beginning of tom’s slides...julie j. mirzabekov, kelly a. zalocusky, aleksandra k. denisin joanna...
TRANSCRIPT
Michael Faraday Inventions
1791-1867
Gauss-Maxwell-Heaviside
1850-1950
1777–1855
1831-1879
Ernest Rutherford
1871-1937
Ruska-Knoll Microscope
1906–1988 1897-1961
Hodgkin-Huxley Model
1914–1998 1917-2012
Hubel-Wiesel Experiments
1925-2013 1924-
Optogenetics
. . .
Kenneth J. Hayworth, C. Shan Xu, Zhiyuan Lu, Graham W. Knott, Richard D. Fetter, Juan Carlos Tapia, Jeff W. Lichtman, and Harald F. Hess. Ultrastructurally smooth thick partitioning and volume stitching for large-scale connectomics. Nature Methods, 12:319-322, 2015.
Ultrastructurally Smooth Thick Partitioning and Volume Stitching for Large-scale Connectomics
Overview• EM material preparation!
– Perfusion-fixation!– Staining and embedding!– Light and EM correspondence!
• Manual serial-sectioning!• Reconstruction!
– Imaging!– Registration and alignment!– Segmentation!
• Combining network structure and function
. . .Davi Bock Clay Reid
. . .
Sebastian SeungViren Jain
Mice! Low Level AnalysisCalcium Physiology
Cell Type Staining / Histology!
ATUM / ATLUM! TEM! Volume Store: Petabyte Scale!
3-D Electron!Micrographs!
!
Database Automated Segmentation
Networks !Functional
Neuroanatomy
Human SegmentationAnnotation Tools
Alignment!Stitching!Circuit!Tracing!
x 104
Online DatabaseComputational!Models:!
Neural Circuits!Algorithmic Insights
Algorithms:!Signal Processing!
Robotics!Computer Vision
Functional Connectomics
Forceps for Grids
Heat Plate
Eye LashesVideo !
Recorder
Draught Shield
Diamond !Knife Boat
Cutting ArmVideo!
Camera
Air Table &!Arm Rests
Specimen
Water Spout
Water Boat
Diamond Knife Edge
Cutting Arm
Resin BlockTrimmed Specimen
22
ATLUM: Automatic Tape-collecting !Lathe Ultra Microtome
TEM Camera Array
CCD Camera Array
Large Scintillator
film chamber vacuum extension
motorized positioner knobs
9” custom scintillator
leaded glass
2 x 2 array of f2.0 optical lenses with 50 MHz 11 MegaPixel CCDs
camera trigger breakout box
lead shielding
cables to four acquisition PCs
camera array + scintillator closeup
Overview• EM material preparation!
– Perfusion-fixation!– Staining and embedding!– Light and EM correspondence!
• Manual serial-sectioning!• Reconstruction!
– Imaging!– Registration and alignment!– Segmentation!
• Combining network structure and function
. . .Davi Bock Clay Reid
. . .
Sebastian SeungViren Jain
Fly EM @ HHMI Janelia
Data from [www.janelia.org/team-project/fly-em] [Thanks to Davi Bock]
Mouse Visual Cortex
From Bock et al., Nature 471(7337), 2011. [www.openconnectomeproject.org/]
raw camera data, c1-4
normalized camera data, c1-4
stitched tile, whole scintillator
stitched tile, inscribed square
moving the motors, raw camera data
moving the motors, normalized
moving the motors, stitched tiles
moving the motors, inscribed squares
zooming in 1
zooming in 2
zooming in 3
Reconstructing 180µ3 of Rat Hippocampus
Kinney JP, Spacek J, Bartol TM, Bajaj CL, Harris KM Sejnowski TJ , Extracellular sheets and tunnels !modulate glutamate diffusion in hippocampal neuropil, Journal Computational Neurology 521: 448-464!
Light Sheet Fluorescent Microscopy
Tsai-Wen Chen, Trevor J. Wardill, Yi Sun, Stefan R. Pulver, Sabine L. Renninger, Amy Baohan, Eric R. Schreiter, !Rex A. Kerr, Michael B. Orger, Vivek Jayaraman, Loren L. Looger, Karel Svoboda and Douglas S. Kim (2013) !Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature 499: 295–300
CLARITY + Immunofluorescent Labeling
Kwanghun Chung, Jenelle Wallace, Sung-Yon Kim, Sandhiya Kalyanasundaram, Aaron S. Andalman, Thomas J. Davidson, Julie J. Mirzabekov, Kelly A. Zalocusky, Aleksandra K. Denisin Joanna Mattis, Sally Pak, Hannah Bernstein, Charu Ramakrishnan, Logan Grosenick, Viviana Gradinaru, and Karl Deisseroth. Structural and molecular interrogation of intact biological systems. Nature, 10, 2013.
Miniature Fluorescence Microscope
Barretto RPJ, Messerschmidt B, Schnitzer MJ. (2009) In vivo fluorescence !imaging with high-resolution microlenses. Nature Methods. 6(7):511-512.
Lobster Stomatogastric Ganglion
Gerhard, Felipe, Kispersky, Tilman, Gutierrez, Gabrielle, Marder, Eve, Kramer, Mark, Eden, Uri. Successful reconstruction of a physiological circuit with known connectivity from spiking activity alone. PLoS Compututational Biology, 9(7):e1003138, 2013.!
Mouse Cortex Thickness
Monte-Carlo Molecular Simulation