BY2201 Colm Basic NeurochemOct11

8/2/2019 BY2201 Colm Basic NeurochemOct11

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Basicneurochemistry

ColmCunningham


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BasicNeurochemistry

[email protected]


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Thebraincontains

neurons


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GLIA la#nglue(Virchow,1850s)

neurons@85billion,about10% Macroglia

Astrocytes(mostabundant55%) Oligodendrocytes30%

Microglia-about5%


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Meetthe

family

1oligodendrocyte

2axonhillock

3neuronalsoma

4myelinsheath

5microglia

6astrocyte

7synapse

8bloodvessel


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GlialfuncUons

Oligodendrocytes

(&Schwanncells)

-myelinaUon

Microglia-immunefuncUon

scavengers

Astrocytes

-NTreuptake

-isolaUon/insulaUon

-BBB

-migraUon(radial)

-NMJsynapUcreg.


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Thecells Cajal,neurons,glia,glialmetabolism

TheNeurons types,projecUons,tracers

TheNeuron structure,regions

Thecontents neurotransmi[ers,metabolism,vesicles,

bioenergeUcs

TheacUon(presynapUc) Na+-K+ATPase,acUonpotenUal,depolarisaUon, Ca2+channels,synapUcrelease

TheacUon(post-synapUc)receptors,signalling(ionotropic,metabotropic)

diversityofreceptorsandresponses

DysfuncUon incidental;paralysis,depression,excitotoxicity, recreaUonaldrugs,disease


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Neuronaltypes


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Neuronsofthe

hippocampus

Cajal1893


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NeuronalcommunicaUon

Structual-cell,axon,synapse

Chemical-neurotransmi[errelease


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GlutamateandGABA

MajorExcitatory,inhibitoryNTsrespecUvely

ShortaxonsandInterneurons-localsignalling

+H3N-CH2

I

CH2

I

CH2I

COO-

COO-I+H3N-C-H

I

CH2

I

CH2I

COO-

GABAglutamate


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Tracttracing (1970s)

Anterograde(soma-terminal) PhaseolusvulgarisleucoaggluUnin(10days) BiocyUn(48hrs)

Retrograde(terminal-soma) Horseradishperoxidase(axons&synapses) WheatgermaggluUnin-HRP Choleratoxin-HRP (morespecific)

Immunohistochemistry BiosyntheUcenzymescharacterisUcofspecificpopulaUons


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Dopaminergicsystem


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NoradrenergicprojecUons


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5HT(serotonin)projecUons


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5criteriafordesignaUonasaneurotransmi[er

Synthesisinneuron Verifiablereleasefromneuron Effectonpost-synapUcneuron

ExogenousapplicaUonshouldhavesameeffectonpost-synapUccellsaselectricalsUmulaUonofthepre-synapUcterminal

Specificpost-synapUcreceptorBlockedbyspecificantagonistsandmimickedbyagonists

AppropriateterminaUonmechanismsReuptake,degradaUon


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Neurotransmi[ers

Biogenicamines Aminoacidneurotransmi[ers Othertypes:Acetylcholine,NO,

D-serine,neuropepUdes

AcetylCoA+choline=CH3-C-O-CH2-CH2-N-(CH3)3

O


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Noradrenaline

&Dopamine

biosynthesis

catecholamines

PD


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Serotonin(5-HT)biosynthesis

indoleamines


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GABA&

glutamate

GABAmadefrom

Glutamateby

decarboxylase

GABAtransaminase

GABA

+

2-oxoglutarate

to

succinicsemialdehyde+

glutamate

if

2

-OOCCCH2CH

2CCOO-=

O

=O

GAD


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Neurotransmi[ervesicles

SSV-smallsynapUcvesicles

-35-60nm

-classicalneurotransmi[ers

-packedanywhereincell-synapse

LDCV -largedensecorevesicles

-largersize

-NTsandneuropepUdes(co-release)

-cellbody-ER-golgi-secretorygranules

-hardertorelease,notreusable(notclassicalNTs)


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Vesicularuptake

BioenergeUcs-protonelectrochemicalgradient

-vesicularATPase

-drivesprotonsin(pH5.5)

-thisdrivesNTagainstitschemicalgradient

-2protonsforoneNT

-upto100mMNT

-lowaffinity(0.3mM)


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ptakeandstorageinhibitors

tetrabenazine

Bindtotransporter

Inhibituptake

anU-psychoUcdrugs

AmphetamineandEcstasyalsodeplete

Tyramine

falsetransmi[er-its

Similarenoughtoget

Packaged,but

Decreaseefficacy

TreatHypertension

PEA


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SynapUcvesiclecycle

1 acUvefillingofvesicles2 ClusteringinregionofacUvezone3 DockingofasubsetoftheseatAZ4 PrimingtomakecompetentforCa 2+-dependentexocytosis5 Ca2+triggeringofexocytosis(fusionporeopening)6 Recycling


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Axonconductance

ResUngmembranepotenUalof-65mVw.r.t.toextracellular

Membranechannelspermeable

toK+soitleaksoutandleaves

localisednegaUvecharge,

LikewiseNa+leaksin

Na+-K+Pumpmaintainshigh

intracellularK+andlowNa+

despitethisleakage(against

electrochemicalgradients

(HydrolysisofATP))

ponsUmulaUon,voltage-gated

Na+channelsopen,Na+flowsin,depolarizingthecell(-65to-55).

ThisopensnearbyNa+channels

andtheacUonpotenUalis

propagatedalongaxon.

Conc

gradient


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SynapUcpotenUal

ResUngpotenUal

=-60to-70mV

3

2

ATP

HighintracellularK+(400mMvs20)

HighextracellularNa+(440mMvs50)

Leakageofbothbypassivediffusiondown

concentraUongradient

AcUve(ATP)pumpingofbothagainst

gradient-electrogenic(3+vs2+)

AcUonpotenUalarrivesandvoltage-gated

Na+channelsopen(Na+in)-

depolarisaUon

CausesvoltagegatedK+channelstoopen

(K+out)-repolarisaUon

IniUallythoughtthatNa+influxwas

thechemicaltriggerforrelease


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Experimentalevidence (chapter14Kandel)

Katz&Miledi,1967

Na+entryisimportant(exptswithtetrodotoxin:Na+channelblocker).Butonlyin

sofarasitdepolarisesthemembranesufficientlytogeneratetheacUonpotenUal

thatfacilitatestransmi[errelease.DirectapplicaUonofasimilaracUonpotenUalproducesnormalrelease

K+channelblockersliketetraethylamoniumdonotpreventrelease(infactextend

releaseperiodbyblockingrepolarizaUon).

Ca2+:increasingextracellularCaincreaseNTrelease(oppfordecrease)10,000foldexcessofCaextracellularlywithrespecttocellular

ThereforeCawouldneedtogetintothecell

Voltage-dependentCachannelsdiscoved(Llinasetal.,1977)


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Neurotransmi[er

release

ResUngpotenUal

=-60to-70mV

3

2

Na+channelsopen

Voltagechangeabove40mV

(depolarisaUon)

Voltage-dependentCa2+

channelsopen-Ca2+flows

downconcentraUongradient

LowaffinityCa2+sensorsin

acUvezone(100m)bindCa2+

Ca2+-dependentexocytosis

AndrapidturnoffasCa2+

diffusesawayfromacUvezone

Ca2+104higher


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Neurotransmi[errelease


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SNAREproteins


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SNAREcomplexandexocytosisSNARE - SNAP receptor proteinsSNAP-soluble NSF attachment

ProtNSF-N-ethylmaleimide-sensitive

fusion protien

The SNARE complex is a four--helix bundle, formed

between the vesicle and cellmembranes. Synaptobrevin (blue) is partlyinside the vesicle (R SNARE) Syntaxin (red) is partly within

the cell membrane (Qa SNARE).

SNAP-25, a third SNARE

complex protein, is shown ingreen and yellow (two SNARE

motifs, Qb & c).


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Vesicledocking:Toxins

TETANS


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SNAREcomplex

Blue-synaptobrevinRed-syntaxin-1AGreen-SNAP-25BToxin cleavage sites are

located:at the peptide bonds

between Gln-76 andPhe-77 (TeNT and

BoNT/B), between Ala-81 and

Ala-82 (BoNT/G) forsynaptobrevin-II, between Lys-253 andAla-254 (BoNT/C) for

syntaxin-1A, and between Gln-197and Arg-198 (BoNT/A)for SNAP-25B. Fasshauer et al, 1998


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Neurotransmi[errelease

PhosphorylaUon

Kinases

DephosphorylaUon

Phosphatases

Sudhof


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Fusion/exocytosis


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ToxinsaffecUngrelease/transmission

Tetrodoxin-pufferfish-Na+channelblocker-death Clostridialtoxins

BotulinumA,B,CcleaveSNAP25,VAMP,syntaxinrespecUvely(BOTOX:ACh) TetanustoxincleavesVAMP(zincendopepUdase) DisruptTHEkeyinteracUoninneurotransmi[errelease(fig14-13)

Dendrotoxin-blackmamba-K +channels Latrotoxin-blackwidowspider-massiveacetylcholinerelease -bungarotoxin-snakevenom-AChnicoUnicreceptoronskeletalmuscle

(paralysis)


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Neurotransmi[errelease-itsfast-motorfuncUon

WholecyclehastoberepeatedrapidlyforrepeatacUons!!!RECYCLING


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SNAREdisassembly


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NTacUons/Receptordiversity

Ionotropic-directgaUngbyNTbinding

Metabotropic-metabolic/signallingchangesleadindirectlytoionflux

-Gproteincoupled-manyNTs

-Receptortyrosinekinases-hormones,neuropepUdes

ACh:nicoUnic(I),muscarinic(M)

Glutamaterecptors:ionotropic:NMDA,KA,AMPA,metabotropicmGLR

D1-D5

Alpha,beta-adrenergic

5HT:manytypes

DifferentreceptorsforthesameNTondifferentneurons:differenteffects


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Acetylcholine

Excitatoryorinhibitory

Receptors:

NicoUnic-ionotropic

Muscarinic-metabotropic

VisibleacUonatNMJ


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NeuromuscularjuncUon-ACh


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NicoUnicAcetylcholinereceptor-MyastheniaGravis

Alpha-bungarotoxin


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Acetylcholinergic

terminal

permeability

ionotropic metabotropic

Metabolism-ChAT,AChE,

no-reuptake

AD,

Learningandmemory,mostlymuscarinic

(metabotropicreceptors)

Q


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Glutamatereceptors


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SignallingGproteinmechanisms cAMP

5HTbinding

ExposureofGproteinbindingdomain

GDPexchangedforGTP

Gsalphasubunitreleased

AcUvaUonofadenylylcyclase

ProducUonofcAMP,HydrolysisofGTP

(toterminatecAMPproducUon)

cAMP-dependentProteinKinase

ReformingofG, , complex

DissociaUonof5HT


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Signalling

mechanisms

GalphaacUvatesPLC

Cleavesphospholipids

MakesDAG&IP3

ReleasesCa2+fromER

Ca2+acUvates:

PKC

Ca2+/calmodulin-dependentPK

PhosphorylaUon&cellularresponse


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Receptordiversity

Q

Q


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Receptordiversity


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Glutamate

uptake

Release

GLT1/EAAT1uptake

Glutaminesynthetase

GLtoGLN

Export

Glutaminaseinneuron


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GABA

transaminase


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GABA

GABAtransaminase

GABA

+

2-oxoglutarate

to

succinicsemialdehyde

+

Glutamate

SSADHdeficiency-GHB

if

2

-OOCCCH2CH

2CCOO-=

O

=OQ


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Catabolismofamineneurotransmi[ers


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COMT


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COMT


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5-HTcatabolism


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MAO-monoamineoxidase

Outermitochondrialmembrane

Neurons,glia,endothelialcells,gut,liver

MAOA-clorgyline(In),5HTselecUve

MAOB-selegiline(deprenyl)(In),PEA,Benzylamine

Both-NA,dopamine,tyramine,tryptamine,adrenaline


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MAO

Inhibitors 2nd

1st

3rd

}

First

anU-depressants


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MAOinhibitors


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Threeclassesof

AnU-depressants


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SerotoninacUon/inhibiUon


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RecreaUonaldrugs


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Noradrenergicsynapse

MAOinhibitor

Tricyclics

COMTinhibitor


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Dopaminergicsynapse


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Ischemicstroke

Bloodsupply

Q


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Ischemicstroke

Bloodsupply

Oxygen,glucose

OxidaUvephosphorylaUon


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Ischemicstroke

Bloodsupply

Oxygen,glucose


ATP, increasedROS(complexIV)


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Ischemicstroke

Bloodsupply

Oxygen,glucose



CollapseofmembranepotenUal,reversalofpumps


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Ischemicstroke

Bloodsupply

Oxygen,glucose




Massivereleaseofglutamate


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Ischemicstroke

Bloodsupply

Oxygen,glucose





Massiveionimbalances,Ca2+stores


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Ischemicstroke

Bloodsupply

Oxygen,glucose






ROS,proteases

Protein,DNA,membranedamage


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Ischemicstroke

Bloodsupply

Oxygen,glucose






ROS,proteases

Protein,DNA,membranedamage

Cellswelling,rupture


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LacUcacidosis-acidificaUonofcytoplasm

Reducingequivalents:NADHOxPhos->ATP

NAD+oxidisingpowerforglycolysis


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Glutamate

uptake

Nadownto80mM

Kupto20mM

ReversalofGLT1/EAAT2

uptake

ElevatedGL]

FurtheracUvaUon

Ca2+influx


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Glutamate


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Glutamate

&GABA

AnUconvulsants

Na+channels

GABAagonists


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Thecells Cajal,neurons,glia,glialmetabolism

TheNeurons types,projecUons,tracers

TheNeuron structure,regions

Thecontents neurotransmi[ers,metabolism,vesicles,

bioenergeUcs

TheacUon(presynapUc) Na+-K+ATPase,acUonpotenUal,depolarisaUon, Ca2+channels,synapUcrelease

TheacUon(post-synapUc)receptors,signalling(ionotropic,metabotropic)

diversityofreceptorsandresponses

DysfuncUon incidental;paralysis,depression,excitotoxicity, recreaUonaldrugs,disease

BY2201 Colm Basic NeurochemOct11

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