Author: Jerry Husak Page 1 of 3

Exercise has many effects on your body, most of which are good, and is why we humans do it to stay healthy. However, some of those changes, especially under very intense regimens, can have unseen consequences that might be bad. Your immune system, for example, responds to different types of exercise (aerobic endurance versus anaerobic resistance) by altering which branch of your immune system is dominant at that time. Both kinds of exercise tend to increase the more specific ‘humoral immunity’ (B-cell immunity below) over the more general ‘cell-mediated immunity (T-cell immunity below), though the routes to get there are very different for the two kinds of exercise. However, most of what we know about exercise-immunity tradeoffs is from humans and rodents. What about in other animals that have limited access to resources? Might simple energy limitation cause overall immunity suppression when energy is diverted to athletic performance?

My former student Andrew Wang and I studied this experimentally with green anoles. We trained lizards for endurance on a treadmill, or for resistance with weights on a racetrack, for 9 weeks, and compared those to a sedentary control group. Both of these types of locomotion are important to anoles in the wild, and the training schedule was meant to simulate the high end of movement patterns in nature. We then subjected them to three immune challenges: (1) swelling response to phytohemagglutinin (cell-mediated immunity), (2) antibody response to sheep red blood cells (humoral immunity), and (3) wound healing ability (integrated response across all parts). We expected that if simple energy limitation explained tradeoffs, all immune measures would decrease, with endurance-trained suffering the most. If protein limitation was the reason for tradeoffs, then we expected all immune measures to decrease, with sprint-trained suffering the worst. Finally, if the response is due to changes in molecular pathways specific to type of exercise, we expected humoral immunity to be favored over cell-mediated in both trained groups.

Figure 1 from Wang and Husak (2020)

Our results did not support only one of our hypotheses. Endurance-trained lizards had the lowest cell-mediated immunity, whereas sprint-trained had the lowest wound healing ability. Antibody production did not differ among treatments. Our hypothesis of sprint-trained lizards (or even endurance-trained) having the lowest overall immune function was not supported, suggesting that energy limitation alone does not explain immune system alteration. For sprint-trained lizards, energy was likely important, since wound healing, an expensive task, went down the most in that group. For endurance-trained lizards, though, the change in T helper cell production favored humoral over cell-mediated immunity. Since both types of exercise favor humoral immunity, it was not too surprising that antibody production did not differ among treatments. Lots of questions remain to be answered, though!

What does this all mean? In nature, individuals vary dramatically in how much, and for how long, they move around their environment. Those that are more active, thus likely have different immune capabilities compared to more sedentary individuals. It would be very interesting to see how natural variation in survival strategies, high-performance versus high-immunity, affected success in nature. This is a wide-open field for anoles and other reptiles!

Source: Wang, A. Z. and J. F. Husak. 2020. Endurance and sprint training affect immune function differently in green anole lizards (Anolis carolinensis). Journal of Experimental Biology 223: jeb232132.

Green anoles were trained, marked, released, and tracked in New Orleans. Photo by Jerry Husak.

In the US, we spend a lot of money trying to stay fit. This isn’t necessarily a bad thing, since there is a major problem with obesity and type II diabetes in the country. In humans, investment in increased performance abilities via the exercise response is also associated with numerous health benefits, such as decreased incidences of metabolic syndrome, cardiovascular disease, obesity, and diabetes, and aerobic capacity is considered to be an important predictor of longevity. However, it is these “side effects” that make exercise so interesting to an evolutionary biologist, because those wide-ranging, multi-system responses can tell us something about the evolution of animal life histories.

Superior locomotor performance has been shown to be advantageous to a variety of organisms in terms of male combat success, survival, and fitness. In addition, one of the most striking aspects of exercise physiology is how similar the response to exercise is across vertebrate animals, suggesting that the response to exercise is both ancient (yes, even fish respond to exercise!) and adaptive. However, until now, no studies have tested whether non-human animals that invest in increased athletic performance through exercise realize a fitness advantage in nature.

Jerry Husak and Simon Lailvaux set out to test whether superior performance after exercise training would increase survival probability in green anole lizards. Previous work with green anoles showed that they respond to different forms of exercise training, and that enhanced performance results in tradeoffs in other systems, such as reproduction and immnuocompetence. Why? Because performance abilities are energetically expensive to build, maintain, and use.

Urban islands in New Orleans where the study was conducted. Photo by Jerry Husak.

Jerry and Simon conducted their study in a New Orleans urban park that they cleared of existing lizards. They trained 30 lizards (15 male, 15 female) for endurance on a treadmill, 30 lizards for sprinting with weights on a racetrack, and had 30 untrained controls. All were released into isolated, urban islands in New Orleans, LA, USA and monitored for survival over an active season, over winter, and through the next active season. They predicted that training would enhance survival during the active season, but that the associated maintenance costs of training would decrease survival overwinter compared to controls.

This male made it a year in the wilds of New Orleans, but it looks like it was a rough year. Photo by Jerry Husak.

Contrary to expectations, they found that sedentary controls realized a significant survivorship advantage over all time periods compared to trained lizards. Trained lizards had reduced immune systems and lower fat stores, suggesting that in an environment with limited resources, it does not pay to exercise too much. These results suggest that locomotor capacity is currently optimized to maximize survival in green anoles, and that forcing additional investment in performance moves them into a suboptimal phenotypic space relative to their current environmental demands. We as humans can get away with it because we are not food limited. On the other hand, this is why doctors suggest consultation before going on a diet and doing intensive exercise training.

Source: Husak, J.F., and S.P. Lailvaux. 2019. Experimentally enhanced performance decreases survival in nature. Biology Letters 15:20190160. doi: doi.org/10.1098/rsbl.2019.0160.

Leptin is made by fat cells and serves as a signal of available energy to lots of systems in the body. Diagram from healthjade.com

When you only have so much money to spend, you have to carefully consider what you’ll use it for. Do you go for instant gratification (dinner at your favorite, but expensive, restaurant!), or do you invest in something with a longer-term return (a needed kitchen appliance that will last years)? Free-living organisms have to make this choice throughout their lives. Of course they don’t cook in a kitchen, but their bodies have to ‘decide’ what to do with precious and limited energy. For our beloved anoles, in what do they invest that hard-earned energy from ingested bugs? Make more and bigger babies right away? Grow more? Invest in their immune system or locomotor performance to survive better?

Animal bodies don’t actually make ‘decisions’ about these things. Instead, hormonal and molecular mechanisms are arranged as networks in the body to make ‘decisions’ under different sets of conditions. In a new paper, Andrew Wang, a recent graduate from Jerry Husak’s lab, was curious how such decisions are made in green anoles. Previous work in the Husak lab showed that when calories are restricted, and lizards are forced to invest in athleticism via exercise training, both reproduction and immune function suffer. Why is that, and is it reversible?

The observation that trained and food-deprived lizards had little to no body fat (imagine elite marathon runners!) suggested that the hormone leptin, produced by fat cells, might be responsible. Leptin affects lots of systems in the body (see figure above), and less fat means less leptin. This means that leptin serves as a direct and convenient signal of energy stores: if you have enough energy, then you can direct organs to get to work. This fact has led to a huge literature on how leptin, as an energy signal, controls tradeoffs among traits. Hopefully you’re seeing a slight paradox here – if more leptin means more energy available, how could it mediate tradeoffs? How do you get more of one trait than another if leptin controls both in the same general direction?

Andrew conducted an experiment to find out. He replicated previous work, training and calorie restricting male and female green anoles to cause suppressed reproduction and immune function. He then gave half supplemental leptin and the other half saline, expecting leptin to ‘rescue’ reproduction, immunity, or both. The results were clear: immunity was ‘rescued,’ but reproduction was not. That is, both sexes were investing in survival-related traits to (hopefully) reproduce later instead of just reproducing right away. These results suggest that either there wasn’t enough energy for reproduction and the signal was moot, or the two traits have different sensitivities to leptin. Future work will help to disentangle these possibilities, but this work gives us more understanding of how anoles allocate energy when it’s limited.

Figure from Wang et al. (2019). Key: U=untrained, T=trained, H=high diet, R=restricted diet, L=leptin injected, S=saline injected. Note here that the swelling response to PHA injection was suppressed with training and caloriee restriction, but it was rescued with leptin (T-R-S vs T-R-L).

Paper: Wang AZ, Husak JF, Lovern M. 2019. Leptin ameliorates the immunity, but not reproduction, trade-off with endurance in lizards. J Comp Physiol B, in press. doi: 10.1007/s00360-019-01202-2

The somatropic axis regulates growth in vertebrates

Our growth during development is controlled by a complex brain-body axis called the somatotropic axis. Put simply as in the photo below, the hypothalamus in the brain signals the pituitary to release growth hormone into the body where it stimulates the liver to produce two forms of insulin-like growth facts (IGF1 and IGF2). Both growth hormone and IGF have different effects on growth of muscle and bone. While we rely on mouse models to study how IGF might impact human development, it turns out that the relative secretion of IGF1 and IGF2 over the course of life is quite different in the two species. In fact, we know little about IGF production and signaling in non-mammals.

Expanded view of the somatotropic axis to show receptors and binding proteins. From Yakar et al. (2018).

Abby Beatty, from Tonia Schwartz’s lab at Auburn University, set out to determine the developmental pattern of the somatotropic axis in brown anoles. Of course, the axis is much more complex than the diagram above, including receptors in various tissues, binding proteins that carry the signals around the body (see below), and proteins in cells that cause responses (IRSs). Abby studied expression of IGF1, IGF2, and five binding proteins during brown anole development, from embryo to hatchling to adult. She expected to find that IGF1 and 2 would be expressed differentially and that the expression patterns would differ across life stages. That’s exactly what she found. IGF1 and 2 were both low and similar in expression early in development, but at hatching IGF increased with IGF2 expressed more than IGF in adults. Surprisingly this is more like a human pattern than a mouse is!

As for the binding proteins, expression was similar for all of them in the brain, gonads, and liver, but BP3 was expressed less in the heart. It’s still unclear what these patterns in binding proteins mean for brown anole development, but they make for some excellent future research questions! Indeed, these results add to the already-long list of things that makes anoles good model systems.

Christine Rohlf from the University of St. Thomas presents her research on immune-performance tradeoffs.

Traveling to SICB is always exciting, but like any trip through crowded airports, hotels, and convention centers, you’re more likely to get sick during your travel if you’re not careful. As we all know, getting a travel cold (or worse) makes you feel terrible and certainly doesn’t make you want to run on a treadmill! The same is likely true in wild animals, including anoles. Mounting immune responses is energetically expensive, but so are other things that lizards have to do, like forage, escape predators, and process food. So, does an increasingly large immune challenge decrease a lizard’s ability to perform? Christine Rohlf, an undergraduate student in Jerry Husak’s lab at the University of St. Thomas, wanted to find out in green anoles.

Christine designed a laboratory experiment to determine whether two types of immune challenge, alone and in combination, decreased bite-force performance, sprint speed, or endurance capacity compared to controls. Some lizards received two sequential injections of lipopolysaccharide (LPS), some received a skin wound with a biopsy punch, and some received both. LPS is a signal on gram-negative bacteria that, when injected, tricks the body into thinking it is infected with bacteria. So, you get an immune response, but you don’t actually get an infection.

Surprisingly, none of the immune challenges affected sprint speed or endurance compared to controls. Although the lizards were not calorie-restricted, they were on a modest diet, meaning that energy was limited, but clearly not enough to make a difference. Apparently these two immune challenges aren’t as costly as we thought. The only effect that Christine found was that the second LPS injection significantly decreased bite force. Because bite force is likely the least energetically expensive trait of those measured (imagine running until you’re exhausted versus biting into a hard piece of French bread), Christine suspects that the decrease in bite force was due to a lack of motivation while feeling sick. Future work with calorie-restricted lizards should tell us if mounting an immune challenge is a significant cost to anoles.

Amy Payne of Trinity University presents her research on tail autotomy in 7 lizard species.

One of the most interesting features of many lizards, including anoles, is that they can willingly, and actively, lose their tails to escape predators. While it might seem counterintuitive to lose a large body part, it’s better than being eaten! Despite the obvious benefit of surviving another day, there are some costs associated with tail autotomy.

Amy Payne, a student in Michele Johnson’s lab at Trinity University of San Antonio, wanted to know whether the frequency of tail loss across seven species was associated with predatory and social use of the tail as well as energetic content of the tail. For those that are anole-inclined (which is why you’re here), Amy included A. cristatellus and A. carolinensis. She caught and measured hundreds of lizards, and made behavioral observations on them as well. She was then able to quantify how many lizards of each species had a lost/regenerated tail, as well as what proportion of each tail was lost.

Surprisingly, frequency of tail loss was not associated with using the tail in a social or predatory context. However, there was an association between these two functions of the tail: species that more often used their tail for predatory use also used their tail in social contexts more. There was no relationship between the frequency of tail loss and the proportion of the tail that was lost on average across species. But she did find some really cool results when looking at energetic content of the tail. Amy found that there was a significant positive relationship between frequency of tail loss and tail energy content. That is, the more energy that lizards have in their tails, the more frequently individuals in that species will have a lost/regenerated tail. While this seems opposite to what one might casually predict, Amy hypothesizes that the predator-distraction to survive function of tail autotomy is more likely to succeed if the tail is larger and more beneficial to the predator. In other words, if a lizard has a scrawny tail and drops it off for a predator, it is more advantageous for the predator to ignore the low-cal tail and just eat the lizard. This would put selection on species with low-energy content tails to be more prudent about when they drop their tails. These really interesting results open up some exciting areas for future research on the costs and benefits of tail autotomy!

Above: Faith Deckard presenting her research on how muscle physiology may explain variation in social behavior among Caribbean anoles.

Marathon runners and elite sprinters, like Usain Bolt, have dramatic differences in their muscle physiology that allow them to specialize in their respective track-and-field events. Whereas sprinters have lots of muscle fibers that produce high force but fatigue quickly, marathon runners have lots of muscle fibers that produce less force but allow much longer activity because of their reliance on aerobic respiration. Might this be true for our beloved Caribbean anoles, too? Faith Deckard of Michele Johnson’s lab at Trinity University tried to answer that very question. She studied six species of anoles in the Dominican Republic to test whether anoles that have higher rates of dewlap extension and extend their dewlap for a longer duration have dewlap muscles with a higher proportion of slow-twitch muscle fibers that can be used for endurance. Surprisingly there was no significant correlation between the two behavioral traits and the proportion of slow-twitch fibers! However, this scrutinizing attendee feels pretty strongly that there is a relationship that is just yet to be teased apart statistically. The raw data Faith presented looked very convincing to me, so we’ll see what the future holds for this question. Faith’s results are a very interesting clue to the still-elusive mechanisms that underlie anole behavioral diversity.

Above: Andrew Wang presenting his research on how leptin may be a mechanism underlying life-history trade-offs in green anoles.

All of the gumbo, Po boys, and beignets consumed by attendees of SICB 2017 have to go somewhere after consumption. Much of the energy contained in those delicious foods is used for very important maintenance functions in your body: metabolism, cell repair and replacement, and your immune system. What’s left over after maintenance costs can then be divided amongst other tasks, such as reproduction, movement, and wide variety of other tasks. Unlike humans, anoles do not have unlimited access to gigantic portions of gumbo, so their energetic investments require much harder decisions. Once energy from a cricket, for example, has been put into the immune system, it can no longer be used for making eggs or patrolling a territory a little bit longer. Andrew Wang of Jerry Husak’s lab at the University of St. Thomas was interested in what mechanisms are involved with anoles making these investment “decisions.” He did this by forcing allocation of resources to an energetically expensive trait (endurance running) by exercise training lizards to see what would happen to everything else that they might invest in.

Previous work showed that exercise training and diet restriction results in dramatic trade-offs with reproduction and the immune system. He suspected that what might explain this suppression was the hormone leptin, which is made by fat cells (yours make it, too). Since bigger fat cells means more leptin in the body, leptin can be thought of as a signal to the brain and body of how much resources are available for investment. Indeed, without sufficient leptin, reproduction grinds to a halt from the brain downward. Much like elite athletes, Andrew’s marathon lizards have little to no fat stores in their body, thus suggesting a role for leptin. To address this question, he supplemented half of the lizards with leptin (the rest got only saline as a control) to see if he could “rescue” immune function and reproduction. Interestingly, he found that leptin did rescue his measure of immunity, but it did not rescue reproduction. He attributes this latter finding to either (1) a lack of energetic resources to produce eggs even if there is a leptin signal or (2) the stress of the leptin injections over-rode the leptin signal in the brain where reproduction is controlled. His results suggest some very complex interactions in physiological pathways that can result in the trade-offs observed in many animal species.

Leptin is best known as a satiety hormone, but it has important roles as a signal to the body of adequate energy stores. Image from wiki.brown.edu.

Green anole image from reptilesmagazine.com.

What does it take be a good sprinter? How about a marathon runner? One might think that the traits responsible for such performance traits would be the same in males and females. If you are a green anole, that just isn’t true. Annie Cespedes, working in Simon Lailvaux’s lab at the University of New Orleans, explored the multivariate predictors of seven performance traits (sprint speed, bite force, cling force, exertion, endurance, jump power, and climbing power) in male and female green anoles. Annie explained how animals in nature rely on lots of different performance traits in their daily lives, and the large difference in body size and shape between male and female anoles might mean that the two sexes use different means to be successful in life. To add to this complexity, some individuals are just better overall at ALL performance traits than others (imagine a couch potato versus a very fit athlete), and one must account for this to understand what shapes anole performance.

Multivariate statistics allowed Annie to show that males and females do indeed differ in performance, but only in clinging ability, sprint speed, bite force, and jump power. Even more interesting, the suites of morphlogical traits that explained performance ability differed substantially between the sexes. For example, small females with large leg muscles were better sprinters and jumpers than females who are smaller and are better biters and endurance runners. What is especially important about Annie’s research is her approach. When considering how animals evolve, one must do so by simultaneously looking at a multitude of traits that might impact their survival and reproduction. By knowing how morphology predicts performance, we can begin to better understand how evolution will shape that morphology when selection acts on those performance traits.