Category Archives: Medicine in general

Could Alzheimer’s disease be a problem in physics rather than chemistry?

Two seemingly unrelated recent papers could turn our attention away from chemistry and toward physics as the basic problem in Alzheimer’s disease. God knows we could use better therapy for Alzheimer’s disease than we have now. Any new way of looking at Alzheimer’s, no matter how bizarre,should be welcome. The approaches via the aBeta peptide, and the enzymes producing it just haven’t worked, and they’ve really been tried — hard.

The first paper [ Proc. Natl. Acad. Sci. vol. 111 pp. 16124 – 16129 ’14 ] made surfaces with arbitrary degrees of roughness, using the microfabrication technology for making computer chips. We’re talking roughness that’s almost smooth — bumps ranging from 320 Angstroms to 800. Surfaces could be made quite regular (as in a diffraction grating) or irregular. Scanning electron microscopic pictures were given of the various degrees of roughness.

Then they plated cultured primitive neuronal cells (PC12 cells) on surfaces of varying degrees of roughness. The optimal roughness for PC12 to act more like neurons was an Rq of 320 Angstroms.. Interestingly, this degree of roughness is identical to that found on healthy astrocytes (assuming that culturing them or getting them out of the brain doesn’t radically change them). Hippocampal neurons in contact with astrocytes of this degree of roughness also began extending neurites. It’s important to note that the roughness was made with something neurons and astrocytes never see — silica colloids of varying sizes and shapes.

Now is when it gets interesting. The plaques of Alzheimer’s disease have surface roughness of around 800 Angstroms. Roughness of the artificial surface of this degree was toxic to hippocampal neurons (lower degrees of roughness were not). Normal brain has a roughness with a median at 340 Angstroms.

So in some way neurons and astrocytes can sense the amount of roughness in surfaces they are in contact with. How do they do this — chemically it comes down to Piezo1 ion channels, a story in themselves [ Science vol. 330 pp. 55 – 60 ’10 ] These are membrane proteins with between 24 and 36 transmembrane segments. Then they form tetramers with a huge molecular mass (1.2 megaDaltons) and 120 or more transmembrane segments. They are huge (2,100 – 4,700 amino acids). They can sense mechanical stress, and are used by endothelial cells to sense how fast blood is flowing (or not flowing) past them. Expression of these genes in mechanically insensitive cells makes them sensitive to mechanical stimuli.

The paper is somewhat ambiguous on whether expressing piezo1 is a function of neuronal health or sickness. The last paragraph appears to have it both ways.

So as we leave paper #1, we note that that neurons can sense the physical characteristics of their environment, even when it’s something as un-natural as a silica colloid. Inhibiting Piezo1 activity by a spider venom toxin (GsMTx4) destroys this ability. The right degree of roughness is healthy for neurons, the wrong degree kills them. Clearly the work should be repeated with other colloids of a different chemical composition.

The next paper [ Science vol. 342 pp. 301, 316 – 317, 373 – 377 ’13 ] Talks about the plumbing system of the brain, which is far more active than I’d ever imaged. The glymphatic system is a network of microscopic fluid filled channels. Cerebrospinal fluid (CSF) bathes the brain. It flows into the substance of the brain (the parenchyma) along arteries, and the fluid between the cellular elements (interstitial fluid) it exchanges with flows out of the brain along the draining veins.

This work was able to measure the amount of flow through the lymphatics by injected tracer into the CSF and/or the brain parenchyma. The important point about this is that during sleep these channels expand by 60%, and beta amyloid is cleared twice as quickly. Arousal of a sleeping mouse decreases the influx of tracer by 95%. So this amazing paper finally comes up with an explanation of why we spend 1/3 of our lives asleep — to flush toxins from the brain.

If you wish to read (a lot) more about this system — see an older post from when this paper first came out —

So what is the implication of these two papers for Alzheimer’s disease?


The surface roughness of the plaques (800 Angstroms roughness) may physically hurt neurons. The plaques are much larger or Alzheimer would never have seen them with the light microscopy at his disposal.


The size of the plaques themselves may gum up the brain’s plumbing system.

The tracer work should certainly be repeated with mouse models of Alzheimer’s, far removed from human pathology though they may be.

I find this extremely appealing because it gives us a new way of thinking about this terrible disorder. In addition it might explain why cognitive decline almost invariably accompanies aging, and why Alzheimer’s disease is a disorder of the elderly.

Next, assume this is true? What would be the therapy? Getting rid of the senile plaques in and of itself might be therapeutic. It is nearly impossible for me to imagine a way that this could be done without harming the surrounding brain.

Before we all get too excited it should be noted that the correlation between senile plaque burden and cognitive function is far from perfect. Some people have a lot of plaque (there are ways to detect them antemortem) and normal cognitive function. The work also leaves out the second pathologic change seen in Alzheimer’s disease, the neurofibrillary tangle which is intracellular, not extracellular. I suppose if it caused the parts of the cell containing them to swell, it too could gum up the plumbing.

As far as I can tell, putting the two papers together conceptually might even be original. Prasad Shastri, the author of the first paper, was very helpful discussing some points about his paper by Email, but had not heard of the second and is looking at it this weekend.


For some readers, this might be the most useful post I’ve ever written. But first; some history. Back in grad school, I was dating a Cliffie. We were out to dinner at a nice (and cheap) restaurant in Cambridge. I’d had the flu and probably should have canceled, but in your early 20s, libido conquers all. So we’re sitting there, and I began to feel really nauseous and said we should pack it in, and I should go home.

She said “Let me try this, my father’s a General Practitioner”. So she ordered a can of coke, opened it and let it sit for a while till it warmed up and the fizz was gone. Then she told me to drink it in slow, small sips. It worked ! The nausea vanished and we continued on.

Fast forward to last night and probable food poisoning (or severe flu). No Coke in the house, but as soon as my wife got to a store opening at 7 this AM, it worked again — no nausea and stomach distress within a few minutes (I’d vomited at least 5 times over the course of the night).

Could this have been a placebo effect, because it had worked in the past and I wanted it to work so desperately? Possibly, but I was generally miserable for a period for a period of 10 hours, and the Coke settled my stomach very quickly. Coke is not an anti-diarrheal, but 10 hours into the illness there was nothing left.

Placebos and Nocebos are very complicated entities and a huge review in Neuron will tell you why. It’s very much worth reading – Neuron vol 84 pp. 623 – 637 ’14 — “Placebo Effects: From the Neurobiological Paradigm to Translational Implications”. The article contains references to studies showing that placebo is as effective as morphine on the third day post-op. In med school I’d heard stories to the effect that in Korea and WWII when they ran out of morphine on the battlefield, saline worked just as well. So probably these aren’t myths. It didn’t happen in Vietnam when I was in the service, as the country is long and thin, and no wounded soldier was more than 20 minutes away by chopper from a fully equipped field hospital (once they got him).

The ingredients in Coke are and were a closely held secret, but most think in the 1880s and 1890s, when it was sold as a medication, that Coke contained cocaine, hence the name. Back then, no one knew the potential of cocaine for addiction. Halstead the great Baltimore surgeon, got into it because cocaine is also a local anesthetic. Freud actually used cocaine to treat morphine addiction. Neither was malevolent, just ignorant.

No longer looking under the lamppost

Time flies. It’s been over 5 years since I wrote, essentially a long complaint that biochemists (and by implication drug chemistry and drug discovery) were looking at the molecules they knew and loved rather than searching for hidden players in the biochemistry and physiology of the cell.

Things are much better now. Here are 3 discoveries from the recent past, some of which should lead to drugable targets.

#1 FAFHA — a possible new way to treat Diabetes. Interested? Take a long chain saturated fatty acid such as stearic acid (C18:0). Now put a hydroxyl group somewhere on the chain (the body has found ways put them at different sites — this gives you a hydroxy fatty acid (HFA). Next esterify this hydroxyl group with another fatty acid and you have a Fatty Acid ester of a Hydroxy Fatty acid (an FAHFA if you will). So what?

Well fat makes them and releases them into the blood, making them yet another adipokine and further cementing fat as an endocrine organ. Once released FAHFAs stimulate insulin release, and increase glucose uptake in the fat cell when they activate GPR120 (the long chain fatty acid receptor).

A variety of fatty acids can form the ester, one of which is palmitic acid (C16:0) forming Palmitic Hydroxy Stearic Acid (PAHSA) which binds to GPR120. if that weren’t enough PAHSAs are anti-inflammatory — interested read more [ Cell vol. 159 pp. 238 239, 318 – 332 ’14 ]. I don’t think the enzymes forming HFA’s are known, and I’m willing to bet that are other HFAs out there.

#2 Maresin1 (7S, 14S dihydroxy docosa 4Z 8E 10E, 12Z 16Z, 19Z hexaenoic acid to you) is the way you start making Specialized Proresolving Mediators (SPMs). Form an epoxide of one of the double bonds and then do an SN2 ring opening with a thiol (glutathione for one) forming what they call a sulfido-conjugate mediator. It appears to be one of the many ways that inflammation is resolved. It helps resolve E. Coli infection in mice at nanoMolar concentration. SPMs further neutrophil recruitment and promote macrophage clearance of apoptotic cells and tissue debris. Wouldn’t you like to make a drug like that? Think of the specificity of the enzyme producing the epoxidation of just one of the 6 double bonds. Also a drug target. For details please see PNAS vol. 111 pp. E4753 – E4761 ’14

#3 Up4A (Uridine Adenosine Tetraphosphate) — as you might expect it’s an agonist at some purinergic receptors (PO2X1, P2Y2, P2Y4) causing vasoconstriction, and vasodilatation at others (P2Y1). It is released into the colon when enteric neurons are stimulated. Another player whose existence we had no idea about. Certainly we have all the GI and vasodilating drugs we need. If nothing else it will be a pharmacological tool. Again the enzyme making it isn’t known — yet another drug target possibly. For details see PNAS vol. 111 pp. 15821 – 15826 ’14.

There is a lot more in these 3 papers than can be summarized here.

Who knows what else is out there, and what it is doing? Glad to see people are starting to look

The way it ought to be

A recent paper described the use of sulforaphane in treating autism [ Proc. Natl. Acad. Sci. vol. 111 pp. 15550 – 15555 ’14 ] A sulforaphane trial (double blind, randomized) on 44 men age 13 – 27 with moderate to severe Autism Spectrum Disorder received sulforaphane (50 – 150 micoMoles) for 18 weeks followed by 4 weeks without treatment. There was no change in the 15 placebo patients, while there was a 33% decline in the Aberrant Behavior Checklist scores. When the sulforaphane was stopped total scores rose toward pretreatment levels.

I had posted on sulforaphane before — see the end of this post. I wrote the lead author asking if some of the therapeutic effects could be due to the anti-androgen activity of sulforaphane. He wrote back in a few days.

“Sorry, I missed your email. Absolutely possible. We did not measure androgen levels, but will do so in the future.
Thank you so much.”

Contrast this to the absent responses on whether the subjects in two functional MRI studies of the default network were asleep. See

Vegetarians are wimps: Science now tells us why

Oh, it started innocently enough. Population studies had shown that men who ate lots of cruciferous vegetables (collard greens, cabbage, brussels sprouts, broccoli, cauliflower, bok choy etc. etc.) had less prostate cancer. Some folks in Oregon decided to find out why [ Proc. Natl. Acad. Sci. vol. 106 pp. 16663 – 16668 ’09 ]. One of the compounds found in all these veggies is sulforaphane. There are all sorts of places to be found on the web that will sell it to you for your health. Sulforaphane is said to fight cancer, improve diabetes and kill bacteria (if you believe Wikipedia). Hosanna.

Prostate cancer is made worse by male hormones (androgens). They produce their effects in cells by binding to a protein (the androgen receptor) which then goes into the nucleus of the cell and turns on the genes which make males male. If there’s no androgen around the receptor just sits there outside the nucleus (e.g. in the cytoplasm), doing nothing. Some forms of prostate cancer have mutations in the receptor which turn it on whether androgen is present or not. This makes the cancer even worse. So one of the mainstays of prostate cancer therapy is lowering androgen levels by a variety of means, none of them pleasant — such as castration and various pills.

The Oregon work shows that sulforaphane decreases the amount of androgen receptor around resulting in less androgenic effects, and presumably less prostate cancer in the long run. How this is thought to occur is pretty interesting, highly technical and is to be found in subsequent paragraphs. It also explains why vegetarians are such wimps.

The androgen receptor sits in the cytoplasm bound to a protein called HSP90 (heat shock protein of 90 kiloDaltons). This protects the androgen receptor from being destroyed. Sulforaphane is a fairly simple molecule — a straight 4 carbon chain with a methyl sulfoxide group at one end and an isothiocyanate (-N=C=S ) group at the other. It should be pretty lipid soluble, meaning it can go everywhere in the body without much trouble. The authors showed that sulforaphane inhibits an enzyme called histone deacetylase 2 (HDAC2). This results in more acetylation of HSP90 on lysine, inhibiting the association of HSP90 with the androgen receptor, leading to increased destruction of the receptor and less androgenic effects in the cell.

The active site of one histone deacetylase that we know about is a tubular pocket containing a zinc binding site and two aspartic acid histidine charge relay systems. My guess is that the business end of sulforaphane is the isothiocyanate, which could react by nucleophilic attack of either the histidine nitrogen or the aspartic acid oxygen on the carbon of the -N=C=S group. Perhaps one of readers knows how it works.

Histone deacetylase inhibitors are presently very ‘hot’ and one of them, SAHA was approved by the FDA for the treatment of T cell cutaneous lymphoma in 2007, and many others are under active investigation. It’s important to remember that although this class of enzymes was discovered by their ability to remove acetyl groups from histones, they also remove acetyl groups from proteins which are not histones (e.g. HSP90).

So veggies are a two-edged sword.

Ebola — an update (25 Oct ’14)

The experiment of nature referred to in a previous post ( when Amber Vinson, a nurse who had helped care for a fatal case of Ebola, took a commercial flight from Cleveland to Dallas the day she became symptomatic with Ebola is almost over. She was diagnosed 14 October, the day she took the flight, and so far no one on the flight has become ill (presumably the 100+ or so are under surveillance).

However, another experiment of Nature has just begun. An M. D. who’d been in Africa treating Ebola victims was diagnosed with it on the 23rd. He had returned to NYC from Africa 14 October and had been up and about in the city. According to the Times he began to feel sluggish the evening of the 21st, went all over the city on the 22nd, including a 3 mile jog on the west side, and noted a mild temperature (100.3 not 103 as initially reported) the morning of the 23rd — reported it immediately and was hospitalized the same day. New York City chastened by the disastrous response to the first case in Texas, sent 3 guys in Hazmat suits to his apartment to pick the doctor up, according to the NYT of 26 October. Some contacts, such as his fiancee are easy to trace, the people he rode with on the subway are not.

The incubation period is said to be no more than 21 days, so neither experiment of nature is truly over. From this case we now know the incubation period can be as short as 7 – 9 days.

As noted in the previous post — The genome of Ebola is RNA which mutates much more rapidly than DNA genomes. It does this so quickly that at death from AIDS (another RNA virus), there are so many viral variants present that the infecting ensemble is called a quasiSpecies. With a large population infected in Africa there is more Ebola virus extant than at any time in the past.

We have a small handle on just how fast the virus is mutating [ Science vol. 345 pp. 1369 – 1372 ’14 (12 Sep ’14) ]. This is a report of 98 virus genomes from 78 patients from Sierra Leone (all this year). The Ebola genome contains 18,959 to 18,961 nucleotides and codes for at least 7 proteins. Compared to all previously known Ebola genome sequences, the virus from Sierra Leone contains 341 fixed changes (e.g. the changes were present in every virus they sequenced). The changes were present in all 7 proteins.

It isn’t clear (to me) from reading the paper how much variation in the viral genome there is (1) in a given individual (2) between individuals. Note that all samples were obtained from late May to early June this year, so the work is a good baseline.

Why is this scary? Because, as is typical for a virus with a genome made of RNA, Ebola is mutating rapidly. This means that we can’t be sure that its incubation characteristics, or its ability to spread from human to human will remain constant.

Producing the paper, required lots of collaboration between people in the USA and Africa, so there are 58 co-authors of the paper. Showing just how bad the disease is five of the fifty-eight co-authors died of Ebola. R. I. P. Mohamed Fullah, Mbalu Fonnie, Alex Moigboi, Alice Kovoma, S. Humarr Khan.

An experiment of nature

Yesterday’s post concerned the fact that 2 nurses taking care of a patient in Texas had been infected (presumably even after taking all the recommended precautions). Given that, I was concerned about the possibility of airborne spread.

Bryan wrote in to say the following:

“It seems doubtful airborne spread was involved. Remember, the Texas patient was initially sent home after showing symptoms, yet none of his family members were infected. Only those health workers directly involved in his care (and thus exposed to infected bodily fluids) have been infected, consistent with the idea that the disease can be transmitted only though contact with infected bodily fluids.”

I certainly hope he is right.

In something right out a novel, the possibility of airborne spread is now going to be empirically tested, as one of the two infected nurses flew to Cleveland, and then back to Texas in the 24 hours prior to her diagnosis. She apparently had a slight fever on boarding. So 100+ people were in a confined space with her for a few hours.

It’s why I don’t read fiction — reality is far more fantastic than anything writers can produce.

One more bizarre development. Here in Massachusetts, legislators today are scheduled to hear about the readiness of the state’s hospitals to handle Ebola. Amazingly, they will only get input from hospital CEOs. No nurses, thank you. Naturally the nurses are pissed as they should be (and so should you if you live in the state). If there were ever a time to hear from boots on the ground about Ebola readiness, it is now.

Addendum 17 Oct ’14

The Obama administration has just appointed a former chief of staff for former vice-president Gore and present vice-president Biden as the “Ebola czar”. Presumably, not for his medical expertise but for his ability to coordinate various governmental agencies, which was hardly the problem in the CDC’s response to the Texas cases. Hopefully, this will not be another case of “Brownie, you’re doing a heck of a job,” but I’m not optimistic —

Now for some molecular biology. The genome of Ebola is RNA which mutates much more rapidly than DNA genomes. It does this so quickly that at death from AIDS (another RNA virus), there are so many viral variants present that the infecting ensemble is called a quasiSpecies. With a large population infected in Africa there is more Ebola virus extant than at any time in the past. There is some reason to hope that natural selection for a more transmissible form of Ebola in the large infected human population will not occur (the AIDS virus hasn’t become more infectious over the years). This is only a hope.


This morning (15 October) it was announced that a second health care worker at the Texas hospital where an ebola patient died has ‘tested positive’ for it. If ebola can spread in a hospital environment where presumably precautions were taken, once it gets out into the populace at large it can spread much faster. This had to be human to human transmission — no other animal vector is involved (as it probably is in Africa).

How does it spread? We don’t know, but the two Texas cases probably imply that airborne spread is possible.

What to do?

In our case it means not getting into a confined space with over 100 people we don’t know from all over the world for an 8 – 16 hour period (e.g. an international flight). Have you ever been on a flight where no one had a cold?

For the USA, it should mean banning all flights from endemic countries. This has been the case in the past. My cousin’s wife has a lot of relatives in Brazil, because the people on the boat had lots of pink eye, and the boat was simply turned away over 100 years ago.

It should mean caring for Ebola patients in specialized facilities where only they are cared for –e.g. not in a general hospital since we don’t know how it spreads.

The greatest way to spread the disease (the Hajj — millions of people from all over the world crowded together for days followed by worldwide dispersal) has mercifully just ended before the disease escaped Africa to any extent.

Will ISIS or Al-Qaeda try to bring Ebola to the USA? Of course.

We live in a society where children have supervised play dates, and where walking unattended to school is almost considered child abuse. What will happen to such a risk-averse society when there is actual risk to going out to (the mall, the school, to work)?

The thermodynamic subtlety of cholera

Who knew that the cholera organism passed a thermodynamics course with flying colors? Consider that it has to function at widely different temperatures (37 C when it infects us, and 20 – 30 C when it’s out in the world). When it infects us it needs to make toxins and build a secretion system to export it. This cost a lot of metabolic money (ATP). Clearly there’s no point in doing this at temperatures outside the body and a lot of reasons not to (at least 60 as turning on toxin production and building the secretion system involves synthesizing at least 60 different proteins).

If some of the following terms are unfamiliar have a look at and follow the links.

How does thermodynamics help the organism turn on these genes at body temperature (37 C in us)? ToxT is a protein which turns on production of the 60 proteins. The mRNA for ToxT is only translated into protein by the ribosome at 37 C.

[ Proc. Natl. Acad. Sci. vol. 111 pp. 14241 – 14246 ’14 ] The mRNA for ToxT has what the authors call an RNA thermometer in its untranslated region. It is just a sequence of nucleotides which binds to the Shine Dalgarno (SD) element ( in the ToxT mRNA tying it up, so the SD element can’t bind to the ribosome, meaning the mRNA for ToxT can’t be transcribed into protein . Guess what? The thermometer only binds to the SD element at low temperatures, at higher temperatures the binding is unstable leaving the SD sequence free, turning on synthesis of ToxT which then turns on the 60 proteins involved in toxin production. Clever no?

Cholera is a terrible disease, afflicting less developed countries causing terrible infant mortality. I can’t resist mentioning a completely avoidable epidemic inflicted in the name of risk reduction years ago.

[ Nature vol. 354 p. 255 ’91 ] An amazing article places the blame for the cholera epidemic sweeping South America starting in Peru on a misguided application of an Environmental Protection Study implicating water chlorination as a cause of cancer. During the 80’s Peruvian officials, citing the EPA study, stopped chlorinating many of the well in Lima. However, others say that the decision might have been more based on economics than data from the EPA.

It is comforting to know that the 3516 who have died so far have been spared a long bout with cancer.

9 Oct ’14 — Emo wrote the following comment today

Story of Peruvian officials stopping chlorinating water supply based on EPA study was debunked in a study published in Lancet one year after the nature news story: Swerdlow et al. “Waterborne Transmission of Epidemic Cholera in Trujillo, Peru: Lessons for a Continent at Risk,” Lancet Vol. 340 No. 8810 (July 4, 1992), pgs. 28-33. They never chlorinated water in Trujillo, second largest city in the country because they didn’t believe deep well water needed disinfection and cost of chlorinator and chlorine was too much

Thanks Emo

Can losing one gene do all that? Yes it can — there’s still hope

The Cancer Genome Atlas has dashed our hopes of finding ‘the’ cause of cancer. It has sequenced the genomes of a large number of cancers — the following paper looked at 21 tumor types sequencing the protein coding parts (exomes) of 4,742 specimens, along with that of normal tissues [ Nature vol. 505 pp. 495 – 501 ’14 ].

The problem is that lots of mutations have been found in every type of cancer studied this way.

The following is typical — 178 cases of lung cancer (squamous cell variety) were studied. Some 360 mutations in exons, 165 genomic rearrangements, and 323 copy number alterations were found — but this doesn’t represent the results for the 178 cases as a whole. This was the average amount of genomic mayhem seen in each individual tumor . How do you find ‘the’ cause of the cancer in this mess? One way might be to find a gene mutated in all 178 cases (e. g. recurrent mutations). This would be the holy grail — the mutation driving cancer formation, the rest being the chaff of the well known genomic instability due to the high mutation rate of cancer cells. They found 11 such genes, but they were far from mutated in all cases. Pretty depressing isn’t it?

A recent paper [ Proc. Natl. Acad. Sci. vol. 111 pp. 14009 – 14010, E4066 – E4075 ’14 ] gave an example of a huge number of changes in the clinical activity of a cancer cell line due to the functional loss of just one gene (called COSMC). Here’s what happened. In a pancreatic cancer cell line, COSMC knockout produced malignant xenografts (e.g. placing the cells in an immunodeficient animal and watching what happens), which could be reversed by reintroduction of COSMC. The changes include (1) increased proliferation, (2)loss of contact inhibition of growth, (3) loss of tissue architecture, (4) less basement membrane adhesion and (5) invasive growth — remarkable that knocking out just one gene could do so much. Perhaps not a driver mutation, but certainly a delicious drug target. Before getting too excited, remember that this occurred in a cell line which was cancerous to begin with.

The quick and dirty explanation of what is going on is that COSMC is a protein chaperone for an enzyme adding a sugar to proteins destined either for secretion or for insertion into the cell membrane. Lose COSMC and the whole pattern of sugar attachments to these proteins changes. There are a lot of proteins modified by adding sugars (glycosylated proteins), actually 446 of them, with 1,471 sites for this to happen.

The rest of the post is for the cognoscenti and concerns the gory details.

From the paper itself — “Neoplastic transformation of human cells is virtually always associated with aberrant glycosylation of proteins and lipids.” The most frequently seen glycophenotype are the Tn and STn carbohydrate epitopes of epithelial cell cancers. They arise when mucin-type O-linked glycans (normally more complex) are truncated so that only a single -N-acetylgalactosamine (Tn) or N-acetylgalactosamine modified with sialic acid (STn) remains attached to the protein by a serine or a threonine. There are ‘up to’ 20 GalNAc transferases adding GalNAc to serine or threonine. Overall there are some 200 glycosyltransferase found in the secretory pathway. In most cases the GalNAc is modified with beta 1 –> 3 galactose by a single enzyme (called C1GalT1). This reaction is dependent on COSMC, a protein chaperone.

Although there weren’t mutations in the glycosyltransferases studied in 46 cases of pancreatic cancer, 40% of them showed hypermethylation of the COSMC (e.g. methylated cytosines in the promoter region, which shut down transcription of COSMC). This correlated with expression of truncated O-Glycans (e.g. the Tn and STn antigens) and loss of C1GalT expression.

Thrust and Parry about memory storage outside neurons.

First the post of 23 Feb ’14 discussing the paper (between *** and &&& in case you’ve read it already)

Then some of the rather severe criticism of the paper.

Then some of the reply to the criticisms

Then a few comments of my own, followed by yet another old post about the chemical insanity neuroscience gets into when they apply concepts like concentration to very small volumes.

Are memories stored outside of neurons?

This may turn out to be a banner year for neuroscience. Work discussed in the following older post is the first convincing explanation of why we need sleep that I’ve seen.

An article in Science (vol. 343 pp. 670 – 675 ’14) on some fairly obscure neurophysiology at the end throws out (almost as an afterthought) an interesting idea of just how chemically and where memories are stored in the brain. I find the idea plausible and extremely surprising.

You won’t find the background material to understand everything that follows in this blog. Hopefully you already know some of it. The subject is simply too vast, but plug away. Here a few, seriously flawed in my opinion, theories of how and where memory is stored in the brain of the past half century.

#1 Reverberating circuits. The early computers had memories made of something called delay lines ( where the same impulse would constantly ricochet around a circuit. The idea was used to explain memory as neuron #1 exciting neuron #2 which excited neuron . … which excited neuron #n which excited #1 again. Plausible in that the nerve impulse is basically electrical. Very implausible, because you can practically shut the whole brain down using general anesthesia without erasing memory.

#2 CaMKII — more plausible. There’s lots of it in brain (2% of all proteins in an area of the brain called the hippocampus — an area known to be important in memory). It’s an enzyme which can add phosphate groups to other proteins. To first start doing so calcium levels inside the neuron must rise. The enzyme is complicated, being comprised of 12 identical subunits. Interestingly, CaMKII can add phosphates to itself (phosphorylate itself) — 2 or 3 for each of the 12 subunits. Once a few phosphates have been added, the enzyme no longer needs calcium to phosphorylate itself, so it becomes essentially a molecular switch existing in two states. One problem is that there are other enzymes which remove the phosphate, and reset the switch (actually there must be). Also proteins are inevitably broken down and new ones made, so it’s hard to see the switch persisting for a lifetime (or even a day).

#3 Synaptic membrane proteins. This is where electrical nerve impulses begin. Synapses contain lots of different proteins in their membranes. They can be chemically modified to make the neuron more or less likely to fire to a given stimulus. Recent work has shown that their number and composition can be changed by experience. The problem is that after a while the synaptic membrane has begun to resemble Grand Central Station — lots of proteins coming and going, but always a number present. It’s hard (for me) to see how memory can be maintained for long periods with such flux continually occurring.

This brings us to the Science paper. We know that about 80% of the neurons in the brain are excitatory — in that when excitatory neuron #1 talks to neuron #2, neuron #2 is more likely to fire an impulse. 20% of the rest are inhibitory. Obviously both are important. While there are lots of other neurotransmitters and neuromodulators in the brains (with probably even more we don’t know about — who would have put carbon monoxide on the list 20 years ago), the major inhibitory neurotransmitter of our brains is something called GABA. At least in adult brains this is true, but in the developing brain it’s excitatory.

So the authors of the paper worked on why this should be. GABA opens channels in the brain to the chloride ion. When it flows into a neuron, the neuron is less likely to fire (in the adult). This work shows that this effect depends on the negative ions (proteins mostly) inside the cell and outside the cell (the extracellular matrix). It’s the balance of the two sets of ions on either side of the largely impermeable neuronal membrane that determines whether GABA is excitatory or inhibitory (chloride flows in either event), and just how excitatory or inhibitory it is. The response is graded.

For the chemists: the negative ions outside the neurons are sulfated proteoglycans. These are much more stable than the proteins inside the neuron or on its membranes. Even better, it has been shown that the concentration of chloride varies locally throughout the neuron. The big negative ions (e.g. proteins) inside the neuron move about but slowly, and their concentration varies from point to point.

Here’s what the authors say (in passing) “the variance in extracellular sulfated proteoglycans composes a potential locus of analog information storage” — translation — that’s where memories might be hiding. Fascinating stuff. A lot of work needs to be done on how fast the extracellular matrix in the brain turns over, and what are the local variations in the concentration of its components, and whether sulfate is added or removed from them and if so by what and how quickly.

We’ve concentrated so much on neurons, that we may have missed something big. In a similar vein, the function of sleep may be to wash neurons free of stuff built up during the day outside of them.


In the 5 September ’14 Science (vol. 345 p. 1130) 6 researchers from Finland, Case Western Reserve and U. California (Davis) basically say the the paper conflicts with fundamental thermodynamics so severely that “Given these theoretical objections to their interpretations, we choose not to comment here on the experimental results”.

In more detail “If Cl− were initially in equilibrium across a membrane, then the mere introduction of im- mobile negative charges (a passive element) at one side of the membrane would, according to their line of thinking, cause a permanent change in the local electrochemical potential of Cl−, there- by leading to a persistent driving force for Cl− fluxes with no input of energy.” This essentially accuses the authors of inventing a perpetual motion machine.

Then in a second letter, two more researchers weigh in (same page) — “The experimental procedures and results in this study are insufficient to support these conclusions. Contradictory results previously published by these authors and other laboratories are not referred to.”

The authors of the original paper don’t take this lying down. On the same page they discuss the notion of the Donnan equilibrium and say they were misinterpreted.

The paper, and the 3 letters all discuss the chloride concentration inside neurons which they call [Cl-]i. The problem with this sort of thinking (if you can call it that) is that it extrapolates the notion of concentration to very small volumes (such as a dendritic spine) where it isn’t meaningful. It goes on all the time in neuroscience. While between any two small rational numbers there is another, matter can be sliced only so thinly without getting down to the discrete atomic level. At this level concentration (which is basically a ratio between two very large numbers of molecules e.g. solute and solvent) simply doesn’t apply.

Here’s a post on the topic from a few months ago. It contains a link to another post showing that even Nobelists have chemical feet of clay.

More chemical insanity from neuroscience

The current issue of PNAS contains a paper (vol. 111 pp. 8961 – 8966, 17 June ’14) which uncritically quotes some work done back in the 80’s and flatly states that synaptic vesicles have a pH of 5.2 – 5.7. Such a value is meaningless. Here’s why.

A pH of 5 means that there are 10^-5 Moles of H+ per liter or 6 x 10^18 actual ions/liter.

Synaptic vesicles have an ‘average diameter’ of 40 nanoMeters (400 Angstroms to the chemist). Most of them are nearly spherical. So each has a volume of

4/3 * pi * (20 * 10^-9)^3 = 33,510 * 10^-27 = 3.4 * 10^-23 liters. 20 rather than 40 because volume involves the radius.

So each vesicle contains 6 * 10^18 * 3.4 * 10^-23 = 20 * 10^-5 = .0002 ions.

This is similar to the chemical blunders on concentration in the nano domain committed by a Nobelist. For details please see —

Didn’t these guys ever take Freshman Chemistry?

Addendum 24 June ’14

Didn’t I ever take it ? John wrote the following this AM

Please check the units in your volume calculation. With r = 10^-9 m, then V is in m^3, and m^3 is not equal to L. There’s 1000 L in a m^3.
Happy Anniversary by the way.

To which I responded

Ouch ! You’re correct of course. However even with the correction, the results come out to .2 free protons (or H30+) per vesicle, a result that still makes no chemical sense. There are many more protons in the vesicle, but they are buffered by the proteins and the transmitters contained within.


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