Hello, my name is Katie Pumayalli and I just finished the second week of my six-week internship at the Ye Lab at Scripps Institute in San Diego, California. I have been working with my amazing mentors Cailynn Wang and Zhengyuan (Ben) Pang. I started my mentorship on Monday, June 17th, and fell in love with the lab and all of the people. My mentor’s research focuses on using a technique called clearing-assisted tissue click chemistry (CATCH) to track hallucinogenic drugs within brain tissue. CATCH involves making brain tissue slices transparent by removing lipids (fats) from the cells, while still preserving the overall cellular structure. This technique allows for precise visualization of where the drugs bind within the brain. This week I mainly focused on learning about the Click chemistry and doing steps of the process to learn for next week’s protocol.
Harvested brains in gelOn Monday, my day began by listening to a presentation of a lab member’s data update. While listening, I learned new things, especially about the different types of fat in our bodies and how to differentiate them. After lunch, my mentor, Cailynn, showed me how to slice a brain to create thin sections for the Click chemistry reaction, in preparation for further analysis such as immunohistochemistry.
Brain Tissue
On Tuesday, I learned the final steps of the Click reaction process. This involved washing the tissue and staining it with a fluorescent dye called DAPI, which specifically targets the nucleus. This step allows us to visualize how the drug affects different neurons by comparing the fluorescence patterns of the drug (with a different dye) and the nucleus.
Brain tissue in overnight solution
Wednesday began with video tutorials and an article on tissue clearing. The article explained the specific chemical reactions that render tissue transparent. After delving deeper into the theoretical aspects of our lab’s process, I began researching the mouse brain to identify my area of interest. Armed with this knowledge, I dissected brain tissue from various mice, allowing me to initiate the Click reaction process from the very beginning.
Full brain tissue (I was even allowed to keep this!)
Image of how the drug affects the Hippocampus
On Thursday, I began by mixing reaction buffer for the Wednesday tissue samples, initiating the final stages of the Click process. After the reaction, I cleaned the tissue with a solvent and stained the nucleus with DAPI for mounting on a slide. Finally, I excitedly took the slide to the imaging room with my mentor to see if the Click chemistry I performed independently was successful. To my relief, the slide showed clear patterns, allowing me to visualize the effects of the PF-04457845(PF) drug.
I am handling the mouse
Brain I harvested
On Friday, to prepare for the day’s mouse perfusion, I started by reviewing the protocol, which outlined the steps for injecting a solution into the mouse’s circulatory system, typically through the heart or a major blood vessel. This process delivers substances throughout the body and allows researchers to manipulate the animal’s physiology for various studies. At eleven, I joined half of the lab to attend a lecture by Akiko Iwasaki on Lung Covid. Later, I was entrusted with my first perfusion! I injected two mice with a serum into the abdominal cavity, to prepare them for the actual perfusion process. Then I performed the actual perfusion and harvested their brains.
Siena Parr
Ferris Wheel at the Fair
Last Sunday Siena and I went to the San Diego County Fair to explore more of what it means to be in San Diego during the summertime. This allowed us to have a much-deserved break to help calm our brains.
Thank you for reading in. Can’t wait for next week’s adventures!
Katie P
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