Week Three

The past week at the Arizona Biomedical Collaborative has been quite a unique and enlightening experience.

But it started normally though.

On Monday, Caitlin, our secondary advisor, and I ran some more Wes plates. Since we had finished running Wes plates testing for Neuroligin-1 and Glypican 4 in the VPM (ventral posteromedial nucleus) region of mice, we then proceeded to run Wes plates for the S1BF region. After running two plates, I then did some housekeeping work and proceeded to call it a day.

On Tuesday, Caitlin was not in the lab, so my main supervisor Sarah Ogle decided that I assist the Research Lab manager, Bret, on his study involving cage dividers and the reduction of aggression on mice in their cages. So, for the rest of the day, I assisted Bret by accounting for the amount of time a specific rat in the cage spent in a tunnel created by the cage divider.

On Wednesday, Murphy’s Law reared its head.

At 9, Sarah told me to gather samples that we needed for protein extraction of sample tissues from certain regions. But the whole process soon turned into a fiasco due to both mistakes on my part as well as misplacement of sample tissues in previous studies. Truth be told, it took us nearly two and a half hours to find all of our samples top run a protein extraction and assay. After a nerve racking rest of the day, I retired at my usual time.

On Thursday, in lieu of the recent developments (see Wednesday), Sarah assigned me the task of noting down which samples we could find for the hippocampus region of the brain. The region had been used by many different people and as a result were scattered across the lab for me to find. After a few hours of noting down, I then had to leave early to make it back to school.

Till next week, signing off.

Week Two

My second week at the Arizona Biomedical Collaborative can be best described as a wacky and unique experience.

The week started normally enough.

On Tuesday, I, with the help my research supervisors Sarah and Caitlin, ran some more Wes plates. That day, I tested for the presence of neuroligin-1 in the prepared samples we were given. This procedure, compared with the procedure for glypican-4, takes considerably less time. The glypican-4 denaturation takes place at 37 degrees Celsius for 30 minutes plus heating up the heating bloc to 95 degrees Celsius for the biotinylated ladder control for 5 minutes with all the other standard procedures in place. In contrast, the neuroligin-1 denaturation process takes place at 95 degrees Celsius along with the ladder for 5 minutes. But technicalities aside, the runs were successful and I have completed quite some progress in two weeks.

On Wednesday, Caitlin did not show up to help us with the Wes procedure. So to pass the time working in the lab, I managed to double down slides by putting slides from one box in the freezer into another box to save space in a 4 degree Celsius room for three hours. It was not a job I expected to do, but one that I learned was vital to the organization of the lab.

But the highlight for the week was Thursday.

On Thursday, the labs hold their weekly lab meetings, where they review orders of business along other items on the agenda. This lab meeting, after all the items were taken care off, one of the lab regulars gave a presentation on an article that a reputable neuroscience journal had published related to the research in the lab. The lab regulars then proceeded, to much hilarity, rip the article into pieces regarding its factual assumptions and premises. Though the experience in general was hilarious, it taught a valuable lesson in what NOT to do in a research study.

On Friday, I hope to resume running Wes plates again, albeit for a different region.

Till next week, signing off.

Week One

My first week at the Arizona Biomedical Collaborative at the Translational Neurotrauma Lab can be wholesomely described as an entirely new and alien experience.

Before joining the research efforts at the ABC, I had a very preconceived notion about the nature of research. I thought that the professor controls everything within the lab and dictates every responsibility to high school interns out of a sense of untrustworthiness. Also, I thought that to pursue research was to throw yourself into the fire immediately with some training and you proceeded under the watchful eye of the professors and lab interns.

All of these notions were proven a mix of truths and untruths.

But anyway, updates on the past week.

On Monday, I learned all about the materials needed to properly run the Wes machine and where they were all stored, which is important information to consider for experiments.

On Wednesday, I ran a sham plate for the samples from the VPM region of the brain for Sprague Dawley rats to ascertain which samples would be appropriate to run with the injured samples.

On Thursday, I participated in a lab meeting with all the personnel in the lab. There, we discussed specific projects that other lab personnel were pursuing as well as addressing problems and solutions to many issues in the lab. Afterwards, we watched a presentation on a study on the reduction of aggression in mice after the addition of a cage divider, where it was ascertained that a cage divider heavily affected aggression levels in mice. Then, lab personnel offered other avenues to pursue as well as improvements to make on the experiment. After the lab meeting, my advisor then again extensively reviewed the steps needed to run the Wes machine again and I wrote the steps in my lab notebook.

I hope that on Friday, I can finally run Wes plates on our own and be well on my way to maximizing our data.

So that’s the update for the week.

Till next week, see you all later.

Project Summary

It is common knowledge that traumatic brain injury (TBI) leads to increased synaptogenesis, which can amplify the traumatic sensations caused by diffuse TBI due to the increase in nerve impulses. The traumatic sensations that can be triggered by increased synaptogenesis initiated by diffuse TBI into memory related areas (Hippocampus, PFC, Amygdala) include loss of memory and difficulty of memory retrieval. Using a streamlined version of the western blot technique, I will measure the impact of diffuse TBI on the levels of neuroligand-1 and glypican4, protein factors essential in the regulation of synaptogenesis in CNS neurons, present in memory related brain areas over a two-month time period. The results can prove to be essential in determining factors that play a vital role in stimulated synaptogenesis while also proactively aid in finding a solution for post -TBI symptoms in the memory related areas of the brain.

Terms to know

Wes machine: A machine that has streamlined the traditional Western blot process in our Translational Neurotrauma Lab. For those who are not familiar with a Western blot procedure, it is a widely used  technique for protein analysis that utilizes both gel electrophoresis to separate out the denatured proteins by the length of the polypeptide as well as the transfer to a membrane that stains the protein with an antibody specific to the target protein. Usually, the Western blot process takes about 2 days to complete, taking into account the  time needed to create the gel among other things. This Wes process shortens the time frame to about 6 hrs to gather data from one plate.

Neuroligand-1: protein factor in the brain that stimulates the production and redirection of synapses after traumatic brain injury using thrombospodin 1.

Glypican-4: astrocyte secreted signals sufficient to induce functional synapses between ganglion cells in the human brain. Have similar effect like neuroligand-1

Sham: Designates a sample from a region from an uninjured animal. Acts as a baseline for comparison to injured animal samples.

Project Goals

• Examine amount of protein factors in select tissue samples from Sprague Dawley rats at specified days after TBI
• Hopefully aid in research into factors controlling increased synaptogenesis after TBI.