Category Archives: Molecular Biology

What can dogs tell us about cancer, and (wait for it) sexually transmitted disease

What can 546 dogs tell us about cancer, and STDs (sexually transmitted diseases)?  An enormous amount ! [ Science vol 365 pp. 440 – 441, 464 3aau9923 1 –> 7 ’19 ].  You may have heard about the transmissible tumor that has reduced the Tasmanian Devil population from its appearance in ’96 by 80%.  The animals bite each other transmitting the tumor.  Only 10 – 100 cells are transferred, but death occurs within a year.  The cells survive because Tasmanian devels have low genetic diversity.

The work concerns a much older transmissible tumor (Canine Transmissible Venereal Tumor — aka CTVT) which appeared in Asia an estimated 6,000 year ago, and began dispersing worldwide 2,000 years ago.   Unlike the Tasmanian devil tumor, the tumor is usually cleared by the immune system.

The Science paper has 80+ authors from all over the world, who sequenced the protein coding part of the dog genome (the exome) to a > 100fold depth.   The exome contains 43.6 megabases.   The tumor is transmitted by sex, and the authors note that this mode of transmission nearly requires a rather indolent clinical course, as the animal must survive long enough to transmit the organism again.  This fits with syphilis, AIDs, gonorrhea.  Contrast this with anthrax, cholera, plague which spread differently and kill much faster.

So what does CTVT tell us about cancer?   Quite a bit.  First some background.  The Cancer Genome Atlas (CGA) was criticized as being a boondoggle, but it at least gave us an idea of how many mutations are present in various cancers– around 100 in colon and breast cancers.

Viewed across all dogs, the CTVT genome is riddled with somatic mutations (as compared to the genome of the dog carrying the tumor) –148,030 single nucleotide variants (3.4/1000 !) 12,177 insertion/deletions.  Of the 20,000 dog genes only 2,000 didn’t contain a mutation.   This implies that most genes in the mammalian genome aren’t needed by the cancer cells.  The CTVTs also show no signs of the high rates of chromosomal instability seen in human tumors.

The work provides evidence that cancer isn’t inherently progressive.  This gives hope that some relatively indolent human cancers (say cancer of the prostate) can be controlled.  This calls for ‘adaptive therapy’  — something that limits tumor  growth rather than trying to kill every cancer cell with curative therapy which, if it fails, essentially selects for more aggressive cancer cells.

Some 14,412 genes have 1 mutation changing the amino acid sequence (nonSynonymous) and 5,704 have protein truncating mutations.  The ratio of synonymous to non synonymous mutations is about 3 implying that the mutations which have arisen haven’t been selected for (after all the triplet code for 20 amino acids and 1 stop codon has 64 possibilities), so the average amino acid has 3 codons for it.  This is called neutral genetic drift.

They also found 5 mutated genes present in all 541 tumors — these are the driver mutations, 3 are well known, MYC, PTEN, and retinoblastoma1.

Tons to think about here.  I’ll be away for a few weeks traveling and playing music, but this work should keep you busy thinking about its implications.




Schizophrenia research, the good news and the bad news

If you are an identical twin whose twin is schizophrenic, your chances of getting schizophrenia is 40%, if you are just a fraternal twin your chance is 15%, amazingly much higher than the 1% chance the rest of us  have, because that’s the incidence in the general population. For what schizophrenia is really like see the old post after the *** at the end

So to find out what causes schizophrenia, study the genes of schizophrenics and compare them to those without it.     [ Neuron vol. 103 pp. 203 – 216 ’19 ] First off —  the Psychiatric Genomics Consortium (PGC) has identified well over 100 (genomic loci) loci with a significant genome-wide association with risk for schizophrenia.  This means that unlike cystic fibrosis (where over 1,700 disease associated mutations have been found in the causative gene),and despite the genetics schizophrenia is not going to be due to one gene.

The good news is that serious money and attention to the genomes of schizophrenics is being paid.The paper reports the latest results from the horribly named BrainSeq.  This is a precompetitive initiative launched by the Liber Institute for Brain Development (LIBD) with heavy big pharma involvement (Eli Lilly, Johnson and Johnson, Hoffman-LaRoche, AstraZenica).    The LIBD has over 1,900 human  postmortem neuropsychiatric disease and control samples.   They are mapping all sorts of genetic information (DNA sequences, RNA sequencing, DNA epigenetics (cytosine methylation) etc. etc.)

The bad news is what this research is telling us.  The paper looked in differences in messenger RNA (mRNA) levels in two areas of the brain in 286 schizophrenics and 265 normal controls.  mRNA levels are a marker for gene expression, levels of the proteins coded for by the mRNA would be better, but is presently beyond our technology (when you are looking at the whole genome, as they were).

Well, out of our 20,000 or so protein coding genes, they found 48 differently expressed (by schizophrenics compared to normals) in one area (the hippocampus) and 245 in another (the dorsolateral prefrontal cortex).  That’s not a big deal, the two areas of the brain have rather different neurons and organization.

The bad news is there was almost no overlap between the 48 and the 245.  So although schizophrenics express their genome differently than normals, the expression varies in brain areas.  It would be great if there was some overlap, so then the genes differentiating schizophrenics from normal could be intensively studied.

The work also casts a shadow over a lot of earlier work, in which gene expression in schizophrenic brain was studied either in one area (or in ground up whole brain), and the results were assumed to be applicable to the brain as a whole.  They aren’t.  Back to the drawing board.


What is schizophrenia really like ?

The recent tragic death of John Nash and his wife warrants reposting the following written 11 October 2009

“I feel that writing to you there I am writing to the source of a ray of light from within a pit of semi-darkness. It is a strange place where you live, where administration is heaped upon administration, and all tremble with fear or abhorrence (in spite of pious phrases) at symptoms of actual non-local thinking. Up the river, slightly better, but still very strange in a certain area with which we are both familiar. And yet, to see this strangeness, the viewer must be strange.”

“I observed the local Romans show a considerable interest in getting into telephone booths and talking on the telephone and one of their favorite words was pronto. So it’s like ping-pong, pinging back again the bell pinged to me.”

Could you paraphrase this? Neither can I, and when, as a neurologist I had occasion to see schizophrenics, the only way to capture their speech was to transcribe it verbatim. It can’t be paraphrased, because it makes no sense, even though it’s reasonably gramatical.

What is a neurologist doing seeing schizophrenics? That’s for shrinks isn’t it? Sometimes in the early stages, the symptoms suggest something neurological. Epilepsy for example. One lady with funny spells was sent to me with her husband. Family history is important in just about all neurological disorders, particularly epilepsy. I asked if anyone in her family had epilepsy. She thought her nephew might have it. Her husband looked puzzled and asked her why. She said she thought so because they had the same birthday.

It’s time for a little history. The board which certifies neurologists, is called the American Board of Psychiatry and Neurology. This is not an accident as the two fields are joined at the hip. Freud himself started out as a neurologist, wrote papers on cerebral palsy, and studied with a great neurologist of the time, Charcot at la Salpetriere in Paris. 6 months of my 3 year residency were spent in Psychiatry, just as psychiatrists spend time learning neurology (and are tested on it when they take their Boards).

Once a month, a psychiatrist friend and I would go to lunch, discussing cases that were neither psychiatric nor neurologic but a mixture of both. We never lacked for new material.

Mental illness is scary as hell. Society deals with it the same way that kids deal with their fears, by romanticizing it, making it somehow more human and less horrible in the process. My kids were always talking about good monsters and bad monsters when they were little. Look at Sesame street. There are some fairly horrible looking characters on it which turn out actually to be pretty nice. Adults have books like “One flew over the Cuckoo’s nest” etc. etc.

The first quote above is from a letter John Nash wrote to Norbert Weiner in 1959. All this, and much much more, can be found in “A Beatiful Mind” by Sylvia Nasar. It is absolutely the best description of schizophrenia I’ve ever come across. No, I haven’t seen the movie, but there’s no way it can be more accurate than the book.

Unfortunately, the book is about a mathematician, which immediately turns off 95% of the populace. But that is exactly its strength. Nash became ill much later than most schizophrenics — around 30 when he had already done great work. So people saved what he wrote, and could describe what went on decades later. Even better, the mathematicians had no theoretical axe to grind (Freudian or otherwise). So there’s no ego, id, superego or penis envy in the book, just page after page of description from well over 100 people interviewed for the book, who just talked about what they saw. The description of Nash at his sickest covers 120 pages or so in the middle of the book. It’s extremely depressing reading, but you’ll never find a better description of what schizophrenia is actually like — e.g. (p. 242) She recalled that “he kept shifting from station to station. We thought he was just being pesky. But he thought that they were broadcasting messages to him. The things he did were mad, but we didn’t really know it.”

Because of his previous mathematical achievments, people saved what he wrote — the second quote above being from a letter written in 1971 and kept by the recipient for decades, the first quote from a letter written in 12 years before that.

There are a few heartening aspects of the book. His wife Alicia is a true saint, and stood by him and tried to help as best she could. The mathematicians also come off very well, in their attempts to shelter him and to get him treatment (they even took up a collection for this at one point).

I was also very pleased to see rather sympathetic portraits of the docs who took care of him. No 20/20 hindsight is to be found. They are described as doing the best for him that they could given the limited knowledge (and therapies) of the time. This is the way medicine has been and always will be practiced — we never really know enough about the diseases we’re treating, and the therapies are almost never optimal. We just try to do our best with what we know and what we have.

I actually ran into Nash shortly after the book came out. The Princeton University Store had a fabulous collection of math books back then — several hundred at least, most of them over $50, so it was a great place to browse, which I did whenever I was in the area. Afterwards, I stopped in a coffee shop in Nassau Square and there he was, carrying a large disheveled bunch of papers with what appeared to be scribbling on them. I couldn’t bring myself to speak to him. He had the eyes of a hunted animal.

Antioxidants — the dark side

There was (and probably still is) quite a vogue for antioxidants.  They were supposed to counteract aging, vascular disease, and prevent cancer.  So much so that 25 years ago, they were given in a trial to prevent lung cancer.  It didn’t work.  Here are the gory details

[ New England J. Med. vol. 330 pp. 1029 – 1035 ’94 ] The Alpha-Tocopherol, Beta-Carotene Trial (ATBC trial)  randomized double blind placebo controlled of daily supplementation with alpha-tocopherol (a form of vitamin E), beta carotene or both to see if it reduced the incidence of lung cancer was done in 29000 Finnish male smokers ages 50 – 69 (when most of the damage had been done).  They received either alpha tocopherol 50 mg/day, beta carotene 20 mg/day or both.   There was a high incidence of lung cancer (876/29000) during the 5 – 8 year period of followup.  Alpha tocopherol didn’t decrease the incidence of lung cancer, and there was a higher incidence among the men receiving beta carotene (by 18%).    Alpha tocopherol had no benefit on mortality (although there were more deaths from hemorrhagic stroke among the men receiving the supplement).   Total mortality was 8% higher among the participants on beta carotene (more deaths from lung cancer and ischemic heart disease).  It is unlikely that the dose was too low, since it was much higher than the estimated intake thought to be protective in the uncontrolled dietaryt studies.   The trial organizers were so baffled by the results that they even wondered whether the beta-carotene pills used in the study had become contaminated with some known carcinogen during the manufacturing process.  However, tests have ruled out that possibility.

Needless to say investigators in other beta carotene clinical trials (the Women’s Health Study, the Carotene and Retinoid Efficacy Trial) are upset.  [ Science vol. 264 pp. 501 – 502 ’94 ]  “In our heart of hearts, we don’t believe [ beta carotene is ] toxic”  says one researcher.

This is not science.

On to the present [ Cell vol. 178 pp. 265 – 267, 316 – 329, 330 – 345 ’19 ] in which the following appears “Recent evidence ‘suggests’ that antioxidants can also promote tumor formation”

The work concerns an animal model of nonsmallcell lung cancer (NSCLC).  I’m always wary of animal models, as they have been so useless in pointing to a useful therapy for stroke.  But the model is worth studying as it provides a mechanism by which antioxidants promote metastases of the primary tumor.  It is also worth studying because it shows the fiendish complexity of cellular biochemistry and physiology.

The only way you can appreciate complexity is by being buried in details. So let’s begin.  The actual details aren’t that important, just the number and the intricacy of them.

30% of humans with NSCLC have mutations in two genes (NFEL2L2, KEAP1).  The mutation in NFEL2L2 produces mutated NRF2 (a transcriptional activator of the antioxidant response gene set). The mutation doesn’t inactivate NRF2, but leaves it in a hyperactivated state.  KEAP1 normally inactivates NRF2, but not the mutated forms found in NSCLC.

One gene turned on by activated NRF2 is HO1 (heme oxidase).  During oxidative stress heme is released from heme containing resulting elevated intracellular heme lever resulting in the creation of free radicals which are inherently oxidative.  HO1 destroys heme. So this is one mechanisms of NRF2’s antioxidative activity.

Heme isn’t all bad, as it destabilizes BACH1 (not the composer)which is a prometastatic transcription factor.  Antioxidants (N-acetyl-cysteine, tocopherol [ vitamin E to you ] reduce heme levels stabilizing BACH1 (hence promoting metastasis).  Genes activated by BACH1 include glycolytic enzymes (hexokinase2, GAPDH).  So what?  Cancer cells use a lot of glycolytic enzymes even when oxygen is available — this is called aerobic glycolysis.  This is the Warburg effect.

I’m sure there’s far more to discover, but this should be enough to convince you that things are pretty complicated inside us.

The wages of inbreeding

Saguenay Lac St. Jean is a beautiful region of Quebec. It’s fairly isolated. Once you get to the top of the lake there is no way that you can drive farther north (no road).  We spent part of our 25th anniversary there.  The population bears a heavy load of genetic disease (through no fault of their own).

The reason is historical. Only 8,000 people emigrated from France to Quebec between 1608 and 1763. After the English victory that year  only 1,000 emigrated in the next 90 years.  In 1992, the population of the Saguenay  region was around 300,000 and Quebec itself 2,000,000.

This means that once the population began expanding with relatively little outside input, recessive genes began to meet each other, as in a large population there are so many more ways to make this happen than in a small one.

To keep the the nonBiologists reading this aboard, here is what recessive means. Our genome has 46 chromosomes.  We all have two sex chromosomes (either X and Y or X and X).  The other 44 chromosomes come in pairs.  This gives you two copies of each gene.  The classic recessive gene is that for sickle cell anemia.  If just one of the pair has the Sickle trait you are OK, if both have it, you have sickle cell anemia (which you definitely don’t want to have).  Actually if you live in Africa it is better if you have one gene with the trait as it makes you more resistant to Malaria.  This is why the trait became so common in Africans.  It’s natural selection in action (and in a human population to boot).  Just one good sickle gene (not carrying the trait) is enough to mask the effects of the bad gene, so the carrier is normal.   This is why sickle cell trait is called a recessive gene.

Here is one example.  The incidence of a muscle disease (myotonic dystrophy) worldwide is 2 – 14/100,000.  In the Saguenay region it is 189/100,000.

Even 20 years ago, the carrier frequency of many genetic disorders up there was quite high [ Proc. Natl. Acad. Sci. vol. 95 pp. 15140 – 15144 ’98 ]

Spastic ataxia 1/21

Type I tyrosinemia 1/22

Sensorimotor polyneuropathy 1/23

Pseudovitamin D deficient rickets 1/26

Cytochrome C oxidase deficiency 1/26

Cystinosis 1/39

Histidase 1/32

Lipoprotein lipase 1/43

Pyruvic kinase 1/64

Then again, there are all sorts of genetic diseases found only in this region.

Similar conditions may apply to the ancestors of today’s native Americans — for details see the previous post —  Incredible as it may sound, the rape and pillage of the conquistadores may have actually been good from a genetic point of view.  Similar considerations may apply to any pair of populations meeting each other for the first time.  Hard stuff indeed, but you can’t repeal biology.

So, from a genetic point of view, it’s good if you reproduce with someone from a different group.  It’s why I’m glad to have a Chinese daughter in law, 2 grand-nephews whose father is Hindu, and a Russian woman about to marry our nephew.



The initial native Americans were quite inbred

From Science vol. 365 pp. 138, eaat 5447 pp.  1 —> 9 ’19  12 July ‘19

“Genetic studies of contemporary Indigenous people and ancient individuals from Asia and the Americas reveal an outline of the ancestry of the first humans to settle the Americas, providing age estimates for the timing of population contact, divergence, and migration. Studies of contemporary mitochondrial DNA (mtDNA) and Y-chromosome DNA lineages gave the first genetic insights into Indigenous American population history (6). These studies demonstrated that the ancestors of all contemporary Indigenous people had descended from only five maternal lineages (haplogroups A, B, C, D, and X) and two paternal lineages (haplogroups C and Q). These lineages also showed that the founding population came from Asia and experienced a severe genetic bottleneck, in which a small number of people with limited genetic diversity gave rise to all Indigenous people who occupied the continent before European arrival.”

Interesting that the authors of the papers discussed below didn’t know this (or weren’t telling) when I wrote them last December asking if there was limited genetic diversity in the ancestors of today’s native Americans (or Indians as they called themselves when we lived in Montana in the 70s and 80s).


Usually when I eMail the author(s) of a paper or a math book with a question or a comment I get a quick response.  My cynical wife says thing this is because mathematicians don’t have much to do.  Not so in this case. Hence the hopefully attention getting title of this post.

I refer to the following papers [ Cell vol. 175 pp. 1173 – 1174, 1185 – 1197 ’18 ]  Nature vol. 563 pp. 303 – 304 ’18,Science vol. 362 pp. 1128 eaav2621  1 –> 11 ’18 ] I’ve sent a bunch letters to the authors and have heard nothing back in a week.

So what is all this about?  It’s about population bottlenecks and founder effects in the ancestors of what are now called ‘native Americans’ — although while living in Montana from ’72 – ’87, if you called an Indian, a Native American, you would have received some strange looks.

I am not a population geneticist, so I wonder just how many people made it over the Bering land bridge during the last ice age, and just how genetically diverse they were.  Northern Siberia today is a rather forbidding place, and I doubt that hordes of genetically different people lived here.  I’m not sure how long the land bridge was open and how many people crossed it.

So modern native Americans may be quite genetically homogeneous.  How to tell?  This is where the papers come in.  They sequenced genomes from a variety of locations in the western hemisphere, all dying over a thousand years ago (before the Europeans came and interbred with them).  It seems that they have around 100 such genomes.

I wrote to ask how similar these genomes are.  No response.  Is it because the answer might be politically incorrect?

I don’t think the question is idiotic.  Possibly we don’t have enough genomes to make a sensible statement, but if they’re all really close (however defined) we could say something.

Anybody out there have any thoughts (or even better)  knowledge about these matters?

How general anesthesia works

People have been theorizing how general anesthesia works since there has been general anesthesia.  The first useful one was diethyl ether (by definition what lipids dissolve in).  Since the brain has the one of the highest fat contents of any organ, the mechanism was obvious to all.  Anesthetics dissolve membranes.  Even the newer anesthetics look quite lipophilic — isoflurane CF3CHCL O CF2H screams (to the chemist) find me a lipid to swim in.  One can show effects of lipids on artificial membranes but the concentrations to do so are so high they would be lethal.

Attention shifted to the GABA[A] receptor, because anesthetics are effective in potentiating responses to GABA  — all the benzodiazepines (valium, librium) which bind to it are sedating.  Further evidence that a protein is involved, is that the optical isomers of enflurane vary in anesthetic potency (but not by very much — only 60%).  Lipids (except cholesterol) just aren’t optically active.  Interestingly, alfaxolone is a steroid and a general anesthetic as well.

Well GABA[A] is an ion channel, meaning that its amino acids form alpha helices which span the membrane (and create a channel for ion flow).  It would be devilishly hard to distinguish binding to the transmembrane part from binding to the membrane near it. [ Science vol. 322 pp. 876 – 880 2008 ] Studied 4 IV anesthetics (propofol, ketamine, etomidate, barbiturate) and 4 gasses (nitrous  oxide, isoflurance, devoflurane, desflurane) and their effects on 11 ion channels — unsurprisingly all sorts of effects were found — but which ones are the relevant.

All this sort of stuff could be irrelevant, if a new paper is actually correct [ Neuron vol. 102 pp. 1053 – 1065 ’19 ].  The following general anesthetics (isoflurane, propofol, ketamine and desmedtomidine) all activate cells in the hypothalamus (before this anesthetics were thought to work by ultimately inhibiting neurons).  They authors call these cells AANs (Anesthesia Activated Neurons).

They are found in the hypothalamus and contain ADH.  Time for some anatomy.  The pituitary gland is really two glands — the adenohypophysis which secretes things like ACTH, TSH, FSH, LH etc. etc, and the neurohypophysis which secretes oxytocin and vasopressin (ADH) directly into the blood (and also into the spinal fluid where it can reach other parts of the brain.  ADH release is actually from the axons of the hypothalamic neurons.  The AANs activated by the anesthetics release ADH.

Of course the workers didn’t stop there — they stimulated the neurons optogenetically and put the animals to sleep. Inhibition of these neurons shortened the duration of general anesthesia.

Fascinating (if true).  The next question is how such chemically disparate molecules can activate the AANs.  Is there a common receptor for them, and if so what is it?

Happy fiscal new year !

A sad (but brilliant) paper about autophagy

Over the past several decades I’ve accumulated a lot of notes on autophagy (> 125,000 characters).  It’s obviously important, but in a given cell or disease (cancer, neurodegeneration) whether it helps a cell die gracefully or is an executioner is far from clear.  Ditto for whether enhancing or inhibiting it in a given situation would be helpful (or hurtful).

A major reason for the lack of clarity despite all the work that’s been done can be found in the following excellent paper [ Cell vol. 177 pp.1682 – 1699 ’19 ].  Some 41 proteins are involved in autophagy in yeast and more in man.  Many are described as ATGnn (AuTophagy Gene nn).

Autophagy is a complicated business: forming a membrane, then engulfing various things, then forming a vacuole,  then fusing with the lysosome so that the engulfees are destroyed.

The problem with previous work is that if a protein was found to be important in autophagy, it was assumed to have that function and that function only.   The paper shows that core autophagy proteins are involved in (at least) 5 other processes (endocytosis, melanocyte formation, cytokinesis, LC3 assisted phagocytosis and translocation of vesicles from the Golgi to the endoplasmic reticulum).

Experiments deleting or  increasing a given ATGnn were assumed to produce their biological effects by affecting autophagy.

The names are unimportant.  Here is a diagram of 6 autophagy proteins forming a complex producing autophagy

1 2 3

4 5 6

So 2 binds to 1, 3 and 5

But in endocytosis

1 2 3


form an important complex

In cytokinesis the complex formed by

2 3


is important.

Well you get the idea.  Knocking out 2 has cellular effects on far more than autophagy.  So a lot of work has to be re-thought and probably repeated.

Notice that all 6 functions involve movement of membranes.  So just regard the 6 proteins as gears of different diameters which can form the guts of different machines as they combine with each other (and proteins specific to the other 5 processes mentioned) to move things around in the cell.

Set points, a mechanism for one at last.

Human biology is full of set points.  Despite our best efforts few can lose weight and keep it off.  Yet few count calories and try to eat so their weight is constant.  Average body temperature is pretty constant (despite daily fluctuations).  Neuroscientists are quite aware of synaptic homeostasis.

And yet until now, despite their obvious existence, all we could do is describe setpoints, not explain the mechanisms behind them.  Most ‘explanations’ of them were really descriptions.

Here is an example:

Endocrinology was pretty simple in med school back in the 60s. All the target endocrine glands (ovary, adrenal, thyroid, etc.) were controlled by the pituitary; a gland about the size of a marble sitting an inch or so directly behind the bridge of your nose. The pituitary released a variety of hormones into the blood (one or more for each target gland) telling the target glands to secrete, and secrete they did. That’s why the pituitary was called the master gland back then.  The master gland ruled.

Things became a bit more complicated when it was found that a small (4 grams or so out of 1500) part of the brain called the hypothalamus sitting just above the pituitary was really in control, telling the pituitary what and when to secrete. Subsequently it was found that the hormones secreted by the target glands (thyroid, ovary, etc.) were getting into the hypothalamus and altering its effects on the pituitary. Estrogen is one example. Any notion of simple control vanished into an ambiguous miasma of setpoints, influences and equilibria. Goodbye linearity and simple notions of causation.

As soon as feedback (or simultaneous influence) enters the picture it becomes like the three body problem in physics, where 3 objects influence each other’s motion at the same time by the gravitational force. As John Gribbin (former science writer at Nature and now prolific author) said in his book ‘Deep Simplicity’, “It’s important to appreciate, though, that the lack of solutions to the three-body problem is not caused by our human deficiencies as mathematicians; it is built into the laws of mathematics.” The physics problem is actually much easier than endocrinology, because we know the exact strength and form of the gravitational force.

A recent paper [ Neuron vol. 102 pp. 908 – 910, 1009 – 1024 ’19 ] is the first to describe a mechanism behind any setpoint and one of particular importance to the brain (and possibly to epilepsy as well).

The work was done at significant remove from the brain — hippocampal neurons grown in culture.  They synapse with each other, action potentials are fired and postsynaptic responses occur.  The firing rate is pretty constant.  Block a neurotransmitter receptor, and the firing rate increases to keep postsynaptic responses the same.  Increase the amount of neurotransmitter released by an action potential (neuronal firing) and the firing rate descreases.  This is what synaptic homeostasis is all about.  It’s back to baseline transmission across the synapse regardless of what we do, but we had no idea how this happened.

Well we still don’t but at least we know what controls the rate at which hippocampal neurons fire in culture (e.g. the setpoint).  It has to do with an enzyme (DHODH) and mitochondrial calcium levels.

DHODH stands for Di Hydro Orotate DeHydrogenase, an enzyme in mitochondria involved in electron transfer (and ultimately energy production).   Inhibit the enzyme (or decrease the amount of DHODH around) and the neurons fire less.  What is interesting about this, that all that is changed is the neuronal firing rate (e.g. the setpoint is changed).  However, there is no change in the intrinsic excitability of the neurons (to external electrical stimulation), the postsynaptic response to transmitter, the number of mitochondria, presynaptic ATP levels etc.

Even better, synaptic homeostasis is preserved.  Manipulations increasing or decreasing the firing rate are never permanent, so that changes back to the baseline rate occur.

Aside from its intrinsic intellectual interest, this work is potentially quite useful.  The firing rate of neurons in people with epilepsy is increased.  It is conceivable that drugs inhibiting DHODH would treat epilepsy.  Such drugs (teriflunomide) are available for the treatment of multiple sclerosis.

The paper has some speculation of how DHODH inhibition would lead to decreased neuronal firing (changes in mitochondrial calcium levels etc. etc) which I won’t go into here as it’s just speculation (but at least plausible spectulation).

Will flickering light treat Alzheimer’s disease ? — Take II

30 months ago, a fascinating paper appeared in which flickering light improved a mouse model of Alzheimer’s disease.  The authors (MIT mostly) have continued to extend their work.   Here is a copy of the post back then.  Their new work is summarized after the ****

Big pharma has spent zillions trying to rid the brain of senile plaques, to no avail. A recent paper shows that light flickering at 40 cycles/second (40 Hertz) can do it — this is not a misprint [ Nature vol. 540 pp. 207 – 208, 230 – 235 ’16 ]. As most know the main component of the senile plaque of Alzheimer’s disease is a fragment (called the aBeta peptide) of the amyloid precursor protein (APP).

The most interesting part of the paper showed that just an hour or so of light flickering at 40 Hertz temporarily reduced the amount of Abeta peptide in visual cortex of aged mice. Nothing invasive about that.

Should we try this in people? How harmful could it be? Unfortunately the visual cortex is relatively unaffected in Alzheimer’s disease — the disease starts deep inside the head in the medial temporal lobe, particularly the hippocampus — the link shows just how deep it is -

You might be able to do this through the squamous portion of the temporal bone which is just in front of and above the ear. It’s very thin, and ultrasound probes placed here can ‘see’ blood flowing in arteries in this region. Another way to do it might be a light source placed in the mouth.

The technical aspects of the paper are fascinating and will be described later.

First, what could go wrong?

The work shows that the flickering light activates the scavenger cells of the brain (microglia) and then eat the extracellular plaques. However that may not be a good thing as microglia could attack normal cells. In particular they are important in the remodeling of the dendritic tree (notably dendritic spines) that occurs during experience and learning.

Second, why wouldn’t it work? So much has been spent on trying to remove abeta, that serious doubt exists as to whether excessive extracellular Abeta causes Alzheimer’s and even if it does, would removing it be helpful.

Now for some fascinating detail on the paper (for the cognoscenti)

They used a mouse model of Alzheimer’s disease (the 5XFAD mouse). This poor creature has 3 different mutations associated with Alzheimer’s disease in the amyloid precursor protein (APP) — these are the Swedish (K670B), Florida (I716V) and London (V717I). If that wasn’t enough there are two Alzheimer associated mutations in one of the enzymes that processes the APP into Abeta (M146L, L286V) — using the single letter amino acid code – Then the whole mess is put under control of a promoter particularly active in mice (the Thy1 promoter). This results in high expression of the two mutant proteins.

So the poor mice get lots of senile plaques (particularly in the hippocampus) at an early age.

The first experiment was even more complicated, as a way was found to put channelrhodopsin into a set of hippocampal interneurons (this is optogenetics and hardly simple). Exposing the channel to light causes it to open the membrane to depolarize and the neuron to fire. Then fiberoptics were used to stimulate these neurons at 40 Hertz and the effects on the plaques were noted. Clearly a lot of work and the authors (and grad students) deserve our thanks.

Light at 8 Hertz did nothing to the plaques. I couldn’t find what other stimulation frequencies were used (assuming they were tried).

It would be wonderful if something so simple could help these people.

For other ideas about Alzheimer’s using physics rather than chemistry please see —


The new work appears in two papers.

First [ Cell vol. 1777 pp. 256 – 271 ’19 ] 7 days of auditory tone stimuli at 40 cycles/second (40 Hertz) for just one hour a day reduced amyloid in the auditory cortex of the same pathetic mice described above (the 5XFAD mice).  They call this GENUS (Gamma ENtrainment Using sensory Stimuli).  Neurologists love to name frequencies in the EEG, and the 40 Hertz is in the gamma range.

The second paper [ Neuron vol. 102 pp. 929 – 943 ’19 ] is even better.  Alzheimer’s disease is characterized by two types of pathology — neurofibrillary tangles inside the remaining neurons and the senile plaque outside them.  The tangles are made of the tau protein, the plaques mostly of fragments of the amyloid precursor protein (APP).  The 5XFAD mouse had 3 separate mutations in the APP and two more in the enzyme that chops it up.

The present work looked at the other half of Alzheimer’s the neurofibrillary tangle.  They had mice with the P301S mutation in the tau protein found in a hereditary form of dementia (not Alzheimer’s) and also with excessive levels of CK-p25 which also results in tangles.

Again chronic visual GENUS worked in this (completely different) model of neurodegeneration.

This is very exciting stuff, but I’d love to see a different group of researchers reproduce it.  Also billions have been spent and lost on promising treatments of Alzheimer’s (all based on animal work).

Probably someone is trying it out on themselves or their spouse.  A EE friend notes that engineers have been trying homebrew transcranial magnetic and current stimulation using themselves or someone close as guineapigs for years.

What is legionella trying to tell us?

10 years out of Med School, a classmate in the Public Health service had to deal with the first recognized outbreak of Legionnaire’s disease, at the Bellevue Stratford hotel in Philly, about one air mile from Penn Med where we went.   The organism wasn’t known at the time and caused 182 cases with 29 deaths.  We’ve learned a lot more about Legionella Pneumophila since 1976 and the organism continues to instruct us.

The most recent lesson concerns one of the 300 or so proteins Legionella injects into a cell it attacks.  This is remarkable in itself.  The organism uses them to hijack various cellular mechanisms to build a home for itself in the cell (the LCV — Legionella Containing Vacuole).  Contrast this with diphtheria which basically uses one protein (diphtheria toxin) to kill the cell.

One of the 300 proteins is called SidJ and looks like a protein kinase (of which our genome has over 500).  However [ Science vol. 364 pp. 787 – 792 ’19 ] shows that SidJ carries out a different different reaction.SidJ is activated by host-cell calmodulin to polyglutamylate the SidE family of ubiquitin (Ub) ligases inhibiting them. Crystal structures of the SidJ-calmodulin complex reveal a protein kinase fold that catalyzes ATP-dependent isopeptide bond formation between the amino group of free glutamate and the gamma carboxyl group in the catalytic center of SidE a ubiquitin ligase.   This, instead of just esterifying the hydroxyl group of serine or threonine or tyrosine with the terminal phosphate of ATP as a kinase is supposed to do.

Why is this important? The only protein known to have polyglutamic acid added to it is tubulin, the protein from which microtubules (neurotubules to the neurologist).  The work is important because some of the 500+ protein kinases in our genome might be doing something else.  Has the chemistry each and every member of the group been studied?  Probably not..

The authors close with “In summary, our results underscore the diversity and catalytic versatility of the protein kinase superfamily. We propose that ATP-dependent ligation reactions may be a common feature among the vast diversity of eukaryotic protein kinase–like enzymes found in nature (25). There are more than 500 protein kinases in humans and our results suggest that they should be ex- amined for alternative activities.”

I couldn’t agree more.