Yesterday (12/14) was our slug lab and neuroscience holiday party. Please ignore the fact that we had food in the lab; focus instead on the festive fun time (complete with a crackling fireplace broadcast on the lab projector… toasty!).

We celebrated the graduation of our mid-term neuroscience majors

• Steven Proutsos, who is graduating with a 4.0! (well, hopefully, depending on how he does on C-J’s molecular biology final) and just scheduled his MCAT
• Christian Gonzalez, who just accepted a job working as a research technician in a neuroscience lab at Rush medical school!

We also celebrated birthdays, off-campus departures, successful TA-ships, the holiday, and anything/everything else worth celebrating about our wonderful lab students and neuroscience majors (a few non-majors even snuck in; the more the merrier!)

Here was the scene:

CJ, Fiona, Stephanie, Me, Gold, Michele, Live, Christian, Jaqueline, Stephen, Brian, Jas, Delaney, Theresa and Zayra!

Yesterday, two Slug lab members had their first chance to present their ongoing research projects, premiering their work at the 2022 PUMA-STEM summer research conference:

• Jaqueline Gutierrez presented work on very long-lasting sensitization in Aplysia
• Monica Lopez presented work on developing earthworms to study long-term sensitization

It was a great event, and both Jaqueline and Monica did themselves proud–they had really nicely designed posters, presented with confidence, and did a fantastic job fielding question.

Go slug lab!

It’s July of 2022 and for the past 2 months the SlugLab has been lurching back into life.

For the first time since 2019, the sluglab welcomed a new cohort of summer research students: a record 10 students!

We’ve so far been confronting the many problems associated with getting the lab back up and running. All previous SlugLab students had graduated, so training had to start from scratch with everything: tank maintenance, siphon-withdrawal reflex measurement, sensitization training, dissections, RNA isolation, qPCR, and data analysis.

Not only has getting everyone up to speed been a challenge, there have also been many challenges to confront from raising up the lab from dormancy. We had challenges with our RNA isolation protocol, an unhealthy batch of animals, a tank that shut down mysteriously over the weekend (and a tank monitoring system that didn’t sound the alarm!), a new file sharing service imposed by the university (which has been terrible), a simulator set up wrong… it’s really been an uphill fight almost every step of the way.

While the list of challenges has been lengthy, it’s turned out to be a lot of fun overcoming them. Our new and large group of sluglab scientists has brought tremendous enthusiasm and camaraderie, a surprisingly deep level of artistic talent, donuts, and a whole lot of fun to the lab. We’ve been knocked down, but we’ve made funny memes about it, and got back up again.

Maybe we’re also smiling because as we finally seem to have kick started the engine in the lab. We switched to hand homogenization and RNA yields have been amazing. We fixed the stimulator, got healthy animals, and doubled-down on training how to measure behavior, and viola–behavioral data has been pretty fantastic. With data starting to roll in we were finally able to have a lab meeting to work through how to analyze qPCR data, and students have been adding plate after plate of new data for us all to ponder.

At this point, it’s late July and things are really cooking! We have developed and pre-registered (https://osf.io/wvx6z/) an experiment to examine the transcriptional correlates of a very long lasting memory, and it looks like we might end the summer with all behavioral data and tissue collection complete (or at least close to it!). This is an exciting experiment. It’s very clear that forming new long-term memories changes gene expression. What is less clear, though, is if these transcriptional changes are needed to help create the memory, or if they are needed both to create and maintain the memory. Neuroscientists have generally assumed an important role in maintenance, and some models specifically imagine transcriptional feed-back loops that help perpetuate transcription to help maintain memory expression^​1​. But this would be a costly way to store a memory. Maybe instead, memories can become transcriptionally independent–perhaps by re-allocating resources within a neuron rather than permanently increasing them.

Our lab has had some hints that transcription might not persist throughout maintenance, at least not for the form of long-term sensitization we study in Aplysia. First, we’ve found that transcriptional changes after sensitization fade within 5 days, 2 days earlier than the memory lasts^​2​. This might mean that transcription isn’t needed for maintenance, but it could also mean that there is a slight lag between gene expression decaying and memory expression decaying (2 days isn’t that much of a gap). A second line of evidence is that we’ve found that re-activating a seemingly forgotten memory requires no new changes in gene expression (at least none we could detect), suggesting an uncoupling between memory expression and transcription^​3​. This is all suggesting, but not at all definitive.

Now we are collecting data that might help illuminate what role (if any) transcription plays in maintaining a long-term sensitization memory. To do this, we’ve cranked up our training protocol to 11– we are training each animal for 4 consecutive days rather than 1. Work in the Byrne lab^​4​ and other labs has suggested that this extended training protocol produces very long-lasting sensitization, and indeed we’re seeing robust behavioral expression 11 days after training (in our typical 1-day training protocol, behavior was almost always back to normal within 7 days). With this longer-lasting training protocol we can examine if transcription also lasts a long time (more than 5 days) or it it still fades quickly. Specifically, we’ll conduct microarray on samples harvested 1 and 5 days after the end of training, and compare the levels of gene regulation at those two time points. If we see that the widespread transcritptional changes at 1 day are still present at day 5, this would suggest a potential role in memory maintenance. However, if we see a decay in transcription at day 5, it would suggest something else is going on…. perhaps transcriptional changes are offset by compensatory mechanisms? Or perhaps memories can be maintained without an ongoing transcriptional change?

At this point we have no idea how the new study will work out… will transcription persist as long as behavior? Will it fade early? We don’t know, but we’re excited to find out. At this point, it looks like we might end the summer with all behavioral data collected and tissue harvested… so it won’t be too much longer now before we have an answer (hopefully).

It’s been a grueling but fantastic summer.

Today we received a shipment of Aplysia–the first shipment we’ve had since February of 2020.

It’s been a long, frustrating, and anxiety-ridden time for the animal colony to be empty. It’s not that the lab has been inactive–in fact, we published what I think is our best paper ever just a few months ago^​1​ . But it has been a long stretch without being able to provide the our typical level of involvement and excitement for our student researchers in the slug lab.

It feels really good to know that we are getting back on track. In fact, in addition to welcoming new slugs we’ve welcomed 5 new lab members: Lucas Eggers, Cynthia Espino, Daniel Mason, Delaney Mcriley, & Steven Proutsos. They join continuing member Melissa Nguyen to round out the Fall 2021 edition of the Sluglab. Let’s kick some a**! (scientifically)

Our first project with this batch of animals will be to explore for epigenetic markers accompanying long-term sensitization.

Over the last summer, C-J has worked like crazy on protocols for measuring methylation. We’ve found that it is surprisingly easy to full yourself, to obtain signals due to non-specific binding. What we’ve settled on is a process to check specificity of primer sets exhaustively by using synthetic DNA that we can manually methylate. Using this approach we’re pretty sure a key CPG island in the CREB1 promoter is *not* methylated in either control or trained animals. And our summer results also identified a CPG island in the egr promoter that seems to be default methylated, but with no change after sensitization. Our goal with these new animals is to now survey other methylation sites in the promoters of highly learning-regulated transcripts. Having lab meetings back in person has been fantastic (masks, of course, and DU has a vaccine mandate which has been very well implemented); very excited to see where research involvement takes our latest batch of slug lab members.

It’s summer and the slug lab is rocking. We have 8 students working in the lab (!), and a number of really exciting projects.

Here’s the lab photo to start the summer.

We knew this spring that we had recruited a special group of students in to the lab. So far the work this summer has confirmed our hunch–we’ve already completed two rounds of behavioral testing, students are making progress learning qPCR, and yesterday we had a great start to learning electrophysiology. I’m sure we’ll have our ups and downs, but it seems like we’re poised for a fun and productive summer.

Projects we’ll be working on include: 1) investigating if savings memories are re-formed or re-covered, 2) investigating the role of the peptide transmitter FMRF-amide in forgetting, 3) exploring the role of methylation in memory maintenance, and 4) some exciting pilot testing with a paradigm for sensitization in fruit fly larva, in collaboration with Scott Kreher in biology.

Our work this summer continues to be supported by the NIH (our current R15 expired at the end of May, but looks like it will be renewed starting July 1). Huzzah.

In addition, Dominican has received a generous donation from Joe Moskal to start the Moskal scholars program. Joe is a professor of biomedical engineering at Northwestern, a biotech entrepreneur, a Dominican trustee, and an all-around amazing guy. He generously helped Irina and me develop pilot data for our first grant and provided a sparkling letter of support… so it is no exaggeration to say he has already helped make the slug lab what it is today.

This year, Joe took the next step in his efforts to develop and broaden the biotech pipeline by funding the Moskal scholars program. Over the next five years this program will fund students interested in careers in the life and health sciences to spend a summer engaged in intensive research. The goal is for students to have the space, mentoring, and encouragement to develop their skills and passions in the science, and to launch them forward to great things.

Our first two Moskal scholars are Annette Garcia and Tania Rosiles. Tania will be spending her second summer in the slug lab–she’s already gained tremendous lab skills and helped co-author our recent paper on the long-term transcriptional response to sensitization ​(Patel et al., 2018)​. Annette is new to the lab, but was a star in Dr. C-J’s neurobiology class and has already been making big strides in the lab.

Neither Irina nor I would be where we are today if we hadn’t been fortunate enough to have amazing summer experiences. For Irina it was a summer working at Loyola Medical School. For me, it was a summer at Carnegie Mellon. In both cases it was generous funding from sponsors that enabled us to forgo our usual summer jobs and spend 3 months in intense and life-altering contemplation and study. We are so excited and proud to pay that forward each summer with a new batch of slug lab recruits, and we’re extremely grateful to Joe Moskal for his generosity and support.

One of our annual summer traditions is having DU photographer Ryan Pagelow come to the lab for a group photo and some science B-roll. As always, he does an amazing job. Here’s this year’s album:

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Graduates of the sluglab have been moving on to amazing careers. So we were excited to get the news that lab alumni and neuroscience major Derek Stek has just been offered a full scholarship to attend medical school at the School of Medicine and Public Health at the University of Wisconsin. Woo hoo!

Derek spent the summer of 2017 working in the lab, and also did summer research programs at the University of Colorado (2016) and UCLA (2018). In the sluglab, Derek learned how to do extract RNA (which does *not* start by vortexing the DNA-ase) and conduct qPCR. He helped track the expression of several transcripts regulated after learning, and was a co-author on the lab’s most recent paper (Patel et al., 2018).

Derek was also a star outside of the classroom, playing a big part on the DU varsity basketball team. This year, as he finishes at DU, Derek has been working with children with autism and learning behavioral therapy.

Congrats, Derek!

Our DU media lab spent a bit of time in the lab this summer talking to students and documenting their work. Here’s the video; really great footage of our amazing students.

Quanta Magazine recently ran a feature article on the nature of forgetting.  The piece covers several new lines of research on forgetting, including the work we’ve been doing with Aplysia.  It’s a great piece, and it’s amazing to see strong public interest in our work.

To Remember, the Brain Must Actively Forget by Dalmeet Singh, Quanta Magazine

Of course, accolades like this would not be possible if it were not for our amazing students.  Here’s the lab enjoying our end-of-summer party.