Scott Lab - Neural circuits and behaviour

Addressing the brain’s complexity

In the Scott Lab, we are interested in the workings of the brain at the level of cells and circuits.  We aim to understand how sensory stimuli are perceived and processed in the brain, and how the brain then interprets these stimuli to produce adaptive behaviours.

Because of the brain’s extraordinary complexity, these questions are difficult to address by looking at individual cells.  The flow of information through the brain relies on the coordinated activity of thousands or millions of cells, and on ensembles of neurons that are active simultaneously.  For this reason, our research involves imaging activity in thousands of cells, and seeking salient patterns of activity across these populations.  In a range of projects, we characterise the neurons and circuits that respond to various visual, auditory, and vestibular stimuli; that play a role in the integration of information from these modalities; and that filter sensory information to produce behaviour.

Optogenetics in zebrafish

We work in the zebrafish model system because of its strengths in genetics, behaviour, microscopy, and optogenetics.  Specifically, we use transgenic techniques to express genetically encoded calcium indicators (GECIs) or optogenetic proteins in specific parts of the zebrafish brain.  We then use selective plane illumination microscopy (SPIM) to observe the GECIs, which reveal activity across our cells and circuits.  We also use optical physics to produce holograms in the brain for optogenetics in our larval fish.  Find more details about our methods, including our house-built SPIM microscopes.

Circuits neuroscience

From a broad perspective, this work is part of a burgeoning field of circuits neuroscience. Over the coming decades, neuroscientists will make the leap from understanding how regions of the brain function to describing, in concrete terms, the cellular circuits that underlie perception and behaviour.  Our group hopes to contribute to this effort, specifically in describing how the senses work, how the brain produces a coherent representation of the outside world, and how information from the outside world is translated into appropriate actions.

Opportunities for new researchers

Our lab spans numerous experimental approaches and techniques, and as a result, we need all kinds of expertise.  Historically, most lab members have been trained as biologists, especially neuroscientists, but recently this has diversified as we have adopted new techniques. 

Specifically, as our microscopes have begun producing vast datasets containing activity from tens of thousands of neurons, we have a great need for people who can code solutions for image analysis, data mining, and the detection of salient patterns among these vast data.  As a result, we are particularly eager to bring in mathematicians and bioinformaticians.  We are also building our own microscopes and performing tricks on them like digital holography (see our methods).  This vastly increases our experimental capacities, but it also means that we need capable optical physicists on the team.  Of course, we are still happy to bring in biologists of all varieties.

Positions are available across our research program for undergraduates, honours students, PhD students, and postdocs, and we are happy to entertain new ideas.  If you are interested in joining the lab, please contact Ethan.

Current projects

Anatomical circuit mapping

We are interested in both the physical and the functional connections that underlie brain function, so we use both anatomical and optogenetic approaches for circuit mapping.  Anatomical approaches include imaging the architectures of individual neurons within a brain region of interest.  This allows us to view these brain regions as collections of discernable cell types, which is a prerequisite for explaining how those cells form functional circuits.

 Tectal periventricular neuron. See text below for more.

A single tectal periventricular neuron labelled the BGUG technique
(adapted from Scott et al., 2009).

Cerebellar projection neurons.  See caption for more explanation.

 Tracings of five cerebellar projection neurons to the tectum, and a spatially registered map of topography between the structures
(adapted from Heap et al, 2013).

 

We also use targeted photoconversions to discern patterns of projections among different brain regions.  By illuminating Kaede-expressing neurons with violet light, we can cause Kaede to change from a green to a red fluorophore.  If we target this photoconversion to particular cells and wait for the red Kaede to diffuse into the axons, we can identify these cells’ targets throughout the brain. 

 

Kaede photoconversions

tectal neuropil from a larva in which both the thalamus and retina are labelled with Kaede

These are images of the tectal neuropil from a larva in which both the thalamus and retina are labelled with Kaede. 
We have photoconverted Kaede in thalamic, but not retinal, projection neurons, allowing us to determine which layers of the tectal neuropil receive inputs from each. 
(Unpublished data from Lucy Heap.)

Sensory coding and integration

The mechanisms by which signals from the outside world are perceived, processed, and integrated are largely mysterious.  We hope to reveal some of these processes with whole-brain imaging of neural activity while our animals are experiencing sensory stimuli.  This involves expression of GCaMP6 (a genetically encoded calcium indicator developed at HHMI Janelia Research Campus) throughout the brain, combined with selective plane illumination microscopy (SPIM).  You can learn about our home-made SPIM microscopes and bioinformatics approaches on our Methods and Machines page.

To investigate a range of sensory modalities, we have designed microscopes that provide open space around the larva.  This permits us to surround the larva with LCD screens for visual stimuli, auditory speakers, and even microfluidics devices for generating water flow across the animal’s body.  We then look at activity across tens of thousands of neurons while these stimuli are presented alone or in combination.  The result is a map of what types of stimuli elicit what types of neural responses, where the responding neurons are, and what their relationships with each other are.  All of this moves us a little closer to understanding how these senses are encoded and integrated by the brain.

Sensory stimulation imaging

Diagram showing imaging setup.  Refer to text below for explanation.

 

Our imaging setup, showing a larva surrounded by two SPIM illumination objectives, in imaging objective, and an LCD screen with visual stimuli.  Auditory and other stimuli can also be presented.  The images resulting from the preparation must be segmented in MATLAB to identify the individual cells, and then these cells can be tracked individually for their responses to each stimulus.  The image on the right shows the tectal cells that respond to a particular visual stimulus. (Figure from Thompson and Scott, 2016)


 

Functional circuit mapping with optogenetics

The anatomical and imaging approaches described above still don’t concretely demonstrate how information passes through the brain circuits.  To address this, we need to test the functional relationships among the cells that we are studying.  For this, we turn to optogenetics.   By driving activity specifically in one brain region that we are interested in, and performing calcium imaging of the neurons in another region that may be connected, we can see whether and in what way one region’s activity affects cells in the downstream region.  Given the complex and interconnected nature of the nervous system, this does not necessarily mean that the stimulated cells and the responsive cells are synaptic partners (anatomical mapping is needed for that), but it does establish a functional relationship between one region and another.

Optogenetic circuit mapping

Image showing optogenetic circuit mapping.  See text below for explanation.

Optogenetic stimulation using sculpted laser light and Channelrhodopsin2 (blue circle, left) can be used to drive activity in a small patch of the cerebellum.  The resulting activation in tectal neurons is shown in green.  The right panel shows a raster plot of individual tectal cells that respond to this cerebellar stimulation and the average response of this population to three consecutive trials. (Unpublished data from Lucy Heap.)

Zebrafish models of Autism Spectrum Disorder

Psychiatric disorders necessarily have their origins in the dysfunction or loss of neural circuitry.  Our lab has recently begun modelling Autism Spectrum Disorder (ASD) by mutating genes linked to ASD in humans.  The strength of the zebrafish model system is that it permits access to the functioning brain, thus allowing analyses of developmental dynamics, population-level neural coding, and synapse dynamics in vivo. 

We are currently in the process of using CRISPRs to mutate a number of zebrafish homologs to ASD-linked human genes.  These mutants will be used to gauge the anatomical, developmental, and physiological mechanisms underlying ASD etiology, at the cellular level. 

2017

Gilles Vanwalleghem, Lucy Heap, and Ethan K. Scott (2017).
A profile of auditory-responsive neurons in the larval zebrafish brain.
Journal of Comparative Neurology, in press.

2016

Andrew W. Thompson and Ethan K. Scott (2016).
Characterisation of sensitivity and orientation tuning for visually responsive ensembles in the zebrafish tectum.
Scientific Reports, 6 (3487): 1-10 
URL: Thompson et al, 2016.

Lilach Avitan, Zac Pujic, Nicholas Hughes, Ethan K. Scott, and Geoffrey J. Goodhill (2016).
Limitations of neural map topography for decoding spatial information.
The Journal of Neuroscience. 36 (19): 5385-5396. 
URL: Avitan et al, 2016.

Andrew W. Thompson, Gilles C. Vanwalleghem, Lucy A. Heap, and Ethan K. Scott (2016).
Functional profiles of visual, auditory, and water flow responsive neurons in the zebrafish tectum. 
Current Biology. 26: 1-12. 
URL: Thompson et al, 2016.

Kelsey Chalmers, Elizabeth M. Kita, Ethan K. Scott, and Geoffrey J. Goodhill (2016).
Quantitative analysis of axonal branch dynamics in the developing nervous system.
PLoS Computational Biology.  12 (3). 
URL: Chalmers et al, 2016.

2015

Itia A. Favre-Bulle, Daryl Preece, Timo A. Nieminen, Lucy A. Heap, Ethan K. Scott*, and Halina Rubinsztein-Dunlop* (*co-corresponding).
Scattering of Sculpted Light in Intact Brain Tissue, with Implications for Optogenetics.
Scientific Reports. 5: 11501. 
URL: Favre-Bulle et al, 2015.

Jacob H. Hines, Andrew R. Ravanelli, Rani Schwindt, Ethan K. Scott, and Bruce Appel (2015).
Activity-dependent competition for axon selection during myelination in vivo.
Nature Neuroscience. 18: 683-689. 
URL: Hines et al, 2015.

Geoffrey J. Goodhill, Richard A. Faville, Daniel J. Sutherland, Brendan A. Bicknell, Andrew W. Thompson, Zac Pujic, Biao Sun, Elizabeth M. Kita, and Ethan K. Scott (2015).
The dynamics of growth cone morphology. 
BMC Biology.  13:10. 
URL: Goodhill et al, 2015.

Elizabeth M. Kita , Ethan K. Scott, and Geoffrey J. Goodhill (2015).
The influence of activity on axon pathfinding in the optic tectum.
Developmental Neurobiology. Published online February 2015.
URL: Kita et al, 2015.

Elizabeth M. Kita , Ethan K. Scott, and Geoffrey J. Goodhill (2015).
Topographic wiring of the retinotectal connection in zebrafish.
Developmental Neurobiology. 75(6): 542-556.
URL: Kita et al, 2015.

2013

Lucy A. Heap, Chi-Ching Goh, Karin S. Kassahn, and Ethan K. Scott (2013).
Cerebellar output in zebrafish: an analysis of spatial patterns and topography in eurydendroid cell projections.
Frontiers in Neural Circuits.  7: 53.
URL: Heap et al. 2013

Hugh D. Simpson, Elizabeth M. Kita, Ethan K. Scott, and Geoffrey J. Goodhill (2013).
A quantitative analysis of branching, growth cone turning and directed growth in zebrafish retinotectal axon guidance.
Journal of Comparative Neurology.  521: 1409-1429.
URL: Simpson et al. 2013

2012

Isabel Formella, Ethan K. Scott, Tom H.J. Burne, Lauren R. Harms, Ashley Liu, Karly M. Turner, Xiaoying Cui, and Darryl W. Eyles (2012).
Transient Knockdown of Tyrosine Hydroxylase during Development Has Persistent Effects on Behaviour in Adult Zebrafish (Danio rerio).
PLoS One. 7 (8).
URL: Formella et al, 2012.

Joshua Simmich, Eric Staykov, and Ethan K. Scott (2012).
Zebrafish as an appealing model system for optogenetics.
Progress in Brain Research. 196: 145-162. 
URL: Simmich et al, 2012.

Phil McClenahan, Michael Troup, and Ethan K. Scott (2012).
Fin-tail Coordination During Escape and Predatory Behavior in Larval Zebrafish. 
PLoS One. 7(2). 
URL: McClenahan et al, 2012

2011

Thomas Burne, Ethan K. Scott, Bruno van Swinderen, Massimo Hilliard, Judith
Reinhard, Charles Claudianos, Darryl Eyles, and John McGrath (2011).
Big ideas for small brains: what can psychiatry learn from worms, flies, bees and fish?
Molecular Psychiatry. 16: 7-16. 
URL: Burne et al, 2011.

2010

Filippo Del Bene, Claire Wyart, Estuardo Robles, Amanda Tran, Loren Looger, Ethan K, Scott, Ehud Y. Isacoff, and Herwig Baier (2010).
Filtering of visual information in the tectum by an identified neural circuit.
Science. 330: 669-673.
URL: Del Bene et al, 2010.

Linda Nevin, Estuardo Robles, Herwig Baier, and Ethan K. Scott (2010).
Focusing on optic tectum circuitry through the lens of genetics.
BMC Biology. 8: 126.
URL: Nevin et al, 2010.

Shuichi Kani, Young-Ki Bae, Takashi Shimizu, Koji Tanabe, Chie Satoh, Mike Parsons,Ethan K. Scott, Shin-ichi Higashijima and Masahiko Hibi (2010).
Proneural gene-linked neurogenesis in zebrafish cerebellum.
Developmental Biology. 343 (1-2): 1-17.
URL: Kani et al, 2010.

2009

Claire Wyart, Filippo Del Bene, Erica Warp, Ethan K. Scott, Dirk Trauner, Herwig Baier and Ehud Isacoff (2009).
Optogenetic dissection of a behavioural module in the vertebrate spinal cord.
Nature. 461: 407-410. 
URL: Wyart et al, 2009.

Herwig Baier and Ethan K. Scott (2009).
Optics and genetics cross paths in the zebrafish nervous system.
Current Opinions in Neurobiology. 19(5): 553-560. 
URL: Baier and Scott, 2009.

Ethan K. Scott and Herwig Baier (2009).
The cellular architecture of the larval zebrafish tectum, as revealed by Gal4 enhancer trap lines.
Frontiers in Neural Circuits. 3(13).
URL: Scott and Baier, 2009.

Ethan K. Scott (2009).
The Gal4/UAS toolbox in zebrafish: New approaches for defining behavioral circuits.
Journal of Neurochemistry. 110(2): 441-456.
URL: Scott, 2009. 

Lisette A. Maddison, Jainjun Lu, Tristan Victoroff, Ethan K. Scott, Herwig Baier, and Wenbiao Chen. (2009).
A gain-of-function screen in zebrafish identifies a guanylate cyclase with a role in neuronal degeneration.
Molecular Genetics and Genomics. 281(5): 551-563.
URL: Maddison et al, 2009.

2007

Stephanie Szobota, Pau Gorostiza, Filippo Del Bene, Claire Wyart, Doris L. Fortin, Kate Kolstad, Orapim Tulyathan, Matthew Volgraf, Rika Numano, Holly Aaron, Ethan K. Scott, Richard Kramer, John Flannery, Herwig Baier, Dirk Trauner and Ehud Isacoff (2007).
Remote control of neuronal activity with a light-gated glutamate receptor.
Neuron. 54(4): 535-545.
URL: Szobota et al, 2007.

Ethan K. Scott, Lindsay Mason, Aristides B. Arrenberg, Limor Ziv, Nathan J. Gosse, Tong Xiao, Neil C. Chi, Kazuhide Asakawa, Koichi Kawakami, and Herwig Baier (2007).
Targeting neural circuitry in zebrafish using GAL4 enhancer trapping.
Nature Methods. 4(4): 323-326.
URL: Scott et al, 2007.

Lab Head

 

Staff

Students

Targeted gene expression

Our lab is interested in neural circuit function, and we use genetically encoded tools to describe the anatomy and patterns of activity in these circuits.  This requires us to express exogenous genes in targeted parts of the zebrafish nervous system. As a postdoc with Herwig Baier at UC-San Francisco, Ethan led a group of researchers in a Gal4 enhancer trap (ET) screen.  The screen generated nearly 200 lines of zebrafish expressing Gal4 in subsets of the nervous system, and these lines are an important part of how our lab targets proteins to circuits of interest.  The lines can be ordered from the Zebrafish International Resource Center, and images can be found at ZFIN.org.  Each line has a four digit identifier (Gal4s1000t, for example) that can be used in searching ZFIN.
 

Gal4 ET lines
A smattering of the lines from the San Francisco ET screen.
More details can be found in:  Scott et al, 2007Scott et al, 2009Scott and Baier, 2009 and  ZFIN.org.


Since Gal4 can be used, in principle, to express any UAS-linked transgene, we have great flexibility with what we can express in the circuits from our Gal4 lines.  Examples include photoconvertible proteins, markers for subcellular compartments, optogenetic indicators of neural activity, and optogenetic proteins for activating or inactivating neurons.  Details of these anatomical and functional approaches are described on the lab’s Projects and Opportunities page.

Selective Plane Illumination Microscopy (SPIM)

SPIM is an approach in which a collimated laser beam is shaped into a thin plane of light, which is projected into a specimen.  Fluorescence then occurs in this plane, and the emitted light is imaged from an orthogonally-oriented imaging objective. Our design is customised for high-speed brain-wide calcium imaging in zebrafish, and incorporates open space around the larva to permit a range of sensory stimuli to be applied. 
 

SPIM diagram

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The SPIM principle - a plane of excitation light is formed in the specimen, and emissions are imaged at 90°. (Image: openspim.org, CC3.0)



Imaging components

Labelled illumination path

The image shows the illumination light path comprising laser launch, beam expander, a 50:50 splitter, and then two plane paths with a mechanical slit, a cylindrical lens, and an illumination objective. 

  • The 3D-printed imaging chamber (centre) has coverslip walls, and holds a fish larva in position to be illuminated by the planes.  A vertical imaging objective captures fluorescent emissions. 
  • Our imaging column (right) is as simple as possible: a 20x water-immersion lens, an emissions filter, a focusing lens, and a camera. 
  • The only additional components are two empty filter cubes.  These allow us to put light into the specimen through the imaging objective and to project holograms into the brain for optogenetics.  

 

A complete parts list for a simpler version of this microscope can be found in the supplemental information accompanying Thompson et al, 2016.  If you have any questions about parts or construction, feel free to contact Ethan.

Optical physics

The filter cubes on the imaging column allow us to apply light to the specimen through the imaging objective.  Spatial Light Modulators (SLMs) are devices that modulate the wavefront of coherent light resulting in a pattern of constructive and destructive interference.  With an SLM, therefore, we can target and sculpt light in the brain while performing calcium imaging.  By applying a Gerchberg-Saxton algorithm, we can produce arbitrary three-dimensional shapes within the brain, allowing for spatially controlled optogenetic manipulations.

sculpted beam focusing through agarose


A schematic (left) for how SLM-sculpted light is projected into the specimen, and an image (right) of sculpted light emerging from the objective.  These images are  adapted from Favre-Bulle et al, 2015, where we describe this technique in greater detail.
 


Find out more about our research environment and how to apply to do a short or long-term research project with us.