Category Archives: soul

Happy Fat Tuesday from Schneider Lab

As Fat Tuesday approaches, my mind turns to cycles of indulgence and moderation. This is not news to women. They are quite familiar with regular, repeated bouts of uncontrollable appetite. Women are more prone to obesity and binge eating, and their binges are more likely to occur at a certain phase of the menstrual cycle. These differences are related, at least in part, to changes in hormones secreted from the ovaries. The ovaries secrete steroid hormones, such as estradiol and progesterone. Changes in estradiol and progesterone secretion alter the steroid environment in the brain and body, so that when hunger strikes, we might feel “just peckish” at one stage of the menstrual cycle or ravenously hungry at another phase.

How does this work? I received a four-year research grant from the National Science Foundation in 2013 to study ovarian hormone effects on appetite.

One clue to understanding estradiol is that it affects sexual desire and hunger for food in the opposite direction. In the middle of the menstrual cycle, when estradiol levels are highest, the appetite for food falls to its lowest level. This is the time when females are most fertile and sexual desire peaks (although sexual activity can occur any time during the menstrual cycle). As women approach menstruation, progesterone levels and the appetite for food rise while sexual desire tends to fall. Similarly, after menopause, as the ovarian hormones wane so does sexual urgency (to different degrees, depending on the individual).

So, changes in hunger for food are correlated with changes ovarian steroid hormones, but correlations cannot tell you what causes what. My work started by looking for brain hormones secreted by cells with steroid receptors, brain hormones that increase the appetite for food and decrease the desire for sex. The problem is, we can’t really muck around in our own brains to study these hormones (neuropeptides). It’s difficult to study human food intake and sexual behavior because people lie about how much and what they eat. Don’t even get me started on measuring their sexual desire. No thanks. I like to study Syrian hamsters because I can precisely control what they eat, and they ovulate like clockwork every 4 days, unlike women who ovulate every 24-32 days. Plus, hamsters have a great way to demonstrate their hunger. After a period of dieting (say, we feed them only 75% of their normal daily food intake for a week), when we give them back their food, they increase their food hoarding. The hungrier they are the more food they carry in their cheek pouches from a distant source to their home cage. We can measure hamster sexual desire and hunger for food quite easily and accurately.

1 SFRR CHAPTER 3 fig hamsters hoarding over cycle

Hamster food hoarding (mean and s.e.m.) over the four days of the ovulatory cycle in food-restricted (open triangles, dotted lines) and food-unlimited (solid circles and lines) females housed with the choice between staying home, visiting a male, or hoarding food. The predominant sex behavior of the food-restricted female is shown in a cartoon above the hoarding data. On day 4 of the cycle, the periovulatory day, the females show mating behavior. On day 3, they show vaginal scent marking but do not mate. On days 1 and 2 they spend more time hoarding food than visiting the male. (Adapted from Klingerman et al., 2010 by Jay Alexander)

            Candice Klingerman (a former grad student in my lab at Lehigh University and now a real professor at Bloomsburg University), found that hamsters on calorie-restricting diets show little interest in males and spend most of their time busily hoarding food, except on the day of ovulation. As ovulation approaches, however, they spend more and more time near the males. What you might find surprising is unrestricted females are obsessed with males throughout the ovulatory cycle! Whether they are ovulating or not, they ignore the food and spend more than 75% of their time leaving vaginal scent marks near the males. Like most rodents, they mate only on the day of ovulation, but the chubbier, calorie-unrestricted females prefer males over food every day of their four-day cycle. The differences between the calorie-restricted and unrestricted females are illustrated in the graph to the right, where you can see that calorie-restricted females do lots of food hoarding on most days of the cycle, with a conspicuous dip at the time of ovulation. The unrestricted females’ hoarding levels are low and flat throughout the cycle because they spend most of their time with the male (Klingerman et al., 2010).

You can see from the figure above that living in an “all-you-can-eat” buffet masks the effects of the ovarian cycle on the appetites for food and sex. Females on the “all-you-can-eat” diet consistently prefer to court males rather than stock their larder with food. Those females that are calorie-limited save all their sexual ardor for the small window of fertility on the day of ovulation. They spend the rest of the ovulatory cycle busily hoarding food. In the wild, this would ensure that there will be plenty of energy available for their offspring if their mating results in a pregnancy. This result makes me wonder whether our understanding of sex hormones has been clouded by studying animals housed in small cages with unlimited food. It makes me wonder how much our own species has diverged from our ancestors, now that we have adopted a sedentary lifestyle with food available in office vending machines, coffee break rooms, fast food restaurants, and well-stocked homes. No wonder we sit around watching Game of Thrones.

What are the brain differences between the hamsters on a limited-calorie diet and the hamsters at the “all-you-can-eat” buffet? I suspected that I would be able to find a brain hormone (neuropeptide) secreted by cells that have steroid receptors. I further suspected that the secretion of this neuropeptide is increased by food restriction. A review of the literature revealed many such chemicals. I have posted a handy table in a previous blog post here.

At the moment, we are interested in gonadotropin inhibiting hormone, GnIH. The figure below shows a hamster brain cell (neuron) that produces GnIH (a neuropeptide), which is stained red. Those GnIH cells that were activated by food restriction are shown in red with a green/yellow dot in the middle. These are cells labeled for GnIH and Fos, a marker for cellular activation. I got interested in GnIH when my colleague, Lance Kriegsfeld at the University of California at Berkeley, showed that GnIH inhibits reproduction in Syrian hamsters.

40x-GnIH-Fos(1)

Brain cells stained for GnIH (red) and Fos (green). The red stain represents GnIH which occurs in the cytoplasm and thus colors a wide area of the cell body. The greed stain represents the proto-oncogene product Fos, a protein that is synthesized upon cellular activation. Fos resides within the cell nucleus. Cells that are red with a green/yellow stained nucleus are double-labeled with GnIH and Fos. These represent GnIH-containing cells that have been activated by food restriction. (Photograph and immunohistochemistry by Noah Benton)

Some of my other great colleagues in Australia (e.g., Iain Clarke) were showing that GnIH increases food intake in sheep, monkeys, and rats. GnIH sounded promising. Thus, I approached Lance about studying the effects of GnIH on the appetites for food and sex in Syrian hamsters. My student Candice Klingerman partnered with a grad student from the Kriegsfeld lab, Wilbur P. Williams. Together, Klingerman and Williams found that the level of calorie restriction was a good predictor of the level of GnIH cell activation (Klingerman, Williams, et al., 2011).

           This suggested that GnIH might be part of the system that orchestrates the appetities for food and sex. This was confirmed by Noah Benton (Lehigh) and David Piekarski (UC-Berkeley). They administered GnIH to the brains of well fed females, and found that the GnIH-treated hamsters acted like they were starving. Their sexual appetites were lowered and their hunger for food was increased by GnIH treatment in the brain.

            Another prediction you can make based on the hoarding data shown above is that GnIH will have different effects depending on the day of the ovualtory cycle (and the levels of estradiol and progesterone secreted from the ovary). Consistent with this idea, my student Noah Benton is finding that in food-restricted females, the activity of GnIH is elevated only during the nonfertile periods of the female cycle. In the figure above, GnIH cells are shown in red, and the activation of those cells is indicated by the central dot stained green for Fos, a protein that shows up in cells that have been activated. Noah double-labeled cells for both Fos and GnIH in food-restricted and food-unlimited females on every day of the ovulatory cycle. On nonfertile days of the cycle, there are significantly more GnIH cells activated in food-restricted compared to food-unlimited females. As you would predict from their sexy behavior, however, on the day of ovulation, GnIH is not elevated by food restriction. Go, Noah!

GnIH activity is usually elevated in food-restricted females, except at ovulation. We think the effects of GnIH are dampened by one of the hormones that is high around the time of ovulation. Noah Benton’s dissertation work will determine which ovarian steroid hormones and receptor are important for these effects. Will it be estradiol, progesterone, or testosterone? Place your bets.

Many obesity researchers think that appetite suppressing hormones are suppose to function to preserve our youthful figures and keep our body weights in fashionable and healthy limits. The work of my students shows that an important function of these hormones is to orchestrate the appetites for food and sex, perhaps to maximize reproductive success in environments where energy availability fluctuates. These effects are short-lived and change rapidly in the small time window of fertility (basically 1 day of the 4-day ovulatory cycle). It is probably unrealistic to expect any one of these neuropeptides to be a long-term or permanent cure obesity. Maybe we should think more broadly about how all this obesity has come about, and put some energy into understanding the link between energy balance (food intake, body fat storage, and energy expenditure) and reproduction.

Meanwhile, happy Mardi Gras!  http://www.youtube.com/watch?v=mcNJpIp8w0Y 

             

           

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March 1, 2014 · 4:40 pm

Taint That Peculiar

English: House mouse, 4 days old.

English: House mouse, 4 days old. (Photo credit: Wikipedia)

Mind blown. I just discovered that data I collected one afternoon during my junior year of college is important and useful, now. I was sitting at the banquet of the annual SBN conference, when former president of the society, Art Arnold, said “Hey, that paper you published 30 some years ago has been so useful in getting researchers to start paying attention to sex differences.” At first I was confused. Was he talking about that old review article I co-authored with my postdoctoral mentor (which keeps getting cited even though we’ve both written much better reviews)? Or maybe he meant the overrated article we published in Science in 1989? No, it turned out he was excited about a paper that emerged from a side project loosely related to my FSU undergraduate honors thesis in which I measured the length of mouse taints

It all started when some grad students and a postdoc decided they wanted to give the googly-eyed, work-study student (me) a “discovery experience.” This, by the way, is the essence of mentoring, and I am forever grateful to those guys. They gave me 50 or so one-day-old mouse pups and said “Figure out which ones are male and which are female; collect some data; and show us how to document the sex difference.” I sat alone in the room, like the miller’s daughter in Rumplestiltskin, staring hopelessly at the seemingly identical embryonic bodies (see pic). I focused on the place where you might expect to see a tiny penis, but where I hoped to see a penis, there was only a small bump and a small hole behind the bump. Each and every mouse had the same bump-and-hole arrangement. Finally, after more staring, I noticed that the bump-to-hole distance was small in some mice and large in others. The hoity toity science name for this is the “anogenital distance.” To the likes of me, the term for this distance is the taint, because it tain’t the genital and it tain’t the other thing. So, I divided the pups into two piles, a short-taint pile and long-taint pile. Now how was the miller’s daughter going to spin mouse taint into scientific gold? My Rumplestiltskin was a professor who studied the visual system, the late Howard Baker. I told him I was trying to measure a very tiny distance that I could barely see with my naked eye. He gave me a reticle, a glass eyepiece for a microscope with a ruler engraved on the lens, which allows measurements accurate to 0.01 mm. I measured all the little mouse taints, and found the mean for each group. I did a t-test, and the difference was highly significant. I guessed that the those mice with the smaller taints were the females. Correct! Chuck and John, the grad students and postdoc in the lab already knew the answer, but since I discovered it without any help, they encouraged me to publish a paper. Hence the 1978 publication in Behavioral Research Methods and Instrumentation, “Determining the sex of neonatal mice, Mus musculus.

Howard Baker never demanded my first-born son, few colleagues since have been so generous regarding authorship, and I rarely think of this one-page publication in a minor journal. Half the time I don’t even list it on my c.v. I just assumed in 1978 that everyone knew the importance of knowing the sex of your experimental animals on the day of birth. Tragically, the majority of scientists stubbornly refuse to look at both sexes in their experiments. The fact is, males and females differ in response to pain, drugs, hormones, and their propensity for many different diseases. Sex hormones have profound effects during early development, both pre- and neonatally, and these hormones masculinize or feminize the individual, determining their adult response to drugs, cancer, infections, pain, diet and exercise. We now have a large body of data on common diseases and biological processes; most of it on males by a ratio of 5:1. In 1993, the importance of sex differences and of early hormonal effects prompted the NIH to mandate the enrollment of women in human clinical trials. This mandate should also apply to the animal research on which the human research is based. Excuses for the male-only bias include “females are too variable due to their estrous cycles,” and even more ridiculous, “I don’t know how to tell the difference between the sexes.” It taint rocket science; even a work-study student at a state school could figure it out.

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