Wednesday, May 11, 2016

Uncoupling and weight loss

I've spent the last three posts making the point that fatty acid oxidation (supplemented by ketosis) increases the amount of ATP (and energy yield of ATP hydrolysis) available per unit oxygen consumed. This is particularly clear under the conditions of extended, intensely hypocaloric eating described by Phinney, where exercise can be sustained for longer, at a lower VO2, than on a mixed diet.

Now, oxygen consumption is a surrogate for caloric output. How many calories you "spend" per unit oxygen consumption is a complex calculation and depends on your fat to carb ratio.

But we don't run on calories. We run on ATP (mostly), or rather we run on the energy yielded from ATP hydrolysis.

To make that absolutely clear: We know, from Phinney, that under pure fat oxidation, we can generate enough ATP energy (physical treadmill load) to sustain moderate exercise by using less calories (ie lower VO2) on fat oxidation than on mixed diet oxidation. The increase in ability shows as a 25% drop in VO2, ie a 25% drop in calories needed to get enough ATP energy to move at 70% VO2 max.

That, to me, is pure survival adaptation. It's elegant, neat, cool etc.

It's not providing a metabolic advantage for weight loss.

Now, once again, I must wander off in to rodent studies.

If you take a rat or a mouse and feed it a genuine ketogenic diet you get some interesting effects. Let's look at this small study in rats. Here's heat output. Red is chow fed, grey is ketogenic:

Day or night, energy output is lower for the ketogenic rats compared to the chow fed rats. Phinney got a 25% drop in VO2 on his treadmill, the rats have calorie output down by an average of 11%, at a similar RQ. Running on fat (+/-ketones) requires less calories to generate adequate ATP levels.

Note, these are not real heat outputs in the rats. No one measured heat flux in any way. They're calculated from the VO2. They're done using the software built in to a CLAMS device around well accepted values of calories used per litre of oxygen consumed. This drop in calculated heat output, in itself, is not a surprise in view of Phinney's work.

What is surprising is that VO2 actually increased to generate this reduced heat output:

The rats should be using less oxygen per minute to produce their whole-body ATP energy requirement running on fat, according to Phinney. And me. And the chart. They're not, they're using more, in absolute terms.

The conclusion here is that the VO2 has gone the wrong way. So we have to ask: What is the difference between a fasting, exercising human on an RQ of 0.66 and a ketogenic rat slumming around its cage with a very similar RQ of 0.7?

The rats are uncoupled. They pump protons through complexes I, III and IV but a significant number of those protons drop straight back in to the mitochondria through open uncoupling proteins. Calories and oxygen are used (at the same RQ as any other more productive oxidation) but no ATP is produced from any protons which do not use ATP synthase. VO2 moves in two directions. It goes down (and so do calories used) due to switch from glucose oxidation to fat oxidation. It goes up due to uncoupling. The overall effect, up or down, on VO2 depends on the relative effects of RQ change, uncoupling, gluconeogenesis, NEAT and actual exercise.

Phinney's treadmill walkers had high FFAs and high ketones but absolutely no suggestion of any sort of uncoupling. Why?

To get any further we have to go to the Protons thread back in 2014.

Uncoupling proteins are kept closed by cytoplasmic ATP. And there is always enough cytoplasmic ATP in a functional cell to keep UCPs closed. There is one particular way (of several) to open them. The inhibition from cytoplasmic ATP can be overcome by an excess of mitochondrial ATP. Mitochondrial ATP, obviously, enters the UCP from the opposite end to cytoplasmic ATP and gets in the way of the latter's binding. Mitochondrial ATP cannot reach far enough into the UCP to induce the closed conformation itself, so the pore opens. The blog post has nice images and a more thorough description. Here's my fave picture:

How do we keep mitochondrial ATP levels low?

Phinney had six week starved humans on a treadmill showing every probability of low mitochondrial ATP and UCPs closed tighter than the proverbial monkey's @rsehole.

On the opposite front we have rats in a cage whose biggest effort is to move over to the hopper of ketogenic pellets and have a munch. These animals uncouple like mad while eating to satiety. They also either maintain low fat reserves or lose fat reserves if previously made obese from fat/sucrose feeding. We've all read this mouse study even if today's rat epic is very inaccessible (thanks Mike).

It seems to me that it is possible to maximise the efficiency of energy usage to ensure survival under near starvation conditions. However fat based your metabolism, you are not going to uncouple your oxidative metabolism unless you have adequate ATP within the mitochondrial matrix.

It's very clear that an ad libitum ketogenic diet allows uncoupling and metabolic inefficiency down to a lean bodyweight, certainly in rodents. This is not arguable. Here's the graph. No mouse was forcibly calorie restricted:

Days 1-4 after switch to ketosis they ate less, by day eight after the switch to ketogenic eating they were eating more calories (ns) than other groups but staying weight stable.

The question to me is: By how much do you have to deliberately restrict the calories of a ketogenic diet fed human to eliminate the uncoupling effect? Or, more simply, turn the question round: How do you get a human to lose weight most effectively on a ketogenic diet? This is easier to answer.

As Amber O'Hearn suggests:

Eat meat.
Not too little.
Mostly fat.

Perhaps someone should tell Dr Hall this. Better still, make it his epitaph as science progresses.



E-S said...

That experiment should be redone with the rats isolated inside a calibrated calorimeter.

Peter said...

That would be interesting...


Zachary said...

"Eat meat. Not too little. Mostly fat."

Michael Pollan just had a heart attack.

raphi said...

"UCPs closed tighter than the proverbial monkey's @rsehole" ==> This is what my dry molecular biology textbooks are missing.

When talking about Phinney's subjects with 'high ATP (efficiency)' needs due to 1) 6 weeks without food & 2) exercise, you say "It's not providing a metabolic advantage for weight loss". That makes sense.

The rats have 'low ATP (efficiency)' needs due to 1) an ad lib ketogenic diet & 2) being sedentary, so are they the corollary to Phinney's subjects, thus exhibiting Dr.Ludwig's (~300kcal) 'metabolic advantage'? (via UCP opening using up kcals as heat)

Just making sure Im not missing the main point.

raphi said...

I ask because you used the term "metabolic inefficiency" when talking about the rats.

Peter said...

raphi, I'm not sure the term "inefficiency" was a very good choice. That uncoupling is there for a reason. You can argue it's to avoid overloading the proton gradient or it could simply be doing the job of thermogenesis, thus making T3 unneeded... Pity we don't have T3 levels for the leto ratties.

Zachary, heehee so be it! Eat plants, bleughch. Though I do admit to eating some.


Peter said...

That should be lab ratties, not "leto"!

annlee said...

Not "Leto Atreides"? ;)

Peter said...

'Twould have been a neat typo, but no, nothing Dune about the ratties...


Kindke said...

Interesting post, im definitely against calorie restriction in all realms. Great insight again to highlight CR/ad lib and the cytoplasmic/mitochondrial ATP levels. I always learn from your posts.

Unfortunately a keto diet doesnt cause obese humans to drift down to their lean normal bodyweights due to excess fat cell number. Pre-adipocyte recruitment is the reason obesity is largely irreversible.

I mean just look at cases of "paradoxical adipose hyperplasia in cryolipolysis" in these cases the patients are not over eating, or eating too many carbs, with high insulin, whats happened is adipogenesis pathway is activated in the pre-adipcoytes, and then the lipogenesis genes in these cells are switched on, then fat accumulates.

Passthecream said...

Peter, what is the fate of the protons that fall down the uncoupling well? Water? Hydrogen? Are they the ultimately primitive mitochondrial anti-oxidants?

karl said...

Great post as always.

I read someplace that rodents are will do more ''Uncoupling'' than humans - was related to lifespan. I think birds wasted little.

- A couple of links - but not where I read about it:

I think it was Nick Lane:

So I would expect that the magnitude of this effect would be somewhat less for humans?

Could very well be that low-carbs causes weight loss by multiple effects.

( I've been speculating lately that low-carb diets tend to reduce LA consumption - a lot of LA in chips and bread products )

There was a diet pill - that worked - and also killed - (DNP or 2-4-Dinitrophenol) by the same method.

It is interesting that the archaic metabolic systems with their ability to provide homeostasis on a cellular level are still with us. The more I continue my quest to know the possible causes of the obesity/T2D pandemic the more I think that we need to understand the history of life to understand why it is as it really is - not easily understandable as we wish it were.

Bacteria multiply mostly asexual ( they can trade genes - even inter species (weird) but mostly asexual ). - while mammals are sexual.

Evolutionary this means that bacteria honed their metabolic tricks without the mix-master effect of sex on their genes. Sex introduces the risk of lethal single point mutations - and I think it explains why the high level systems are so complex - that nature selects for redundant overlapping control systems.

Such complex high level systems are not likely to be easily fitted to an easily understandable human meme.

Ash Simmonds said...


It looks like there's a lot of contradictions in this area.

"Uncoupled and surviving: individual mice with high metabolism have greater mitochondrial uncoupling and live longer" -

I wonder the difference between a mouse vs a bat, both have similer size and metabolism (ish), but mice become decrepit as they age and die in about 2 years, whereas bats remain negligbly senescent and retain their "youthfulness" until they nearly die - in 10-30 years.

Zachary said...

Peter, I have a random question. Thought i'd ask since you are a vet! Why does 90% of the internet say that a dog can not digest much fat? I'm even seeing people say that it causes pancreatic problems. This makes no sense to me, as an animal that is evolved from carnivorous ancestors. How on earth would a dog lack the enzymes sufficient to hydrolyze and digest fatty acids?

js290 said...

Thanks for pointing out that there are no metabolic processes that biochemically use "calorie" as a substrate.

Calorie counters fail to understand that there is a mathematical coupling between CI and CO. One affects the other, and in unintuitive ways when they ignore the hormonal effects. Decreasing CI may cause a subsequent and larger decrease in CO. Or, it may require an increase in CI to increase CO.

raphi said...


'Inefficiency' may not be quite apt but neither is metabolic 'advantage'. I never bought the Metabolic Advantage argument as i had always understood it to imply Wastefulness. Apparently, this is not the case. What Im gradually understanding it to mean is that the sum total of metabolic dials are tweaked such that adiposity is normalized without negative effects on lean body mass or other things important to living a good life. So somewhat paradoxically, the 300kcal not going towards ATP synthesis (or fat mass accumulation) is quite parsimonious.


You said "The more I continue my quest to know the possible causes of the obesity/T2D pandemic the more I think that we need to understand the history of life to understand why it is as it really is" ====> I couldnt agree more! Biological complexity is such that we must get as many hints as we can. This entails studying (amongst other things) our industrial, neolithic & paleolithic history...not only, we're helped by going all the way back to the beginning of life (Phanerozoic eon Nick Lane is a good time machine for that).
I hear a beautiful hum when there is consilience between findings from biochemistry & paleoanthropology :)

@Ash Simmonds,

The mouse vs bat comparison is a great example. A nice head scratcher...the first thing that comes to my mind is that evolution doesnt care how long you need to live to propagate your genes; this cuts both ways. If short is good enough for a species, than that's that. If long lives are needed or at least dont interfere, then great.

Tomasz said...

To make sure I get this right, what this all suggests is that if someone wants to lose fat on a ketogenic diet, restricting calories or increasing the energy output is not a good idea because this will block the uncoupling (vide Phinney's runners). Is that correct?

If so, does it imply that any study that attempts to measure the effectiveness of a LCHF diet, should not use calorie restriction, which would mean that the recent Hall's study was ill-constructed and in a way doomed to failure?

ItsTheWooo said...

Brilliant Peter, brilliant.

"On the opposite front we have rats in a cage whose biggest effort is to move over to the hopper of ketogenic pellets and have a munch. These animals uncouple like mad while eating to satiety. They also either maintain low fat reserves or lose fat reserves if previously made obese from fat/sucrose feeding."

Me IRL haha.

Anecdote; i was in a leptin study, a giant woo lab rat. They gave me a darth vader hat and measured my breath after an 8 hr fast. This was many years ago.
Results were i was consistently @ RQ of 0.7 that was true leptin naive and leptin treated.
However, my metabolic rate was quite low at 1040ish calories and this as well didnt change much with drug or naive state. Now i know for a FACT i was eating more than that, and i DEFINITELY inhale food now. Your blog entry seems to address these paradoxes (low caloric maintenance yet inability to gain much fat eating 2000 calories/day, with little activity to justify that)

I generate more ATP per calorie because I am on a fat based metabolism...
Similarly, due to uncoupling, should I feed to satiety (Or "excessively"), energy is inefficient and wasted.

Makes sense and explains absolutely all aspects of my physiological metabolic state.

Peter said...

Hi all, might not get much time to comment but just to E-S; respirometry is crap for assessing caloric output, certainly in Bl/6 mice at resting metabolic rate. Of course, it under estimates. Direct calorimeters are not available commercially, you have to build your own. i.e. you need need to be a meticulous engineer doing science. Nutrition "science" should be incapable of this level of engineering and, if they managed to bodge something together, it would undoubtedly be be adjustable to give the politically correct numbers.


Peter said...

Woo, you're such a marvellous lab rat!


JohnN said...

Agree that neither "inefficiency" nor "metabolic advantage" is an apt term. Uncoupling is what made us hot-blooded mammal and adaptable.

JohnN said...

Regarding adaptability: in the Vital Life Nick Lane expands on his thinking of long-lived bird and the power of flight. Migratory birds, not Dodo, have low reproductive rate due to high threshold for matched genes between nucleus and mt and low threshold for adaptability (that's why they have to migrate.)

Anonymous said...

I enjoyed the post, but wouldn't a simpler explanation of the increase in RMR seen by Hall be the cost of GNG + the cost of ketogenesis?

Making ketones doesn't come for free, and GNG is well known to be inefficient. It's also highest in the beginning of keto-adaptation, and it correlates with the protein catabolism seen by Hall.

Anonymous said...

Oops, couldn't delete comment after I realized the post was about the OTHER Hall study....

Peter said...

Hi normalcarb,

Yes, there are a number of other things which could have been said about either of the Hall studies. I've tried to stick to metabolic situation on a long term basis and why LC might win, as it normally does in calorie unlimited LC vs calorie restricted "normal" diets. That's really why we have to wait for Hall's funeral.


Peter said...

Kindke, yes, I’d agree that adipose hypertrophy and hyperplasia are different problems. Oddly enough, while putting mice on F366 produces the graph, putting them on rodent chow doesn’t, weight drops for 2 weeks then the climb in inezorable… BTW even Lutz mentioned that LC doesn’t work for all. Compliance is part but adipose hypertrophy might well be another big chunk.

Pass, They just dissipate the proton gradient. The lost energy shows up as heat. They’re antioxidant in so far as they reduce any back pressure from ATP synthase not being able to cope with a given proton gradient vs ATP need for the cell.

Ash and karl, Nick Lane was basing the difference of mitochondrial numbers needed for flight, I think. Excess mitocs mean they run at tick over much of the time and random ROS are kept to a minimum…

Zachary, very few dogs have normal digestion. Just measure a few B12’s, The number which are subnormal is remarkably high. My assumption: They all have non coeliac gluten intolerance with SIBO as a complication. There aren’t the villi or enzymes to cope with high fat. This is very real, Hills crapinacan ND (which works) had a very low acceptance rate when I was a clinician and used to use it. Diarrhoea ++++ in quite a lot of dogs. They’re pre-f*cked! Gradual introduction and complete cereal free helps. Bone meal binds some of them rather nicely too.

Tomasz, pretty much so. You still lose weight but more on a calorie counting basis. Al Sears did a similar study and medium carb beat LC once they increased the carbs on LC. It was run at 30% below expected calorie intake. One of the worst written diet studies I read. Information gaps all over the place.


Passthecream said...

Peter:"They just dissipate the proton gradient. "

Nick Lane suggests that uncoupled protons suck electrons from the e.t.c. (thereby acting as oxidizers wrt the electron carriers) so presumably by driving it harder would reduce any reverse transport and minimise free radical generation.
(Proton(s) plus electron(s) => hydrogen?) Is this futile cycling?

Peter said...

Pass, hadn't picked that up and the structure of UCPs doesn't suggest it but...

The REAL uncoupler is, of course, superoxide. Each electron which hops from an NADH directly to O2 is going to not-pump a whole ETC of protons!


Passthecream said...

That makes sense!

Either way i wouldn't have thought that protons would last long as charged entities inside the (pH basic?) negative interior of a mitochondrial 'battery'. Small distances => large attractive forces. The heat generated comes of course from electrons losing energy as they fall into some lower potentialed arrangements with protons.

... is potentialed a word??

Peter said...

Why not????

Passthecream said...

PsS a passing thought, the cristae in mt are reminiscent of the ribbed sections of high voltage insulators.

Passthecream said...
This comment has been removed by the author.
Rattus said...

@ Ash,

i remember reading that longevity in bats and molerats relative to other rodents was in part explainable by the high CO levels in their dwellings, which coincidentally has a mild mitochondrial uncoupling effect:

Molerats evolved to be immune to the pain caused by acids, because the CO builds up so much underground that it accumulates in their tissues.

cavenewt said...

@Rattus "i remember reading that longevity in bats and molerats relative to other rodents was in part explainable by the high CO levels in their dwellings,"

CO or CO2?

Peter said...

"Carbon monoxide and mitochondria—modulation of cell metabolism, redox response and cell death" is the link, it's CO. Where does this come from in the mole rat burrows? Apart from via hypoxia and carbon oxidation. I don't see how the TCA can generate CO though...