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Mashimo Lecture
Regulatory Mechanisms of Synaptic Transmission in the Neural Network
Shigetada Nakanishi
Kyoto University, Kyoto, Japan
 
  • The Basal Ganglia Neurocircuit
  • Drug Addiction and Brain Function
  • Paraneoplastic Cerebellar Ataxia Due to Autoantibody Against a Glutamate Receptor


  • Neurotransmission systems in the brain are one focus of the basic science research of Nakanishi and colleagues. This lecture reviews the types of receptors in the brain, one aspect of the development of the glutamate receptors, and some clinical studies.

    Neurotransmission occurs via neurotransmitters; from the terminus of the nerve the neurotransmitter is released and interacts with the receptors to cause various functions.
    The brain has more than 50 neurotransmitters or substances modulating transmission, and these may be excitatory, inhibitory, or modifying transmitters. Additionally, there are various receptors for the neurotransmitters.

    To understand the function of the brain, they elucidated the receptors. About 15 years ago, they were able to clone different receptors using a method they developed that combined genetic engineering and electrophysiology to isolate receptors.

    Glutamate receptors were cloned by this group to clarify their fundamental function. At least two types of glutamate receptors exist: ionotropic and metabotropic. The ionotropic glutamate receptor has an ion channel that opens when the glutamate response is seen to allow the ion to flow in and quick excitation of the nerve to occur.

    The ionotropic receptor has an AMPA, kinate, and NMDA receptor. The NMDA type is important for memory and learning, and the AMPA receptor for very rapid excitation of the nerve. All of the receptors have G proteins, so depending on the condition of the glutamate, a metabolic-type receptor, metabotropic glutamate receptor (mGluR), is present. Their work to clone the mGluR led to the understanding there are different types of receptors. They have cloned the NMDA receptor and the metabotropic receptor.

    The glutamate receptors comprise more than 20 genes, and as expected G protein coupled-type ion channels are present. Presently they are investigating the role of these receptors in various sites of the brain and how they act on specific neurons. There are 8 types of G protein-coupled receptors, which are divided into 3 groupings based on functionality.





    The Basal Ganglia Neurocircuit


    To understand one aspect of the brain, they are researching the basal ganglia neurocircuit. The basal ganglia, comprised by the striatum and nucleus accumbens, is a key site for human disease. For example, Parkinson's disease is due to a reduced level of dopamine.
    Basically, in this circuit the glutamate transmission from the cerebral cortex is received by the basal ganglia. A control mechanism feeds the information back to the cerebral cortex to control motor movement. Under normal conditions, there is always a positive and negative control of this feedback to control motor movement, a direct and indirect pathway. In this positive and negative pathway, dopamine positively controls movement; a dopamine deficiency, as seen in Parkinson’s disease, results in a decline in motor movement.

    Acetylcholine, produced by about 1% of the cells in the basal ganglia, is a physiologic antagonistic transmitter against dopamine. To study the role and function of acetylcholine, which is not well understood, they used cell targeting to create a transgenic mouse expressing human interleukin-2 receptor (IL-2R). They injected into the transgenic mouse an anti-IL-2R antibody fused to pseudomonas toxin, an immunotoxin. The antibody reacted to the human IL-2R, was taken into the complex, and the pseudomonas toxin killed the cell. In this model, the human IL-2R responds, but the mouse IL-2R does not, and thus the neurons expressing human IL-2R are eliminated specifically.

    Then, they generated transgenic mice expressing IL-2R/GFP under the control of the mGluR-2 promoter. By using GFP, they showed that this model specifically expresses human IL-2R in the ACH-producing cells (choline acetyl transferase; ChAT). Then, using immunotoxin-mediated cell targeting, they selectively eliminated the ChAT cells.

    After the selective elimination of the ChAT cells, the ACh levels are selectively reduced; the dopamine level is not affected. Further, the elimination of the ChAT cells results in changes in movement. Specifically, contralateral rotation towards the site where the cells were eliminated occurs in about 3 days, a finding that was somewhat expected. ACh acts antagonistically against dopamine, and the removal of dopamine results in ipsilateral turning in the mice. Interestingly, after the acute phase, movement returns to normal. Brain function is somehow compensated and results in recovery.

    Why did contralateral abnormal rotation occur in the acute phase and recovery in the chronic phase? In a subsequent experiment, they showed that a decrease in ACh results in dopamine becoming relatively more dominant; the positive effect of dopamine became stronger and the negative effect weaker. Thus, in this setting, the stimulus to the cerebral cortex becomes stronger. The information from the basal ganglia acted stronger on the eliminated site.

    In the chronic phase, downregulation occurs at the site of the relatively stronger action of dopamine. As a result, dopamine becomes stronger because there is no ACh. But, there is a decrease in the number of receptors and thus the left and the right sides maintain a certain balance; compensation occurs.

    To treat Parkinson's disease dopamine is administered. In some cases there is simultaneous administration of an ACh antagonist. But, administering an ACh antagonist in the chronic phase is tantamount to reducing ACh. Therefore, there is some amount of compensation, perhaps resulting in some amount of imbalance remaining overall. Their work supports the fact that a change in one site results in a reaction in another site.

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    Drug Addiction and Brain Function


    In the mesolimbic dopaminergic pathway of ACh, dopamine and ACh are antagonistic. The nucleus accumbens, found in the anterior portion of the basal ganglia, is closely and critically related to drug addiction. Globally 30 million people are addicted to substances.

    Cocaine and morphine addiction is closely related to brain function. The role of ACh in persons with drug addiction can be studied because the nucleus accumbens is present in both the mesolimbic dopaminergic pathway and the basal ganglia neurotransmitting pathway.

    Cocaine blocks the dopamine transporter, so the dopamine level at the nucleus accumbens is potentiated on a longer-term basis. But the mechanism for this has not been elucidated. Morphine also elevates the dopamine level, because of the effect on the nucleus accumbens, and results in chronic poisoning or intoxication from morphine. Because dopamine and ACh are antagonists, they presume that an ACh antagonist will suppress the excessive dopamine.

    This group developed an experimental animal lacking ACh-producing cells in the nucleus accumbens, because cocaine works on and binds strongly to the nucleus accumbens, to study the action of cocaine and morphine in this model. Other work had shown that the ACh antagonist and agonist work promiscuously. Thus the role of ACh is not clearly understood. In their work, they specifically eliminated ACh-producing cells only at the nucleus accumbens.

    Cocaine was strongly potentiated in this mouse model. To study this further, they used the conditioned place preference test. In this test, a saline-injected animal is placed in one chamber and a drug-injected animal in another chamber. Each chamber is given a different appearance and colors. The mice are conditioned to prefer one room over to the other, because when cocaine or morphine is given, the animal has a sense of pleasure and comfort.

    Is it possible to suppress the effects of cocaine and morphine if the animal can preserve ACh production, despite lacking ACh-producing cells? Various pharmacologic drugs have been developed to potentiate ACh production. For example, to suppress the progression of Alzheimer's disease, an acetylcholinesterase inhibitor was developed, which suppresses degradation of ACh in the brain. Two drugs that are being clinically are donepezil and galanthamine.

    In the animal model, pre-treatment with donepezil completely suppresses the action of cocaine. Further work showed that the presence of ACh at the nucleus accumbens is very important to suppress the action of morphine and cocaine.

    Typically, the continued use of cocaine results in continued hyperactivity. But, even after the action of cocaine is established, pre-treatment with donepezil suppresses this action in their animal model. Although the safety of donepezil in humans has already been established, whether it can be used for drug addiction requires further research.

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    Paraneoplastic Cerebellar Ataxia Due to Autoantibody Against a Glutamate Receptor


    In work in paraneoplastic cerebellar ataxia, this group found that with some neoplasms there is some brain disorder. Although this is known to be some sort of autoimmune disease, it is not understood.

    The autoantibody production may work on the cerebellum and cause motor disorder, specifically gait disorder in some patients. The presence of a specific anti-MgluR1 antibody has been shown in some patients, causing cerebellar ataxia and memory and motor disorders.

    This group showed that in knockout mice, immunohistochemical staining strongly suggested that a specific antibody against mGluR-1. Further study showed that although mGluR1 and mGluR5 are similar, immunoreactivity was only observed for mGluR1, but not mGluR5. Inositol phosphate 3 production was reduced also by this antibody. This antibody suppresses the action of the mGluR1 and its receptors.

    To produce cerebellar ataxia in the mouse model, antibody serum was injected into mouse cerebellum. This injection produced gait staggering in the mice and convulsion-like symptoms. A rota-rod test, using a rotating bar, was used to quantify the motor disorder.  Normally the mouse can stay on the bar while it rotates. Injection of serum from a patient with Hodgkin’s disease and cerebellar ataxia into the mice cerebella suppressed mGluR1 function and motor ataxia resulted. The mice fell off the rotating rod. With time, the function returned to normal.

    So, although the exact mechanism is not understood, after the onset of Hodgkin's disease, an mGluR-1 antibody was produced in the two patients studied. Its high level of expression in the cerebellum caused ataxia. When the serum is given to mice, similar symptoms can be reproduced. Therefore, paraneoplastic cerebellar ataxia is partly caused by selective autoantibody production against mGluR1. The metabotropic receptors are important for the function and regulation of the cerebellum.

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