Instructor: Ryuichi Shigemoto, Sandra Siegert
Teaching Assistant: Felipe Fredes, David Kleindienst, Marijo Jevtic, Marco Benevento
At the beginning of the course, each student is asked to select a brain region of interest.
Theoretical part: Students are introduced into classical concepts, recent experimental strategies and technologies in two brain regions, hippocampus and the retina. In parallel, students will analyze whether these principles are also applicable in their chosen brain region. This requires from the student to perform literature search and to develop strategies how to reveal information and critically access them.
Practical part: Students are introduced into the principles and caveats of commonly used morphological methods, develop research and analysis skills, and cultivate critical eyes for light microscopic images.
In detail, students will
- design a cloning strategy and perform it in silico.
- design an immunofluorescent staining and are introduced to all the caveats of a staining.
- perform stereotactic injection of adeno-associated virus (AAV) expressing GFP/tdTomato or channelrhodopsin-GFP into the region of interest. The students will learn the basics of virus design (which virus to use (antereo-/ retrograde), how to strategize to perform animal experiments (anesthesia/ analgesia), and how to target a distinct brain region by identifying stereotactic coordinates. Six weeks after the injection, we will examine whether the students could successfully target the brain region-of interest site and reveal the projection areas. We will perform sectioning of the brain and perform immunolabeling for cell-type specific markers to analyze target cells.
A major goal is that students will be capable to develop a research proposal outlining the following:
This course is for all students, who wish to affiliate to a neuroscience group as well as it is open to all life scientists. However, the course pre-requests a solid knowledge in neuroscience e.g. terms such as “neuronal circuit”, “spike”, “synaptic release”, “excitatory and inhibitory neurons” should be known.
Audit of the lecture is permitted and welcomed. However, only students, who are taking the course for credit, will be allowed to perform the practical part. These students have to proof that they are allowed to perform animal experiments. They have either to to show their certificate confirming their successful attendance of the "Animal handling course at IST Austria" or have to attend the Neuro Preperatory Course on the 22nd-23rd February. In doubt, please contact Joana ALMEIDA (email@example.com).
On Wednesday, February 15th, the course instructors will hold a general preliminary meeting (14:00-15:00, Lab Building East seminar room ground floor). Every student, who takes the course for credit, has to attend to this pre-meeting. Here we will go over the syllabus and will discuss organisatory details.
- Active participation of the student is required as well as full attendance to each class (exceptions have to be discussed with the course instructors).
- Several lectures have reading assignments and it is expected that each student read the paper(s) before the lecture.
- The students are asked to write a 2-3 page research proposal about their brain region-of-interest and submit it by the end of the course. Before submission, they will present the proposal in a 5-min chalk-talk with a 10-min questions-and-answer session. To prepare for this research proposal, each student should prepare after each lecture a summary of the topic/ new findings and identify similar studies in their brain region-of-interest. The results may be presented in the recitation (Depending on the amount of time of discussion: random selection of the people to present).
|15. Feb (Wed)||Pre-meeting to discuss course syllabus||14:00-15:00, LBE Seminar room (ground floor)||RS+SS|
|If not otherwise indicated:||Tue/Thu (10:15-11.30), Recitation Thu (11.45-12.35)|
|28. Feb (Tue)||Introduction to neuronal cell types, Part I: Brain circuits consisiting of different cell types; Functional analysis of identified neuronal cell types||LBE Seminar room (ground floor)||Lecture 1||RS|
|02. Mar (Thu)||Introduction to neuronal cell types, Part II: Strategies to identify cell types anatomically||LBE Seminar room (ground floor)||Lecture 2||SS|
|02. Mar (Thu/ Recitation)||Students will design an immunohistochemical experiment and identify common pitfalls||LBE Seminar room (ground floor)||Practical 1||SS||Bring your laptop|
|07. Mar (Tue)||Introduction to neuronal cell types, Part III: Strategies to Identify cell types electrophysiologically||LBE Seminar room (ground floor)||Lecture 3||SS|
|09. Mar (Thu)||Basic principles of cloning: Promoter selection, Linkers, Resources (Students will perform in silico cloning of their future construct for the practical part for the virus injection)||LBE Seminar room (ground floor)||Lecture 4||SS||Bring your laptop. Have "SerialCloner" installed (http://serialbasics.free.fr/Serial_Cloner.html)|
|09. Mar (Thu/ Recitation)||Student plans for discussing their brain region of interest||LBE Seminar room (ground floor)||Recitation||RS|
|14. Mar (Tue)||Principles of classical dissection of neuronal circuits, Part I: Classical and virus-mediated tracing, Cre-lox-system, Trans-synaptic tracing using Rabies virus, Allen Brain Atlas (Students learn to use web sites for planning tracing and optogenetic experiments||LBE Seminar room (ground floor)||Lecture 5||RS|
|16. Mar (Thu)||Principles of classical dissection of neuronal circuits, Part II: Transgenic mice, 2-photon whole cell patch clamping, Drug-induced circuit manipulation||LBE Seminar room (ground floor)||Lecture 6||SS|
|16. Mar (Thu/ Recitation)||Discussion about brain region of interest||LBE Seminar room (ground floor)||Recitation||SS+TA|
|21. Mar (Tue)||Hippocampal cell types and circuits: Optogenetics, Pharmagogenetics||LBE Seminar room (ground floor)||Lecture 7||RS|
|23. Mar (Thu)||Novel strategies to dissect neuronal circuits, Part I||LBE Seminar room (ground floor)||Lecture 8||SS|
|23. Mar (Thu/ Recitation)||Discussion about students plan for practical||LBE Seminar room (ground floor)||Recitation||RS+TA|
|28. Mar (Thu)||Novel strategies to dissect neuronal circuits, Part II||LBE Seminar room (ground floor)||Lecture 9||SS|
|30. Mar (Thu)||Detection of neuronal activity: in vivo electrophysiology, Ca imaging, IEG expression, Mouse behavior experiments||LBE Seminar room (ground floor)||Lecture 10||RS|
|30. Mar (Thu/ Recitation)||Planning of sterotaxic injection experiment, Paper discussion||LBE Seminar room (ground floor)||Recitation||RS|
|31. Mar (Fri)||AAV injection||TBA||Practical 3||RS|
|04. Apr (Tue)||Molecular neurobiology Part I - Epigenetics matters: miRNA, environmental factors influencing gene function||LBE Seminar room (ground floor)||Lecture 11||SS|
|06. Apr (Tue)||Molecular neurobiology Part II - Genetic matters: Gene profiling of cell types, Gene networks, Single cells||LBE Seminar room (ground floor)||Lecture 12||SS|
|06. Apr (Thu/ Recitation)||Discussion about "how to write a research proposal"||LBE Seminar room (ground floor)||Recitation||SS+TA|
|02. May (Tue)||Molecular neurobiology Part III - Gene level modification and genome editing, Part A, iRNA||LBE Seminar room (ground floor)||Lecture 13||SS|
|04. May (Thu)||Molecular neurobiology Part IV - Gene level modification and genome editing, Part B, CRISPR/Cas9||LBE Seminar room (ground floor)||Lecture 14||SS|
|04. May (Thu/ Recitation)||Design your own CRISPR/Cas9 mediated genome editing experiment in silico||LBE Seminar room (ground floor)||Recitation||SS||Bring your laptop|
|09. May (Tue)||Molecular neurobiology Part V - Gene level modification and genome editing, Part C, CRISPRi/CRISPRa||LBE Seminar room (ground floor)||Lecture 15||SS|
|11. May (Thu)||Correlation and causal link - is optogenetics useful? Critical assessment of selected papers||LBE Seminar room (ground floor)||Lecture 16||RS|
|04. May (Thu/ Recitation)||Planning experiments||LBE Seminar room (ground floor)||Recitation+Practical||RS|
|12./ 13. May (Fri/Sat)||Optogenetics and perfusion||TBA||Practical 4||RS+TA|
|16. May (Tue)||Identification of neuronal cell types: immunohistochemistry (Students apply knowledge for Friday for their sections).||LBE Seminar room (ground floor)||Lecture 17||RS|
|18. May (Thu)||Back-up||LBE Seminar room (ground floor)||Lecture 18||RS|
|18. May (Thu/ Recitation)||How to take good images/ pictures?||LBE Seminar room (ground floor)||Recitation||TA|
|19. May (Fri)||Observe fluorescence expression, collect data (In case of immunlabeling, secondary antibody, then viewing and collecting data)||TBA||Practical 5||RS+TA|
|23. May (Tue)||Back up for any additional experiments or discussion on the results||LBE Seminar room (ground floor)||Lecture 19||RS|
|30. May (Tue)||Do cell types matter or is this concept overstated?||LBE Seminar room (ground floor)||Lecture 20||SS|
|01. Jun (Thu)||Image Data Analysis||Meeting at workstations in 04. Eg.035||Practical 6||RS+SS|
|01. Jun (Thu/ Recitation)||Troubleshooting of Image Data Analysis||TBA||Practical||RS+SS|
|02. Jun (Fri)||DEADLINE for PROPOSAL SUBMISSION|
|08. Jun (Thu)||Exam||TBA||RS+SS|
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