Category Archives: Neurology & Psychiatry

Just when you thought you understood neurotransmission

Back in the day, the discovery of neurotransmission allowed us to think we understood how the brain worked. I remember explaining to medical students in the early 70s, that the one way flow of information from the presynaptic neuron to the post-synaptic one was just like the flow of current in a vacuum tube — yes a vacuum tube, assuming anyone reading knows what one is. Later I changed this to transistor when integrated circuits became available.

Also the Dale hypothesis as it was taught to me, was that a given neuron released the same neurotransmitter at all its endings. As it was taught back in the 60s this meant that just one transmitter was released by a given neuron.

Retrograde transmission was just a glimmer in the mind’s eye back then. We now know that the post-synaptic neuron releases compounds which affect the presynaptic neuron, the supposed controller of the postsynaptic neuron. Among them are carbon monoxide, and the endocannabinoids (e. g. what marihuana is trying to mimic).

In addition there are neurotransmitter receptors on the presynaptic neuron, which respond to what it and other neurons are releasing to control its activity. These are outside the synapse itself. These events occur more slowly than the millisecond responses in the synapse to the main excitatory neurotransmitter of the brain (glutamic acid) and the main inhibitory neurotransmitter (gamma amino butyric acid — aka GABA). Receptors on the presynaptic neuron for the transmitter it’s releasing are called autoreceptors, but the presynaptic terminal also contains receptors for other neurotransmitters.

Well at least, neurotransmitters aren’t released by the presynaptic neuron without an action potential which depolarizes the presynaptic terminal, or so we thought until [ Neuron vol. 82 pp. 63 - 70 '14 ]. The report involves a structure near and dear to the neurologist the striatum (caudate and putamen — which is striated because the myelinated axons of the internal capsule go through its anterior end giving it a striated appearance).

It is the death of the dopamine containing neurons in the substantial nigra which cause Parkinsonism. They project some of their axons to the striatum. The striatum gets input elsewhere (from the cortex using glutamic acid) and from neurons intrinsic to itself (some of which use acetyl choline as their neurotransmitter — these are called cholinergic interneurons).

The paper makes the claim that the dopamine neurons projecting to the striatum also contain the inhibitory neurotransmitter GABA.

The paper also says that the cholinergic interneurons cause release of GABA by the dopamine neurons — they bind to a type of acetyl choline receptor called nicotinic (similar but not identical to the nicotinic receptors which allow our muscles to contract) in the presynaptic terminals of the dopamine neurons of the substantial nigra residing in the striatum. Isn’t medicine and neuroanatomy a festival of terms? It’s why you need a good memory to survive medical school.

These used optogenetics (something I don’t have time to explain — but see http://en.wikipedia.org/wiki/Optogenetics ) to selectively stimulate the 1 – 2% of striatal neurons which use acetyl choline as a neurotransmitter. What they found was that only GABA (and not dopamine) was released by the dopamine neurons in response to stimulating this small subset of neurons. Even more amazing, the GABA release occurred without an action potential depolarizing the presynaptic terminal.

This literally stands everything I thought I knew about neurotransmission on its ear. How widespread this phenomenon actually is, isn’t known at this point. Clearly, the work needs to be replicated — extreme claims require extreme evidence.

Unfortunately I’ve never provided much background on neurotransmission for the hapless chemists and medicinal chemists reading this (if there are any), but medicinal chemists must at least have a smattering of knowledge about this, since neurotransmission is involved in how large classes of CNS active drugs work — antidepressants, antipsychotics, anticonvulsants, migraine therapy. There is some background on this here — http://luysii.wordpress.com/2010/08/29/some-basic-pharmacology-for-the-college-student/

The prions within us

Head for the hills. All of us have prions within us sayeth [ Cell vol. 156 pp. 1127 - 1129, 1193 - 1206, 1206 - 1222 '14 ]. They are part of the innate immune system and help us fight infection. But aren’t all sorts of horrible disease (Bovine Spongiform Encephalopathy aka BSE, Jakob Creutzfeldt disease aka JC disease, Familial Fatal Insomnia etc. etc.) due to prions? Yes they are.

If you’re a bit shaky on just what a prion is see the previous post which should get you up to speed — https://luysii.wordpress.com/2014/03/30/a-primer-on-prions/.

Initially there was an enormous amount of contention when Stanley Prusiner proposed that Jakob Creutzfeldt disease was due to a protein forming an unusual conformation, which made other copies of the same protein adopt it. It was heredity without DNA or RNA (although this was hotly contended at the time), but the evidence accumulating over the years has convinced pretty much everyone except Laura Manuelidis (about whom more later). It convinced the Nobel Prize committee at any rate.

JC disease is a rapidly progressive dementia which kills people within a year. Fortunately rare (attack rate 1 per million per year) it is due to misfolded protein called PrP (unfortunately initially called ‘the’ prion protein although we now know of many more). Trust me, the few cases I saw over the years were horrible. Despite decades of study, we have no idea what PrP does, and mice totally lacking a functional Prp gene are normal. It is found on the surface of neurons. Bovine Spongiform Encephalopathy was a real scare for a time, because it was feared that you could get it from eating meat from a cow which had it. Fortunately there have been under 200 cases, and none recently.

If you cut your teeth on the immune system being made of antibodies and white cells and little else, you’re seriously out of date. The innate immune system is really the front line against infection by viruses and bacteria, long before antibodies against them can be made. There are all sorts of receptors inside and outside the cell for chemicals found in bacteria and viruses but not in us. Once the receptors have found something suspicious inside the cell, a large protein aggregate forms which activates an enzyme called caspase1 which cleaves the precursor of a protein called interleukin 1Beta, which is then released from some immune cells (no one ever thought the immune system would be simple given all that it has to do). Interleukin1beta acts on all sorts of cells to cause inflammation.

There are different types of inflammasomes and the nomenclature of their components is maddening. Two of the sensors for bacterial products (AIM, NLRP2) induce a polymerization of an inflammasome adaptor protein called ASC producing a platform for the rest of the inflammasome, which contains other proteins bound to it, along with caspase1 whose binding to the other proteins activates it. (Terrible sentence, but things really are that complicated).

ASC, like most platform proteins (scaffold proteins), is made of many different modules. One module in particular is called pyrin (because one of the cardinal signs of inflammation is fever). Here’s where it gets really interesting — the human pyrin domain in ASC can replace the prion domain of the first yeast prion to be discovered (Sup35 aka [ PSI+ ] — see the above link if you don’t know what these are) and still have it function as a prion in yeast. Even more amazing, is the fact that the yeast prion domain can functionally replace ASC modules in our inflammasomes and have them work (read the references above if you don’t believe this — I agree that it’s paradigm destroying). Evidence for human prions just doesn’t get any better than this. Fortunately, our inflammasome prions are totally unrelated to PrP which can cause such havoc with the nervous system.

Historical note: Stanley Prusiner was a year behind me at Penn Med graduating in ’67. Even worse, he was a member of my med school fraternity (which was more a place to get a decent meal than a social organization). Although I doubtless ate lunch and dinner with him before marrying in my Junior year, I have absolutely no recollection of him. I do remember our class’s medical Nobel — Mike Brown. Had I gone to Yale med instead of Penn, Laura Manuelidis would have been my classmate. Small world

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.https://luysii.wordpress.com/2013/10/21/is-sleep-deprivation-like-alzheimers-and-why-we-need-sleep-in-the-first-place/

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 (http://en.wikipedia.org/wiki/Delay_line_memory) 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.

Here’s just how poor the research on Autism Spectrum Disorder has been

A stroke 50 years ago would be a stroke today. Cancer is still cancer. Not so with Autism, Autism Spectrum Disorder (ASD), Asperger’s syndrome. The Diagnostic and Statistical Manual of the American Psychiatric Association (DSM to you) has gone through 6 editions (DSM-I through DSM-V, including DSM-IV-TR). The terms are defined differently in all (or not even defined). For the gory details see:

http://www.autismconsortium.org/symposium-files/WalterKaufmannAC2012Symposium.pdf

Here’s a summary.

DSM-I (1952) & DSM-II (1968)
No term Autism or Pervasive Developmental Disorder Closest term: Schizophrenic Reaction (Childhood Type)
1980 DSM-III
Pervasive Developmental Disorders (PDD):
Childhood Onset PDD, Infantile Autism, Atypical Autism
1987 DSM-III-R
Pervasive Developmental Disorders (PDD):
PDD-NOS, Autistic Disorder
1994 DSM-IV
Pervasive Developmental Disorders (PDD):
PDD-NOS, Autistic Disorder, Asperger Disorder, Childhood Disintegrative Disorder, Rett syndrome
2000 DSM-IV-TR
Same diagnoses, text correction for PDD-NOS

Needless to say, DSM-V (out since May 2013) has it differently.

The following paper [ Proc. Natl. Acad. Sci. vol. 111 pp. 1981 - 1986 '14 ] should have neuroscientific researchers on Autism Spectrum Disorder (ASD) of the DSM-IV hanging their heads in shame.

The criteria for ASD are obviously a wastebasket. Here they are from the DSM-IV-TR

Autistic disorder (classic autism)
Asperger’s disorder (Asperger syndrome)
Pervasive developmental disorder not otherwise specified (PDD-NOS)
Rett’s disorder (Rett syndrome)
Childhood disintegrative disorder (CDD).

Just about any kid not doing well cognitively fits in to #3. Hopefully the DSM-V makes things better — it’s too early to tell. The link has the details and a lot of the reasoning behind yet another change.

All the work cited in the PNAS paper concerns ASD as diagnosed by DSM-IV-TR. DSM-V is too new to have papers out using its criteria.

If you take 100 kids developing normally, do various types of magnetic resonance imaging (MRI) on their brains, and compare then with 100 kids not doing well, you are certain to find more structural abnormalities of the brain in the second group, regardless of how they were diagnosed, or what they were diagnosed with.

Dozens of papers were written on MRIs in ASD kids. 40% had fewer than 15 subjects. The most replicated finding (up to the PNAS paper) was poor connections between various parts of the brain, manifest as abnormalities of the white matter. Exactly what they were measuring, and how the measurement requires a tensor and not a vector is really quite interesting and can teach you some math. I’ll save this for the end.

Only 2 of the dozens of papers controlled for data quality. MRIs have been around long enough, that most know that to get a decent study the subject has to be quite still, something very difficult for kids, and even harder for autistic kids.

When head motion was controlled for in this large (52 ASDs 73 normals to start) all the abnormalities disappeared (save one, and they looked at 18 different white matter tracts in the brain). They had to throw out the studies on 12/52 of the autistics and 2/73 normals because of motion– showing how suspect the previous data really was. The head motion was producing the abnormalities.

Is this terrible research or what (PNAS paper excepted)?

Perhaps the new criteria in DSM-V will result in a more homogenous group.

What does diffusion tensor imaging actually measure? Imagine the nerve fibers (axons) of the brain in the deep white matter as a bundle of wires, most of them going in the same direction in any small volume. Assume they are bathed in water. Now add some colored water at one end, and see how quickly the color diffuses in various directions in the bundle. The color will diffuse faster along the bundle, than in the two directions perpendicular to it. This is what the diffusion tensor measures — diffusion of tissue water in a variety of directions. If the bundle is loose, or disorganized, or some wires are missing, than the diffusion in the various directions won’t differ as much — this is called lack of anisotropy. Take out the wires and the diffusion is the same in all directions (no anisotropy at all). This was the finding in ASD — less white matter anisotropy in diffusion tensor imaging — implying that there is something wrong with it.

Why wouldn’t an overall vector summing up the diffusion in the major direction be enough. One can add vectors together after all. Because you’d lose all the information about anisotropy. The tensor preserves it. It’s why tensors are used to measure the stress in a given material. Slick. Now you understand (something) about tensors. However it should be noted that vectors are tensors too. There’s a lot more too it (particularly indices).

Who chose Ariel Sharon’s MDs, Arafat?

The MDs who anti coagulated Ariel Sharon after his first stroke when he was known to have cerebral amyloid antipathy (CAA) effectively killed him. Sometimes you don’t know a patient has CAA, although lobar hemorrhage is a clue. CAA replaces the normal lining of cerebral blood vessels by structureless proteinaceous gunk, making them incredibly fragile.

I know that most of you are not histologists, but look at the picture anyway

http://www.google.com/search?q=Cerebral+amyloid+angiopathy&client=safari&rls=en&tbm=isch&source=iu&imgil=D2fj9YvUZUYDAM%253A%253Bhttp%253A%252F%252Ft3.gstatic.com%252Fimages%253Fq%253Dtbn%253AANd9GcSOHKNeKuYiaeRctet7kj3Zy-pnyfKgCcXEzoK11WyySNRZIECK%253B4272%253B2848%253B84Gvq0qSBo4RNM%253Bhttp%25253A%25252F%25252Fen.wikipedia.org%25252Fwiki%25252FCerebral_amyloid_angiopathy&sa=X&ei=hUnTUqX6CMLnsASkgYHwCw&ved=0CEkQ9QEwAw&biw=1550&bih=933#facrc=_&imgrc=D2fj9YvUZUYDAM%253A%3B84Gvq0qSBo4RNM%3Bhttp%253A%252F%252Fupload.wikimedia.org%252Fwikipedia%252Fcommons%252Fd%252Fd5%252FCerebral_amyloid_angiopathy_-_very_high_mag.jpg%3Bhttp%253A%252F%252Fen.wikipedia.org%252Fwiki%252FCerebral_amyloid_angiopathy%3B4272%3B2848

The distorted oval with the relatively clear center in the middle of the picture is a blood vessel within the brain. At 3 o’clock it looks fairly normal, and you can see dark cell nuclei within purplish cytoplasm. At 11 to 2 o’clock the red stuff is amyloid. Note how it distorts the normal anatomy, thickening the vessels wall and decreasing the number of cells. This is where these vessels break.

Using anticoagulants on a patient with known CAA is ghastly malpractice. What were they thinking? Unsurprisingly he had a massive brain hemorrhage shortly thereafter, which kept him in a vegetative state until his passing this month.

Son of a nonAddicting Painkiller

I’ve decided to republish yesterday’s post with some added material to show exactly how to find one of the world’s most useful, not to say lucrative drugs. The response of a friend led me do this. If you’ve read yesterday’s post — start reading after the ****

Synthetic organic chemists, molecular modelers and X-ray crystallographers fire up your engines. A great target is available, discovered by the humble centipede of all things.

Several times during my career as a neurologist, a new drug was ballyhoo’d as a non-addicting narcotic — Talwin springs to mind, among others. The hope was that a molecule with both agonist and antagonist properties wouldn’t be addicting. All basically inhibited neurotransmitter release (like the opiates), to which the synapse responded by making more receptors for the neurotransmitter, explaining tolerance, addiction to the new drug and why junkies can take doses of morphine that would kill you and I.

Have a look at Proc. Natl. Acad. Sci. vol. 110 pp. 17534 – 17539 ’13. Here’s some background. To conduct a nerve impulse, neurons have to let sodium ions flow through their membranes. This is done through proteins called sodium channels. We have 9 distinct genes for them, and most neurons express more than one.

Consider the humble pufferfish. It sometimes makes a beautiful organic molecule called tetrodotoxin (http://en.wikipedia.org/wiki/Tetrodotoxin) which blocks all but 2 of our sodium channels stopping them from conducing sodium ions. It can kill you and is 100x more potent than cyanide. Even so, it’s a very pretty molecule (to an organic chemist) with some resemblance to adamantane.

The centipede makes a 42 amino acid protein which essentially blocks only one sodium channel (NaV1.7). It’s quite potent, doing this at a concentration of 25 nanoMolar. Clinically, it is even more potent than morphine (well, in animal models anyway)

So why get excited? Because as far as we can tell, its action is on peripheral nerve fibers, not the brain. For some reason nerve fibers carrying painful impulses (a philosophic conundrum — impulses themselves are no more painful than a fire feels hot to itself) from the periphery to the spinal cord and brain use lots NaV1.7. So addiction shouldn’t be a problem.

There were some clues already — there is a disorder called congenital insensitivity to pain [Nature vol. 444 pp. 831 - 832 '06], due to loss of function mutations in NaV1.7. Other mutations here cause several painful syndromes — erythromelalgia [J. Med. Genet. vol. 41 pp. 171 - 174 '04 ], chronic rectal pain. These mutations cause a hyper functioning sodium channel which stays open too much.

Total absence of pain isn’t good, as we need it to warn us that we’re stressing our joints excessively. In the bad old days when there was a lot of syphilis around, it sometimes caused peripheral nerve degeneration, resulting in something called Charcot joints — http://en.wikipedia.org/wiki/Neuropathic_arthropathy.

So get cracking guys, if tetrodotoxin, a small compact molecule can nonspecifically block most sodium channels, surely you should be able to find something smaller then a 42 amino acid protein to block NaV1.7 (selectively of course). I’m not sure that we have a structure of NaV1.7, but others are known, along with all their amino acid sequences, so it should be possible to model binding sites by analogy.

****

I received the following from a computational chemist friend. ” Although good luck finding a peptidomimetic for a 42 amino acid peptide…”

I don’t think we’ll need a peptide at all. Likely, it will be a modified tetrodotoxin-like molecule. So we know both where to look and how to look.

First some more background, which I should have put in the original post. People with mutations in NaV1.7 and congenital insensitivity to pain, are cognitively normal (except for pain sensation). This implies that just blocking the NaV1.7 with even a drug getting into the brain won’t have cognitive and/or addictive side effects (and any medicinal chemist will tell you how hard it is to get drugs into the brain).

Consider the Grasshopper mouse of the Arizona desert [ Science vol. 342 pp. 428 - 429, 441 - 446 '13 ]. It eats the fearsome Bark scorpion, without suffering pain. The venom slows the inactivation of the NaV1.7, effectively activating it and causing pain in every other animal. Mutations in another sodium channel (NaV1.8) cause binding of the toxin, so it doesn’t get near NaV1.7. The variants are very near the pore of the channel (where the ions go through). Just a switch in the sequence order of two amino acids (Glutamic acid and glutamine) in NaV1.8 is enough to make NaV1.8 inhibited by the venom.

What does all this have to do with finding a drug to inhibit NaV1.7?

We have the amino acid sequences of all 9 of our sodium channels? NaV1.7 is sensitive to tetrodotoxin (e.g blocked by it), NaV1.8 is Insensitive. The Xray crystallographers need to give us structures of both channels, with and without tetrodotoxin bound. The computational chemist has to dock a tetrodotoxin analog into NaV1.7 similar to the way it fits into NaV1.8 blocking it. Not easy, but we know the target, and we have an excellent candidate to start with. There is no need to mess with 42 amino acid proteins, even though nature and the Grasshopper mouse did. The synthetic organic chemist will be kept busy making tetrodotoxin variants, perhaps those suggested by the computational chemists.

The really hard part in all this will be finding a drug that isn’t a super tetrodotoxin, blocking NaV1.7 and everything else. It should be doable.

The rewards for the successful company doing this are enormous. I’ve got enough money and am too lazy to suit up and go back into the lab (assuming anyone would have me). The reward of pointing the way will be enough. See http://luysii.wordpress.com/2011/04/13/an-attaboy/

Addendum 8 Nov ’13 — Tetrodotoxin should be particularly easy to model, as its stereochemistry should be relatively simple. The adamantine-like structure doesn’t move at all, being locked in place. There is a CH2OH side chain which can flop about, and a cyclohexane ring fused to the rigid part. Even there, the guanido group embedded there is likely to be planar. It’s easy to see why it’s water soluble with all those hydroxyls, and the fact that it is a zwitterion. Modeling solvent interactions is never easy, but modeling hydrogen bonds to amino acid side chains is probably easier, which is what’s important here.

A non-addicting painkiller?

Synthetic organic chemists, molecular modelers and X-ray crystallographers fire up your engines. A great target is available, discovered by the humble centipede of all things.

Several times during my career as a neurologist, a new drug was ballyhoo’d as a non-addicting narcotic — Talwin springs to mind, among others. The hope was that a molecule with both agonist and antagonist properties wouldn’t be addicting. All basically inhibited neurotransmitter release (like the opiates), to which the synapse responded by making more receptors for the neurotransmitter, explaining tolerance, addiction to the new drug and why junkies can take doses of morphine that would kill you and I.

Have a look at Proc. Natl. Acad. Sci. vol. 110 pp. 17534 – 17539 ’13. Here’s some background. To conduct a nerve impulse, neurons have to let sodium ions flow through their membranes. This is done through proteins called sodium channels. We have 9 distinct genes for them, and most neurons express more than one.

Consider the humble pufferfish. It sometimes makes a beautiful organic molecule called tetrodotoxin (http://en.wikipedia.org/wiki/Tetrodotoxin) which blocks all but 2 of our sodium channels stopping them from conducing sodium ions. It can kill you and is 100x more potent than cyanide. Even so, it’s a very pretty molecule (to an organic chemist) with some resemblance to adamantane.

The centipede makes a 42 amino acid protein which essentially blocks only one sodium channel (NaV1.7). It’s quite potent, doing this at a concentration of 25 nanoMolar. Clinically, it is even more potent than morphine (well, in animal models anyway)

So why get excited? Because as far as we can tell, its action is on peripheral nerve fibers, not the brain. For some reason nerve fibers carrying painful impulses (a philosophic conundrum — impulses themselves are no more painful than a fire feels hot to itself) from the periphery to the spinal cord and brain use lots NaV1.7. So addiction shouldn’t be a problem.

There were some clues already — there is a disorder called congenital insensitivity to pain [Nature vol. 444 pp. 831 - 832 '06], due to loss of function mutations in NaV1.7. Other mutations here cause several painful syndromes — erythromelalgia [J. Med. Genet. vol. 41 pp. 171 - 174 '04 ], chronic rectal pain. These mutations cause a hyper functioning sodium channel which stays open too much.

Total absence of pain isn’t good, as we need it to warn us that we’re stressing our joints excessively. In the bad old days when there was a lot of syphilis around, it sometimes caused peripheral nerve degeneration, resulting in something called Charcot joints — http://en.wikipedia.org/wiki/Neuropathic_arthropathy.

So get cracking guys, if tetrodotoxin, a small compact molecule can nonspecifically block most sodium channels, surely you should be able to find something smaller a 42 amino acid protein to block NaV1.7 (selectively of course). I’m not sure that we have a structure of NaV1.7, but others are known, along with all their amino acid sequences, so it should be possible to model binding sites by analogy.

Is sleep deprivation like Alzheimer’s and why we need sleep in the first place

Ask a cardiologist why the heart needs to pump and you’ll get a strange look. Ask any neuroscientist why the brain needs to sleep, and they’ll scratch their head — until now perhaps. A paper in Science a few days ago may have the answer [ Science vol. 342 pp. 316 - 317, 373 - 377 '13 ] Essentially the brain gets washed out during sleep.

First — a bit of history. The tissue of the brain is so tightly packed that it is impossible to see the cells that make it up with the usual stains used by light microscopists. People saw nuclei all right but they thought the brain was a mass of tissue with nuclei embedded in it (like a slime mold). Muscle is like that — long fibers with hundreds of nuclei here and there. It wasn’t until that late 1800′s that Camillo Golgi developed a stain which would now and then outline a neuron with all its processes. Another anatomist (Ramon Santiago y Cajal) used Golgi’s technique and argued with Golgi that yes the brain was made of cells. Fascinating that Golgi, the man responsible for showing nerve cells, didn’t buy it. This was a very hot issue at the time, and the two received a joint Nobel prize in 1906 (only 5 years after the prizes began).

How tightly packed are the cells in the brain? The shortest wavelength of visible light is 4000 Angstroms. Cells in the brain are packed far more tightly. To see the space between the brain cell external membranes you need an electron microscope (EM). Just preparing a sample for EM really fries the tissue. Neurons are packed together with less than 1000 Angstroms between them. So how much of this is artifact of preparation for electron microscopy has never been clear to me. One study injected a series of quantum dots of known diameter into the cerebral spinal fluid (CSF) to see the smallest sized dot that could insinuate itself between neurons [ Proc. Natl. Acad. Sci. vol. 103 pp. 5567 - 5572 '06 ]. The upper limit was around 350 Angstroms. No wonder the issue was contentious when all they had was light microscopy.

Surprisingly, the PNAS paper comes up with an estimate that brain extracellular space comprises 20% of brain volume. I find this hard to accept given the above. So how does the brain get rid of waste products? It turns out that there is a circulation of cerebrospinal fluid (CSF) of sorts. Inject a tracer that you can follow into the CSF. After a period of time the tracer enters the brain along arteries (not veins) and after still more time it leaves the brain along the veins (not the arteries). How the tracer gets to veins isn’t discussed in the Science papers. This has been called by the horrible name of the glymphatic system (don’t ask).

Using a great deal of ingenuity, experimental finesse and some very cooperative mice, the flow of CSF into, through and out of the brain was studied. Several findings are striking — the extracellular space (aka interstitial volume) dearly doubles (from 14% to 23%) during sleep. More importantly, the flow into the brain decreases by 95% when you wake the mouse up. Presumably flow out of the brain decreases by the same amount during wake. CSF flow into the brain was present only in the surface exposed to bulk CSF when the animals were awake.

So what? The Abeta peptide is held by many to be the culprit in Alzheimer’s disease. When injected into the mouse cerebral cortex (hardly a physiologic procedure) Abeta peptide is cleared twice as fast from the brain during sleep. We all know that you don’t think as well when sleep deprived, and this may be why. The current thinking on Alzheimer’s is that it isn’t the visible plaques that you can see under the microscope (made largely of Abeta peptide aggregates), but the soluble form of Abeta which you can’t see which causes the trouble. This always struck me as a cop out similar to the way docs would say that labyrinthitis was due to a virus (not that anyone every isolated one). You might as well say both are due to angels (or devils).

So the difficulty thinking with sleep deprivation may be similar to Alzheimer’s disease, if similar goings on occur in our brain. Distinguish this from the sleepiness due to sleep deprivation –Alzheimer patients often have disturbed sleep patterns, but they aren’t particularly sleepy when they’re awake.

The sleepiness may be due to the build up of something else. Bulk flow of fluid is incredibly nonspecific, and will carry anything soluble along with it. Adenosine has been mentioned as one metabolite building up which makes us sleepy. Probably looking for a single compound washed out by CSF as ‘the’ cause of sleepiness or cognitive problems, is like looking for ‘the’ single compound in kidney failure causing similar symptoms. It’s everything the kidney/brain filters and gets rid of.

So, at very long last, we may have found out why we spend 1/3 of our lives asleep.

Is concentration meaningful in a nanoDomain? A Nobel is no guarantee against chemical idiocy

The chemist can be excused for not knowing what a nanodomain is. They are beloved by neuroscientists, and defined as the part of the neuron directly under an ion channel in the neuronal membrane. Ion flows in and out of ion channels are crucial to the workings of the nervous system. Tetrodotoxin, which blocks one of them, is 100 times more poisonous than cyanide. 25 milliGrams (roughly 1/3 of a baby aspirin) will kill you.

Nanodomains are quite small, and Proc. Natl. Acad. Sci. vol. 110 pp. 15794 – 15799 ’13 defines them as hemispheres having a radius of 10 nanoMeters from channel (a nanoMeter is 10^-9 meter — I want to get everyone on board for what follows, I’m not trying to insult your intelligence). The paper talks about measuring concentrations of calcium ions in such a nanodomain. Previous work by a Nobelist (Neher) came up with 100 microMolar elevations of calcium in nanodomains when one of the channels was opened. Yes, evolution has produced ion channels permeable to calcium and not much else, sodium and not much else, potassium and not much else. For details read the papers of Roderick MacKinnon (another Nobelist). The mechanisms behind this selectivity are incredibly elegant — and I can tell you that no one figured out just what they were until we had the actual structures of channels in hand. As chemists you’re sure to get a kick out of them.

The neuroscientist (including Neher the Nobelist) cannot be excused for not understanding the concept of concentration and its limits.

So at a concentration of 100 microMolar (10^-4 molar) how many calcium ions does a nanoDomain contain? Well a liter has 1000 milliliters and each milliliter is 1 cubic centimeter (cc.). So each cubic centimeter is 10^7 nanoMeters on a side, giving it a volume of 10^21 cubic nanoMeters. How many cubic nanoMeters are in a hemisphere of radius 10 nanoMeters — it’s 1/2 * 4/3 * pi * 10^3 = 2095. So there are (roughly) 5 * 10^17 such hemispheres in each cc.

How many ions are in a cc. of a 1 molar solution of calcium — 6 * 10^21 (Avogadro’s #/1000). How many in a 10^-4 molar solution (100 microMolar) — 6 * 10^17. How many calcium ions in a nanoDomain at this concentration? Just (6 * 10^17)/(5 * 10^17) e.g. just over one ion/nanodomain.

Does any chemist out there think that speaking of a 100 microMolar concentration in a volume this small is meaningful? I’d love to be shown how my calculation is wrong, if anyone would care to post a comment.

They do talk about nanodomains of radius 30 nanoMeters, which still would result in under 10 calcium ions/nanoDomain.

Addendum 10 Oct ’13

My face is red ! ! ! “6 * 10^21 (Avogadro’s #/1000)” should be 6 * 10^20 (Avogadro’s #/1000), making everything worse. Here’s how the paragraph should read.

How many ions are in a cc. of a 1 molar solution of calcium — 6 * 10^20 (Avogadro’s #/1000). How many in a 10^-4 molar solution (100 microMolar) — 6 * 10^16. How many calcium ions in a nanoDomain at this concentration? Just (6 * 10^16)/(5 * 10^17) e.g. just over .1 ion/nanodomain. As Bishop Berkeley would say this is the ghost a departed ion.

Even if we increased the size of the nanoDomain by an order of magnitude (making it a hemisphere of 100 nanoMeters radius), this would give us just over 10 ions/nanodomain.

A new parameter for ladies to measure before choosing a mate — testicular volume

I’m amazed that they actually did this work [ Proc. Natl. Acad. Sci. vol. 110 pp. 15746 - 15751 '13 ] but they did. From Atlanta Georgia, the home of the Southern Gentleman. You do have to wonder what sort of wimps would permit this type of work. 70 such individuals were found, still cohabiting with the mother. Clearly a skewed distribution as 65/70 were actually married. No mention of any effect of the sex of the offspring on what they found.

Here’s what they did

Testosterone levels and testicular volume predicted how much parenting a male actually did (based on self-reports from the two parents). Functional MRI on viewing a picture of the offspring also predicted the degree of male parenting.

So which way do you think it went?

The bigger the testicles and the higher the testosterone, the less parenting the father did. Similarly the less activation of one area of the brain in response to a picture of their chile, the less parenting.

So ladies, you may get a macho dude for a mate, but don’t expect much help.

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