Tuesday, August 12, 2014

Veterinary super heroes!

V-FORCE from the AVMA


I was tickled to discover that the AVMA published a comic book. V-Force! Veterinarians to the Rescue! Did you know that veterinary superheroes have x-ray vision and use it to diagnose leptospirosis? Did you know that some of them (us? can I say some of us?) even have microscopic vision so that they can see microbes and diagnose infection with West Nile Virus? Why didn't I get these super powers when I graduated?

My favorite scene bar none is the one in which the doctor (that is doctor-of-humans not doctor-of-animals) says "I'm just glad the veterinarian was around to help link the cause of the disease." Words spoken by no doctor ever. More fantastical than teleporting veterinarians?

The point of the comic is to leave it lying around in veterinary waiting rooms for small children to pick up, so that they learn a) some of the alternate careers vets can have besides working as small animal clinicians and b) that veterinary medicine isn't just about helping animals, it's about helping people, too. (OneHealth to the rescue! I want a super hero named OneHealth Man!)

I like this effort by the AVMA to communicate some important information about veterinary careers, but I do wonder if a somewhat heavy-handed comic is the best way to do it. You're not going to get a lot of bang for your buck with it (although on the other hand it probably didn't cost all that many bucks to produce, relatively). Hey, AVMA, are you listening to me? (AVMA reps have commented on my blog posts in the past when I've made enough noise, so consider my voice raised now.) I have an idea.

Did you see the recent SyFy show Helix? Which actually starred a veterinary pathologist, and yet didn't do a great job of portraying what veterinary pathology is really like or why it's an exciting career. I think the AVMA, if they are serious about this goal of getting the public to better understand what veterinarians are capable of, should be trying to get more veterinarian characters on TV shows and in movies, and make sure they are well characterized. I suggest one way to do it is to offer a liaison service to Hollywood: the AVMA would provide a veterinarian consultant appropriate for the particular role, and would pay their salary and expenses for the job. The consultant would help the writers make the role realistic.

I'm completely serious about this: I think Hollywood (or part of it) is starting to realize that a realistic depiction of scientists is something its audience is actually interested in, but it's got to be hard to find someone who can speak to the daily life of a veterinary pathologist. Make it easy for them. Because a favorite character on a TV show is something that makes kids, and even adults, consider their future career options in a different light, not a comic book with a clunky story.

Wednesday, August 6, 2014

Can a father's stressful experiences affect his offspring's stress system?

I am again indebted to my APDT students, who asked a very interesting question which turned into a blog post. This time it was: “Can a father’s stress be passed on epigenetically to his offspring through sperm?” Warning: epigenetic geekery ahead. If you are not in the mood for some technical terminology, this may not be the post for you.

Source: Wikipedia


I’ve blogged about epigenetics before (on the epigenetics of fear and of stress) and there are summaries of what epigenetics is in those two posts, but basically it’s changes in the DNA that don’t involve the sequence of bases. We’ve been so focused on the importance of DNA sequence as we’ve learned more and more about genetics, but in recent years the importance of other factors has started to become obvious. It’s like saying that the content of a book isn’t the only thing controlling whether the book gets read or not — it also matters whether the cover is appealing, how much it costs, and whether it’s shelved where people can find it.

One of my students tried to answer her own question and found this fascinating article:

Rodgers A.B., S. L. Bronson, S. Revello & T. L. Bale (2013). Paternal Stress Exposure Alters Sperm MicroRNA Content and Reprograms Offspring HPA Stress Axis Regulation, Journal of Neuroscience, 33 (21) 9003-9012. DOI: http://dx.doi.org/10.1523/jneurosci.0914-13.2013

Essentially, Rodgers et al stressed out some male mice and then tested their offspring six ways from Sunday to see if the effects had been passed on. And they had, but in some surprising ways.

The mice

Male mice were chosen because we have evidence that environmental effects can be passed on epigenetically through the sperm. Because sperm are made throughout the male’s lifetime, they can easily serve as messengers to pass information about the environment on from fathers to their offspring. Mothers can’t pass this information on through their eggs (so far as we know), because eggs are made before a female is born, and can’t easily be changed later. Of course, a mother has plenty of other chances to pass on information about the environment to her offspring: while they’re in utero and while they’re dependent on her. But for dad, sometimes he just has that one chance; he may never interact with his offspring in any other way than through the information in his sperm.

(Why do parents want to give their offspring information about the  environment? To let them know, at formative times in their lives, how to develop. If the world is a safe one, you don’t need a highly reactive stress system. But if it’s a dangerous place, you need your store of cortisol ready to go. It’s easiest for these sorts of developmental decisions about how to tune the stress system to be made early in development — in utero or post-natally — so that’s why parents have systems to pass the information on early, early, early.)

Male mice, then, were chosen for this study. In addition to the control group of unstressed mice, there were two groups of stressed mice: mice who were stressed in adolescence, and mice who were stressed as adults. Epidemiological research in humans suggests that adolescence is an important time for epigenetic changes in sperm.

The stressors

The males were subjected to a variety of stressors. In reading the list, I was torn between sympathy for the mice, and bemusement at the entries:

Stressors, selected because they are nonhabituating, do not induce pain, and do not affect food or water intake, included the following: 36 h constant light, 15 min exposure to fox odor, novel object (marbles) overnight, 15 min restraint in a 50 ml conical tube, multiple cage changes, novel 100 dB white noise (Sleep Machine; Brookstone) overnight, and saturated bedding overnight.

Wet beds! Scary white noise! Scary marbles! And yet yes, probably very stressful to a mouse, and I should not make fun.

Breeding
The males were given time to recover from the stress and then bred. They were removed from the cage as soon as they had mated with the female, which took 1-3 days, to minimize their interactions with her, so that their stress levels could not affect her. (However, the smart reviewers at F1000 [warning: not open content] note that a stressed male might have been more aggressive in mating, which could cause the female to alter her care of her offspring.)

Offspring stress response
The offspring of stressed males, it turned out, had a less responsive stress response than the offspring of unstressed males. In other words, when these mice were stressed by being restrained in a conical tube for 15 minutes, the ones whose fathers had undergone the variety of stressors had a smaller cortisol response compared to the ones whose fathers had not been stressed. The result was almost exactly the same whether the fathers had been stressed as adolescents or as adults, which surprised the researchers.

Now, if a mouse receives information from his father (or his father’s sperm, but you know what I mean) that the world he’s going to live in is a stressful place, I would have expected that that mouse would develop a more reactive stress system, not less. Worried that terrifying marbles or a wet bed are going to attack you at any moment? Then you had better have your stress response at the ready, right?

The stress system is, of course, much more complicated than that. We don’t understand yet why some models of stress system dysregulation show less reactive responses and some show more reactive responses. For example, humans with depression or PTSD can both show either more or less reactive stress responses than mentally healthy humans. So what exactly does this mean for these particular mice? The next thing I would do is to look at their behavior. Do they act more stressed?

Offspring behavior
The offspring were subjected to quite a few and quite varied tests to see if their stress behaviors were different. The researchers tested things like how much the offspring startled in response to a loud sound; how fearful they were of being in a brightly lit box versus a dark box (mice feel safer in darkness); how long they struggled when suspended by their tails; and more. Really surprisingly (to me, at least), there were no behavioral differences between the offspring of the stressed fathers and the offspring of unstressed fathers, despite this significant difference in stress system responsiveness. So what does that mean?

The researchers tested a bunch of other stuff that left them empty handed as well, like gene expression differences in the brains and adrenals of the different sets of mice. All nothing. But what did they find that was different? They found an epigenetic difference in the fathers’ sperm.

microRNA changes in sperm
Epigenetics is all about gene expression: determining which genes are used frequently to make their associated proteins, and which are left to lie dormant. The two best understood epigenetic mechanisms, acetylation and methylation, affect how much messenger RNA (mRNA) is transcribed from a particular gene. If there is more messenger RNA for a particular gene, then it’s easier to go the next step and make more of the protein that that gene codes for, and that gene’s expression increases. The mechanism that these researchers looked at is different. Instead of methylation and acetylation, they looked at microRNAs (miRNA) in the sperm of these mice. Where methylation and acetylation affect how much messenger RNA is generated, microRNAs attach to the messenger RNA itself after it has been created, and silence it.

The way it works is this: since RNA is basically half of a double strand of DNA, it’s really sticky. It wants to find something that looks like its complement and stick to it. So microRNAs are little bits of RNA that stick to particular messenger RNAs. Then when the cell takes those messenger RNAs and tries to use them to make a protein — it can’t. Because there’s this microRNA stuck to it, blocking the sequence of the message. So microRNAs reduce the expression of a gene, but they do it one step later on in the gene to protein pathway than methylation or acetylation does.

Back to our book example, it’s like if you have a cookbook (the DNA). You copy out a recipe on a piece of paper for later use (the RNA). Then you use the recipe to make cookies (the protein). Methylation puts big rocks in front of the bookshelf so you can't get to it and get at the cookbook. Acetylation glues the pages of the book together so you can’t read it. But microRNAs are your obnoxious husband who draws in marker all over your copied recipe, so you have to go back and copy it out again. (Disclaimer: while my husband is quite capable of being obnoxious, he has never defaced any of my recipes. He has scribbled notes on the medication list for my dogs in the face of my express requests to the contrary, however. Rosie hasn’t been on ciprofloxacin for six months but it still says “cipro” on her meds list. It’s like he’s incapable of thinking ahead.)

There is a lot we don’t know about microRNAs. The whole epigenetics field is like this: we are getting to the point where we can detect these changes, but we still don’t really know what they mean. So in this study, they found that 9 microRNAs were expressed at different levels in sperm of the stressed mice versus the unstressed mice. We can make some predictions, using computer algorithms, about which messenger RNAs these microRNAs were going to stick to and silence, but we don’t know for sure that that’s what they were actually going to do.

Still, the predicted list is pretty interesting, because it contains the messenger RNA for the enzyme which controls methylation. Methylation! Another epigenetic mechanism! So is there some epigenetic chain going on here? The dad passes on microRNAs which will result in the DNA of the offspring being more or less methylated. It’s so hard to know what that means, because methylation has very different effects depending on which gene is affected, and this change is a more global change. But it’s a really intriguing finding, isn’t it?

Conclusions
This study is exciting, but I still felt a bit of disappointment as I read it. No behavior changes? Really? Is it really significant without the behavior changes? I mean, do we really care about stress system changes if there are no behavior changes? Of course we do, and I wonder if future studies will investigate different behaviors, or behaviors at different points in the mouse’s life, and then we’ll understand this system a little better.

What does it mean for dogs? Of course it is immediately applicable to the question: if a male dog is stressed, will this stress affect his offspring? The answer is a nice solid maybe. In some way that we can’t really predict or define.

But at another level, this is another step in our progress towards understanding how genes and the environment interact. Stressful situations change gene expression in the stressed individual and possibly their offspring. How, why? How can we measure it? How can we use our knowledge to help an animal who has been traumatized, or undersocialized? Watching the field of epigenetics unfold is so much fun: everything is new, we understand so little, but the new technologies are coming so fast that we’re learning more and more.

Monday, July 28, 2014

What percent nature? What percent nurture?

The Nature versus Nurture debate is over: we no longer ask if genetics governs personality or if environment does. They work together, and it’s hard to pick their effects apart. But surely we can pick their effects apart a little? For example, if a dog trainer is trying to impress upon their students the importance of getting a puppy from a good breeder who takes behavior into account — or conversely, the importance of bringing a new puppy to a puppy class: what should she tell them? 50/50? 60/40? Surely there are some numbers we can cite?

It’s a tough question, but one that researchers have tackled. The concept is called heritability: the measurement of how much of a trait is due to genetic influences, and how much is due to environment.

Human researchers have it easier than dog researchers, because humans sometimes produce identical twins, and twin and adoption studies form the basis of human heritability studies. Some twins are identical (100% identical genetics), some are fraternal (around 50% similar genetics); some are raised in the same home, and some are adopted out and raised separately. You can do some complex math to all of these situations and come out with conclusions about particular traits. Identical twins more similar than fraternal twins for a particular trait? Strong genetic component. Raised together twins more similar than raised apart twins? Strong environmental component.

These studies have given us some numbers: IQ (how someone scores on a particular standardized test) is about 40-50% heritable. Environment does the rest.

Dog studies are harder. Dogs don’t have identical twins. Theoretically, the best way to study the heritability of personality traits in dogs would be to breed parents who do or do not show the trait in question and assess the puppies, then rinse, wash, and repeat for several generations. But this is expensive and somewhat ethically fraught to do in a laboratory, so we fall back on finding populations of dogs whose personality traits have been well measured and whose pedigrees are well known.

How often does that happen? Not very. But there is a test, the Swedish Dog Mentality Assessment (DMA), which is given to a large percentage of dogs in Sweden and some other European countries. Those crazy, overly-responsible Europeans measure their dogs’ personalities before breeding them, to make sure they're breeding stable dogs. Researchers have mined this resource repeatedly to learn more about the heritability of a variety of personality traits.

As lucky as we are to have this resource, it’s not an ideal one. The DMA is a suite of behavioral assessments which are given to a dog on a particular day in a strange environment by a judge who doesn’t know the dog well. Ideally, personality is best measured over time, by someone who knows the animal very well — its owner. And, in fact, every study I read that evaluated heritability of personality using the DMA noted that one of the most important factors was not genetics but the identity of the judge who gave the test. Did some judges tend to judge more severely than others? Did dogs respond differently (more or less fearfully, perhaps) to different judges? Hard to say, but we know that the reliability of the test suffered as a result.

Perhaps more alarmingly, we’re not really sure about the validity of the test, either. What are these assessments actually measuring? They’re measuring the response of a dog to a particular stimulus in a particular situation. Can this response be generalized to a personality trait? If the dog reacts fearfully to a person wearing a sheet over his head so he looks like a ghost, does that mean the dog is fearful or just that this was a particularly surprising experience? The DMA asserts that it measures playfulness, chase-proneness, curiosity/fearlessness, and most interestingly, aggressiveness. But does it? Studies of the validity of behavioral assessments in shelter dogs — a similar situation in which a series of small tests are given to a dog by a stranger in a strange situation — have repeatedly shown that the subtleties of personality are really hard to measure in this way.

Ideally, a personality heritability study would be designed using the canine behavioral assessment and research questionnaire (C-BARQ), a questionnaire which relies on the dog's owner to assess the dog’s personality through 101 questions. This test has been found to be valid and reliable. And the University of Pennsylvania has a database of the results of this test when given to thousands of different dogs. Except... they don’t have the pedigree information for many (or perhaps not for any) of these dogs. So this isn’t a practical solution, either.

So it’s hard, and I don’t really trust the studies that are out there as a result. What do these studies find? Most studies out there use the DMA or tests like it, and find roughly 20%-50% heritability for most personality traits studied. These numbers might be artificially low, though, because the tests may not be testing real traits — behavior that is stable over time.

I was able to find one study using the C-BARQ, which had much higher heritabilities, around 70%-100%. It's a dramatic difference, but I would hesitate to assign the responsibility for that difference entirely to the C-BARQ. This study used a non-random set of samples, selecting aggressive golden retrievers and dogs related to them. With no control set of non-aggressive goldens and unrelated animals, it’s hard to know how to interpret the study’s results.

So what are the real numbers? I still want to wriggle away from an answer. I don’t think we really know. I’d love to see a C-BARQ study using a random sample — maybe by finding pedigrees for dogs already in their database, if that’s possible. Until then, I’ll guess that the real answer falls in the 30%-60% range for most traits. But, in the end, does it really matter? Genetics are important and environment is important. The best genetics can fail in the face of a poor environment, and the best environment can fail in the face of poor genetics. We can’t predict everything about our next dog; we can just do our best to make a good decision, and then provide the best possible environment for whoever comes home with us.

I owe the inspiration for this post to my students in APDT's Canine Behavioral Genetics course, who asked about the balance of nature versus nurture and would not be satisfied with vague answers.

References
  • Strandberg E. & Peter Saetre (2005). Direct genetic, maternal and litter effects on behaviour in German shepherd dogs in Sweden, Livestock Production Science, 93 (1) 33-42. DOI: http://dx.doi.org/10.1016/j.livprodsci.2004.11.004
  • Liinamo A.E., Peter A.J. Leegwater, Matthijs B.H. Schilder, Johan A.M. van Arendonk & Bernard A. van Oost (2007). Genetic variation in aggression-related traits in Golden Retriever dogs, Applied Animal Behaviour Science, 104 (1-2) 95-106. DOI: http://dx.doi.org/10.1016/j.applanim.2006.04.025

Thursday, June 5, 2014

Why genetics and dog training?

I won’t lie to you. When I first started thinking about teaching genetics courses for the Association of Professional Dog Trainers, I was mostly excited about the second class, which covers behavioral genetics of dogs. The first class was just something we had to do in order to get everyone up to speed on the basics of genetics, to have the information they needed to understand the second course.

But, of course, basic genetics is relevant to every day life with dogs and is interesting on its own. I don’t blog about genetics much, because I’m shoulders deep in highly technical stuff in my PhD program which is hard to communicate to people who aren’t equally immersed in the field. But when I stop to think, it’s not hard to come up with questions about dogs that you can’t answer without basic genetics.

  • In the past decade, new advances in technology have enabled the discoveries of more and more genes in both humans and dogs. These discoveries get reported in the popular press, such as the gene for small size in dogs (discovered in 2007). What exactly is a gene? What does it do? What does it mean to have different “versions” of a gene? It’s hard to understand these news tidbits if you don’t really get some of these basic concepts.
  • When you breed a lab and a poodle, you get a labradoodle with very predictable appearance. But if you breed two labradoodles, you can't predict what the puppies will look like. Some will look more like labs, others more like poodles. They're all the same genes, so why is one generation so different from another?
  • Why is blue merle color associated with deafness in dogs, so that if you breed two blue merles to each other, you're almost certainly going to have some deaf puppies? 
It’s easy to get caught up in the details of a field and forget that that’s not all there is. I’m trying to remember to get my nose out of the books (or PDFs of articles) once in a while and look around me.

(Genetics is beautiful and fascinating and I’m extremely lucky to have the chance to talk about it, through the lens of a shared love of dogs, in my upcoming classes with the APDT.)

Saturday, May 24, 2014

Fish personalities?!

My mobile buzzed: I had a text message from my husband. I’m bored. Call me. He was driving to New England and stuck in traffic.

I called. He asked how my day had been. How was that boring meeting? It was great, I said. I got to talk to a fellow grad student about a project of his during the coffee break. We were talking about a new way of studying fox personalities, using a method he had applied in his study of fish personalities.

ARKive photo - Male three-spined stickleback attacking pregnant female Husband: Wait. What personalities?

Me: Fish.

Husband: Did you say fish?

Me (wondering if the connection is bad): Fish.

Husband: The things with scales that swim?

Me: Fish! Yes!

Husband: ...have personalities?

Oh. Right. Sometimes I forget that my world is not other peoples’ world.

Me: Yes! Some are shy and some are bold.

Husband: Oh right. Continue.

I mean, what’s personality, really?  We make it sound like a big deal when we say that fish have them. My boss doesn’t even like me to say that our foxes have them when I am writing grant applications.

When you break it down to these small traits, like shyness and boldness, it makes more sense, though, right? Some fish are shy: when you put food in their tank, they hide a little bit longer before they will come out to eat. Some are bold: not only do they explore more and hide less, they are more likely to attack other fish who try to take their fishy belongings. If you haven’t observed these differences, I assume it's because you haven’t kept fish.


Different personalities are better (“more adaptive,” if we’re speaking Science instead of English) in different environments. An environment with lots of predators? Better to be shy, more cautious, and check out the surroundings before going for some food that's floating out there in the open. An environment with fewer predators, but lots of other fish of the same species as you? You had better go get that food fast before someone else does, rather than waiting to see if the coast is clear.

So it makes sense for a species to have a reservoir of personality types. This way, when an environment changes (there’s a new predator, or increased population density), that variation is there to be drawn upon. Lots more birds around to eat the fish all of a sudden? The fish with shyer personalities will do better, the ones with bolder personalities will do worse, and the population will gradually come to have more shy fish in it, so that the population as a whole can survive the change in environment.

For sure, human personality is a lot more complex than fish personality. But that is exactly why my friend’s lab studies fish: better to try to understand a simple system first before tackling the more complex one. A lesson I don’t seem to have learned, jumping right in with my questions about dog personality. Oh well.

[If you’re a dog trainer or just interested in dog genetics, you can learn about the genetics of dog behavior with me this summer in an online course with the APDT!]

Sunday, May 4, 2014

On nature and nurture and their interactions to make a personality

My mom called me yesterday because she had experienced some Science and was excited about it. She was watching a TV episode about aggression and how it appears in nearly every species. She called me to say that she thought my lab should look for the gene for aggression. “It should be easy,” she said, “because it should be the same gene in every animal.”

Aggressive silver fox


Yeah, you’d think that there would be single genes controlling bits of our personalities (human and dog — I think dogs are much more interesting, but in this case it’s much the same problem). Only ten or twenty years ago we thought we were in the endgame to find these genes: once the human genome was sequenced, we expected to be able to do a series of big studies to find these answers. Take a few hundred humans and sort them into “violent“ and “not violent.” Then look at markers in their genomes and use computers to find associations: all the violent people should share one marker, which will tell you where the gene for violence is. Done.

But we did those studies and we found, again and again, that these sorts of personality traits don’t give up their answers this way. In fact, in the case of zero personality traits have we found one (or even two or three) genes that control that trait. Sometimes we find genes that we think control a solid chunk of a trait, only to find that it was a statistical error — if you ask enough questions, you’ll find an interesting set of data just by chance. But if you ask the same question of another set of data (in other words, do another study), you’ll see that the first one was wrong. And this is what we have seen, for trait after trait.

Now, occasionally we’ll find a personality trait for which a little bit of it can be explained by one gene. When I say a little bit, I mean that if there is a normal amount of variety in this trait — say, in how violent a person is, ranging all the way from a pacifist to a psychopath — then the genes we find will explain about 0.1 percent of that variety. The rest is — what? Chance? Environment?

It’s a bunch of things, probably. For one thing, it’s surprisingly hard to define a personality trait. What’s violence? In dogs, we diagnose different kinds of aggression: territorial aggression, owner-directed aggression, dog-dog aggression, fear aggression. Are these all the same thing? Probably not. So instead of looking for one trait, “aggression,” should we look for four traits? Maybe. But do we actually know that those are the right four? Maybe there are six. Maybe there are ten. Maybe there are a hundred. We need to understand the traits we study better, and ask more detailed questions about them.

For another thing, yes, environment is important! Genes are important, but they are nowhere near the whole story. And environment is complicated. Certainly the difference between a pet store puppyhood and early life with a responsible breeder is huge. But can you lump early life experience into two bins, “good” versus “bad”? There are all kinds of variables. In the pet store, what kind of crate was the puppy kept in, how much interaction did it get, how young was it when it arrived? At the breeder’s, were there other adult dogs besides the mother to interact with, were there any small children, were there any bad interactions with other dogs or people? And a hundred, a thousand more questions.

I read recently about a pair of conjoined twins with very different personalities. These two had the same genes, because they came from the same embryo originally. And they had the same environment, because due to being conjoined they had to spend their lives in each other’s company. So how could their personalities differ? The article theorized that they reacted to each other, with one taking a bold, outgoing role and the other becoming shy and retiring in compensation.

And finally, the most interesting idea, in my opinion as a genomics researcher: what if we aren’t going to find the answer by looking at the sequences of DNA that make up genes? What if we are going to find the answer by looking at how the genes are regulated? If it isn’t that my dog is more fearful because some gene is a little broken, but she is more fearful because some gene is getting turned on much more or much less often than it should? It’s hard to investigate gene regulation when you have questions about the brain, because to do it you kind of have to get inside the brain, and it’s hard to do that without killing the person you’re studying. But I think looking at regulation is where things are going to have to go, and researchers are working on finding non-lethal ways of doing it.

So, nature and nurture: both important. Personality: super, super complicated. But also wicked interesting.

[If you’re a dog trainer or just interested in dog genetics, you can learn about the genetics of dog behavior with me this summer in an online course with the APDT!]

Thursday, April 24, 2014

I'm not dead

Dear everybody: I'm not dead, I'm just spending all my writing energy writing other things than blog posts. I keep asking my brain for an inspiration for a blog post and it keeps telling me "but you just wrote that long story for that magazine!" Ah well. I will keep asking.