughput, en toto confocal imaging platform combined with automated image analysis software for single cell analysis of genetically defined subsets of neurons within larvae. Using this technology, we demonstrate the ability to image neurodegeneration within live larvae, providing longitudinal single cell analysis. In conjunction, we demonstrate the ability to measure turnover of mutant Tau, a pathological protein involved in multiple neurodegenerative diseases such as Alzheimer’s disease and Parkinson’s disease.

High-throughput, High-resolution Neural Circuit Mapping
Ben Shababo, Shizhe Chen, Xinyi Deng, Alex Naka, Liam Paninski, Hillel Adesnik

UC Berkeley

We present a novel experimental and computational method for mapping large portions of local neural circuits. Our method combines whole-cell electrophysiology, two-photon holographic optogenetics, and statistical modelling to infer the monosynaptic connectivity between hundreds to thousands of neurons in a single circuit. We propose solutions for two main challenges to mapping neural circuits at a fine-scale over large volumes. One, the biological variability in the response of individual neurons to optical stimulation, the limits of the spatial resolution of optical stimulation, and the variability in the postsynaptic features (e.g. amplitude, success rate) of individual connections make confident inference of unitary monosynaptic inputs difficult. Two, the maps must be inferred with limited data because often times the preparations are short-lived and in general the amount of data one can collect is paltry compared to the extent of neural circuits. We demonstrate our method in simulation and on data from the somatosensory cortex of mice.

Hippocampal spiking sequences during a working memory task
Jiannis Taxidis, Apoorva Mylavarapu, Kian Samadian, Emily Hoffberg, Nima Saboori, Melody Bedrossian, Tiffany Taimoorazy, Jonathan Sadik, Peyman Golshani

UCLA

How does the brain retain information in memory, for short amounts of time, in order to execute a function? Recent studies described spiking sequences in cortex and dorsal hippocampus during behavioral tasks that involve working-memory. But the link between such sequences and working-memory is still not well understood. Are they associated with the actual memory of the stimulus? What is their relationship with learning and performance? Do they evolve over days? What is the role of inhibition in sustaining them? We applied in vivo two-photon calcium imaging in the mouse dorsal CA1, during a delayed non-match-to-sample task, involving odor stimuli and requiring working memory activation. We observed stable spiking sequences, involving both pyramidal cells and interneurons, that encode the olfactory stimulus and the delay period following it. We describe how these sequences emerge during learning and how they adjust to increasing memory load.

How altered dendritic morphology influences neural computations in the retinal direction selective circuit
Ryan D. Morrie, Marla B. Feller

UC Berkeley

Direction selective ganglion cells in the retina fire action potentials maximally to objects moving in a preferred direction, but minimally to objects moving in the opposite direction. How does this asymmetry emerge?

Starburst amacrine cells are required for direction selectivity and characterized by their radially symmetric dendritic arbors. The ends of these dendrites provide asymmetric GABAergic inhibition via: 1) greater GABA release for motion away from than towards the starburst cell soma and 2) a selective wiring process between starburst cells and direction selective cells.

To determine how the starburst morphology of these cells enables and contributes to direction selectivity, we utilized mice lacking Sema6A, whose starburst cells no longer display radial symmetry, and whose ganglion cells have reduced direction selectivity. Using paired recordings and subcellular 2-photon Ca2+ imaging, I will present results describing how morphology impacts direction selectivity at multiple levels within the retina.

Imaging Extracellular Dopamine Dynamics Using Optical Nanosensors
Abraham G. Beyene, Kristen Delevich, Jackson Travis Del Bonis-O’Donnell, Linda Wilbrecht, Markita P. Landry

UC Berkeley

The brain, an organ so central to human existence and experience, is incredibly complex and the dearth of tools that can probe it is one manifestation this complexity. Therefore, successful research into elucidating brain function will be dependent in part on whether concomitant advances in investigative methods can be made. In particular, tools that operate at the nanoscale can provide the type of spatial and temporal resolution needed to explore the brain and hold considerable promise in making this complex organ more accessible to research. To this end, carbon nanotubes have distinct photophysical properties well suited to imaging optically dense and complex organs like the brain. When functionalized with polymers, carbon nanotubes assume conformational states that display unique fluorescent modulation in the presence of certain analytes. Here, we present results from imaging the dynamics of evoked dopamine release in ECS in acute brain slice preparations using such a nanosensor.

Molecular Recognition of Dopamine with Dual Excitation-Emission Near Infrared Two-Photon Microscopy
Jackson Travis Del Bonis-O’Donnell, Ralph H. Page, Abraham G. Beyene, Eric G. Tindall, Ian R. McFarlane, Markita P. Landry

UC Berkeley

The development of non-invasive, optical methods to directly quantify neuromodulatory neurotransmitters in the living brain are vital to the study and understanding of neurological disorders. Recently, functionalized single walled carbon nanotubes (SWNTs) have emerged as promising near-infrared (NIR) fluorescent nanosensors for the sensitive and rapid detection of neurotransmitter dopamine. These nanosensors fluoresce in the NIR-II window (1000-1700 nm) where absorption and scattering of light is minimal, making them optimally suited for non-invasive imaging in brain tissue. Using a femtosecond pulsed erbium laser with peak wavelength in the NIR-II window, we demonstrate SWNT-based molecular recognition of dopamine using two-photon excitation, confirming that the molecular recognition principle of our nanosensors is compatible with non-linear excitation. Furthermore, we show significantly improved fluorescence spatial resolution attained by two-photon excitation when imaging in strongly scattering tissue phantoms, motivating future work using these nanosensors for in vivo, real-time sensing of dopamine in brain tissue.

Multi-Scale & Multi-Modal Correlative Imaging: Leveraging Knife- Edge Scanning Microscopy for the Generation of Tissue Atlases
Huffman TM, Farahani N

3Scan

Comprehensive cytoarchitectural understandings of human tissue are essential for unraveling structure-function relationships, yet current anatomical and histological atlases harbor major limitations. Large-scale histological reference atlases of the human brain, for example, demonstrate marked variability in their degree of whole-organ coverage, information content, and structural annotation. Furthermore, data collection is usually restricted to a few manually defined regions of interest (ROIs) within a few two-dimensional (2D) tissue sections, drastically limiting the scope of analysis. Lastly, since histological processing relies on tissue sectioning, direct spatial correlation of destructive 2D cross sections to non-destructive three-dimensional (3D) imaging (e.g. magnetic resonance imaging [MRI], positron emission tomography [PET], etc.) is arduous, resource intensive, and generally lacking.

Destructive 3D imaging techniques, like serial-section and serial-block face microscopy, can be combined with non-invasive 3D imaging methods, and are thus also well-suited for the purposes of correlative imaging whereby a ROI is identified for sequential investigation. Herein we sought to explore the power of multi-scale and multi-modal correlative imaging.

Probing the relationship between Amyloid-beta (1-42) and neuronal activity using voltage imaging
Alison S. Walker, Kaveh Karbasi, Albert Lee, Patrick Zhang & Evan W. Miller

UC Berkeley

Recent studies of Alzheimer’s Disease (AD) patients reveal a high incidence of subclinical epileptiform activity when compared to non-demented controls. However, it is currently unknown how changes in neuronal activity are expressed, or whether aberrant neuronal activity is a driver or a consequence of the disease. Here, we assess the interplay between Amyloid-beta which aggregates into plaques during AD progression, and neuronal activity using voltage imaging. We treat primary neuronal cultures with varying concentrations of Amyloid-beta (1-42) peptide and optically assess spontaneous action potential rates and patterns 48 hours later. Preliminary data reveals a complicated relationship between neuronal activity and Amyloid-beta (1-42) concentration and aggregation state. Recording neuronal activity with voltage imaging enables mapping of neuronal activity with spatial and temporal resolution inaccessible to either traditional electrophysiology or calcium imaging, and as such adds an important overview of the functional landscape of challenged neuronal networks.

The connectome of alpha1-containg GABA(A) receptor in cortical inhibitory microcircuit
Ming-Chi Tsai, Wan-Chen Lin, Richard Kramer

UC Berkeley

Dozens of GABA(A) receptors isoforms mediate synaptic inhibition in the brain. They differ in their kinetics and expression, and are the primary targets of many pharmacologies. While the diversity of GABAergic interneurons has been recognized in playing different roles within neural circuit, the role of GABA(A) receptor diversity at circuit level remains elusive. Here we present the first functional survey of “receptor connectome” of alpha1-containing GABA(A) receptor, the most abundant one in the brain, in cortical inhibitory microcircuit. Using Optogenetic Pharmacology (Lin, Tsai et. al. (2015)), we observed a synapse-specific presence of alpha1-GABA(A) receptors that is contingent to the presynaptic interneuron classes. Unexpectedly, we found that this specificity seems to be predominantly established by the postsynaptic neurons. The result suggests that alpha1-containing GABA(A) receptor may contribute its fast kinetics differentially at different motifs of inhibitory microcircuit to influence synaptic integration within a neuron and therefore network circuit dynamics.

Sensing light through a ciliary non-visual opsin in the developing vertebrate spinal cord
Amy Winans, Drew Friedmann, Tong Xiao, Ehud Isacoff

HWNI, UC Berkeley

Many human diseases, collectively called “ciliopathies”, are caused by the dysregulation of cilia and ciliary signaling and often present with neurological defects. Recent evidence suggests that cilia and ciliary signaling respond to chemical and mechanical cues as environmental “ciliopathies”, and serve has crucial signaling hubs for developmental signaling pathways such as Hedgehog and WNT. Here, we study a unique function of cilia in zebrafish spinal cord development. The Isacoff laboratory recently discovered that Valopa, a non-visual G-protein linked opsin, expresses in the zebrafish spinal cord, suppresses neural activity and locomotive movement when activated, and, surprisingly, localizes to cilia. To further elucidate Valopa’s mechanism of action, we use semi-high-throughput behavior assays, and live and fixed cell confocal imaging to probe from where in the circuit Valopa inhibits locomotion and to test the necessity of ciliary localization in Valopa activity.

Simultaneous fast measurement of circuit dynamics at multiple sites across the mammalian brain
Kim CK, Yang SJ, Pichamoorthy N, Young NP, Kauvar I, Jennings JH, Lerner TN, Berndt A, Lee SY, Ramakrishan C, Davidson TJ, Inoue M, Bito H, and Deisseroth K

Stanford University

Real-time access to activity signals within many simultaneously-assessed cell-type-specific populations and projections will likely be important for understanding the operation of the brain as a dynamical system. Here we describe high-speed, multi-site observation of genetically-specific neural activity traffic across the adult mammalian brain, suitable for quantifying experience-triggered joint activity relationships among multiple brainwide projections and populations. The frame-projected independent-fiber photometry (FIP) microscope can record fluorescence activity from at least seven brain regions or projections simultaneously with a single fast sCMOS camera, and is also suitable for dual-color activity tracking and combined optical observation/perturbation of neural activity in defined cells and projections. We demonstrate performance and sensitivity of this system with simultaneous recording of multiple independent axonal activity signals representing diverse projections of ventral tegmental area dopamine (VTA-DA) neurons, tracking previously inaccessible activity relationships among these circuit elements during distinct salient sensory experiences.

Surround integration organizes a spatial map during active sensation
Evan H. Lyall, Scott R. Pluta, Greg I. Telian, Elena Ryapolova-Webb, Hillel Adesnik

UC Berkeley

During active sensation, sensors scan space in order to generate a representation of the outside world. However, since spatial coding in sensory systems is typically addressed by measuring receptive fields in a fixed, sensor-based coordinate frame, the cortical representation of scanned space is poorly understood. Furthermore, while spatial summation across the sensory array is critical for coding in passive systems, its contribution to active sensation is essentially unknown. To address these questions, we probed spatial coding in the rodent whisker system using a combination of two photon imaging and electrophysiology during active touch. We found that surround whiskers powerfully transform the cortical representation of scanned space. On the single neuron level, surround input profoundly alters response amplitude and modulates spatial preference in the cortex. On the population level, surround input organizes the spatial preference of neurons into a continuous map of the space swept out by the whiskers. These data demonstrate how spatial summation over a moving sensor array is critical to generating population codes of sensory space.

The coding of temperature and thermal pain in the spinal cord
Ran, C., Kamalani, G., Hoon, M. and Chen, X.

Stanford University

In vivo two photon calcium imaging study of the spinal cord thermosensory neurons.

Understanding Resolution Limits for Microscopes with Shift-Varying Point Spread Functions
Michael Broxton

Stanford University

The optical transfer function (OTF) has long been used in light microscopy to characterize the performance of 2D and 3D imaging methods where the point spread function (PSF) is shift-invariant. In this case, image formation can be modeled via convolution, and the OTF can be understood as a low-pass filter that imposes a uniform resolution limit on the volume.

We have explored single-shot 3D imaging methods, including light field microscopy, in which the PSF is not shift-invariant. Here, image formation must be modeled more generally as a linear superposition integral, and “resolution” typically varies as a function of position in the volume. There is no single OTF in this case. We will show that the resolution limit of microscopes with shift-variant PSFs can instead be understood by studying Fisher Information, singular value decomposition of the microscope’s measurement operator, and the Fourier (or wavelet) crosstalk matrix.

Unmasking Direction Selective Computations in the Retina with Complex Stimuli
Summers, M.T., Feller, M.B.

UC Berkeley

Great strides have been made toward cracking sensory circuits by using reduced stimuli to get at the key features of circuit computations. However, in order to achieve a deeper understanding of how the brain intakes and processes information from the natural world, more sophisticated stimuli will be necessary to recreate the rich patterns of circuit and population activity that organisms experience in vivo. Here, I use complex stimuli to study direction selectivity in the retina. A great deal of work has been done studying direction selective ganglion cells (DSGCs) and the mechanisms by which they encode image motion, but much of this has been done with simple stimuli such as moving bars, drifting gratings, or two spot apparent motion. Recently, more complex stimuli have unmasked additional circuit components that are critical for DSGC computation in naturalistic settings. I am using complex visual stimuli to understand mechanisms of DSGC computation.

Global Representations of Goal-Directed Behavior in Distinct Cell Types of Mouse Neocortex
Will Allen, Isaac Kauvar, Karl Deisseroth

Stanford University

The successful planning and execution of adaptive behaviors in mammals may require long-range coordination of neural networks throughout cerebral cortex. The neuronal implementation of signals that could orchestrate cortex-wide activity remains unclear. Here, we develop and apply methods for cortex-wide Ca2+ imaging in mice performing decision-making behavior and identify a global cortical representation of task engagement encoded in the activity dynamics of both single cells and superficial neuropil distributed across the majority of dorsal cortex. The activity of multiple molecularly defined cell types was found to reflect this representation with type-specific dynamics. Focal optogenetic inhibition tiled across cortex revealed a crucial role for frontal cortex in triggering this cortex-wide phenomenon; local inhibition of this region blocked both the cortex-wide response to task-initiating cues and the voluntary behavior. These findings reveal cell-type-specific processes in cortex for globally representing goal-directed behavior and identify a major cortical node that gates the global broadcast of task-related information.

3D Scanless Holographic Optogenetics with Temporal Focusing
Nicolas C. Pégard, Alan R. Mardinly, Ian Antón Oldenburg, Savitha Sridharan, Laura Waller & Hillel Adesnik

UC Berkeley

Optical methods capable of manipulating neural activity with cellular resolution and millisecond precision in three dimensions will accelerate the pace of neuroscience research. Existing approaches for targeting individual neurons, however, fall short of these requirements. Here we present a new multiphoton photo-excitation method, termed 3D-Scanless Holographic Optogenetics with Temporal focusing (3D-SHOT), that allows precise, simultaneous photo-activation of arbitrary sets of neurons in any number of focal planes. This technique uses point-cloud holography to place multiple copies of a temporally focused disc matching the dimensions of a neuron’s cell body anywhere within the operating volume of the microscope. Experiments in cultured cells, brain slices, and in living mice demonstrate single-neuron spatial resolution even when optically targeting up to 50 cells distributed randomly in 3D. This approach opens new avenues for mapping and manipulating neural circuits, allowing a real-time, cellular resolution interface to the brain.

Precise bidirectional control of custom neural activity patterns
Alan R. Mardinly, Ian Antón Oldenburg, Nicolas C. Pégard, Savitha Sridharan, Evan Lyall, Kirill Chesnov, Stephen Brohawn, Laura Waller and Hillel Adesnik

UC Berkeley

Understanding brain function requires technologies that can control neural activity with high fidelity in space and time. We developed a new experimental approach to bidirectionally control neural activity with cellular resolution and sub-millisecond precision. We engineered powerful new soma-targeted (ST) optogenetic tools, ST-ChroME and a stabilized version of GtACR1, optimized for multiphoton activation. We then employed a novel all-optical paradigm that achieves three-dimensional optogenetic control of neural activity at cellular resolution. This approach enables the synthesis and editing of complex neural activity patterns in distributed neural circuits in behaving animals. Combined with three-dimensional (3D) calcium imaging, this new system achieves the necessary breakthrough in performance needed to gain insight into the fundamental principles of the neural codes underlying perception and behavior.