Epilepsy

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NEUROTRANSMITTERS

• M.Prasad Naidu
• MSc Medical Biochemistry,
• Ph.D.Research Scholar

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• Definition - The chemical substance helpful for
  signal transmission in central nervous system
  peripheral nervous system (via) the chemical
  synapses is neurotransmitters.
• Synaptic transmission is the predominant means by
  which neurons communicate with each other.

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• The criteria for Chemical neurotransmitter
• 1) found in presynaptic axon terminal.
• 2) enzymes necessary for synthesis are present in
  presynaptic neuron .
• 3) stimulation under physiological conditions
  results in release.
• 4) mechanism exist for rapid termination of
  action.
• 5) direct application to postsynaptic terminal
  mimics the activation of nerve stimulus.

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• 6) drugs that modify metabolism of the
  neurotransmitter should have predictable
  physiological effects invivo assuming that the
  drug is transported to the site where
  neurotransmitter acts.
• Not all neuron to neuron transmission is by
  neurotransmitters , gap junctions provides direct
  neuron to neuron electrical conduction.

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• Neurotransmitter is stored in synaptic vesicle
  released in response to nerve impulse controled
  by calcium influx.
• Release of neurotransmitter is quantal event ,
  that is a nerve impulse reaching presynaptic
  terminal result in release of transmitter from a
  fixed number of synaptic vesicle.
• Neurotransmitter action is terminated by
  metabolic degradation , reuptake , or diffusion
  into other cell types.

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• Class - 1 acetylcholine
• Class -2 The biogenic amines
• norepinephrine , epinephrine
• dopamine , serotonin .
• Class - 3 amino acids
• gamma amino butyric acid (GABA) ,
• glycine , glutamate , aspartate.
• Class - 4 nitric acid (NO)
• carbonmonoxide ( co )

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• In addition to classical neurotransmitters many
  neuropetides are identified as definite or
  probable neurotransmitters,
• eg - substance p , neurotensin ,
  enkephalin , ß endorphin , histamine,
• vasoactive intestinal polypeptide,
• cholecystokinin , neuropeptide Y
• somatostatin.

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• Neurotransmitters modulate the function of post
  synaptic cells by binding to specific receptors
  of 2 types
• 1) ionotropic receptors ( direct ion channels
  that open after binding of neurotransmitters. )
• 2) metabotropic receptors ( interact with G
  proteins stimulating production of second
  messengers activating protein kinases , which
  modulate the cellular events. )

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• G proteins couple several receptors to intra
  cellular signaling system , linking neuronal
  excitability to energy metabolism second
  messenger systems.
• G protein binding receptors include adenosine ,
  Ach ( muscarnic ), norepinephrine , dopamine ,
  serotonin

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• Kinetics of ionotropic receptors are fast , (lt 1
  ms ) , because neurotransmitters directly alter
  the electrical property of the postsynaptic cell.
• Kinetics of metabotropic receptors functions over
  longer time periods.
• This contributes to the potential for
  selective finely modulated signaling by
  neurotransmitters

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• The membrane of neuronal cell maintains an
  asymmetry of inside outside voltage , is
  electrically excitable.
• Neuronal membranes are polarized to a potential
  of -90 mV by the activity of Na_k ATPase
  transport system.

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• Factors that control the neuroexcitability
• 1)voltage gated ion channels
• 2) neurotransmitter activated ion
  channels.
• 3)neuromodulators
• 4)second messenger system.
• The control of neuronal activity within normal
  limits is by the modulation of excitatory
  inhibotory events simultaneously.

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• Ligand gated channels are responsible for
  communication between cells.
• Voltage gated sodium channels are involved in
  propagation of action potential , rapid
  activation is at -60mV due to opening of fast
  transient channels.
• 
  
  
  Voltage gated
  potassium channels contribute to repolarization
  ,this regulate repeated firing of action
  potential by prolonging after spike
  repolarization.

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• Voltage dependent calcium channels trigger
  neurotransmitter release , at rapid activation is
  around -70mV.
• Autoantibodies to ca channels in motor nerve
  terminal leads to decreased release of Ach from
  nerve terminal , this is seen in eaton lambert
  myasthenic syndrome.
• Voltage gated channels determine how inhibitory
  excitatory influences are integrated .

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Acetyl choline

• Acetyl choline is the neurotransmitter used by
  all motor axons that arise from spinal cord, that
  is at neuromuscular junction.
• Junction consist of a single nerve terminal
  separated from post synaptic region by synaptic
  cleft.
• Motor end plate is the specialized portion of the
  muscle membrane involved in the junction.

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• Junctional folds are prominent they contain high
  density of Ach receptors.
• Synthesis of Ach takes place in cytosol of nerve
  terminal .
• choline acetyl
  transferase
• acetyl coA choline Ach coA
• Ach is incorporated into membrane bound particle
  called synaptic vesicles.
• Assembly of synaptic vesicle with cell membrane
  resembles assembly of transport vesicle involving
  SNAREs.

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• Release of Ach into synaptic cleft occurs by
  exocytosis , which involves fusion of vesicle
  with presynaptic membrane.
• Nerve ending is depolarized by transmission of
  nerve impulse this opens the voltage gated Ca
  channels , permitting influx of Ca from
  synaptic cleft to nerve terminal , this Ca
  plays a role in exocytosis of Ach vesicle.

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• Approximately 200 vesicles are released into
  synaptic space.
• Each vesicle contains 10000 molecules of Ach.
• Ach binds Ach receptor , receptor undergoes
  conformational change opening the channel in the
  receptor that allows entry of Na, k resulting
  in depolarization of muscle membrane.

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• Properties of Ach receptor of NMJ
• nicotinic receptor (nicotine is an agonist
  for the receptor)
• a membrane glycoprotein containing 5 subunits.
  ( 2aß?d subunits).
• only a subunit binds Ach with high affinity.
• 2 molecules of Ach binds receptor to open the
  ion channel which permits Na , K the receptor
  is thus transmitter gated ion channel.
• autoantibodies to receptors are implicated in
  causation of myasthenia gravis

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• Snake venom a bungarotoxin binds tightly to the a
  subunit can used to label the receptor .
• Formation of autoantibodies to Ach
  receptors in NMJ
• damage to receptors by autoantibodies
• reduction in number of receptors
• Episodic weekness of muscles supplied by cranial
  nerves

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• When the channel closes Ach dissociates it is
  hydrolyzed by acetyl choline esterase.
• acetyl choline esterase
• Ach H2O Acetate
  choline
• Choline is recycled into nerve terminal by active
  transport , it can be used for synthesis of Ach.

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• The classical neurotransmitter of autonomic
  ganglia whether sympathetic or parasympathic is
  acetyl choline.
• 2 classes of receptors are present in autonomic
  nervous system.
• 1) nicotinic eceptors ,
• 2) muscarnic recptors.
• Nicotinic receptors in autonomic ganglia are
  different from those on skeletal muscle.

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• Nicotinc muscarnic receptors mediate excitatory
  postsynaptic potentials (EPSP) , but these
  potential have different time course.
• Stimulation of presynaptic neuron elicits a fast
  EPSP followed by a slow EPSP.
• Fast EPSP results from activation of nicotinic
  receptors which cause of ion channels to open.

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• Slow EPSP is mediated by activation of muscarnic
  receptors that inhibit the M current , a current
  that is produced by K conductance.
• Besides acetyl choline sympathetic preganglion
  neurons may release enkephalin , substance p ,
  LHRH , neurotensin or somatostatin.

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• Neurotransmitter in parasympathetic
  postganglionic neurons is acetyl choline.
• Actions are mediated by 3 types of muscarnic
  receptors.
• 1) M1 receptor (neural ) produces slow excitation
  of ganglia.
• 2) M2 receptor (cardiac) activation slows the
  heart.
• 3) M3 receptor (glandular) , causing secretion,
  contraction of visceral smooth muscle , vascular
  relaxation.

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• Muscarnic Ach receptors act by way of inosine
  triphosphate system they may also inhibit
  adenyl cyclase thus decreasing cAMP synthesis.
• Muscarnic recptors also open or close ion
  channels particularly K or Ca this action
  occurs through G proteins.
• Muscarnic receptors relax smooth muscle by an
  effect on endothelial cells which produces nitric
  oxide (NO) .

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• Nitric oxide ( NO ) relaxes smooth muscles by
  stimulating guanylate cyclase there by
  increasing levels of cGMP which in turn
  activates cGMP dependent protein kinases.
• The number of muscarnic receptors are regulated
  exposure to muscarnic agonist decreases the
  number of receptors by internalization of
  rceptor.

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• The betz cells of motor cortex uses acetyl
  choline as their neurotransmitter.
• Acetyl choline probably acts as an imporatant
  neurotransmitter in basal ganglia which is
  involved in control of movements.
• Deficits in cholinergic path way in the brain
  implicated in some form of Alzheimer's disease.

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• GABA major fast inhibitory neurotransmitter in
  the fore brain. 30 synapses of C.N.S contain
  GABA.
• Glutamic acid dehydrogenase synthesizes
• GABA from glutamate in nerve terminal .
• 3 types of receptors GABA a
• GABA b
• GABA c

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• GABA a GABA c are ionotropic receptors are
  post synaptic linked to chloride channel.
• GABA b receptors are metabotropic may be pre or
  post synaptic are coupled to ca or k ion
  channels via GTP proteins.
• Presynaptic GABA b receptors serve autoreceptors
  to inhibit release from nerve terminal.

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• Binding of GABA leads to an opening of chloride
  channels resultant hyperpolarization.
• Glycine is inhibitory neurotransmitter in brain
  stem spinal cord.
• Post synaptic receptor for glycine is ligand
  gated chloride channel that allows influx of Cl-
  to hyperpolarize the postsynaptic neuron

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• Glutamate aspartate are excitatory
  neurotransmitters.
• Glutamate is responsible for 75 of excitatory
  neurotransmission in brain.
• Synthesis of glutamate aspartate within central
  neuron glial cells is from carbohydrates
  involved in TCA cycle.

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• Mitochondrial enzyme aspartate transaminase
  interconverts glutamate aspartate.
• Glia contains glutamine synthase which converts
  glutamate to glutamine.
• Glutamine is subsequently transferred to neuron
  where it is deaminated to glutamate by
  glutaminase.

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• Glial inactivation specific uptake systems for
  glutamate reduces interstitial glutamate levels
  to terminate neurotransmitter action prevent
  excitotoxic damage.
• Monosodium glutamate produces migrainous head
  ache.
• Excessive glutamate can result in neurotoxicity ,
  celldeath neurodegeration seen alzheimers
  disease.

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• The receptors are subdivided into 5 classes.
• 1 )NMDA (N methyl D aspartate )
• 2 )AMPA (a amino 3 hydroxy 5 methyl 4 isoxazole
  propionic acid )
• 3 )The kainate recptor ( isolated from sea weed)
• 4 )L AP 4 ( synthetic agonist )
• 5 )Metabotropic receptors.
• First four receptors are cation channels .

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• Metebotropic receptors are linked to
  intracellular production of diacylglycerol,
  inositol triphosphate by phosphoinositide path
  way.
• NMDA is receptor is complex contains 5 distinct
  sites for binding
• 1 ) site for transmitter binding glutamate
• 2 ) a regulatory site that binds glycine.
• 3 ) a voltage dependent Mg binding site
• 4 ) a site that binds phencyclidine
• 5 ) a site that binds Zn.

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• NMDA receptor opens when glutamate binds allows
  influx of Ca Na into the cell.
• Mg , zn , poly amines , steroids can also
  modulate NMDA.
• one of the most important controls on the ionic
  conductance through the NMDA receptor is voltage
  sensitive blocking by Mg

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• Activation of AMPA receptor channels may
  depolarize the neuron sufficiently to remove the
  voltage dependent Mg block activate NMDA
  channels.
• AMPA NMDA are co activated are present on the
  same part of the neuron.

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• A separate site that modulates the gating of NMDA
  channel binds polyamines such as spermine
  spermidine which are synthesized by
  neurons.different concentration dependent effects
  have observed.
• Endogenous Zn reduces NMDA activated current.
• Zinc is present in high concentrations in the
  hippocampus released with some neurotransmitter
  in nervous system.

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• Hydrogen ions also modulate the ion conductance
  which is maximal at slightly alkaline pH ,
  reduces with increasing acidity.
• During hypoxic ischemic injury , progressive
  acidification resulting from glycolytic
  metabolism , may turn off the NMDA receptor
  channel.

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• AMPA receptor is coupled to both Na , K
  channels. its activation opens the above
  channels, depolarizes the neuronal cell rapidly,
  it is responsible for the majority of rapid
  excitatory neurotransmission.
• Kainate receptor is also coupled to Na , k
  channels.
• Kainate receptor has slower rate of depolarizing
  capacity than AMPA receptor.

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• Excitatory aminoacids are also able to interact
  with metabotropic recptors that activate the
  second messenger system, these receptors are
  found both pre post synaptically.
• Activation result in presynaptic inhibition
  post synaptic excitation.

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• The spectrum of neurological disorders mediated
  by excitotoxicity include epilepsy, stroke ,
  neurodegerative disorders ( parkinsons disease ,
  amyotropic lateral sclerosis , AIDS dementia )
• Most strokes are caused by thromboembolic events
  causing diminished perfusion resulting in
  reduction of supply of oxygen glucose.

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• 3 subsequent stages are there in the development
  of brain damage caused by ischemia.
• 1)induction ,
• 2)amplification ,
• 3)expression.
• Induction ischemia causes depolarization of the
  neuronal membrane leading to release of
  glutamate.

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• Glutamate overexcites the NMDA receptors in
  adjacent neuron , leading to abnormally large
  influxes of Ca Na and resultant cell injury
  or death .
• In addition glutamate stimulate AMPA kainate
  receptor ( leading to additional influx of Na )
  also metabotropic receptors , causing the
  release of ITP diacylgycerol.

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• Amplification further build up of intra
  cellular calcium occurs by following mechanism ,
• 1) increased intracellular Na activates Na -
  Ca transporters.
• 2)voltage gated Ca channels are activated by
  depolarozation.
• 3) ITP release Ca into cytosol from within
  endoplasmic reticulum.

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• Expression high levels of intra cellular Ca
  activates Ca dependent nucleases , proteases ,
  phospholipases.
• Degradation of phospholipids
• formation of platelet activating factor
  (PAF) release of arachidonic acid
• eicosanoids
  (vasoconstriction)
• damage by oxygen free radicals
• This is called glutamate cascade.

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• Huntington disease characterized by selective
  neuronal death in corpus striatum glial
  proliferation .
• Apoptosis , protein aggregation , excitotoxins
  may all contribute cell death in huntington
  disease.
• Excitotoxicity is by glutamate cascade.

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• The dopaminergic neurons are found in
  nigrostrital , mesolimbic , mesocortical
  tuberohypophysial systems.
• Dopamine synthesis occurs from tyrosine ,
  tyrosine hydroxylase is rate limiting enzyme in
  formation.

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• Entry of dopamine into synaptic vesicle is is
  driven by pH gradient established by a protein in
  vesicular membrane that pumps protons into
  vesicle at the expense of ATP
• Release of dopamine involves exocytosis.
• Dopamine has 5 post synaptic recptors
• D 1 receptor family(D1 D5)
• D 2 receptor family(D2, D3, D4)
• D4 receptor exhibits 5 polymorphic variants.

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• The effect of dopamine is to increase direct path
  way by D 1 recptor, supress indirect path way
  by D 2 receptor.
• D 1 receptor activation augments adenylate
  cyclase ( linked to stimulatory G protein).
• D 2 receptor activation decreases the activity of
  adenylate cyclase ( linked to inhibitory G
  protein ).

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• ATP dependent reuptake of dopamine achieved by a
  high affinity transporter in presynatic membrane
  , this is incorporated into vesicles reused
  again.
• Degradation of dopamine occurs within synaptic
  cleft or following reuptake ,within presynatic
  terminal.
• Mono amino oxidase B present in the outer
  membrane of mitochondria also in synaptic
  cleft.

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• MAO B MAO A are distinguished from each
  other by preference for different substrates
  by their different susceptibility to various
  inhibitors.
• Both the above enzymes acts on dopamine to
  produce 3 hydroxyphenyl acetaldehyde (DOPAC).
• DOPAC converted to homovanillic acid by the
  action of catechol o methyl transferase.

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• parkinson disease is due to loss of dopaminergic
  activity excessive cholinergic activity in
  basal ganglia.
• Signs of parkinson disease reflects a deficiency
  of dopamine in the substantia nigra , corpus
  striatum ( caudate nucleus putamen )

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• Basal ganglia are important for motor control
  they include putamen
• caudate
  nucleus
• globus pallidum
• substantia nigra
• subthalamic nucleus
  .
• All circuits in basal ganglia are inhibitory
  utilizing GABA except glutamatergic subthalamic
  input to globus pallidum internum(GPi) which is
  excitatory.

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• Cell damage in parkinson disease reflect a
  process of ageing , 13 of cells of substantia
  nigra are lost per decade from 25 age onwards . (
  parkinson disease rarely occurs befor 40 years )
• Mutattions in gene encoding a synuclein , a
  presynaptic protein involved in neuronal
  plasticity is associated with parkinson disease.
• Lewy bodies are found strongly stained with
  antibodies of a synuclein .

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• Signs of parkinson disease appear when the level
  of dopamine is droped in nigrosriatal system by
  80.
• Exposure to high levels of Manganese
• ( miners) leads to parkinson disease.
• Reserpine inhibit dopamine storage many
  neuroleptics block dopamine receptors.

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• Schizophrenia is a manifestation of
  hyperdopaminergia .
• Measurement of dopamine metabolite homovanillic
  acid in CSF is high in schizophrenics.
• Level of D2 receptors appears to be increased in
  the brains of schizophrenics.
• Dopamine mimetic drugs ( L dopa) induces
  schizophrenia

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• Low dopamine activity in prefrontal cortex of the
  brain of schizophrenics correlate well with the
  negative symptoms .
• Low dopamine activity in prefrontal cortex
  releases the inhibitory action on subcortical
  dopamine neurons resulting in elevated
  dopaminergic activity.

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• Adrenergic neurotransmission is by norepinephrine
  epinephrine.
• The adrenergic neurons of locus ceruleus , pons ,
  medulla project to every area of brain spinal
  cord.
• Sympathetic postganglionic neurons typically
  release norepinephrine.
• NE E serve important role in the regulation of
  blood volume blood pressure.

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• Norepinephrine is synthesized from tyrosine .
• dopamine ß hydroxylase
• dopamine
  norepinephrine
• cu
• dopamine ß hydroxylase is bound to inner
  membrane of synaptic vesicle release
  norepinephrine in a tetrameric glycoprotein form.

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• The overall system of epinephrine synthesis ,
  storage secretion from adrenal medulla are
  regulated by neuronal controls also by
  glucocorticoid hormones synthesized in secreted
  from adrenal cortex in response stress.
• Secretion of epinephrine is signaled by neural
  response to stress , which is transmitted to
  adrenal medulla by way of a preganglionic acetyl
  cholinergic neuron.

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• A small number of neurons in the medulla contain
  phenyl ethenolamine N- methyl tranferase enzyme
  that converts norepinephrine to epinephrine with
  SAM as methyl donor.
• These neurons project to the thalamus, brainstem
  , spinal cord.
• Concentration of epinephrine secreting terminals
  in the paraventricular nucleus suggests a role in
  secreton of oxytocin vasopressin.

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• Dense innervation of dorsal motor nucleus of
  vagus , nucleus solitarius suggets role in
  regulating cardiovascular respiratory reflexes.
• Receptors on target cells may be either a or ß
  adrenergic receptors.
• Receptors are further sub divided into a1, a2, ß1
  , ß2 , ß3.

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• a1 receptor are located postsynaptically but a2
  receptors may be either pre or postsynaptic .
• Receptors located presynaptically are
  autoreceptors inhibit release of
  neurotransmitter.
• The effects of a1 receptors are mediated by
  activation of ITP/ diacyl glycerol second
  messenger system.
• ß receptors can be antagonized by action of a1
  receptor.

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• a2 receptors decease the rate of synthesis of
  cAMP through an action on inhibitory G protein.
• ß1ß2 receptors activates stimulatory G protein
  to increase cellular cAMP levels.
• Activation of ß receptor result in coactivation
  of ß adrenergic receptor kinase (BARK), this
  phosphorylates the receptor.
• Phophorylation is prominent mechanism of receptor
  desensitization.

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• Number of ß receptors is regulated .
• ß receptor is phosphorylated desensitized
  their number also decreased if they become
  internalized.
• ß receptors can also be increased by
  denervation.
• The number of a receptor is also regulated.

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• ß1 adrenergic receptors principally found in
  heart cerebral cortex.
• ß2 receptors principally found in lung
  cerebellum.
• ß1 receptors equally prefer NE E as agonist.
• ß2 receptors prefer epinephrine to
  norepinephrine.

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• In synaptic neurons norepinephrine decreases the
  amplitude of calcium spikes.
• Excitatory effects of norepinephrine in various
  parts of CNS sympathetic ganglion neuron
  results from a1 receptor activation. This
  activity primarily depends on blockade of a
  resting K conductance as a result neuron
  depolarizes firing rate increases.

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• Inhibitory effects of norepinephrine results from
  a2 receptor activation , which results in
  increase of K conductance this hyperpolarizes
  the neuron decreases its firing rates.
• NE acting at a2 receptor also block Ca current.
• Both the above inhibitory mechanisms account for
  the autoreceptor function of a2 receptors which
  decreases neurotransmitter release.

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• a1 adreno receptor activation results in ,
  1)vasoconstriction ,
• 2) relaxation of gastrointestinal smooth
  muscle,
• 3) salivary secretion ,
• 4)hepatic glcogenolysis.

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• a2 adreno receptor activation results in
• 1) inhibition of transmitter release,
  (including NE Ach from autonomic nerves)
• 2) platelet aggregation
• 3) contraction of vascular smooth muscle.
• 4) inhibition of insulin release.

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• ß1 adreno receptors activation results in
• 1) increased cardiac rate force
• ß2 adreno receptors activation results in 1)
  bronchodilatation ,
• 2) vasodilation,
• 3) relaxation of visceral smooth muscle,
• 4) hepatic glycogenolysis,
• 5) muscle tremor.
• ß3 adreno receptor activation results in
  lipolysis.

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• The action of catecholamine neurotransmitters is
  terminated by reuptake into presynaptic neuron by
  specific transporters.
• Enzymes involved in metabolism are catechol 0
  methyl transferase monoamino oxidase .
• End product of norepinephrine epinephrine
  metabolism is 3 methoxy 4 hydroxy mandelic acid.

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serotonin

• More than 95 of bodys serotonin is stored in
  platelets GI tract , only 5 is seen in brain.
• Serotonin is distributed in brain regions that
  affect behaviour especially the hypothalamus
  limbic system.

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• Availability of tryptophan is the main factor
  regulating synthesis of tryptophan.
• Process of synthesis , storage , release ,
  reuptake degradation are similar to
  catecholmines.
• Urinary 5 HIAA provides a measure of 5 HT
  turn over.

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• Functions associated with 5-HT path ways
• 1)hallucination behavioural changes,
• 2)sleep, wakefulness mood ,
• 3) feeding behaviour,
• 4)control of sensory pathway including
  nociception,
• 5) vomiting.
• Serotonin receptors are metabotropic.
• Melatonin a derived product of 5-HT has role in
  establishing circadian rhythm.

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• Histamine has neurotransmitter role in brain.
• Acts on metabotropic receptors.
• H1 receptors are excitatory H2 , H3 receptors
  are inhibitory.
• H1 receptors in cortex RAS contributes to
  arousal wakefulness.
• Has role in food water intake, thermoregulation

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• Nitric oxide synthase is present in many CNS
  neurons.
• NO production is increased by mechanisms that
  raise intracellular Ca concentration( eg
  transmitter action).
• NO affects neuronal functions by increasing cGMP
  formation ,producing both inhibitory excitatory
  effects on neurons.

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• ATP functions as neurotransmitter , it acts via
  ionotropic receptors as fast excitatory
  transmitter , via metabotropic receptors acts as
  neuro modulator.
• Adenosine exerts inhibitory effects through
  metabotropic receptors.
• Neurons contain CO generating enzyme, heme
  oxygenase , have role in cerebellum olfactory
  neurons which have cGMP sensitive ion channels.

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