It might come to no surprise that kids that live in difficult environments can develop anxiety and other psychological disorders. But why is that? What is the biology governing this type of development? Can we avoid it? Can we give a better future to kids that have already suffered enough?

Dr. Jennifer Honeycutt, Assistant Professor at Bowdoin College, might be helping us do exactly this. She studies how rats develop if they are subjected to early life adversity, like being separated from their mom and litter-mates. The rats represent a great model and an opportunity to get a better understanding of humans too! In many human studies, the only thing scientists can observe is a correlation, meaning that they see how two things can happen very often together, but it is difficult to say that one thing caused the other. Not only rats models can be used to determine causality, but they can provide so much biological data (obtained by directly looking into their brain) to help researchers pinpoint exactly how their brain change when things go bad in their childhood.

But Jennifer, whom I have interviewed as part of the series “Draw My Science”, cautions us that to be able to trace these findings in rats back to humans, it is important to run tests for rats that are analogous to the humans’ tests. If you want to know more about she successfully made one of these analogous tests, and invented tiny rat-sized earbuds in the process, read on the interview extract below or come back next week to listen to the full recording on Youtube.

The Interview

Welcome, Jennifer, to “Draw My Science”! — tell us what you do.

For my research, I am really interested in how early life experiences influence the trajectory of brain development and the development of mental health issues. I do my research in a rat model: early life adversity via maternal separation. We are modeling very severe caregiver deprivations similar to what you would see in an understaffed orphanage, and how that early experience actually changes the connectivity between the limbic region of the brain and the prefrontal cortex. And that can either make you more vulnerable to later risks and induce the development of anxiety and schizophrenia, or, in some cases, it can make you a little bit more resilient. Most of what we do is looking at males and females, trying to understand sex differences within the entire realm of early adversities. Most of the outcomes are sex-specific: for example, schizophrenia is more likely to occur in males versus anxiety is more likely to occur in females. So, what is it about this early life risk period that can diverge and lead to different outcomes?

What does it mean for the limbic and prefrontal cortex to have different connectivity? And what technique do you use to measure that?

Primarily I use neuroanatomical measures. We do neural tract tracing and we look at axonal tracks from regions of interest -for example, the amygdala- and we follow how many of those axons, and when, are at the prefrontal cortex. These axons are responsible to signal potential threats. One of the experimental evidence I see is that especially in females that have experienced early adversity, those connections arrive too early. What looks like adults’ patterns of innervation arrive when the female rats are still juvenile. The same patterns are delayed in male rats and do not form until adolescence. I also do a little bit of fMRI work (ed. functional Magnetic Resonance Imaging). So that we can see what the functional connectivity of those brain regions is and if it is in line with the trajectories that we are seeing from the neuroanatomy. In fact, the axons can get there (as we see in the neuroanatomical measures) but we do not know if they are functionally integrated yet. So the fMRI scan analysis is to see if they are functionally integrated.

When rats do not experience early life adversity, like being separated from their litter-mates, they are less at risk of developing psychological disorders, like anxiety or schizophrenia – illustration by Valentina Ferro

How did you come up with the idea for this study?

I started the study about the fMRI connectivity when I came in for my postdoc — and that was almost 5 years ago now! It was a very long study with lots of animals, lots of surgeries to do the tracing, a lot of behavior observation and, of course, all of the fMRI. All of that was part of my mentor’s R01 (ed. the R01 is a type of NIH grant), but I have done an off-shoot of that one and have my own funding in the lab now through a Brain and Behavior Research Foundation NARSAD grant. I do something similar where I still look at cortical limbic circuitry, looking at amygdala reactivity as well as the prefrontal cortex, but I do it in awake animals in the fMRI (whereas with my mentor we used sedated rats). I am looking specifically at whether we have an increased likelihood of hyper-vigilant behavior following exposure to a potential threat.

For this particular study, I was inspired by a lot of work that is being done in Dr. Nim Tottenham’s group at Columbia. They did a study a few years back that looked at kids that have been raised in orphanages and then adopted. They followed these kids and then they did what is called the “fearful face” task. It is a really cool test you can do with people: you are in the MRI and the experimenters will show you a picture of a fearful face, a happy face, a neutral face and then they can look at what your brain is doing when you see these faces — even if they are just flashed, we are really good at looking at faces and determine whether we should be worried. In their study, kids with early life adversity, when they were shown a fearful face, had connectivity patterns that look like the ones in older kids. They are almost wired to recognize that threats really quickly. 

I asked myself, “How do we do the same study in rats?” — I could not just show faces to rats in the MRI machine, of course. I had to get a little bit creative, if I wanted to test that same paradigm . So, I built these ultrasonic headphones. They are tiny rat-size earbuds that go into the fMRI with the rats so that I can playback ultrasonic vocalizations. Rats make these ultrasonic sounds, that we cannot hear, and each of them represents something; some might be alarm calls that indicate a potential predator threat, others are like giggles that you get if you tickle them. I play these sounds back while the rats are in the fMRI, and I look at what’s different in terms of brain activity, what areas of the brain are active if I play an alarm call versus the giggle.

So far, we’ve gotten the legwork down, where we developed the technique and tested that indeed we do see different brain regions are active depending on which sounds were playing that are congruent with what we would expect to see. In fact, the amygdala “fires up” when we playback the alarm calls but you don’t see any change in the amygdala with the giggles. We’re really excited and so now it’s just about implementing the early life adversity part. I’m excited to see whether we’ll see differences in those animals. I think we will see that they’re a little bit more reactive to the alarm call sounds if they’ve experienced adversity.

What’s one thing in your study that really surprised you? Either because it was unexpected, or just really fun.

What probably was most surprising for me is realizing that a lot of times there is a lot of unexpected “development” to do, at least when trying a new method for the first time. For example, I was learning fMRI for the first time and trying to figure out how to get the animals into the scanner — how to load them, how to get the fMRI working, how to get the scans. etc… We did that initially while the animals were in a resting state, meaning they had been low-level anesthetized. But for my new experiment, I needed the rats to be awake, but I could not give them directions to follow or reassure them about the fMRI being really loud. Awake imaging brings a whole other thing to learn, where you have to habituate them to the machine (we used a mock scanner with MRI sounds for that) so that they’re not stressed out when you put them in the actual MRI. To get a good picture, they also have to be restrained so that they do not move around. After a couple of days, they are okay with that: “Oh, this is happening again. So I just chill out in this box, in this restrainer for like 20 minutes, and I’ll be fine”. I think that the learning curve was interesting and it was also weird! I now read some similar papers, and I can see what they did, but in the meantime, I am thinking “how do they actually get that to work?”.

Building the headphones was also pretty comical. I contacted a bunch of people who do human headphones and nobody wanted to make rat headphones. Then I found this one guy that would; he obviously didn’t have access to the rats, but he had gotten a stuffed rat, just to show me where they would be positioned. I would get all these pictures of a stuffed rat wearing tiny earbuds in my inbox. That was fun!

That must have been a fun challenge. Talking about the challenges, how are you managing animal research in the time of the covid-19 emergency?

We had a couple of litters that were already almost old enough to start doing the measurement with earbuds in the fMRI. But I had to cross this off my calendar because of the COVID-19 pandemic. The two graduate students in the group do not own a car, and we did not want them to get the bus and put themselves in a dangerous situation. Unfortunately, the PI and I alone did not have the man-power to run all the operations on our own. So in the end, I decided to put everything on hold. Thankfully the students had a lot of their data already collected, so they could work on data analysis and drafting papers. I am in a weird position. I just published the eLife paper in January, and that was a big thing, and I had started a new study and was just in the middle of another one. It is stressful to be in a standstill collecting data, but I have been trying to use this time to catch up on other things like reading literature. I am also teaching a class this semester, so that is keeping me pretty busy!

How do you think is it going to be when you will be back?

I do not know how it is going to look. I’ll be finishing my postdoc at the end of July as I got a faculty job that will be moving into. I was hoping to actually have all this data to take with me to analyze while my new lab will get set up. At this point, I am trying to think of contingencies for when things will go back to normal: maybe I will come back down to Boston from Maine (where I got the faculty position) to finish up some of this work. My wife and I are also expecting a baby, so I will also have to figure out how maternity leave is going to look this summer.

I think that flexibility is the key right now. Everybody’s dealing with something. I know how challenging it is for me, therefore I know that I have to be flexible with deadlines for my students. I think we all need to be a little bit more flexible and compassionate. Students are stressed out too; they’ve got multiple classes -I’m teaching only one class and I can’t imagine taking like three or four classes, all online, always synchronous, and trying to complete exams, all of that while they are also dealing with being at home. And not everybody’s back in a great space. So, that’s why flexibility has been a kind of go-to number one value in this period!

You mentioned that you’re expecting a baby. Do you think your research is going to affect the way you’re gonna handle maternity?

Oh, yes! It has already had an effect! We actually did reciprocal IVF, meaning that my wife is carrying my egg and we got a donor. One of the reasons that we had this specific arrangement came from my research. Originally the plan was that I was going to carry her egg, but things didn’t work out on the job market for me two years ago and I was back looking for a job this past fall. I was very stressed and did not want to have to carry the baby because this stress would cause me to have high levels of cortisol all the time: I’ve done a lot of research in the past on prenatal stress, and I wanted to avoid any sort of prenatal stress as much as possible. We chose that my wife would carry because she had a really chill semester. Of course now with everything going on, we are all just stressed anyway so it did not really matter but at least the beginning of the pregnancy was pretty relaxed.

Doing developmental research, I’m maybe too acutely aware of risk factors for different things. I know that we’re always thinking about that, and we’re very careful. Before my wife Christina got pregnant, she got her flu shot and during the pregnancy, we did not go out much so that we could limit the risk of her getting sick – and that’s because I knew all the risks associated with gestational immune activation and later mental illnesses in the baby. We were actually very concerned because we were hearing some initial things where in some cases if the mother was giving birth and she was positive for Coronavirus, they would take the baby from her for two weeks or limiting contact. Given that I do caregiver deprivation work, that of course raised every red flag in my head: I know that avoiding separation is important even in those early days, especially in the first few hours! Sure, it would be good not to get the baby infected, but what about the psychosocial effects later in their life. Last night I was reading that luckily it seems possible to avoid the separation, and at a hospital in Brooklyn, a woman who had COVID was still able to breastfeed wearing a mask and taking appropriate measures. This situation of uncertainty, and knowing what the long-term consequences can be, has definitely been stressful for me.

Rats that have being separated from their litter-mates develop brain connections much earlier than they should and they are more susceptible to threats. Similarly children coming from understaffed orphanages, develop brain patterns of risk recognition that are more similar to that of adults. — illustrated by the Valentina Ferro.

Trying to understand what the underlying reason for that risk is, what is the biology that’s under that, is fundamental to inform treatment for humans. If we’ve identified somebody who’s at risk, is it possible to intervene later in life? Or do we need to be thinking about it even sooner?

Dr. Jennifer Honeycutt

I typically ask the researchers I interview why people should care about their research. It seems fairly obvious to me in your case, as I can see how your research can have such dramatic repercussions in childcare. So, while giving me your pitch of “why people should care”, can you also comment on how you think your contribution is going to change the result we’ve seen in humans? What do you think is going to be the next step?

Anytime there’s a child growing up we have to consider that things are vastly different for different people around the world and that environment is incredibly important. Caregiver interactions — whether it’s a mom and a dad, a single mom or a single dad, two moms, two dads, grandparents — are critical for accurate and timely brain development and the lack of that, or even other forms of trauma, is going to present a big risk for later on in life. For me, trying to understand what the underlying reason for that risk is, what is the biology that’s under that, is fundamental to inform treatment for humans. If we’ve identified somebody who’s at risk, is it possible to intervene later in life? Or do we need to be thinking about it even sooner?

By doing the research in rodents we’re able to do systematic manipulations to pinpoint and try and find causality as opposed to in the human work where it is almost purely correlational. I am working so that we can compare the two studies (rodents and humans) as much as possible. There are studies with kids that have experienced adversity and tested the fearful face task; now we can do use the same paradigm (or at least an analogous one) in rats and we can ask the same question. That is going to allow us to be able to predict what we might see in human scans later based on what we had seen in rats. The early work that I’ve been doing with the headphones, a rat fearful face task analog, would be helpful bridging this big gap between what we’re doing in the animals to understand human disorders and what’s happening in human disorders. Historically, we have everything in rats here, everything in humans there, but can we compare them? And I think finding analogous tasks and tests is the way we can do that!

You mentioned that you’re at a turning point in your career, where you’re starting to develop your own funding. What’s the biggest challenge you experienced, and what advice do you have for young researchers that want to work in your field?

I think for me the biggest challenge is that I’m a first-generation college student. From very early on, my parents thought college was expensive and that I did not really need to go to college. I was hell-bent at going to college, but at the start, I was stumbling through it. Especially at the beginning, I was trying to figure out what I wanted to do. I started as an art major and then switched to psychology and neuroscience. And then I struggled again with what my next step would be after college. I decided I would get my Ph.D. because I was still really curious about the neurobiological side of things. But not having anybody at all, not even in my extended family, who’s gotten a Ph.D. or has had to deal with student loan payments or applying for academic jobs or anything like that, it was hard! I had to build my own academic family that could help me navigate the application and the publishing processes. There are lots of silent rules in Academia that you don’t really know unless you’re already from that world. I’m constantly questioning myself even now.

I’ve been incredibly lucky to have met a lot of amazing mentors on Twitter. World-famous scientists are literally at your fingertips on Twitter and it’s been kind of wild to be able to have an idea and have those people support you or ask you if you need help with something. You can reach out to them and it doesn’t feel and as intimidating as other ways of contact. I found that to be very helpful, especially needing that extra support. Twitter has been insanely amazing for that — I guess you wouldn’t normally think of a social media platform to be an academic tool.I’ve been incredibly lucky to have met a lot of amazing mentors on Twitter.

Read more about Jennifer

In the chat with Jennifer, we discussed in a few more details her career path, a few more suggestions she gave to other researchers and students, and even how to say if a rat is schizophrenic! I will publish the whole recording on Youtube next week — stay tuned.

In the meantime, if you want to check out more of Jennifer’s work, visit her website at www.jenniferhoneycuttphd.com or reach out to her on twitter @Ohambiguity.


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