Pandas are lazy!

Pandas are closely related to carnivorous mammals (like all the other bears), but they consume mostly bamboo.  Their digestive tracts are short and adapted for digesting meat, not cellulose that is found in plants.  In fact, they only digest about 20% of all the bamboo they eat, and they eat a lot of bamboo (30-60 pounds a day)!  How are these large, adorable bears able to get enough energy to function from their inefficient digestion of bamboo?  Researchers in China and Scotland addressed this question by studying captive and wild pandas, described in a recent Science article.

Low energy expenditure

Nie et al. measured the daily energy expenditure of the pandas and found that they used an unusually low amount of energy, only 37.7% of the predicted value based on their body mass.  In fact, pandas are expending energy at levels similar to the three-toed sloth, the epitome of a low-energy mammal.  The measly amount of nutrients they get from all that bamboo would be able to sustain such a low energy expenditure, so that’s how the panda is able to get by with such a maladapted digestive system.

How do the pandas manage to spend so little energy?  There must be some adaptations that are allowing the panda to survive without expending so much energy.  The authors found a number of these adaptations:

1) Pandas have a thick layer of fur, so they can maintain their internal body temperature with less heat loss through the skin.  The researchers measured temperature at the surface of various animals and the pandas consistently were cooler than other mammals (like a cow or dog).  Their internal body temperature would be considerably warmer because the fur helps insulate them, so they don’t have to spend as much energy on maintaining their body temperature.

2) Pandas are lazy.  No surprise: pandas spend more time inactive and when they do move, it is slowly.  So that is less energy needed for muscle contractions.

3) Pandas have small brains, livers and kidneys, so their organs need less energy.

4) Pandas have a low resting metabolic rate, which is driven by the thyroid hormones, T3 and T4.  In fact, levels of these two hormones were considerably lower than for other mammals of the same body mass, even lower than a hibernating bear.  The thyroid hormones regulate protein, carbohydrate and fat metabolism, as well as growth and development.  If the pandas don’t need to produce as much heat or energy, then there is no reason to have a high metabolic rate.

Interestingly, pandas have a single mutation in a gene called DUOX2, which is not found in any other mammals.  DUOX2 encodes for a protein that is necessary for the production of T3 and T4.  The mutation causes a premature “stop” in the protein, so it likely affects the function of DUOX2. 

In other words, pandas cannot synthesize T3 and T4 as well because of this mutation, so they have a reduced metabolic rate.  But that’s okay, because they are good at maintaining their body temperature and they have developed an enjoyable lifestyle of relaxing and eating.  The fact that their digestive tracts have not evolved for plant digestion is alright given the fact that they don’t really need that much energy from their food.  So it all works out: pandas are able to survive on their diet of bamboo and we can watch them sit around.

Probiotics for autism

The human microbiome is a hot topic in biology these days.  It is becoming clear that the microbes living in and on our body can have major consequences for our health and happiness.  In fact, abnormalities in the gut microbiome may underlie one of the great medical mysteries of our time: autism.   That some bacteria in our intestines could affect our behaviors and brain development is mind blowing.

Hsiao et al. recently published a study in the journal Cell that investigated the connection between the gut microbiome and autism using a mouse model of autism.  They were drawn to this subject based on the fact that individuals with autism spectrum disorder (ASD) often have gastrointestinal abnormalities, like irritable bowel syndrome and increased intestine permeability.

Autistic mice?

Apparently you can produce mice that exhibit the “core communicative, social and stereotyped impairments” associated with ASD, by injecting their pregnant mothers with a molecule that stimulates an immune response.  In humans, maternal infection is linked to increased risk of autism in their children.  The production of these mice was the most questionable part of the paper in my opinion.  They never call these mice autistic, and the mice do show impairments associated with neurological diseases.  So perhaps we should think of it as a model of a generic neurological disorder.  For the sake of simplicity, though, I will refer to them as “autistic mice”, but remember that it is not a perfect model system.

They find that the autistic mice have various defects in their gastrointestinal (GI) tract.  For instance, their intestinal walls are leaky, so molecules that are not supposed to be absorbed can cross from the gut into the blood stream.  This problem seems to be caused by the fact that these mice express less of the proteins that make the tight junctions between cells.  Think of these as fences between cells, so molecules can’t sneak through there into the body.  In an ideal situation, all molecules that are absorbed from the gut must go through the cells, a process which is highly regulated. 

Tight junctions prevent molecules from passing from the gut into the blood.  Image adapted from dbriers.com

They find a number of metabolites that are produced in the intestine from bacteria, which end up in the blood of autistic mice, but not in the normal mice.  In other words, these are potentially toxic molecules that they need to get rid of, but the toxins are leaking into the blood of the autistic mice.  That’s not good.  In fact, if you inject one of these molecules into a normal mouse, it will become more anxious, similar to the autistic mice.  They couldn’t reproduce all of the behaviors of the autistic mice just with this one molecule, but it’s a good proof of principle.  Presumably it’s the build up of all of these metabolites in the blood that cause impairments of the nervous system.

Dysbiosis of the intestinal flora

I love that word “dysbiosis”.  It means that the intestinal microbiome is out of whack.  The wrong types of bacteria are in there messing stuff up.  Hsiao et al. found a number of species present in the autistic mice that were not in normal mice and vice versa.  Presumably this imbalance in the microbiome is what is making the gut leaky. 

To prove this, the authors fed the autistic mice a probiotic (a “good” type of bacteria) called Bacteroides fragilis (B. frag).  Interestingly, B. frag never actually colonized the guts of the mice, but just having it pass through helped to restore the normal microbiome.  Some of the species that were only present in autistic mice disappeared after they consumed B. frag.  The leakiness of the gut was almost completely reversed, including expression of tight junction proteins.  It wasn’t a perfect reversal, but a number of those metabolites in the blood decreased back to normal.

Behavior affected by microbiome

To review: when a pregnant mouse has an infection, her offspring show signs of autism (a mouse-version).  Somehow this infection causes the wrong bacteria to colonize the guts of the offspring.  The dysbiosis leads to changes in gene expression and a leaky gut that allows toxic molecules into the blood stream, thus affecting the development of the nervous system.  Consumption of a probiotic at weaning age fixes a lot of the gut issues.  Does it also reverse some of the behavior impairments associated with autism?

The short answer is yes!  Autistic mice fed B. frag were less anxious, less obsessive, more communicative and interacted more with other mice.  The test for obsessive behavior was kind of cute.  The mice were put in a cage filled with sand with marbles sitting on top.  The autistic-like mice bury a greater percentage of the marbles, demonstrating a stereotyped behavior.

Yogurt from everyone!

If I had an autistic child and read this paper, I would start them on probiotics right away.  I mean probiotics are good for everyone, right, so it definitely seems worth trying.  In fact, the authors say that B. fragilis is depleted in human ASD children compared to matched controls.  Furthermore, probiotics have already been shown to be beneficial in treating chronic fatigue syndrome.   

The authors end their paper with this bold statement: “We propose the transformative concept that autism, and likely other behavioral conditions, are potentially diseases involving the gut that ultimately impact the immune, metabolic, and nervous systems, and that microbiome-mediated therapies may be a safe and effective treatment for these neurodevelopmental disorders.”

Throw another adipocyte on the fire

Humans are able to live in so many different climates, in a wide range of temperatures and yet our inner core body temperature remains nearly constant.  This ability to thermoregulate has something to do, of course, with clothing and the ability to cool and heat our living spaces, but our bodies also offer many adaptations to regulate body temperature.  If it’s too hot, we sweat, releasing excess heat through evaporative cooling.  If it’s too cold, we shiver, producing heat in our working muscles.  The production of heat through physiological mechanisms is called thermogenesis and also includes a non-shivering version.  Today’s paper is about non-shivering thermogenesis, which is when our fat cells produce heat.

Non-shivering thermogenesis

To understand how non-shivering thermogenesis works, we need to take a step back and discuss cellular respiration.  The cells of our body store energy from food in the chemical bonds of a molecule called ATP.  During cellular respiration, a cell will convert glucose or fat into carbon dioxide, while slowly tapping into the energy in those food molecules in order to make ATP.  The final step of cellular respiration is that the energy from the electrons in glucose are passed from protein to protein, releasing energy that is used to pump protons into a membrane-bound cellular space.  You can think of these protons as a form of potential energy, like stuffing a closet full of balls.  When you open up the closet door, all the balls come tumbling out, releasing their potential energy in the process.  During cellular respiration, this potential energy is used by an enzyme to make ATP.  During non-shivering thermogenesis, though, the potential energy stored in all those protons stuffed into a small space is released by the cell as heat.  Thus, the energy from food is used to heat the body rather than being stored in ATP.

The main type of cell that does non-shivering thermogenesis is brown adipocytes, or fat cells.  Brown fat is very common in infants, but is also found in adult humans in the upper chest and neck.  The purpose of brown fat is to provide heat for the body.  Thus, non-shivering thermogenesis is activated by a drop in body temperature.  The cold temperature is sensed by the brain, which activates the sympathetic nervous system (the “fight or flight” response), which signals to the brown fat cells to express the genes necessary to bypass ATP production and release heat instead.  In a recent paper published in PNAS, Ye et al. describe how a different type of fat cell is able to skip all the nervous system steps and sense the cold directly (red arrow in diagram).  It is pretty cool that the fat cells are able to sense temperature, as if they were neurons, and can act autonomously to remedy the situation.  No need for a brain here!

Independent thermogenesis

Through a series of experiments, the authors demonstrate that a particular type of fat cell will express genes necessary for non-shivering thermogenesis when exposed to cold, independent of sympathetic nervous system activation.

In one experiment, they grew fat cells at different temperatures and measured gene expression using a technique called quantitative PCR (qPCR).  The idea behind this technique is that if a gene is highly expressed, there will be a lot of mRNA in the cell (remember the “central dogma” of molecular biology) and qPCR is a method for measuring the concentration of mRNA for a particular gene.  They focused their measurements on thermogenic genes that are known to be part of the non-shivering thermogenesis mechanism, such as Ucp1, which is the enzyme that actually allows the protons to fall back across the membrane, thereby releasing their energy as heat.  They found that these fat cells that were exposed to the cold expressed more Ucp1 mRNA, even in the absence of any nervous system.  These are just cells in a dish, so this must be an intrinsic property of fat cells.

It wasn’t just any fat cell that had this response.  In fact, brown adipocytes did not express more Ucp1 in the cold.  It was a different type of fat cell called a white adipocyte.  What is white fat?  The majority of fat in our body is white fat and its purpose is to store fat for energy (for cellular respiration) and to act as a thermal insulator, so we don’t lose as much heat through our skin.  There is one subtype of white fat that has been shown to do non-shivering thermogenesis and it was this type that could express thermogenic genes, like Ucp1, in the cold, independent of the nervous system.

Okay, so these white fat cells don’t need input from the nervous system, but do they still use the same intracellular pathway to turn on expression of these genes?  Normally, when a fat cell is activated by the sympathetic nervous system, it sets off a molecular cascade of events inside the cell, which involves activation of molecules in a pathway called the cAMP pathway (as shown in the diagram).  The authors inhibited this pathway in various ways and found that the cells could still respond to the cold as before, so this effect must use a different pathway.

There are still a number of open questions, such as: how do fat cells sense temperature?  Do they use the same types of receptors as temperature-sensitive neurons?  Why are some white fat cells independent, but brown fat cells need the nervous system to activate thermogenesis?  One thing that is clear, however, is that white fat cells are clearly important for temperature regulation as well as fat storage.  The authors suggest that tapping into thermogenesis might be a good way to help obese patients get rid of excess energy storage by releasing it as heat.  This pathway that is independent of the sympathetic nervous system could allow medications to target only the fat cells without involving the sympathetic nervous system which controls so many other functions in the body.

Something to think about as the cold Bay Area summer sets in.

Swapping eggs

This week’s paper describes a new technique that could be used to manipulate human oocytes (i.e. eggs) to prevent a group of diseases called mitochondrial diseases.  The paper was presented by Tachibana et al. in Nature along with a similar paper by Paull et al.  For the sake of brevity, I will only discuss the findings from the first paper.

Mitochondria

So what are mitochondria?  Mitochondria are little compartments in the cell that make cellular energy.  They convert the energy stored in food into an energy source that the cell can use to drive chemical reactions.  In other words, they are absolutely essential for our survival.  The oxygen that we breathe in goes to the mitochondria to aid in this energy conversion, and we all know how vital oxygen is. 

There are two other interesting facts about mitochondria that relate to our story:

1) All the mitochondria in our body are duplicates of the mitochondria that were in our mother’s egg.  In other words, embryonic mitochondria are not made from our genomic DNA (gDNA) or from sperm contributions.

2) Mitochondria have their own DNA , which directs the synthesis of proteins that are necessary for their function.  This DNA is known as mitochondrial DNA (mtDNA) and it is only inherited from the mother, since all mitochondria originate from the egg.

If there are mutations in the mtDNA, then this can lead to problems with the synthesis of cellular energy, which can lead to human diseases known as mitochondrial diseases.  There are different types of mutations, which can affect people in different ways and with differing severities.  In this paper, the authors propose a way to prevent mitochondrial diseases from being inherited from generation to generation.  Let’s see how that works.

Nuclear transplantation

Let’s say you have a female patient with a mitochondrial disease, who wants to have a healthy child.  She is guaranteed to pass this disease on to her child via the mitochondria in her oocytes.  However, most of what makes the child “hers” is what lies in the mother’s genomic DNA, not in the mitochondrial DNA.  What if you could take the mother’s genomic DNA (plus the DNA from the father) and stick it into a healthy “enucleated” oocyte from a donor who has good, functioning mitochondria?  All the genomic DNA will have to be cleared out of the donated oocyte first, creating an enucleated egg.  The embryo that results from this nuclear transplantation will have genomic DNA from its mother and father, but its mitochondria will originate from the donor oocyte.  This would circumvent the mutated mtDNA that is in the real mother’s oocyte.

Tachibana et al. obtained human oocytes from volunteers and transfered the genomic DNA from one into another.  They then injected these oocytes with sperm DNA (like during real fertilization) and observed what happened.  Some oocytes failed to be fertilized and others died soon after, but a handful of oocytes survived into the blastula stage of development.  You can’t really grow a human embryo in a dish beyond the blastula stage and they are not allowed (yet) to implant these into women, so we don’t know what would happen to a child born from this procedure. 

They did carry out the above scenario with monkeys.  They transplanted the genomic DNA from one oocyte into another and implanted the blastula into another female monkey who carried the embryo to term.  The monkey youths are 3 years old now and doing just fine.  Their maternal genomic DNA is from one mother and their mitochondria are from a different oocyte donor. 

Isn’t this amazing?  I seriously doubt this procedure will be approved for human use anytime soon, because it’s too much like cloning, which basically follows the same procedure of putting genomic DNA into an enucleated egg.  It's a cool idea, though. 

Burning carbs in the Andes

I am finally back from a very busy semester.  I taught physiology classes at Mills College and UC Berkeley this semester, so I have been interested in new topics in human physiology.  This week’s paper by Schippers et al. came out recently in Current Biology and describes adaptations that mice must make in order to live at high altitude.  They compared the metabolism of mice that live at 4000m above sea level in the Andes where the oxygen content of air is about 13%, to mice at sea level, which contains 21% oxygen.  We all know from experience that we need oxygen to survive and it’s harder to exercise at high altitudes, but why do our bodies actually need oxygen?

Oxygen in cellular respiration

Most all physiology can be explained by the following equation, which describes the process of cellular respiration:

Glucose + 6 Oxygen (O2) --> 6 Carbon dioxide (CO2) + 6 Water (H2O) + 34 ATP

Glucose is a simple carbohydrate (sugar) that we use as a direct energy source.  Glucose, which is 6 carbons long, gets broken down step by step in a series of chemical reactions.  At each step, a little bit of energy is released by the reaction and that is stored in carrier molecules.  These carriers then donate the energy in the form of electrons, which is then harnessed to make another molecule called ATP.  ATP is cellular energy.  The chemical bonds in ATP store high energy and can be used to drive other cellular reactions, like pumping ions, or the process that causes muscle contraction.  Without ATP we die. 

But what does oxygen have to do with this?  Well, the electrons that are donated by the carrier molecules must hop from protein to protein in what is known as the “electron transport chain”.  The final electron acceptor is oxygen (O2), which forms water with that extra electron.  That’s it.  That’s why we breathe, that’s why our heart pumps blood— our tissues need energy (ATP) to perform cellular tasks and in order to get energy from glucose, we need oxygen to accept the final electron.  Carbon dioxide is produced as a byproduct and is removed from the body during exhalation.

Glycolysis is anaerobic respiration and does not use oxygen. Note that oxygen is used as the last step of the electron transport chain to make the majority of the ATP. (www.phschool.com/science/biology_place/biocoach/cellresp)

I should also mention here that the more our tissues are active and working, the more ATP they need and the more O2 needs to get to the cells.  When we exercise our muscles are very active, so that’s why the heart rate and breathing rate increase; our body needs to intake more oxygen and distribute it faster to our muscles.

You can see that oxygen plays a critical role in our cells, so the mice at high altitudes are going to have a harder time getting their cellular energy.  How do they manage to run around when there is so little oxygen?

Energy sources

As I mentioned above, we can make ATP directly from glucose (a carbohydrate).  We can also make ATP by using fats as an energy source.  There are two differences between these two energy sources:

1) When we use fats as an energy source, it always requires oxygen.  Glucose, on the other hand, can make a limited amount of ATP without oxygen, which is called anaerobic respiration.  This is useful during short vigorous activity, but we cannot make enough ATP by anaerobic respiration for sustained exercise.

2) For a given amount of oxygen, more ATP is produced from carbohydrates, like glucose, than from fats.  However, the amount of ATP created from a single fat molecule is greater than from a glucose molecule.  In other words, if you have plenty of oxygen, you should be burning fats.  But once oxygen becomes limiting, either because you’re working so hard, or because you’re at a high altitude, then carbohydrates should be used.

Given this information, the authors hypothesized that the mice at high altitudes will burn more carbohydrates than mice at sea level.  They have a limited amount of oxygen in the air, so they need to use it in the most efficient way to produce the energy they need.

High altitude mice burn more carbohydrates, but fatigue sooner

The authors did all their tests in the same experimental conditions, with the same amount of oxygen in the air for both sets of mice.  Under normal oxygen conditions and when there was low oxygen content, the high altitude mice burned more carbohydrates than the other mice during moderate exercise.  At rest, they also burned more carbohydrates under low oxygen conditions.  The authors found that the activity of enzymes associated with breaking down carbohydrates were greater in the high altitude mice, specifically in the heart muscles.  The heart has to work harder at high altitude to get enough oxygen to the tissues, so it makes sense that these muscles, in particular, would be burning carbohydrates preferentially. 

The high altitude mice, therefore, have adapted to the low oxygen environment by having more active enzymes to break down carbohydrates rather than fats.  One problem with that, though, is that the carbohydrate storage is less extensive than fat storage.  The researchers found that high altitude mice fatigued more quickly than the sea level mice, which burned more fats.  They suggest that the fast fatigue is a result of using up all the carbohydrate stores.  These mice, though, don’t travel long distances and just need short bursts of speed to escape predators.

I really like how so many physiological processes can be explained through understanding cellular respiration.  It’s so logical that animals at high altitude need to use oxygen more efficiently, so they use carbohydrates more for energy.  It’s simple.

A biological reason for aging weight gain

There is a growing epidemic of obesity in the aging population.  Of course a lot of this has to do with our cultural lifestyle, but could there also be a biological explanation related to the way our bodies age?  One clue comes from the fact that older lab mice have a tendency to become obese, without the added influence of fast food restaurants.  Although there certainly are differences in metabolism as you age, the older mice also intake more food; it’s as if their body isn’t telling them the “I’m full” signal.  Yang et al examine the biological mechanism underlying this age-dependent obesity in the newest edition of Neuron. 

The Hypothalamus

There is a region towards the interior of the brain called the hypothalamus which controls all sorts of basic physiological parameters.  For instance, it sets the body temperature, monitors blood pressure and the water content of the blood, and initiates the feeling of thirst and hunger.  There are a group of neurons in the hypothalamus called POMC neurons, which release a hormone, called a-MSH, which decreases appetite (the feeling of “I’m done eating”).  Could it be that these neurons don’t function properly in older mice, so they aren’t getting enough a-MSH to signal them to stop eating?

Problems with the POMC neurons

The authors find that as the mice get older, their POMC neurons get more negative inside.  Remember that active neurons fire action potentials, which are basically short bursts of positive ions rushing into the cell.  If the POMC neurons are more negative than usual, they will have further to go to fire an action potential and will be less active.  The older POMC neurons are in fact much less active and therefore release less a-MSH.

What makes the older POMC neurons more negative?  The authors find that the neurons are overexpressing a potassium channel (K channel), which will mean there are more open pores in the membrane for K to escape the cell.  As the positive K ions leave the neuron, it will make the inside more negative.  Okay, but why are K channels overexpressed in older neurons?  Turns out this whole cascade is initiated by a key signaling protein called TOR.  Increased TOR levels have been associated with various aspects of aging before, and an inhibitor of TOR (called rapamycin) can increase the life span of mice and other animals.  Check out the diagram below, which puts all these steps together.

Summary: For whatever reason, POMC neurons overexpress TOR, which makes these neurons less active.  They release less of the a-MSH hormone, so the mice don’t get the “stop eating” signal and continue to intake food, leading to obesity.

Two complimentary experiments

To test that this pathway is actually correct, the authors did two complimentary experiments.

1) If TOR is artificially increased in young mice (to mimic older mice), will they intake more food and gain weight?

2) If TOR levels are decreased in older mice (to mimic younger mice), will they lose weight?

For experiment #1, they raised TOR levels in young mice by knocking out an upstream inhibitor of TOR.  TOR levels are normally controlled within a certain range by inhibitors, so if you get rid of that inhibition, there will be more TOR present.  Over many weeks, these mutant mice did in fact get fatter than the controls and their POMC neurons were too negative and didn’t function properly, just like older mice.

For experiment #2, they wanted to decrease TOR in older mice.  This is actually pretty easy to do by injecting the older mice with the drug rapamycin, which inhibits TOR (TOR actually stands for Target Of Rapamycin).  Rapamycin is made by bacteria, which were first discovered in soil samples from Easter Island.  It is currently approved for human use, as an immunosuppressant for organ transplant patients.  When the older mice were injected with rapamycin for a few weeks, the POMC neurons came back to life and fired many action potentials.  The cells weren’t so negative because there were less K channels being expressed.  And yes, rapamycin caused the older mice to eat less and lose a considerable amount of weight.

So there you have it: rapamycin is the wonder drug—it will make you live longer and healthier, but there will be a price to pay with a lowered immune system.  There may be other ways to tap into this dysfunction in the older POMC neurons to help prevent midlife obesity.