Category Archives: Guest Post

Asthma and Animal Research: A Public Health Perspective

As a public health researcher with a focus on behavior change and complex interventions, I am more interested in studying how to get children to adhere to their asthma medication regimen rather than the mechanisms of inflammatory asthma. I am currently studying the risk factors associated with asthma attacks in children, which include among others, sub-optimal medication use, poverty, and access to healthcare. The aim of this research is to understand what risk factors for severe exacerbations – such as asthma attacks that send children to the emergency room – exist, thereby enabling healthcare and public health professionals to mitigate the risks of these ‘at-risk’ children.

My interests have nearly always been in applied in nature, however I understand that basic research underpins everything thing that we do in public health. Animal research is foundational to what we do as public health professionals. Without animal research, we would not be able to mitigate the risk factors these children have as we would not have the asthma medications we do today.

It seems that the sphere of public health shies away from discussing and supporting animal research; I’ve had colleagues tell me to be careful of talking too openly about my experiences in animal research outreach, for fear of alienating others – and potentially hindering my career. However, I strongly believe that public health professionals should be more open to discussing and supporting animal research. It is imperative to the continuation of both public health research and its application.

To illustrate this point, let’s use asthma as an example. The most effective medications for managing asthma are aptly named preventer and reliever medications. Preventer medications contain glucocorticosteriods and they work to prevent symptoms by reducing swelling, sensitivity, and inflammation in the airways. On the other hand, Reliever medications, or bronchodilators, work to open the airways and rapidly relieve symptoms.

Animal research has played an important role in the discovery of both glucocorticosteriods and bronchodilators. Glucocorticosteriods were developed using mouse models and the derived biomedical pathways. Bronchodilators were developed the 1960s, as a result of Otto Loewi’s research on adrenaline and other neurotransmitters.  Loewi used two beating frog hearts, aligned near each other, to demonstrated that slowing the pulse of the one heart and then circulating that perfusate through the other heart that it caused the other unaltered heart to also slow. He found that the same was true when he repeated the experiment, this time increasing the heart rate. This discovery proved that nerve cell communication is chemical rather than electrical, which led to the discovery of the neurotransmitter acetylcholine, and provided the foundation for future neurotransmitter research.

Glucocorticosteroids were developed using mouse models.

In relation to asthma, bronchodilators (beta2 agonists in particular) mimic the sympathetic nervous system discovered through Loewi’s famous experiment and allow health professionals to synthetically relieve the symptoms of asthma. Other studies using mice models have also elucidated the biomolecular mechanisms of airway hyperresponsiveness in asthma. Without Loewi’s initial experiment relying animal animal research, we would not be able to treat asthma as well as we do today. Without animal research, asthma management would likely rely on alternative medications that offer little in the way in relief; without effective treatment applied asthma research would focus only on prevention.

One of the reasons I was drawn towards public health and applied research was the focus on environmental, cultural, and large system-level factors that influence health, but this can come at the expense of ignoring the wealth of basic research that allows us to study these upper-level factors. When we forget the foundational work that lets us pursue our passions, everyone suffers. Public health professionals, at the very least, need to acknowledge–if not actively advocate for—the value animal research has in improving the health of the broader public and  should actively advocate for.

In writing this post, I had to research on how asthma medications came into being. Skimming through the biomedical literature was daunting (and confusing at times), but there are great resources already created to help clarify points for the those less familiar with biomedical research, such as myself – Understanding Animal Research, Animal Research.Info, and this website, Speaking of Research are great resources. I encourage public health professionals to educate themselves in how animal research allows them to do the work they do today. Then share that knowledge, – be that over Twitter, a blog, an email to colleagues, the options are endless. Support well-evidenced and humane animal research, because our work depends on it.

Audrey Buelo, M.P.H.

Of mice and mint: Animal research uncovers a previously unknown role for menthol in tobacco addiction

Cigarette addiction remains one of the common forms of drug addiction, worldwide; it is associated with remarkably elevated risk for heart disease, stroke and multiple types of cancer, explaining why nearly half a million Americans die each year from complications of smoking. A recent study by Brandon Henderson and his colleagues at the California Institute of Technology (Caltech) have revealed a startling new finding – that one of the most common flavorants added to some cigarettes may actually accelerate tobacco addiction. Dr. Henderson, now an Assistant Professor at Marshall University, answered some questions from Speaking of Research about his most recent discoveries.

 

What is menthol and why is it important to study its effects?

Menthol is the most popular flavor additive in tobacco products and the only non-banned flavor in traditional cigarettes (not e-cigarettes) sold in the US. Over time, reports have indicated that smokers of menthol cigarettes quit smoking at lower rates than those that smoke non-menthol cigarettes. Therefore, we are trying to identify the changes in the brain that occur with menthol that may indicate if menthol increases the addictive potential of nicotine (the primary addictive component in tobacco products). As an aside, menthol cigarettes are heavily used by African American smokers (75 – 90%). As a black scientist, this was another reason why I was attracted to studying menthol.

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Menthol is a naturally occurring chemical found in a number of plants, including mints

 

Can you give us a thumbnail sketch of your study and your findings?

Our goal was to examine well-known effects of nicotine on the brain and determine if they are enhanced by menthol. Two of the ‘well-known’ effects of nicotine are an increase in the number of nicotinic receptors (the proteins that bind nicotine) and an increase in the activity of neurons that release the neurotransmitter dopamine. These two events typically occur following long-term exposure (7 – 10 days) to nicotine, contributing directly to nicotine addiction.

Neuroscientists have known that nicotine, by itself, alters the brain to release more dopamine and that is one of the reasons why it is addictive. In this study, we focused on dopamine neurons of the ventral tegmental area, a part of the brain stem. The dopamine neurons that originate in this region are part of the mesolimbic dopamine system. This region is well-characterized for its importance in mediating the rewarding effects of drugs.

When we examined the combination of menthol with nicotine, we found that the combination increases the number of nicotinic receptors to a level that is significantly greater than is produced by nicotine alone. We also found that the combination of menthol and nicotine increased the activity of dopamine neurons to a degree that was significantly greater than what we observed with nicotine alone.

Since we observed that menthol promotes the pro-addictive effects of nicotine in the brain, we believe this may provide part of an answer to why smokers of menthol cigarettes have a much harder time quitting.

 

Mice were involved in your studies. In what ways are they similar and/or dissimilar from human smokers?

The mesolimbic dopamine system of mice is very similar to humans. Like humans, mice will self-administer (voluntarily consume) many drugs of abuse, including cocaine, opioids, and nicotine. Given that mice experience drug reward, similar to humans, they make an excellent model for studying addiction. Two experimental methods are commonly used in rodent models: intravenous self-administration and conditioned place preference, both of which reveal the degree to which nicotine is rewarding in the animal.

One hurdle associated with using mice to study drugs of abuse is that their bodies break down many drugs much more rapidly than humans. Therefore, proper dosing becomes a big concern so that our studies are relevant to humans. For nicotine, the guidelines for dosing were established long before I started science so this was a great benefit to my work with mice.

 

What are the medical or societal implications of your results?

Years ago, the FDA issued a request of information regarding menthol to determine if it should be banned similarly to other flavors that were banned following the Family Smoking Prevention and Tobacco Control Act (2009). This, and other scientific reports, will hopefully be examined by the FDA in determining the future regulations of menthol in cigarettes and e-cigarettes.

 

You just started a new lab as an Assistant Professor at Marshall University. Can you tell us about your future plans for your research and career?

I will continue to study menthol for a few more years because there is still much we need to understand. In the tristate area of Kentucky, West Virginia, and Ohio (where I am originally from), there is a large opioid addiction problem. Most opiate addicts (~80%) also are heavy smokers. I intend to begin studying how opioids and tobacco act together in the brain to promote addictive behaviors.

Henderson_Brandon_Marshall-headshot

Dr. Brian Henderson is an Assistant Professor in the Joan C. Edwards School of Medicine within Marshall University in Huntington, West Virginia

Dr. Brandon Henderson can be found on Twitter at @Dr_BHenderson and on the web at https://www.hendersonlab.org/

Q & A With a Primatologist: Why Studying Rank Changes is Good for Primate Welfare

In this Q&A post, we talk with Lauren Wooddell, a laboratory assistant at the California National Primate Research Center. Lauren is the first author of a research paper about to be published in the May 2017 issue of the Journal of the American Association for Laboratory Animal Science (JAALAS) titled, “Elo-rating for Tracking Rank Fluctuations after Demographic Changes Involving Semi-free ranging Rhesus Macaques (Macaca mulatta).” The paper is available in early view online now. Here, Lauren describes this study and its implications for animal welfare, and also for a better understanding of primate societies.  

 

Lauren.jpg

Lauren Wooddell

 

Speaking of Research (SR): How did you get into this field of research?

Lauren Wooddell (LW): I’ve always loved animals. I wanted to become a veterinarian ever since I was a toddler, but I realized in college that I’m more interested in animal behavior. I ended up taking some courses on animal welfare research and completely fell in love with it. There’s research to better the lives of animals? Count me in. I originally wanted to work with pigs, but the opportunities were, and still are, scarce. Instead, I worked with cattle, chickens, gibbons, parrots, and dolphins, until I began working with primates at NIH. When my rhesus macaque troop at NIH endured an overthrow, where monkeys with lower dominance ranks displaced the alpha ranked monkey, it was a very devastating event. To be honest, I kicked myself for not seeing it coming. But I then became passionate about understanding why it occurred so that hopefully we can prevent overthrows in captive primates in the future.

SR: Can you explain dominance hierarchies to us?

LW: Most monkeys (and many other animals) naturally form what is called a dominance hierarchy, to gain social order and determine who has priority access to resources. Within a dominance hierarchy animals are “ranked”, usually based on their wins or losses in aggressive interactions. High-ranking monkeys may fight and win against most other members of the social group, but rarely receive threats from others. Low-ranking monkeys, however, generally receive threats  by most other monkeys, but start fights with  others. Within dominance hierarchies high-ranking monkeys will have greater access to food and mates — it “pays” to be high-ranking.

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Rhesus monkeys in an aggressive chase interaction.

SR:  Why is it important to study changes in social rank in rhesus monkeys?

LW: Rhesus monkeys have remarkably rigid hierarchies that can remain stable for decades. This means that monkeys will often have the same social rank for their entire lives. Changes in social rank therefore are generally rare and when they occur, they can occur violently — resulting in devastating consequences for the monkeys. These drastic changes occur both in captivity and in the wild. For example, our lab at NIH found that social upheavals, or intense fights, can result in reproductive consequences for the involved families, whether the monkeys won or lost. These violent upheavals are called “overthrows”, because usually a lower-ranking individual or family will take over a higher-ranking one. By understanding more about how and when these “overthrows” happen, we could learn to predict, and potentially prevent, these violent events from happening. Doing so would greatly benefit the care of rhesus monkeys in captivity —where similar breakdowns in the hierarchy happen.

SR: Can you briefly describe the nature of your study, and what results you discovered? Was there anything surprising that emerged?

LW: Hierarchical stability, or dominance stability, refers to the idea that when we construct dominance hierarchies, individuals hold the same ranks, or positions, over time. If I construct a dominance hierarchy today with individual X as rank #7 for example, I would expect to see that individual as rank #7 (or very close to that) a few months from now as well. The goal of the study was to examine how changes to the social group would influence this stability: how would troop stability change animals had to be removed for health or colony management reasons, or if an overthrow occurred? The important thing to note is that this study was retrospective, meaning that none of the changes were implemented as part of a study. Importantly, in the case of the overthrows, we knew who was involved. This allowed us to go back retrospectively and analyze their behavior before the overthrow to examine potential indicators.

To measure social rank, we used Elo-rating, which is a method originally devised to rank and compare chess players that has since been modified for use in other sports (i.e. soccer and basketball) and now in animal contests. Essentially, Elo-rating allows us to predict the outcome of a potential competition between two individuals, before the competition takes place. Each animal gets a score that reflects its wins and losses. Animals can go up and down in their rating, depending on whom they compete against. So a high-ranking animal won’t increase in Elo-rating for winning against a low-ranking animal, because this is expected. But a low-ranking animal would increase significantly in Elo-rating for winning against a high-ranking animal. In general, high Elo-ratings refer to highly ranked animals, whereas low Elo-ratings refer to lowly ranked animals. The advantage of Elo-rating is that scores can change over time and reflect potential challenges to higher-ranking monkeys.

Unsurprisingly, we found that changes in a social group do indeed affect the stability of the dominance hierarchy. For instance, when we removed a large group of natal males – males that are born into the social group – dominance stability improved. This is generally because young males will attempt to rise in rank as they age, and their mothers will support them (especially if they are high-ranking). The increased stability may explain why, in the wild, natal males naturally emigrate from their troop around puberty. On the contrary, removing top-ranking females resulted in unstable dominance relationships because the females ranked immediately below the removed females increased in Elo-rating. The rising females were also involved in the overthrows of the top-ranking females. Therefore, increases in Elo-rating over time for the #2 ranking families, especially following the removal of females in the #1 ranking family, may be an indicator of an imminent challenge to the hierarchy.

FamilyGrimace

A family of rhesus monkeys indicates submission to an adult male via fear grimaces.

Perhaps the most surprising result from this study was that after overthrows, dominance stability improved. This improvement indicates that overthrows may actually be a stabilizing mechanism for a troop. This poses a real conundrum for captivity though: if overthrows potentially stabilize a troop, should we prevent them from occurring? In terms of animal welfare, I would argue yes. Perhaps the better question is: how can we stabilize the troop before an overthrow occurs? That is what I hope future research will tackle.

SR: How did you study the animals? Can you tell us what this involved, day-to-day?

LW: We studied a troop of rhesus monkeys that lived in a naturalistic, yet captive, 5-acre habitat. The troop was structured around large family groups that totaled 80 monkeys comprised of three major families. On a daily basis, we observed and recorded dominance interactions by standing in the habitat with the animals and recording every time we saw monkeys threaten, chase, or bite each other. However, monkeys use submissive gestures as well, such as moving away from dominant individuals or using facial signals like a fear grimace, which we recorded as well. We recorded the identities of the monkeys involved, and from these interactions, we could then determine a dominance hierarchy by using Elo-rating. Tutorials and software code for Elo-rating using R Stats software package are freely available.

FearGrimace

Rhesus monkey fear grimace indicating subordinate status.

SR: What are the most important implications of this study in terms of animal welfare? And in terms of understanding primate societies? 

LW: In terms of animal welfare, I think this study effectively shows that with careful observation, we can potentially predict some of these major upheavals, which would collectively enhance the well-being of rhesus monkeys in captivity. In terms of understanding primate societies, I think this study shows that certain individuals, which we call keystone individuals, can have a large impact on the overall social stability of the troop. Understanding the presence of keystone individuals can allow us to understand the development, and breakdown, of social groups both in the wild and in captivity.

SR: Are there any implications for the social behavior of wild monkeys?

LW: Absolutely! Wild monkeys also exhibit dominance hierarchies that can fluctuate over time, and changes in their social groups can occur regularly. For example, males commonly transfer into and out of social groups. Males born into a group will leave that group as they mature as to not breed with kin. So how do social groups change in their stability following the emigration or immigration of males? Another potential application would be in the study of group fission events, which occur when a social group becomes so large that it breaks apart into two or more groups. These are rarely observed and poorly understood. A relatively recent group fission on Cayo Santiago, an island off of Puerto Rico that has approximately 1,000 rhesus macaques, occurred following the death of the troop’s alpha female. This resulted in an overthrow and a group fission. The research indicated that there were changes in the monkeys’ social behaviors, like grooming tendencies, before the event, and I would assume that prior to group fissions, there was probably some degree of dominance instability as well. Perhaps with time, researchers in the wild could predict these events, which would allow researchers to have a better understanding of why and when these events occur. In general, our results suggest that any major event could be analyzed to examine how the hierarchy changes in response to or even prior to the event.

 

Displacement

Rhesus monkey dominance is evident in subtle ways, such as the displacement occurring here.

SR: What are some of the most rewarding and some of the most difficult aspects of your job?

LW: The most rewarding aspect of the research is gaining an understanding of the highly complex social lives of the monkeys. It’s a constant soap opera, full of drama and plot-thickening twists. When you work that closely with the monkeys, you get to know them on a very individual level and you become inevitably intertwined in their stories as well, going through all of the changes with them. However, the most difficult aspect of this work is when the hierarchy is challenged in an overthrow, which, thankfully, is rare. There’s a notion in the general public that researchers are data-driven machines that only care about the bottom line: results. That’s simply not the case. I’ve lost monkeys that I’ve come to know on a very intimate level to overthrows, and it has been devastating. We deeply care about the monkeys we work with, more than just the data they provide. Researchers have emotions too, and I am willing to admit that. I hope other researchers will do the same.

SR: Thank you for sharing the important work that you do with us!

LW: Thank you for inviting me to discuss this research. I hope it will inspire future research!

Why I am proud to be a Registered Veterinary Technician in animal research

Christine Archer is a registered veterinary technician at the University of Ottawa in Ontario, Canada.  She has worked in animal research for over seven years.  She currently works with aquatic animals and reptiles in biological research. In this post, Christine looks at the interests and motivations that led her to become a laboratory animal technician, and her interest and love of aquatic animals. Fish account for 43% of research animals in Canada, with amphibians adding a further 3%. 

christine-archer-with-fish-ottawa

While I can’t say that I was ever a typical kid growing up in rural Canada, my surroundings definitely shaped my interests as I grew. I was always interested in the “gross” animals, from fish to frogs, and throw some snakes in there, too. I’d drive my mother crazy, catching animals in dubiously secure containers and bringing them home, only to have her insist that I take them right back where they came from. Just about the only exception was this big female wolf spider I rescued from a trough that ended up having an egg sac, which eventually resulted in many tiny spiders all over our house. I remember I’d also saved some newts from a tiny marsh that was being bulldozed for a new house. I hand fed them and kept them for years. They even went on vacation with us, in their not so dubiously secure critter keeper. All the while, I was also keeping many, many aquariums in my parents’ house, and breeding all manner of tropical fish. We had tanks in just about every room, even the bathrooms.

After a false start in engineering, I ended up studying biology in university, but I was so enamored with all of the sciences that I couldn’t decide what I really wanted to do with myself. When I was finishing up my undergrad, my cat Monty got very sick. The process of his treatment and recovery got me very interested in veterinary medicine. While I was feeling rather burnt out by my university studies at this point, I looked into taking a college program that would offer me practical hands-on skills in addition to the science of veterinary medicine. I went to college for veterinary technology, eager to consume all of the veterinary medical knowledge I could, especially everything that pertained to those “weird” animals that I loved. In the middle of my program, I took an internship at a large medical research facility. This is where I found what I was meant to do with my life. Marrying my love for science and the scientific method with my newfound love for veterinary medicine. The only thing missing (so far) were the weird animals I am so fond of. I learned about rodents and rabbits and the nuances of their biology. I learned about the 3 Rs and how essential good animal welfare is to doing good science. Throughout my life, I’d always felt like the weird kid who stood up for the weird animals that everyone didn’t like. I was a voice for them. Now, in research, I realized that I can continue to speak for my charges, no matter what species they are.

Racks of zebrafish at the University of Ottawa

Racks of zebrafish at the University of Ottawa

I finished school and became a Registered Veterinary Technician. I worked in cardiovascular research in a number of roles, working with traditional research animals like rodents and rabbits, and the occasional pig. It was great to be surrounded by colleagues who shared my interest in animal welfare and working hard to ensure that our charges’ welfare needs were being met every day. And then, one day, I got a call. The university’s aquatics and reptile facility needed an RVT for the summer, and my supervisor wondered if I would be interested. I couldn’t believe it. The opportunity to take part in veterinary nursing and the husbandry of all of those “weird” animals I’m still totally obsessed with? I’m there. However, I was reminded that it was only for the summer. Still, totally worth it. I said my tearful goodbyes to my colleagues at the rodent facility and started my position working with zebrafish, goldfish, trout, frogs, lizards, and even the occasional snake. That was more than three summers ago. I am still proudly caring for the veterinary and welfare needs of these animals today, as I was made a permanent staff member of the facility. I am so honored to be able to work with the animals I love, surrounded by passionate people working on everything from CRISPR research with zebrafish, to biomechanics work with fish that can walk on land. We still have so much to learn about these animals and their specific welfare needs, and I am thankful every day to be on the ground floor, working with colleagues who want to advance science while ensuring these animals, these weird, wonderful creatures, get the best possible care from the humans that depend on them.

Fish Facility at the University of Ottawa

Frogs (Silurana Tropicalis) at the University of Ottawa

On an average day in my facility, I can be found setting up zebrafish breedings and collecting embryos, or culturing live food like rotifers and brine shrimp for the fish to eat. My job requires me to be adept at multiple skills, from understanding the husbandry and welfare needs for our many diverse animals, to working with our staff veterinarians on developing and improving methods to anesthetize fish and frogs. Animal welfare is very important to me, and I strongly believe that the quality of my work has a direct impact on the quality of life of the animals I care for, which in turn has an impact on the quality of the research that my colleagues can perform. I work every day with multiple researchers to help ensure they are able to do the best work possible thanks to animals which are healthy, happy, and leading enriched lives.

I am proud to be a Registered Veterinary Technician in animal research, because I care very much about animal welfare and having the opportunity to speak for those who can’t speak for themselves is something I will never tire of.

Christine Archer

SYR: How sheep can help us understand why girls are reaching puberty at younger ages

michelle-bedenbaughThis guest post is the second written by Michelle Bedenbaugh, a Ph.D. student in the Physiology and Pharmacology Department at West Virginia University. Check out her first post on the benefits of using large animal models to study reproduction. It is also part of our Speaking of Your Research series of posts where scientists discuss their own research. In this post, Michelle discusses some of the cells and signaling pathways that are important for controlling the timing of puberty and how the use of sheep as a model is beneficial for this type of research. If you would be willing to write a guest article for Speaking of Research, please contact us here.

For those of you who have been watching the news in the United States over the past 5-10 years, you have probably heard a few discussions about the fact that girls are reaching puberty at younger ages.  In the 1980s, girls normally reached puberty around the age of 13.  In 2010, the average age of girls reaching puberty had dropped to 11 and has since continued to decline.  Reaching puberty at earlier ages is associated with several adverse health outcomes, including polycystic ovary syndrome (PCOS), metabolic syndrome, obesity, osteoporosis, several reproductive cancers and psychosocial distress.  The public and researchers have pointed fingers at several potential culprits, including an unhealthy diet, chemicals that disrupt the body’s normal hormonal environment, and an individual’s genetic predisposition to disease.  In reality, a combination of factors have probably led to the decrease in the age at which girls reach puberty, but I don’t want get into a discussion about these factors today.  Instead, I want to talk about some of the signaling molecules in the body that these factors may be influencing to affect the initiation of puberty.

As with many processes in the human body, the brain plays a critical role in the control of reproduction and the timing of puberty.  Within a specific area of the brain called the hypothalamus, several populations of neurons (specialized cells in the brain) exist that control reproduction.  The activity of these neurons is influenced by various factors that are communicated from other parts of the body and outside environment to the brain, including nutritional status, concentrations of sex steroids (like estrogen and testosterone), genetics, and many other external factors.  All of these factors tell the brain when an individual has obtained the qualities necessary to successfully reproduce and therefore undergo pubertal maturation.  Gonadotropin-releasing hormone (GnRH) neurons found in the hypothalamus are the final step in this chain of communication and are essential for the initiation of puberty.

A GnRH neuron present in the hypothalamus.

A GnRH neuron present in the hypothalamus.

Most of these nutritional, hormonal, genetic and environmental signals are not directly communicated to GnRH neurons.  Instead, they are conveyed through other types of neurons that then relay this information to GnRH neurons which either stimulates or inhibits the release of GnRH.  Because GnRH is a signaling molecule that ultimately stimulates the maturation of male (sperm) and female (egg) gametes, stimulating GnRH in turn stimulates reproductive processes while inhibiting GnRH inhibits reproductive processes.  The perfect balance of stimulatory and inhibitory inputs is needed for GnRH to be released and for puberty to be initiated.  Consequently, if stimulatory inputs signal to increase GnRH prematurely, puberty will occur earlier, which may result in several of the health concerns that were mentioned above later in life, including reproductive cancers and psychosocial distress.  In contrast, if inhibitory inputs block the release of GnRH, puberty will never occur and result in infertility.

My research looks at some of these stimulatory and inhibitory inputs and how they communicate with each other, as well as with GnRH neurons.  Two of the stimulatory signaling molecules that we research are kisspeptin and neurokinin B (funny names, I know).  We also study dynorphin (another funny name), a molecule that inhibits GnRH release.  These three molecules can all individually affect GnRH release and therefore reproduction.  However, the really cool thing about these three molecules are that they are actually present together in a special type of neuron that is only found in one small and highly specific area of the hypothalamus.  Because these neurons contain kisspeptin, neurokinin B, and dynorphin, they are often called KNDy (pronounced “candy”) neurons.  The fact that kisspeptin, neurokinin B, and dynorphin are all present in these KNDy neurons together allows for them to communicate directly and affect each other’s release.  This communication then ultimately affects the release of GnRH.  Before puberty, inhibitory inputs, like dynorphin, dominate and don’t allow for adequate amounts of GnRH to be released to stimulate reproduction.  As an individual matures, stimulatory inputs, like kisspeptin and neurokinin B, begin to outweigh inhibitory inputs, and GnRH can be released in adequate amounts to support reproductive processes.  Below is a figure that summarizes how we think all of this works within the body.  However, there is still a lot that we don’t know about how kisspeptin, neurokinin B and dynorphin interact with each other that is waiting to be discovered!

Hypothesized model for the initiation of puberty. (1) Internal and external factors are communicated to the body. (2) Next, these factors are relayed through various signaling pathways to stimulatory and inhibitory molecules present in neurons located in the hypothalamus. (3) Stimulatory and inhibitory molecules travel to GnRH neurons and affect the release of GnRH. (4) GnRH stimulates reproductive processes that are critical for the initiation of puberty. (5) Once all of the proper conditions are met, reproductive maturity is attained.

Hypothesized model for the initiation of puberty. (1) Internal and external factors are communicated to the body. (2) Next, these factors are relayed through various signaling pathways to stimulatory and inhibitory molecules present in neurons located in the hypothalamus. (3) Stimulatory and inhibitory molecules travel to GnRH neurons and affect the release of GnRH. (4) GnRH stimulates reproductive processes that are critical for the initiation of puberty. (5) Once all of the proper conditions are met, reproductive maturity is attained.

To complete all of these studies, we use sheep as our model.  I know what some of you are thinking.  “How in the world would sheep serve as a good model for how puberty is initiated in humans?  I don’t think I am similar to a sheep at all!”  In fact, sheep are actually an excellent model in which to do this research.  The signaling pathways that affect the release of GnRH in sheep are very similar to the signaling pathways in humans, and in some cases, are even more similar to the human pathways than the pathways present in mice or rats.  In humans and sheep, neurokinin B has only been found to stimulate GnRH release.  However, in rodents, there have been reports of neurokinin B both stimulating and inhibiting GnRH release.  Since neurokinin B is one of the main signaling molecules that we study, using sheep instead of mice or rats is more beneficial for modeling what is occurring in humans.

sheep-in-reproduction-research

Because we have to collect several blood samples from the sheep in order to measure hormone concentrations, having an animal with a larger blood volume is also advantageous.  Several hormones in the body (including GnRH) are released in a pulsatile manner, meaning one minute GnRH concentrations are high and a few minutes later they are low.  Therefore, in order to appropriately measure GnRH, blood samples need to be taken every 10-12 minutes for several hours.  This is not feasible in rodents.  If you took blood samples as frequently in rodents as is possible in sheep, you would risk killing the animal.  Some scientists who use rodents as their research model attempt to get around this issue by taking blood samples less frequently.  However, this means their hormone measurements are less accurate.

These are just a few of the many reasons why we conduct our research in sheep (to learn more about the advantages of using sheep and other large animal models to conduct research involving reproduction, see my previous post).

While most people (including myself) do not look back fondly on our awkward pubertal years, I absolutely love studying the signaling pathways the body uses to determine when it is ready to successfully reproduce.  We have discovered quite a bit over the past few decades concerning how different internal and external factors affect pubertal maturation, but there are still so many unknowns left to be determined.  I look forward to hopefully discovering some of these unknowns and improving our understanding of how puberty is initiated in both humans and livestock species.

Michelle Bedenbaugh

SYR: The case for using large animal models to study reproduction

michelle-bedenbaughThis guest post is written by Michelle Bedenbaugh, a Ph.D. student in the Physiology and Pharmacology Department at West Virginia University. It is part of our Speaking of Your Research series of posts where scientists discuss their own research. Michelle’s research involves examining the brain’s role in the initiation of puberty.  In this post, Michelle discusses the benefits of using large animal models to study reproduction.  If you would be willing to write a guest article for Speaking of Research, please contact us here.

With the increasing pressure to publish papers and the decreasing amount of funds made available to conduct experiments, it has become more difficult for researchers to survive and thrive in an academic setting (see here, here, and here). Scientists have had to adapt, and in many situations, this has led to a significant amount of research that relies heavily on small animal models, including rodents and invertebrates.  In addition to being less expensive than large animal models (sheep, pigs, cows, horses, etc.) there are also more genetic tools and techniques available to use in small animal models.  For example, transgenic mice, where certain genes can be either deleted or overexpressed, are used commonly by researchers worldwide.  Other cutting edge techniques, like optogenetics, where light can be used to control the activity of cells in the brain, are also being used on a more routine basis in rodent models and currently don’t exist in large animal models.

Optogenetics involved using light to control genetically modified cells inside the body

Optogenetics involved using light to turn off or on cells in the brain

While it is most likely easier, cheaper, and faster to conduct experiments using small animal models, in certain situations they are not always the most comparable to humans.  When modeling certain diseases or understanding certain physiological processes, larger animals, like sheep, pigs, and cows, provide a better model for scientists.  This post aims to look at some areas where larger mammals can provide important knowledge or understanding.

A few of the more obvious benefits to using large animal models when compared to small animal models are that large animals are more analogous to humans in regards to body size, organ size, and lifespan.  In addition to these similarities, animals like sheep, cows, and pigs are much less inbred when compared to rodents.  Some would argue that it is advantageous to use animals that are highly inbred because this decreases the amount of variability in an experiment.  However, each human has a unique genetic makeup, and sometimes solutions for problems in inbred rodents cannot be translated for use in humans.  Therefore, in these instances, it is probably more beneficial to use a less inbred large animal model.  Most large animal models also have the added benefit of being an economically important species.  The majority of researchers who use large animal models are attempting to find solutions to health issues that are present in humans.  However, successful experiments in large animal models have the ability to affect both human and animal health.  For example, if a researcher made an important discovery about the way food intake is controlled in cows, it would have the possibility of improving human health, as well as increasing profitability for cattle producers.  Because cows are very similar to sheep, it may also benefit sheep production as well.  Rodents are not an economically important species that provides food, fiber, or other essential products used by the human population.  Consequently, discoveries made in rodents and other small animal models may only benefit humans if the results are translatable.

My particular research focuses on furthering our understanding of how puberty is initiated in girls, and we use sheep as our animal model.  I won’t get into the specific benefits of using sheep to conduct puberty research today because I will discuss this more in my next post.  However, I did want to touch briefly on some of the advantages of using large animals to perform research used to study reproduction in a broader sense.

The brain plays an essential role in controlling reproductive processes.  The brain structure of large animals is more closely related to humans than small animals because large species have a sulcated cortex (meaning the surface of the brain is wrinkly) as opposed to small animal species which have a smooth cortex.

Comparison between mouse (smooth cortex) and human (sulcated cortex) brain. [Credit: Elizabeth Atkinson, Washington University in St. Louis]

Comparison between mouse (smooth cortex) and human (sulcated cortex) brain. [Credit: Elizabeth Atkinson, Washington University in St. Louis]

Sheep also have the advantage of their brain and the cellular pathways present within it being similarly organized to what is observed in non-human primates.  Hormones serve a major role in relaying information from the brain to reproductive organs and vice versa.  The actions of several hormones that aid in controlling reproduction in female sheep (like estrogen and progesterone) parallel the actions of these hormones in humans.  Older sheep also have a similar response to estrogen replacement therapy when compared to post-menopausal women.  The development and function of several structures on the ovary of sheep is also similar to that which is observed in women.  These structures have a major influence on the reproductive cycle and are critical for the maturation of female gametes (sometimes referred to as eggs).  Assisted reproductive technologies, many of which are used for in vitro fertilization (IVF) protocols in women who are having trouble conceiving, have been adapted from procedures used in livestock species.  For example, artificial insemination, where semen is collected from a male and usually frozen so that it can be used to inseminate a female at a later time, is commonly used in cows, sheep, horses and pigs and is similar to procedures conducted in humans.

Credit: Livestock Breeding Services - http://www.livestockbreedingservices.com.au/images/servicesai.jpg

A laparoscopic procedure is used to artificially inseminate sheep

 

Embryo transfer, where embryos from one female are placed into the uterus of another female, are also used in livestock species and humans.  In addition, sheep are also an excellent animal model for studying pregnancy.  Sheep are used often to examine how stress, maternal nutrition, and exposure to excess hormones or toxins affect the development of a fetus.

These are just a few examples that display reproductive processes occurring in many large animal species are easily relatable to those same processes which also occur in humans.  I only touched on a few species today, but there are many more animal models that are underused in research and would serve as great models for humans.  In addition, I only discussed some of the ways these animals can be used to study reproduction when in fact they can be used to mimic many other biological processes that occur in humans.  Depending on the subject matter being researched, the use of some animal models is more appropriate than others.  Regardless of cost or time, researchers should always consider which animal model may be the most appropriate for their experiments.  I believe conducting research in a variety of species as opposed to just one or two species will always be more advantageous and will aid us in solving health issues in humans more quickly.

Michelle Bedenbaugh

Reigniting My Fire for Animal Research

lisa-headshotThis guest post is written by Lisa Stanislawczyk, a Veterinary Scientist at a pharmaceutical company. She plays a key role in ensuring the standards of animal care are always improving at her institution. Having been introduced to Speaking of Research through a committee member, Lisa kindly agreed to share her experiences. In this post, Lisa explains her passion for innovation in the field of animal welfare and her experiences, positive and negative, in delivering animal care at numerous institutions in the US. If you would like to write for Speaking of Research please contact us here.

When I started out after college working as an animal care technician at a contract research organization (CRO), I never thought I would want to perform the procedures I saw being done to the animals. I didn’t want to make them uncomfortable or scared. I loved animals and had always wanted to be a vet (like so many others in the field of animal research). While working at the CRO I began to see the care and attention that the technicians took in performing these procedures and how careful they were to make the animals comfortable and at ease. I realized they too cared for the animals as much as I did and we all wanted nothing more than to take the best possible care of these animals.

lisa-veterinary-scientist

Later, after 15 years in the animal research field, I found myself looking for a new role. I was always proud of what I did and left work each day with a sense of accomplishment. However, I was finding it difficult to find work, a common problem for so many in the world we live in today.

I realized that in order to stay in the field and get a good job I was going to have to move outside of my comfort zone, away from everything and everyone familiar. It was scary, but I moved to another part of the country, away from my family and all my friends, to pursue a new job. I was anxious and felt isolated. I came to the harsh realization that not everyone holds themselves or others to the same standards I had been taught, or was accustomed to. This realization almost made me stop doing the work that I had grown to enjoy and get a huge sense of accomplishment from.

I didn’t quite know how to deal with what I perceived as poor animal welfare in my new job. This feeling was not from the technicians doing the work, they were doing the best they knew how with what they were taught. There just seemed to be a lack of knowledge of the regulations which one should have working in a vivarium. It was the management that needed to be held accountable. I spoke with the Chair of the Institutional Animal Care and Use Committee (IACUC) in order get a better understanding of what I felt was just not good research. After our conversation, I still felt there was a lack of accountability from the IACUC Committee. I was at a loss and felt drained and hopeless because there continued to be mistakes and mis-steps which could have been avoided.

I spoke with the veterinarian and was told, “I didn’t understand the field that I was in and I was too soft”. I didn’t believe that. I believed I was there to be an advocate for the animals in my charge. I was told there was not a “magic ball” to know outcomes of certain studies, I knew there were humane endpoints that should be followed. I did my best to make things better. We began a better training program so the people performing the procedures had a better understanding of the Animal Welfare Act and the Guide. We updated procedures and SOPs (standard operating procedures.)

It took its toll. I found myself working long hours to make sure the studies I was to oversee were executed correctly and at the same time educating the personnel working with me. I was exhausted and overworked. So were my technicians. I began to become so emotional about some of the things I was seeing that I would spend what free time I had at home, crying myself to sleep. Just thinking about it now, makes my eyes water. We all began seeing things that we could not bear any longer and more people began to have concerns and fill out whistleblower forms. It was heartbreaking and I just didn’t feel like I could do it any longer. Then the day came, I was laid off. It was a blessing!

Thankfully my negative experience is not common and the facility I worked at was taken over by another company. I have heard that they are still overworked (many of us can sympathize) but that things regarding the animals have definitely improved.

Image of macaques for illustrative purposes.  Image courtesy of: Understanding Animal Research

Image of macaques for illustrative purposes.
Image courtesy of: Understanding Animal Research

I moved back to my family and friends. I needed the moral support from them. Still, I didn’t want to go back to it. I was burnt out. I worked at a home improvement contracting office fielding phone calls and organizing the office. It just wasn’t what I could see myself doing long term. I needed a challenge. I missed the animals. I held guilt for not doing more for them even though I still don’t know what more I could have done at the time.

A previous boss of mine who happened to be a veterinarian reached out to me about a job. Again it was a big pharmaceutical company. I was skeptical but I needed to give it one last chance and it was only a temporary position. It was great to experience the investigators working with the animal care technicians to communicate how the animals did while on study and this empowered everyone to know exactly what was going on with each and every animal on a daily basis. The communication between all the investigators, technicians and veterinary staff truly improved the welfare of the animals. The veterinary staff really cared for the animals and the animal care technicians knew every animal’s quirks, likes, and dislikes. Everyone would make sure the animals that were on study got some extra favorites whether it be food enrichment, human contact, or toys. The people there renewed my faith. I could see the ethical behaviors and integrity of each and every person there. It gave me the desire to stay in the industry. This was what I was accustomed to. I felt like I had a “place” again.

Once the temporary position was over, I moved to another company also working with the veterinary technical staff. There I was allowed to attend ILAM (Institute for Laboratory Animal Management). It is a 2 year program and the information, relationships, and contacts you come away with are immeasurable. I shared my story with others I met there (from all over the world) and I realized we all shared in the desire to deeply care for the animals. We go to work every day to make sure everyone does their best to take care of every need of all the animals in their charge. For some time, I have passively been in the industry, not really wanting to be a part of all the external committees and public outreach opportunities available. After attending ILAM, all that changed. Experiencing the love and desire to improve and do better within our industry and making connections and friendships with people with this common thread has re-ignited my passion for the industry. My company encourages people to innovate and strive for better animal welfare. I am so proud to be a part of a program that has refined techniques performed on multiple species to make it easier for both the animals and the technicians. This is how it should be. This is the industry we are in.  Change is key. Once again I am so proud of what I do and the program I am a part of everyday. I flourish when someone asks me what I do, instead of talking vaguely so they won’t understand or want to hear more about it. I am happy to explain why what we do is so important and necessary.

We make miracles happen and improve the lives of humans and animals every day! This is what we do for a living! This is why people and their pets are living longer, happier lives. This is the reason I am proud to be in animal research. I urge my fellow technicians to speak out, be proud, and get involved explaining what you do and why you do it!

Lisa Stanislawczyk