SOLOMON SNYDER WINS PRESTIGIOUS MEDAL OF SCIENCE

July 25, 2005

This is a temporary webpage to permit readers of The Maryland Psychiatrist to see the full text of remarks by Dr Snyder in interviews I conducted for the Maryland Psychiatric Society. Dr Snyder has given me permission to publish it on the web for that purpose..

The following is in two parts, both featuring Dr. Snyder. Part I, immediately below is an uncut version of the article that will appear in shorter form (because of space limitations) in the Fall 2005 issue of The Maryland Psychiatrist, a publication of The Maryland Psychiatric Society. It contains Snyder's description of some of the current work going on in his laboratory, plus some discussion.

Part II is a mostly unedited transcript of a recording of Snyder's detailed response when I asked him to talk about his lab's “advances that opened up the modern field of treatment of neuropsychiatric diseases” and led to his being awarded the Medal of Science in March 2005. Readers should understand that there are still some gaps and possibly some uncorrected errors in the text.

This material may not be copied or republished. Gerald D. Klee, MD

Part I

Solomon Snyder, MD Wins Prestigious Medal of Science

“Snyder is undoubtedly one of the most original, prolific and influential intellectuals of our time." Pasko Rakic, MD, PhD(1)

An interview with Solomon Snyder, MD by Gerald D. Klee, MD

Solomon Snyder, MD, Professor of Psychiatry, Neuroscience and Pharmacology and Director of the Department of Neuroscience at Johns Hopkins University School of Medicine, recipient of the 2005 Medal of Science from the President of the United States for his discoveries of the molecular basis of psychotropic drug action.

According to Professor Pasko Rakic, MD, PhD(1) “Snyder is undoubtedly one of the most original, prolific and influential intellectuals of our time. Some of the major advances that opened up the modern field of treatment of neuropsychiatric diseases have stemmed from his work. His pioneering work has led to novel generations of therapeutic drugs and potential treatments for drug abuse… His work contributed to understanding the major actions of the drugs and differentiating agonists and antagonists, permitting the development of novel, less addicting opiates. Snyder then identified receptors for the major neurotransmitters, establishing the mechanism of the anti-psychotic and side-effect actions of neuroleptics and differentiating serotonin receptor subtypes that led to new classes of anti-psychotic, anti-migraine and anti-emetic drugs. Snyder extended receptor strategies to intracellular signal transduction, identifying and purifying the receptors and showing that they contain the calcium ion channels that mediate their actions… He and his colleagues were involved in the discovery of the dopamine receptors and their role in schizophrenia. Snyder's more recent discovery of novel neurotransmitters such as nitric oxide, carbon monoxide and D-serine portend novel therapies in mental illness, stroke and sexual dysfunction.”

I (Gerald D Klee) interviewed Dr Snyder in June at his laboratory. Before our meeting I asked for a tour of his lab, where I was greeted by many of the happiest and most energetic investigators and office staff I’ve seen anywhere. I was enchanted to hear their enthusiastic descriptions of their work and their fondness of Dr Snyder. I’ve heard similar remarks from colleagues who trained with Snyder and later went on to outstanding careers of their own. This atmosphere, an obvious product of Snyder’s personality, must contribute a lot to the exceptional scientific productivity of the laboratory.

I began by asking Dr. Snyder to describe his “advances that opened up the modern field of treatment of neuropsychiatric diseases”. With only a moment’s initial hesitation to organize his thoughts, he delivered a detailed and elegant account of years of complex neurobiological research, t ak ing pains to credit each of his students for their contributions. It was a singular demonstration of his ability to communicate. Additional discussion of his current work took place by email.

It is impossible to describe all of Snyder’s achievements in anything shorter than a volume. This report will focus on some of the ongoing work in his laboratory. The full text of Snyder’s summary of his work can be seen on the Internet at http://www.letreb.com/solomon_snyder_wins_prestigious_.htm

Snyder: “You asked me to provide a list of projects ongoing in the laboratory, based, at least in part, on the interview. Here are some of them.

1. “D-serine as a neurotransmitter:

Glutamate is the major excitatory neurotransmitter in the brain and acts via several receptors of which the NMDA subtype is best known. Psychotomimetic drugs such as PCP act by blocking NMDA receptors and elicit a psychosis which resembles schizophrenia more than any other drug psychosis. Administration to schizophrenics of D-serine in itself as well as related agents alleviates schizophrenic symptoms. The French neuroscientists Phillipe Ascher discovered some years ago that the NMDA receptor must be activated by two agents and show that glycine can work together with glutamate to activate the receptor. Excess activation of NMDA receptors leads to neurotoxicity and stroke so that Ascher reasoned that the requirement of two neurotransmitters for one receptor (something which is unprecedented) functions as a safety mechanism – two keys for the lock – so that one might not get a stroke from eating a steak dinner. However, glycine, like glutamate, is an abundant dietary constituent. D-serine is the unnatural isomer whose existence in biology was discovered accidentally some years ago with D-serine occurring in the brain and virtually no other D-amino acid existing in all mammalian biology. In fundamental studies of NMDA receptors, it was already known that D-serine was substantially more active then glycine, but at that time no one knew that D-serine even exists in biology. We have established that D-serine is the primary endogenous stimulus to the NMDA receptor, working together with glutamate. One direct item of evidence involved utilizing D-amino acid oxidase, an enzyme discovered by Hans Krebs in 1935 and long neglected as a meaningless protein since their were no D-amino acids in mammals. We showed that, under physiologic conditions, D-amino acid oxidase is extremely selective for D-serine. We used it to selectively degrade D-serine and show that NMDA neurotransmission was abolished even though glycine levels were completely normal. We discovered an enzyme, serine racemase, which converts L-serine to D-serine. We are currently examining mice with targeted deletion of serine racemase. Hopefully, characterization of the behavior of these gene knockout mice provide will provide insights into the normal functions of D-serine. This work is being carried out by Paul Kim, who made it his Ph.D. thesis. Paul is now a Hopkins Medical Student. Asif Mustafa, a M.D. Ph.D. student is now working on the project.

2. Gaseous Neurotransmitters (H2S)

We continue to work on gasses as neurotransmitters with much evidence for nitric oxide and carbon monoxide as signaling molecules. Very recently, we have evidence that hydrogen sulfide (H2S) the foul smelling rotten egg odorant is a neurotransmitter. Our collaborator, Dr. Rui Wang, created mice with a knockout of the gene for cystathione-gamma-lyase, the enzyme which we suspected might physiologically generate H2S from the amino acid cysteine. We have shown that H2S formation is virtually abolished in lyase knockout mice. Moreover, neurotransmission in the intestine, which underlies the relaxation phase of peristalsis, is substantially reduced in the knockout mice, indicating that H2S is a neurotransmitter of this process. Interestingly, formerly we showed that nitric oxide and carbon monoxide are also neurotransmitters mediating peristalsis. Our next challenge will be to work out the function for H2S neurotransmission in the brain by mapping the H2S neurons as well as by evaluating the behavior of the gene knockout mice. Crystal Watkins, a M.D. Ph.D. student who is starting psychiatry residency at Hopkins, did this work.

3. Bilirubin as a Cytoprotectant:

We are studying mechanisms of cell death and cell protection. Bilirubin, long thought to be a toxic breakdown product of heme, appears to be a major cellular protectant. This explains the paradox that bilirubin exists at all. If one wanted to get rid of heme, the enzyme heme oxygenase breaks open the heme ring to generate the green pigment biliverdin which can be excreted in the bile. Surprisingly, biliverdin reductase then reduces biliverdin to the very insoluble bilirubin which then must be conjugated to glucuronide for excretion. Why? It is well known that accumulation of bilirubin in the brain is neurotoxic. Why would nature mAke bilirubin which puts so large a portion of newborn babies at risk for kernicterus? We found that bilirubin very potently prevents neurotoxicity by acting as an endogenous antioxidant. However, because bilirubin is toxic, the body cannot produce large amounts of it despite the need to combat high concentrations of oxygen free radicals. Instead, an ingenious cycle regenerates bilirubin. Thus, whenever a molecule of bilirubin acts as an antioxidant, it itself is oxidized back to biliverdin which is immediately converted by biliverdin reductase back to bilirubin. When we made these findings, we then explored the clinical literature and discovered protective effects of bilirubin that had not been appreciated because they didn’t make sense.” Individuals with Gilbert’s Syndrome, who are less capable of inactivating bilirubin and have moderately elevated levels, have only 1/5^th the incidence of cardiovascular disease of matched controls. In other studies screening large populations, individuals with higher bilirubin levels have a higher incidence of cancer or heart disease. This work is being carried out by Tom Sedlak , an M.D. Ph.D. psychiatrist doing postdoctoral training in our lab.

“You asked about my involvement with patients. I wanted to be a psychiatrist long before I had any interest in research. Indeed, I decided to be a psychiatrist while in high school and never wavered. I enjoyed dynamically oriented psychotherapy and, according to my supervisors, was pretty darned good. For many years I continued taking time out of my other activities to see patients a few hours a week.

You asked about connections between my clinical psychiatric experience and what I did in the laboratory. Once we identified opiate receptors, we were able to transfer this technology to studies of neurotransmitter receptors and focus with particular emphasis on dopamine receptors in order to address directly if neuroleptics might act by blocking dopamine receptors. Because of my clinical background, I was aware of critical questions dealing with side effects of neuroleptics, such as extrapyramidal actions which we showed to be correlated inversely with anticholinergic effects. The ability to quantify potencies of both therapeutic and adverse effects of drugs at various neurotransmitter receptors monitored by ligand binding led to a new way of drug development in the pharmaceutical industry.

You asked how advances in neurobiology will shape clinical psychiatry. The most direct and important impact will probably be in identifying genetic propensities for the major illnesses, schizophrenia and bipolar disorder. Already, one gene has been directly implicated in a small group of patients with familial schizophrenia. This gene, DISC1, has been shown by my colleague in the psychiatry and neuroscience departments, Dr. Akira Sawa, to regulate early brain development fitting with abundant evidence that schizophrenia is primarily a developmental rather than a neurodegenerative disorder. Once we find genes that mediate psychiatric disturbance, it might be possible to develop novel and more selective therapies.

You ask about the future of psychotherapy and allude to your discussions with Eric Kandel who suggested that imaging techniques might be employed to quantify therapeutic benefit. Eric has persuaded the Dana Foundation to sponsor research in this area. It would be interesting to compare effects of psychotherapy, drug treatment and the combination on all of these measures. As a mode of dealing with emotional distress, common sense tells us there will always be an important role for psychotherapy.

In the context of my own musical background you ask about the potential influence of music upon scientific research. I have long been impressed with the link between artistic creativity and scientific discovery. Writing a song, drawing a picture, producing a poem all involve thinking new thoughts. The more these thoughts are “out of the box,” the greater the creative advance. Similarly, the more novel a scientific concept, the greater chance that it represents a major step forward. In my own case, I love to write new songs – especially for my grandchildren – original melodies as well as words. Similarly, in going over scientific projects with my students, I become impatient with dry analysis, in depth, of experimental details. What really turns me on is coming up with a totally novel conceptualization that can clarify seemingly contradictory findings and provide insight into some important question.”

Born in 1938 in Washington , DC , Dr Snyder worked at NIH in the laboratory of Seymour Kety, MD while he was still in high school. He later worked with Julius Axelrod, PhD, who was awarded the Nobel Prize for Physiology or Medicine in 1970. A graduate of Georgetown School of Medicine, Snyder spent several more years working with Axelrod at NIH before starting a psychiatric residency at Hopkins in 1965. He became a full professor by l970 and in l980 he became Distinguished Service Professor of Neuroscience, Pharmacology and Psychiatry and director of the Department of Neuroscience. He gained fame in 1972 with the discovery of opiate receptors in the brain. Since then, he and his students have produced an uninterrupted series of blockbuster neurobiological discoveries. Over 150 of his former students occupy important scientific positions throughout the world.

Regarding his personal life, Dr Snyder said: “My wife, Elaine and I have three grandchildren, Abigail 8, Emily 6, and Leo 3. The grandchildren are from our older daughter Judy, a psychiatrist in Philadelphia . Our younger daughter Debbie is a screenwriter in Manhattan , unattached. I do play the classical guitar and seriously considered a concert career, having performed publicly a good bit while a high school student. Elaine completed the JHMI Mental Health Counselor program many years ago and is now retired after a productive career.”

March 2005, Dr. Solomon Snyder at the Award Ceremony with granddaughters Abigail Kastenberg, age 8, and Emily Kastenberg, age 6 -

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(1) http://www.acnp.org/Bulletin_March_Web_2005.htm

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Part II Part II contains a lot of details for which there was insufficient space in the published interview. It is a mostly unedited transcript of a recording of Snyder's detailed response when I asked him to talk about his lab's “advances that opened up the modern field of treatment of neuropsychiatric diseases” and led to his being awarded the Medal of Science in March 2005. Readers should understand that there are still some gaps and possibly some uncorrected errors in the text. This may not be copied or republished.

Some raw material from an interview with Solomon Snyder, MD by Gerald D. Klee, MD, for The Maryland Psychiatrist, June 15, 2005

“Snyder is undoubtedly one of the most original, prolific and influential intellectuals of our time.” Professor Pasko Rakic, MD, PhD of the American College of Neuropsychopharmacology http://www.acnp.org/Bulletin_March_Web_2005.htm

Dr. Klee asked Dr. Snyder to describe his work that earned the eulogy about him by Professor Pasko R ak ic, MD, PhD of the American College of Neuropsychopharmacology. See link http://www.acnp.org/Bulletin_March_Web_2005.htm

According to Professor Pasko R ak ic, MD, PhD of the American College of Neuropsychopharmacology, “Snyder is undoubtedly one of the most original, prolific and influential intellectuals of our time. Some of the major advances that opened up the modern field of treatment of neuropsychiatric diseases have stemmed from his work. His pioneering work has led to novel generations of therapeutic drugs and potential treatments for drug abuse. He published the initial paper identifying and characterizing opiate receptors; and, subsequently, established the binding sites of the pharmacologically relevant opiate receptors that are localized on discrete brain sites. His work contributed to understanding the major actions of the drugs and differentiating agonists and antagonists, permitting the development of novel, less addicting opiates. Snyder then identified receptors for the major neurotransmitters, establishing the mechanism of the anti-psychotic and side-effect actions of neuroleptics and differentiating serotonin receptor subtypes that led to new classes of anti-psychotic, anti-migraine and anti-emetic drugs. Snyder extended receptor strategies to intracellular signal transduction, identifying and purifying the receptors and showing that they contain the calcium ion channels that mediate their actions. The development of new agents in the pharmaceutical industry has been greatly enhanced by the now universal application of Snyder's techniques for rapid screening of large numbers of candidate drugs. For example, Snyder applied receptor techniques to clarify second messenger systems, especially the inositol 1,4,5-trisphosphate (IP3) receptors of the phosphoinositide cycle that regulate intracellular calcium deposition and the immunophilins that mediate numerous signal transduction events in the brain. He and his colleagues were involved in the discovery of the dopamine receptors and their role in schizophrenia. Snyder's more recent discovery of novel neurotransmitters such as nitric oxide, carbon monoxide and D-serine portend novel therapies in mental illness, stroke and sexual dysfunction.”

Snyder: It was generally assumed, on theoretical grounds that drugs acted on receptors, but the existence of receptors had not been demonstrated. In the 1970’s nobody had been able to monitor a receptor for a drug or neurotransmitter in a test tube, even though by that time biochemically you could measure the biosynthesis of neurotransmitters, their inactivation by reupt ak e and their metabolism and that sort of thing. The reason was that to measure a receptor you’d presumably monitor the binding of the radioactive drug or neurotransmitter to it but the numbers of receptors in the brain would be expected to be only about 1 millionth by weight of the brain and the radioactive drugs you would use to measure the binding in brain membranes could bind to all sorts of sites – proteins, lipids, carbohydrates and these non-specific binding sites would vastly exceed by tens of thousands the numbers of true physiologic receptors. It was very simple techniques of washing very extensively, but very rapidly so that the radioactive drug bound to the true receptor wouldn’t wash away where as non-specific binding would wash away and of course, utilizing radiolabeled drugs of very high affinity for the receptor so they would preferentially bind to the receptor over non-specific sites. Using those techniques, in 1973, we were able identify opiate receptors, learn a great deal about them, how many drugs interact. How for instance, codeine doesn’t even bind to the opiate receptor but must be converted to morphine where it acts. The same is true of heroin, which itself is technically not an opiate – it has to be converted to monoacetyl morphine before it acts and then of course, the reason that man wasn’t born with morphine in him – there must be some neurotransmitter that mimics morphine and that led to ourselves and others identifying opiate-like neurotransmitters. Jon Hughes and Hans Kosterlitz first isolated the enkephalins as the major endorphins and we soon thereafter found the same sorts of things. The fact that the opiate receptors were clearly receptors for neurotransmitters led us to try to use similar technology for monitoring other neurotransmitter receptors and by 1976-77 we had success with identified receptors for many of the major neurotransmitters in the brain. From the perspective of psychiatry, dopamine receptors were of particular interest. As far back as 1963, Arvid Carlsson had speculated that neuroleptics might act by blocking dopamine receptors, but he based the speculation on very limited indirect data specifically that administering to rats neuroleptic drugs changed the level of dopamine bre ak down products which suggested to him that the drugs blocked dopamine receptors and by a feedback mechanism caused the dopamine neuron to release more dopamine giving more metabolic products very, very highly speculative. In 1973 Paul Greengard found that neuroleptics block effects of dopamine on cyclic AMP formation but actually his data really didn’t even fit with the actions of the drugs. Finally, in 1976 when we were able to measure dopamine receptors by the binding of radioactive neuroleptics we could test the relative potencies of neuroleptics and their ability to block the binding sites correlated magnificently with their clinical efficacy. Those receptors we now know to be one subtype of dopamine receptor – the dopamine D2 receptor.

The neuroleptic drugs by competing for radioactive neuroleptics for binding to dopamine receptors gave us evidence that they interacted with the receptors and clearly they were blocking the actions of dopamine. The neuroleptics are dopamine receptor antagonists specifically at the dopamine D2 receptor. A number of things ______ into psychiatry came from similar binding studies, for instance neuroleptics are very sedating and they cause orthostatic hypotension. Their ability to do those things correlated perfectly with their blockade of alpha adrenergic receptors measured in our binding studies. The same held true for similar side effects of tricyclic antidepressants. So it was clear that just by monitoring receptors you could learn both about their therapeutic actions and side effect actions and in fact the technology we develop could be set up in a high-throughput model so that in the drug companies they could do easily 1,000 test tubes a day and screen drugs, in this case with their screening neuroleptics they’d be screening to find drugs that would be more potent in blocking dopamine receptors and were less potent in blocking alpha adrenergic receptors. About the same time in our lab we were able to monitor serotonin receptors. We found at least two distinct subtypes of serotonin receptors and this was of importance because up to that time people had difficulty pinning down the notion that there might exist multiple subtypes for receptors. It was certainly known by then that for adrenergic receptors you had separate alpha and beta receptors and that was elucidated on pharmacologic grounds to be able to do this at a receptor binding level turned out to be quite powerful. In the case of serotonin receptors with the various subtypes, my M.D. Ph.D. student Steve Peroutka was able to discriminate a variety of serotonin receptor subtypes and the work he did after he left our lab, he was able pin down the specific receptor subtype at which anti-migraine drugs like Imitrex act and those techniques of course permitted the drug industry to get all of the tryptan to screen for actions of the tryptan anti-migraine drugs.

Klee: I remember the excitement back in the 1950s over the interactions between serotonin and LSD. Some people jumped to the conclusion that they were about to discover the cause of schizophrenia.

Snyder: The question of how psychedelic drugs act is certainly closely linked to the history of serotonin. Serotonin as a chemical was only discovered in the mid-1950’s. The name serotonin was coined by Irvin Page at the Cleveland Clinic because he found it existed in blood elements in the platelets and so that’s were the sero comes from. The tonin which means contraction, came from the fact that it contracted smooth muscle. Serotonin was called 5Ht because chemically, it’s 5 hydroxytryptomine being generated from the amino acid tryptophane. Indeed a number of workers in the 1950’s found LSD to influence L-serotonin induced muscle contractions. I think Wooley at Rockefeller Institute was one of the key people. There were all sorts of speculation about how LSD acts and it turns out that those speculations, based on very indirect evidence, turned out to be more or less correct. Now there is an overwhelming body of evidence indicating that the major psychedelic drugs even amphetamine related agents, like ecstasy, act by mimicking serotonin at one subtype of serotonin receptor. In fact one of the biggest themes of all neurotransmitter receptor research nowadays is the existence of large numbers of subtypes of receptors. There are roughly 12 subtypes of serotonin receptors and their selective actions at one or another of these sites underlies the selectivity of actions of a number of drugs. The ability to do receptor binding, as we did including brain membranes, gave us the first ability to differentiate subtypes of neurotransmitter receptors at a molecular level and since the ability to clone the genes for neurotransmitter receptors followed by the human genome project, which we know every single gene enables us to have definitive evidence about the exact number of receptor subtypes for every neurotransmitter which has kept the drug companies pretty busy. It has enabled us to have pretty definitive evidence about how drugs act at subtypes of serotonin receptors.

Trying to translate receptor research into diseases is tricky because we don’t know the molecular cause of any major psychiatric illness.

Our best way of understanding molecular underpinnings of major psychiatric disorders at the present time involves learning the mechanism of action of drugs that are therapeutic in these conditions. Hopefully, it won’t be many years before molecular genetic studies underway will pin down definitively genes that are responsible for major mental illnesses. Right now, we know for instance that dopamine is involved somehow – maybe secondarily, maybe primarily in the pathophysiology of schizophrenia because its clear antipsychotic actions of neuroleptics stem, at least in major part, from blocking dopamine receptors and augmenting synaptic dopamine with drugs like cocaine, amphetamine selectively worsens schizophrenic symptoms. In the case of serotonin in schizophrenia, one of the strongest lines of evidence comes from the actions of drugs like clozapine, the atypical neuroleptics. We don’t know what is the magic about why clozapine is such a superb drug and why the other atypical neuroleptic seem to be very good drugs though none of them quite stand up to clozapine in terms of pure therapeutic efficacy. Clozapine is a fascinating drug because it is very potent in blocking a large number of neurotransmitter receptors. Very, very potent antihistamine. It is also quite potent in blocking serotonin 5HT2 receptors and many people have been speculating that the relative balance of its actions on 5HT2 receptors and dopamine D2 receptors determines the therapeutic actions of atypical neuroleptics like clozapine with the actions at the serotonin receptor, preferentially being somewhat more potent than the actions at the dopamine receptor. This theory hasn’t yet been definitively established but of course receptor binding techniques have enabled the drug companies to screen for agents that will act with reasonable selectivity and with the right degree of balance at these different receptor subtypes.

The main evidence we have for a role of serotonin in schizophrenia, as far as I can see is really just the action of neuroleptics, especially atypical neuroleptics at serotonin receptors. LSD was originally proposed to mimic schizophrenic systems but most people would say that the psychosis with LSD doesn’t mimic schizophrenic symptoms nearly so well as psychotic symptoms elicited by PCP and related drugs or even amphetamine and cocaine. Historically after people became disillusioned with the notion that psychedelics provide a model of schizophrenia, there was a lot of focus on amphetamines and cocaine mimicking especially acute paranoid schizophrenia as well as the ability of these drugs in low doses to very selectively exacerbate schizophrenic symptoms. If you couple those data with the fact that those drugs are acting by releasing dopamine or facilitating its synaptic action. If we combine the fact that amphetamines and cocaine worsen schizophrenic symptoms and in high doses can elicit a drug psychosis resembling acute schizophrenia and the fact that these drugs we know act by increasing synaptic actions of dopamine. If we couple those facts with the fact that the antipsychotic actions of neuroleptics block dopamine receptors we have a stronger case for an important role of dopamine in schizophrenia than we have for serotinin but both might be certainly involved and in terms of the pathophysiology of schizophrenia. Today, there is a strong body of work implicating that glutamate acting at a subtype receptor called NMDA receptor and this way of thinking, the glutamate NMDA receptor hypothesis is related in major part to the fact that many people feel that the psychosis elicited by PCP more faithfully mimics schizophrenic state than any other drug psychosis and the fact that we know that the molecular action of PCP and related drugs is to block the NMDA subtype of glutamate receptor. Additionally, based on that line of thinking a number of investigators have looked at the predictions of that model. The predictions of the model that psychotomimetic actions come from blocking NMDA receptors would be that stimulation of NMDA receptors will alleviate schizophrenic symptoms. Joe Coyle, my very first student in my lab who began working with me as a second year Hopkins medical student, and his colleagues have done clinical studies showing that if you give agents that stimulate NMDA receptors this alleviates schizophrenic symptoms. They don’t give glutamate, what they give is D-serine or D-cycloserine or glycine and this brings us to an interesting story about glutamate NMDA receptors and how they are regulated which bears on a lot of recent research in our own laboratory. Let me tell you about that research.

The NMDA receptor is sort of unique among neurotransmitter receptors in that it is the only one in which it t ak es two neurotransmitters to stimulate it. This discovery was made by the French neuropharmacologist Phillipe Ashair in 1989 when he was monitoring glutamate neurotransmission by NMDA receptors in brain preparations that were being perfused. When he speeded up the perfusion he noticed no more NMDA neurotransmission and figured he had washed away something important. When he added back amino acids he found that the amino acid glycine could restore the responses to glutamate so he said we need two neurotransmitters at the NMDA receptor – glutamate and glycine. His reasoning for why this might be done by nature to m ak e such a unique synapse where you need two neurotransmitters was based on the fact that excess release of glutamate acting at NMDA receptors is very toxic. Glutamate is an excitatory neurotransmitter - too much of it literally excites neurons to death. We know that during stroke, for instance in a rat, cut off the middle cerebral artery you get a 50-fold increase in glutamate release and that glutamate acting at NMDA receptors is a major cause of stroke damage because drugs that block NMDA receptors alleviate that stroke damage and NMDA antagonists have been studied extensively in stroke and Alzheimer’s Disease in which the destruction of neurons seems to be related to excess glutamate. In fact, there is a NMDA antagonist on the market for treatment of Alzheimer’s Disease today, Memantine. Because excess glutamate is what we call excitotoxic and because glutamate is a dietary constituent. Dr. Ashair reasoned as follows: You eat a ste ak dinner you might get a stroke. Therefore nature had to have a failsafe mechanism. It t ak es two keys to undo the lock. You need glutamate and glycine. The trouble with that line of reasoning is that glycine is also a dietary amino acid so the ste ak dinner – you have too much glycine and too much glutamate to excite the NMDA receptor. The solution to this riddle came up some years ago when we noticed an article by some Japanese workers that decides L-amino acids which previously had been thought to be the only form of any kind of amino acids. Our bodies are stereospecific, we only have D-sugars – like D-glucose and we only have L-amino acids which enables the efficiency of our bodies biochemical systems to t ak e place. Now, these Japanese workers discovered that the body contains D-serine and a little bit D-aspartate. Those are the only two D-immuno acids. In our laboratory, my student Michael Schell became fascinated by this and developed an antibody selective for D-serine and showed that it occurs selectively in parts of the brain that are loaded with NMDA receptors. Whereas glycine has very different localizations. Moreover, there was already literature that D-serine is much more potent, 10-100 times more potent then glycine at the so-called glycine site of the NMDA receptor. We began exploring the possibility that D-serine might be a novel neurotransmitter that is the co-agonist with glutamate at the NMDA receptors. We tested this by treating brain preparations with a purified enzyme called D-amino acid oxidase. D-amino acid oxidase had been discovered by Hans Krebs, father of the Kreb Cycle, in 1935 as an enzyme that selectively degraded D-amino acids and it seemed sort of silly, since at that time there were no known D-amino acids. We purified and examined D-amino acid oxidase and found that at physiologic Ph it in fact only degrades D-serine. So, we added it to brain preparations, we destroyed D-serine and nothing else and NMDA neurotransmission vanished. We decided this is a very important system. We found the enzyme that conversed L-serine to D-serine. We localized that enzyme, which we called serine racemase and D-serine and not only are they both exactly where NMDA receptors are but more remarkably their not in neurons at all – their in glia – supporting cells that ensheath the synapse right around the area of NMDA receptors astrocytes. Now we know that what actually happens is when a neuron fires, it releases glutamate. That glutamate, some of it, goes directly to the NMDA receptor and some of it goes to the astrocyte that’s ensheathing the synapse and triggers the release of D-serine which then joins glutamate at the NDMA receptor. Our ability to now monitor and measure serine racemase enables us to try now to look for drugs that will selectively increase the formation of D-serine so that we could get even better drugs to alleviate schizophrenic symptoms in this fashion – far better than one could do by just administering D-serine itself which did alleviate schizophrenic symptoms but had to be given in enormous doses to penetrate the blood-brain barrier.

Nitric oxide is a fascinating neurotransmitter. One of the most recently discovered neurotransmitters, nitric oxide was discovered in the late 1980’s by several investigators who show that it accounts for the ability of blood vessels to relax. Physiologic relaxation of blood vessels is primarily mediated by nitric oxide. When we read about that work in the 1980’s and the fact that nitric oxide is a gas, very unprecedented for any kind of biologic molecule, we decided to look in the brain. David Bredt, an extraordinary M.D. Ph.D. student of mine discovered that nitric oxide is actually responsible for many actions of glutamate acting through NMDA receptors. When you activate NMDA receptors you admit calcium into the cell and that calcium binds to calmodulin which directly activates the enzyme that m ak es nitric oxide. This enzyme, called nitric oxide synthase or NOS, t ak es the amino acid arginine which has a guanidino group and bre ak s off and oxidizes that nitrogen into nitric oxide and some of the actions of glutamate NMDA receptors involve nitric oxide. Others do not. The exact functions of nitric oxide – we don’t know all of them. We know some of them because one of the things that David Bredt did in our laboratory was identified the enzyme that m ak es nitric oxide – nitric oxide synthase or NOS, purified it to homogeneity, cloned the gene for it and was able to characterize it in very great detail.

(1) http://www.acnp.org/Bulletin_March_Web_2005.htm