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Endocrinology

The lifecycle of epithelial cells, and a most wonderful coincidence (#30/31)

(This is a reworking of a post I originally wrote in 2007.)

So here's a wonderful example of modeling physiological processes across species: Schutte [1] details the reproductive cycle of the dog, Durrant adapts that information and applies it to the panda, and then Durrant's group and ours uses that panda information to study other species of bear--specifically, sun bears.

Anyway, this is a presentation slide that I adapted (animated, colored) from Schutte's 1967 original drawing [2,3,4] for a talk on using informatics to aid in the reproduction of endangered species at the 2004 American Veterinary Medical Association conference:
 



What you are seeing, as you read left to right, is the change over time in the shape and color of the vaginal epithelial cells, collected by swabbing the animal (like taking a Pap smear).

Shape changes (from more round to more irregular and from a high nucleus-to-cell size ratio to a much lower one, approaching or reaching zero in some cases), and color changes between pink/red (acidophilic) and blue (basophilic) can be seen as the cycle progresses. Although the hormone level peaks and the white and red blood cells (leukocytes and erythrocytes) are shown as well, we'll ignore those for the moment to concentrate on the changes in the colored cells in the middle of the diagram.

With a break in between, the cycle resumes at the left, and the whole process is repeated over and over again through the animal's reproductive life.

The important take-away points from this slide are the irregular borders and small nuclei of the aging cell, and the colors blue and pink (depicted above), and golden (which is also significant in pandas, but which Schutte did not address in dogs in his work).

So keeping those factors in mind, you can see why I was practically dumbstruck when I walked into a glass art studio in Fremont (a Seattle neighborhood) and noticed the piece by James Curtis titled "Little Bang":
 



I mean, it's all there! Large, irregular cell borders, tiny nuclei, and pink (cranberry), blue (teal), and golden color. Even the rack on which it is mounted looks like a graph over time of a cycle.

I swear, if I had commissioned the artist to render mature vaginal epithelial cells in glass, he could not have carried it out more faithfully.

A huge shout-out to James and his assistant Tara for patiently working with me while I arranged financing and donation of the work to our sun bear reproductive project for auction at a later date. If you're looking for glass art, I would recommend The Edge of Glass in Seattle unconditionally for their quality, their vision, and their customer service. On several subsequent occasions, they have donated pieces to support my research, and I am deeply grateful.

James told me he probably will not use the teal again, or if he does, it won't be very often--it is simply so hard to work with that it's not practical. So the bears and I did indeed get very lucky to get in there before someone else bought the piece, and we would never had known about it.

I also thank my husband, Iain, for deciding to surprise me with a drop-in visit there; it worked out so much more beautifully than I could ever have imagined.

References

  1. Durrant B, Czekala N, Olson M, Anderson A, Amodeo D, Campos-Morales R, Gual-Sill F, Ramos-Garza J. Papanicolaou staining of exfoliated vaginal epithelial cells facilitates the prediction of ovulation in the giant panda. Theriogenology. 2002 Apr 15;57(7):1855-64.
  2. Schutte AP. Canine vaginal cytology. I. Technique and cytological morphology. J Small Anim Pract. 1967 Jun;8(6):301-6.
  3. Schutte AP. Canine vaginal cytology. II. Cyclic changes. J Small Anim Pract. 1967 Jun;8(6):307-11.
  4. Schutte AP. Canine vaginal cytology. III. Compilation and evaluation of cellular indices. J Small Anim Pract. 1967 Jun;8(6):313-7.

 

Littoral interpretation (#18/31)

Water, water, every where,

And all the boards did shrink;

Water, water, every where,

Nor any drop to drink.

--Wikipedia, "The Rime of the Ancient Mariner", Samuel Taylor Coleridge, 1798 accessed 18 August 2012

 


The poem on the surface explores violation of nature and its resulting psychological effects on the Mariner, who interprets the fates of his crew to be a direct result of his having shot down an albatross.----Wikipedia, "The Rime of the Ancient Mariner: Interpretations", Samuel Taylor Coleridge, 1798 accessed 18 August 2012

 

Spoiler alert! (although there's a decent chance you already had to read this poem in elementary school, so in that case, you already know what happens to the Ancient Mariner and his crew).

The Ancient Mariner is a sailor who commits an unnecessary act of cruelty, even a crime in Coleridge's estimation--with his bow and arrow, he shoots the albatross (a water bird, like a seagull) who had led his lost ship out of dangerous waters.

With his cruel bow he laid full low
The harmless Albatross.

 

After the killing of the bird, more bad things continue to happen to the ship and crew. All the other crew members are eventually killed, but his punishment is to remain alive, tormented, and to wander the earth telling his story as a warning to others.

He partially redeems himself when--seeing the sea creatures that he had earlier despised, he recognizes how beautiful they really are. He wanders the earth eternally, trying to reach others with his lesson before it's too late for them to learn from it.

There is a metric boatload of things we could discuss about this poem, anywhere from conservation biology, history, psychology, and literature aspects, just to scratch the surface--but I want to talk about my trip to Padilla Bay estuary today, and what living in a littoral environment means for kidney function in animals--and Coleridge's observation about water is the perfect jumping-off point for that discussion.

 

 

 

 


An estuary is a body of water where fresh water from rivers and oceanic saltwater come together and mingle. So it's a complex transition zone, and organism functions that work one way in fresh water and another way in saltwater have to be covered in both cases there.

The littoral zone is the area of a body of water closest to the shoreline. So in an estuary, the littoral zone is where the fresh water and the saltwater mix--it's saltier than the fresh water is, but it's also less salty than the ocean water.

You probably know that, if you're ever lost on a boat on the ocean, you shouldn't drink seawater--not even a little bit, not even if you're very, very thirsty.

The reason is that your body tries to maintain a balance between the concentration of solutions (like dissolved salt) inside your cells and outside of them. Globetrooper has some good diagrams of what the process looks like, both when it's going right and when it's not.

Globetrooper's diagram shows what it looks like when things are balanced inside the cell and outside of it: a situation called "isotonic": the "same pressure" by water on the cell membrane from both sides.

Source: http://globetrooper.com/notes/wp-content/uploads/isotonic-state.png accessed 18 August 2012

 

The large gray circles represent salt ions (to be more specific, sodium ions and chloride ions), and the small blue circles represent water. The proportion of salt dissolved in the water--the concentration--inside the cell is about the same as the proportion of salt dissolved in the water outside the cells. The inside and the outside of the cell are in equilibrium (isotonic), and there is no pressure from the water either to leave or to enter the cell.

Globetrooper's next illustration shows a situation that is no longer isotonic. Salt ions (the gray circles) are dissolved in the water (the blue circles) inside the cell, but outside the cell, there are no salt ions--the water outside the cell is pure water, with no salt in it. This situation is called a hypotonic solution.

Source: http://globetrooper.com/notes/wp-content/uploads/too-much-fresh-water.png accessed 18 August 2012

 

Since the inside and the outside of the cell are no longer in balance, a process called osmosis occurs--the movement of fluid (in this case, water) across a semipermeable membrane (a membrane that substances can move through) from an area of lower concentration (in this case, of salt) to a area of higher concentration.

The effect is to bring the concentrations more into balance (isotonic).

The green arrow in this figure shows the movement of water (its osmotic pressure) from the area of low salt concentration outside of the cell to the area of higher salt concentration inside the cell. That movement of water into the cell dilutes the salt concentration, making it lower inside the cell.

This is what happens when we drink fresh water.

The opposite situation occurs when the water outside the cell is more salty (has a higher salt concentration: lots of gray circles, very few little blue circles) than the water inside the cell. This situation is called a hypertonic solution.

Source: http://globetrooper.com/notes/wp-content/uploads/too-much-salt-water.png accessed 18 August 2012

 

In a hypertonic solution, water flows out of the cells--the osmotic pressure of the water is toward the higher concentration of salt.

So you see what would happen? If you drank even a little salt water, it would draw water out of your cells. Instead of quenching your thirst, salt water would leave you more dehydrated than you were when you started.

At the cellular level, this is what it would look like:

Source: http://upload.wikimedia.org/wikipedia/commons/7/76/Osmotic_pressure_on_blood_cells_diagram.svg accessed 18 August 2012

 

In the hypertonic solution, the osmotic pressure comes from the water leaving the cells, leaving behind shriveled, badly dehydrated, cells (left side of image).

In the isotonic solution, the osmotic pressure balances out to zero, as equal amounts of water enter and leave the cells. The red blood cells in an isotonic state look normal and healthy with the indentation in the center that is typical of them.

In the hypotonic solution, the osmotic pressure pushes water into the cells, stuffing and waterlogging them--the normal indentation starts to disappear as the cell is too full of water.

These microphotographs of real red blood cells show how they really look, as they react to the different solutions we just described.

Source: http://upload.wikimedia.org/wikipedia/commons/6/62/Human_Erythrocytes_OsmoticPressure_PhaseContrast_Plain.svg accessed 18 August 2012

 

So now that we've discussed how cells behave in different kinds of solutions (hypertonic, isotonic, and hypotonic), does Coleridge's verse

And all the boards did shrink;

 

make even more sense now?

Why is that?

What is he describing, and why did the boards shrink as a result?

 

 


Osmoregulation (the control of osmotic pressure in body fluids, such as blood) is controlled in humans and other vertebrates by the kidneys--that's an important function they have in addition to production and excretion of urine.

Hormones produced by the pituitary gland and the adrenal glands, among others, signal to the kidneys what state the solutions in the blood are--hypertonic, isotonic, or hypotonic--and the kidney responds accordingly by reserving water or by releasing it to restore the balance.

As we are land animals, our kidneys have to respond to fluids we drink or take in in other ways (like an IV solution in the hospital, for example), but that's basically it. Fish, on the other hand, are surrounded by fluid, and their kidneys have to respond to that fluid and balance the water that their cells take in.

Too much water, and the cells swell up and get waterlogged--the hypotonic solution in the previous pictures.

Too little water, and the cells shrink and dehydrate and rupture--the hypertonic solution in the previous pictures.

The fish kidneys have to get the solution just right, and in a situation where the fish is surrounded by fluid of a different concentration.

Freshwater fish adapt to this situation by not drinking very much, and by urinating a lot; ocean fish (except sharks, which are a whole different story for another time) adapt by drinking a lot and not urinating very much.

So freshwater fish and ocean fish have adapted to this problem in pretty much opposite ways--that would seem to make a lot of sense.

But salmon live part of their lives in fresh water, and part of the time in salt water--so how can they have adapted to both, when the adaptations are the opposite of each other?

Salmon have adapted to both lifestyles--they can barely drink and urinate a lot when they live in fresh water, and then change to drinking a lot and barely urinating when they're out in the ocean.

Specialized cells in their body can work in opposite ways, depending on what they need at which stage they are in their lives.

But they can't turn it on a dime--they need days or weeks to make the transition between fresh water and ocean water.

And that's where the importance of the estuarine environment, like Padilla Bay, comes in--as an intermediate zone between the two other environments, it provides a place where salmon can make the transition.

In a region where the salmon can move around in the littoral zone to find the right amount of salt concentration they need, estuaries ensure the survival of those salmon leaving the fresh water where they were born, to go out and spend a large part of their lives in the ocean.

And they also ensure a place where--when it's time to go back up the freshwater river and breed--salmon have a place to adjust back from the ocean to the river environment, so that they can give birth to the next generation, and continue the cycle.

So often, in massage school, we don't have time to teach anatomy this deeply, and that's a real shame. If you just have time to memorize the fact that the kidneys control osmoregulation, so that you can recognize it when the MBLEX or the NCBTMB/NCBTM asks you about it, then that doesn't give you any particular preparation for clinical practice.

But if, at a deeper level, you understand what is going on, and you can draw a line from how the kidneys are involved in salt balance to what happens when that balance gets out of control one way or another, then you can understand what is going on with people living with renal failure or other kidney disease, and you are better equipped to know whether or not it's safe for you to provide massage under the circumstances.

Foundational concepts: Neurotransmitters and hormones

  1. What is the difference between a hormone and an neurotransmitter?
  2. Is noradrenaline (norepinephrine), a chemical compound produced by our bodies and involved in the "fight or flight" response, as well as in controlling Alzheimer's-related inflammation in the brain, a neurotransmitter or a hormone?

To figure out the answers, let's review what neurotransmitters are, and what hormones are.

Criterion Neurotransmitters Hormones
Compound is released from: pre-synaptic neuron endocrine gland
Compound is released into: synapse bloodstream
Release mechanism: electrical impulse secretion
Speed of action: fast slow
Target: post-synaptic neuron somatic cell
Distance from target: very close (distance across a synapse) any distance accessible via bloodstream

 

Let's check what norepinephrine does against that list to determine which it is.

When the (endocrine) adrenal glands release norepinephrine secretions into the bloodstream and it travels to the heart and lungs, causing those organs to work faster and getting more oxygen to the muscles to support either fight or flight, it is functioning as a stress hormone.

When norepinephrine-producing neurons in the locus coeruleus area of the brain release norepinephrine into synapses, from where they suppress inflammation in other nervous tissue (the glia)--an effect which appears to protect against Alzheimer's disease--it is functioning as a neurotransmitter.

So there are two important points to remember:

  1. There is not any chemical difference between a hormone and a neurotransmitter: the difference lies in where and how they are released, how fast and how far they have to travel to their target cells, and what their targets are.
  2. Examples such as norepinephrine reinforce that there is no chemical difference between the two--depending on all the factors listed previously, norepinephrine can be either a neurotransmitter or a stress hormone.
    Not all chemical compounds can be both--some compounds are only hormones, some compounds are only neurotransmitters, and some can be both, depending on the situation. But there is nothing chemical about the difference--only the factors listed previously tell you whether the chemical compound you are examining functions as a hormone or as a neurotransmitter.

Source: http://upload.wikimedia.org/wikipedia/commons/e/e8/Epinephrine_structure_with_descriptor.svg

Foundational concepts: Neurotransmitters and hormones

  1. What is the difference between a hormone and an neurotransmitter?
  2. Is noradrenaline (norepinephrine), a chemical compound produced by our bodies and involved in the "fight or flight" response, as well as in controlling Alzheimer's-related inflammation in the brain, a neurotransmitter or a hormone?

Click here to review the relationship between neurotransmitters and hormones.

Review of massage effects in Experience L!fe Magazine

Catherine Guthrie has written a review of massage research aimed at a non-specialist audience, examining its effects on anxiety, low back pain, tension headaches, sleep, depression, and blood pressure.

Click here to read Experience L!fe Magazine's review of research on massage.

P.S. If you've been following the first discussion in Journal Club, you'll know why this particular statement is backwards:

Although Moyer is yet to be convinced of the cortisol connection, both he and Field agree that massage is potentially very therapeutic for what’s known as “state” anxiety.

 

Even so, the article's definitely worth a read.

Free access to Sapolsky's Stanford lectures on biology and behavior

"Who thinks there a magnificent fascinating nuanced interaction between nature and nurture?"

Click here to check out Robert Sapolsky's Stanford lectures on biology and behavior. Although the links to the following lectures below the first video are in Chinese, Chinese uses the same numbers as English, so you can find lectures 2, 3, and so forth, to follow the lecture series for as long as you interested in doing so.


Update from Diane Jacobs:

Here is a link to all the lectures in that series, in English. Great course. http://www.youtube.com/playlist?p=848F2368C90DDC3D .

 

Cheers, Diane!

 

 

Does massage therapy reduce cortisol? Moyer article transcription, part 1

0:01 Greetings, this is Christopher Moyer from University of Wisconsin-Stout. I want to thank you for your interest in my work that was presented at the May 2010 Research in MT and CIM conference.
 
And without any further delay, I'm going to talk about the presentation entitled Cortisol Reductions in Response to Massage Therapy: A Comprehensive Quantitative Review.
 
 
 
0:30 As the audience at this conference and you will surely know, massage therapy is a large and growing industry, many people are using massage therapy for a variety of reasons, including in the expectation that it will benefit their health.
 
 
 
0:48 And as you will also undoubtedly know, massage has been shown to be consistently better than control treatments for certain things, including reducing heart rate and blood pressure, reducing anxiety and depression, and also reducing some specific forms of pain.
 
 
1:07 Now there are several theories for how MT might cause these effects, and it is of course an open question also whether it causes those effects through one or through several different mechanisms, but note here that Field and her colleagues have been unambiguous in their support for the theory that massage therapy first reduces the stress hormone cortisol and that this reduction is a underlying causal mechanism for these other effects, reductions of heart rate, blood pressure, anxiety, depression, and pain, and this is shown here in some quotes from their studies across a period of about 9 years.
 
 
1:58 Now, we must point out that the results that I and my colleagues have gotten have not been consistent with this theory.
 
We have in two quantitative reviews included cortisol as an outcome that we've assessed, and in both cases, whether we were looking at adult recipients or pediatric recipients, we have not found a statistically-significant effect for cortisol, even though we have found statistically-significant effects for those other clinical outcomes: depression, anxiety, pain, heart rate, and blood pressure. This leads us to believe that cortisol just cannot be a causal mechanism.
 
Now why are Field and colleagues getting different results than we are getting, even though we are largely looking at the same set of studies? Well, 
 
 
2:50 It really matters a lot how you analyze the data.
 
Many people who are not researchers would assume that the way to measure the effect of MT on something like cortisol is to take a before-treatment measure which we can picture as a bell curve such as this. We can expect that we'll get a normal distribution if we sample a bunch of people's cortisol levels before massage therapy. And 
 
 
3:15 if we then measure their cortisol levels again after treatment, whether that's a single session or multiple sessions, depending on the research design. this before-and-after change, logically, to most people would be massage therapy's effect on cortisol. 
 
But researchers know that it's not as simple as this--that if we take a before-and-after measure such as this, we don't know that the treatment that we administered in between those measurements is the cause of any difference we see.
 
It could be due to treatment, but it also could be due simply to the passage of time that is, the same change might have happened even if treatment hadn't been done.
 
It could be due to a placebo effect, that is, the person's expectation that the treatment would work, but not directly the effect of treatment itself. 
 
It could be due to spontaneous healing processes, or it could even be due to a statistical phenomenon known as statistical regression, which is a little too complicated for me to go into here, but if that's a new concept to you, the Wikipedia file on that concept is actually quite good, so an Internet search can probably clarify "regression to the mean", what that means.
 
Ok, so what do we do about this? 
 
 
4:32 Well, we do in fact start with before-treatment measurements, but what we must do is assign some people to get massage and some people to not get massage.
 
 
4:44 So that when we're done with our research, we have not one end-point measurement, but two--at least two,
 
That is we have some people who have gotten massage, and some people who have not gotten massage, but have undergone the same passage of time, and hopefully the same amount of attention, and if possible have been convinced that maybe they're receiving a treatment, even if they're not, to control for placebo effect.
 
Now, what we want to look at to best measure the effect of treatment itself, is the difference in those two endpoints, pictured here as a purple distribution and a green distribution.
 
If MT does better than a control treatment, then we've got much better confidence that the difference we see is the result of treatment, and not of one of those confounds, time, placebo effect, statistical regression, and so on.
 
So this is at the root of why Field and her colleagues and we are getting different results. 
 
 
5:48 Actually, one more slide, so we've got our between-groups effect picture here.
 
 
5:52 On to the next slide, which reads "Two approaches to measuring the effect of MT on cortisol". 
 
Touch Research Institute has emphasized very consistently within-groups effects in their studies, and this is despite the fact that they typically conduct between-group studies with control groups, but then they inexplicably ignore the control group data.
 
We have not done this, we, when we look at these effects, always look at the between-groups analysis, so that we can control for the confounds that the control group is intended to control for.
 
And this is universally recognized by methodologists as the optimal way to proceed in clinical research. 
 
So, this is the approach that we emphasize in the quantitative review that we are about to show you. All our analyses are between-group effects on randomized controlled trials.
 
 
End of Part 1 
 

Can contradicting what we've always been taught actually be a win-win proposition?

In addition to the specific discussions around particular research pieces, I'd like to discuss some issues that apply to research in general. Over at the Journal Club, we're discussing Christopher Moyer's work on reviewing studies about the effect of massage on cortisol.

 

 

If you cue up the YouTube video to start at 47 seconds into the talk and let it run from there, you'll see the presentation slide pictured above. At 0:50, you'll hear Moyer say:

0:50 And as you will also undoubtedly know, massage has been shown to be consistently better than control treatments for certain things, including reducing heart rate and blood pressure, reducing anxiety and depression, and also reducing some specific forms of pain.
 
 
These claims are also pictured on the slide. If you look at the sources for these claims on the slide, you'll see that he's referencing two articles written about his own work:
  1. Moyer CA, Rounds J, Hannum JW. A meta-analysis of massage therapy research. Psychological Bulletin. 2004 Jan;130(1):3-18. PMID: 14717648
  2. Beider S, Moyer CA. Randomized controlled trials of pediatric massage: a review. Evidence Based Complementary and Alternative Medicine. 2007 Mar;4(1):23-34. PMID: 17342238
 
I'd like to put out the following questions for discussion in the comments:
  1. When Moyer says, based on his own research, that massage is consistently better than control treatments for:
  • reducing heart rate

  • reducing blood pressure

  • reducing anxiety

  • reducing depression

  • reducing some specific forms of pain

is that a positive endorsement of massage? Why or why not?

  1. When Moyer says, based on his own research, that massage does not significantly lower cortisol, and that cortisol cannot therefore be a cause for these results, is that an attack on massage? Why or why not?
  2. When Moyer says, based on his own research, that Field used an incorrect analysis technique to conclude that massage reduced cortisol, is that an attack on Field and her team? Why or why not?
  3. Is accepting a new explanation to replace what we have traditionally been taught an attack on our teachers? Why or why not?
  4. Is there a way that contradicting something that we've always been taught can actually be a win-win proposition? If so, how could that work?

 

 

 
 
 
 

Can contradicting what we've always been taught actually be a win-win proposition?

As the title of this post indicates, let's explore what it means to examine and change ideas about massage that have been widely taught and accepted. Click here to join the discussion.

 

 

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