January 13 - January 25, 2017
This is an intensive and comprehensive laboratory-oriented course focusing on applying imaging techniques to neuroscience research. The objective of this imaging course is to gain exposure to modern imaging tools from principle optics to applications in modern neuroscience.
The course has been formatted to include lectures in the morning and laboratory sections in the afternoon and evening. Students will rotate through all laboratory sections, and focus on one section (and one optional section) for the remainder of the course.
This course features a dedicated CLEM Electron Microscopy Techniques Module. If you wish to work on a CLEM project, please be sure to select the EM Course track in the application.
This course is intended for anyone interested in using imaging in their neuroscience research, including pre-doctoral students, postdoctoral or clinical researchers and young investigators in the beginning of independent research careers.
When you click on the “APPLY” button, you will be redirected to an online application form. You must complete all of the information on this form before you submit, as you will not be able to re-enter the form.
Please be aware that the form will request the following (be sure to upload all documents in pdf format):
- Research Statement: In approximately 500 words, please tell us why you are interested in this course, and how this course will contribute to your ongoing or future research.
- Contact information for 2 referees (name, institutional affiliation, and email address). Please instruct referees to submit letters via email directly to Jessica Herbst (firstname.lastname@example.org) no later than the application deadline.
- For CLEM EM Module applicants, please include answers to the following questions:
- Why are you interested in learning CLEM?
- What kind of specimens (e.g. brain tissue, slice culture, cells, etc.) do you want to work on?
- List previous EM experience
If you are chosen to participate in the course, you will receive an email that contains detailed registration and payment instructions. At this time, information will also be available to assist you in making travel plans.Fees:
Registration fees include all aspects of course participation and all meals. Those who register for a “shared room” will be housed in a double-occupancy hotel room and be provided transportation to and from the course location. Those who register for a “single room” will be housed in a single-occupancy hotel room and be provided transportation to and from the course location.
- Registration only: $2,000
- Registration Fee + Single Room: $3,755
- Registration Fee + Shared Room: $2,878
- Imaging CA1 neurons and Spines in live animals with the high NA GRIN lens in combination with 2-photon microscopy.
- Surgical preparation for deep brain imaging will be described and performed.
Reference: Impermanence of dendritic spines in live adult CA1 hippocampus. Attardo A, Fitzgerald JE, Schnitzer MJ. Nature. 2015 Jun 22. doi: 10.1038/nature14467
- Understand principle and setup of temporal focused two-photon microscopy.
- Use diffraction grating to control the lateral and axial profiles of a pulsed laser beam.
- Single cell resolution circuit mapping by temporal focused two-photon excitation in combination with novel spatially-restricted channel rhodopsin in brain slices.
- Understand basic principle of 2-photon deep tissue imaging.
- In vivo functional imaging using GCamp-6 under 2-photon in vivo imaging.
- Volumetric imaging of populations of neurons in layer 2/3 of ferret visual cortex at single cell resolution.
- Image neurons labeled with GFP in electron microscopy using the automatic tape collecting ultramicrotome (ATUM) in combination with scanning electron microscopy (SEM).
- Learn shuttle-and-find technique to correlate light and electron microscopy images.
- Demonstration of high-pressure freezing / freeze-substitution and embedding.
- Demonstrate how to study and analyze neuronal plasticity at individual synapses.
- Simultaneous two-photon imaging and two-photon glutamate and/or GABA uncaging on individual excitatory and inhibitory synapses visualized with specialized structures.
- Preparation of slice culture, biolistic gene gun transfection, whole-cell patch clamp recording, imaging tissues under two-photon microscope, and photolysis of caged neurotransmitters at single synapse while monitoring calcium or EPSCs/IPSCs.
Reference: Kwon HB and Sabatini BL., (2011) Glutamate induces de novo growth of functional spines in developing cortex. Nature 474 (7349):100-4.
- Understand principle of FRET and FLIM.
- Learn design principle of new FLIM-FRET sensors.
- Image activity of proteins and protein-protein interaction in single dendritic spines in slice culture / in vivo using 2-photon FLIM.
Reference: H. Murakoshi, H. Wang, R. Yasuda. Localized, persistent activation of Rho GTPases during long-term structural plasticity induced in single dendritic spines. Nature, 472:100-4 (2011)
- Understand principle and setup of 2-photon microscopy to visualize single dendritic spines in live animals.
- Image structural plasticity of dendritic spines in live animals over days.
- Demonstrate thin-scull and cranial window preparations.
Reference: Fu M, Yu X, Lu J and Zuo Y (2012) Repetitive motor learning induces coordinated formation of clustered dendritic spines in vivo. Nature 483(7387):92-95
- Filter, convolution
- Handling background
- Simple programing with ImageJ and MatLab
- Learn basic optic alignment technique: walk the beam, set up beam expander.
- Detailed discussion on 2-photon/confocal system components (optical, hardware and software)/troubleshooting.
- Learn system performance tests such as PSF test and FOV test.
- Building a two-photon microscope.
- Drawing an optical ray
- Designing a scanning microscope