Biochemistryofneurotransmitters

Prof.MamounAhramNeuroscience2020

References

ThislectureMark’sBasicMedicalBiochemistry,4th ed,pp.908-918http://what-when-how.com/neuroscience/neurotransmitters-the-neuron-part-1/

Definitionofaneurotransmitter

Aneurotransmitterisachemicalsubstancethatis:synthesizedinaneuron,releasedatasynapsefollowingdepolarizationofthenerveterminal(usuallydependentoninfluxofcalciumions),whichbindstoreceptorsonthepostsynapticcelland/orpresynapticterminaltoelicitaspecificresponse.

Characteristicsofaneurotransmitter

Achemicalsubstancethat:Issynthesizedandstoredinapresynapticneuron(theenzymesneededforitssynthesismustbepresentintheneuron),Isreleasedatasynapsefollowingdepolarizationofthenerveterminal(usuallydependentoninfluxofcalciumions),bindstoreceptorsonthepostsynapticcelland/orpresynapticterminal,elicitsrapid-onsetandrapidlyreversibleresponsesinthetargetcell,Isremovedorinactivatedfromthesynapticcleft.

Typesofneurotransmitters

Small-moleculeneurotransmittersBiogenicamines(epinepherine,dopamine,histmaine,serotonin)Aminoacids(GABA,glutamate,aspartate,glycine)AcetylcholinePurines(ATP)

NeuropeptidesGases(nitricoxide,carbonmonoxide)

Twoormoretransmitters(usuallyasmall-moleculetransmitterandaneuroactivepeptide)coexistinmanymatureneurons(e.g.,mostspinalmotorneuronscontainacetylcholineandcalcitoningene-relatedpeptide).

Structuresofneurotransmitters

NEUROPEPTIDES

Introduction

Morethan50neuropeptideshavebeendescribedBehaviorPainperceptionMemoryAppetiteThirstTemperatureHomeostasisSleep

Neuropeptides:neurohormonesorneurotransmitters?

Neurohormones: whenreleasedbyneuronsintothehaemolymphandexertitseffectsondistantperipheraltargets.

Peptidesactivatetheirreceptorsatlow(nMtouM)concentrationscomparedtotheconcentrationsrequiredtoactivatereceptorsforsmall-moleculeneurotransmitters.Thesepropertiesallowthepostsynaptictargetsofpeptidestobequitefarremovedfrompresynapticterminals.

Neurotransmitter:whenreleasedfromaneuronataspecializedjunctionanddiffusesacrossanarrowclefttoaffectoneortwopostsynapticneurons,amusclecell,oranothereffectorcell.

Classificationofneuropeptides

Peptidescanbegroupedbystructuralandfunctionalsimilarity.Neuropeptide Families

Tachykinins:substanceP,bombesin,substanceKInsulins:insulin,insulin-likegrowthfactorsSomatostatins:somatostatin,pancreaticpolypeptideGastrins:gastrin,cholecystokininOpioids:opiocortins,enkephalins,dynorphin

OpiateFamily

Name AminoAcidSequenceLeu-

enkephalin Tyr-Gly-Gly-Phe-Leu-OH

Met-enkephalin Tyr-Gly-Gly-Phe-Met-OH

Beta-endorphin

Tyr-Gly-Gly-Phe-Met-Thr-Ser-Glu-Lys-Ser-Gln-Thr-Pro-Leu-Val-Thr-Leu-

Phe-Lys-Asn-Ala-Ile-Val-Lys-Asn-Ala-His-Lys-Gly-Gln-His-OH

Dynorphin Tyr-Gly-Gly-Phe-Leu-Arg-Arg-Ile-Arg-Pro-Lys-Leu-Lys-Trp-Asp-Asn-Gln-OH

• Vasopressin and oxytocin share 7 of 9 amino acids, but have different functions.

• Opiate peptides share a common sequence, but are receptor-selective.

• The three glycoprotein hormones from the anterior pituitary, TSH, LH, and FSH, share a common α subunit, but have distinct β subunits.

StagesofactionSynthesis(ER(1)aspre-propeptidesthenpropeptides andtheninGolgiapparatus(2))Packagingintolarge-densecorevesicles(withmodifyingenzymes)Transport(3)(fast-axonaltransport)

Duringthetransport,proteasescleavetheprecursorneuropeptideintothefinalmatureform(4).

Release(5)Releaseisgradualovertimeinresponsetogeneralincreasesinthelevelofintracellularcalcium.

Action(prolonged)MainlyviaGPCR

Terminationbydiffusionanddegradation(6)

Diversity:alternativesplicing

AlternativesplicingofmRNAleadstotranslationofdistinctprecursors,andsubsequentprocessingleadstouniquematurepeptides.

ExampleisthesubstancePmRNAthatnormallyalsoincludesmRNAencodingsubstanceK.

Diversity:proteolytic,differential,sequentialprocessingNeuropeptidesareproducedfromalongerprecursorproteinby

ProteolyticprocessingVesicularpackagingofdifferentproteasesthatrecognizedifferentcleavagesequencesHidingaproteolyticsitebypost-translationalmodifications(example:additionofacarbohydratesidechain).Tissue-specific

Processing of the pro-opiomelanocortin(POMC) precursor proceeds in an ordered, stepwise fashion. Some of the reactions are tissue specific. ACTH, adrenocorticotropichormone; CLIP, corticotropin-like intermediate lobe peptide; JP, joining peptide; LPH, lipotropin; MSH, melanocyte-stimulating hormone; PC, prohormoneconvertase.

RoleofCa2+ions

• Vesicles are located further away from the presynaptic membrane and away from area of Ca2+ ions influx

• Ca2+ ion influx can be from external of internal sources and at higher concentrations than required for small-molecule neurotrasmitters.

Thelevelsofregulationofneuropeptide expression

Diffusion

Proteolysis

Proteases may or may not be released

SMALL-MOLECULENEUROTRANSMITTERS

StagesofactionSynthesisofenzymesinER(1)andGolgiapparatus(2)wheretheyaremodified(3).Transportofsoluble enzymesviaslowaxonaltransport(4)Nurotransmittersprecursorsaretakenupviatransporterproteinslocatedintheplasmamembraneofthenerveterminal(5),andtheneurotransmitterissynthesizedinthepresynapticnerveterminalthenpackagedinsmallsynapticvesicles(6).Releaseisstimulatedbybriefpulseseachtimeanactionpotentialtriggerstheinfluxofcalcium.Action(short)Terminationbydiffusion,re-uptake,orinactivation

[Ca+] = 2 mM

[Ca+] = 0.1 uM

100 uM-[Ca+] = 50

Proteinsandexocytosis

TheinfluxofCa2+ionsthemtointeractwithsynaptotagminleadingtofusionofthevesicularandpresynapticmembranes.

http://www.sumanasinc.com/webcontent/animations/content/synaptictransmission.html

TYROSINE-DERIVEDNEUROTRANSMITTERS

Dopamine,norepinephrine,andepinephrine

Notes

RoleofcofactorsS-adenosylmethionine(methyltransfer)Pyrodoxalphosphate(vitaminB6):transamination,decarboxylationTetrahydrobiopterin(BH4)

Rate-limitingstep

Pyridoxal phosphate vesicular

Vitamin B12 or folate

Diet/liver

phenylalanine hydroxylase

cytoplasm

Tyrosine (hydroxy-phenylalanine) is present in all food products and synthesized from phenylalanine. It enters the neuron by active transport.

50%

10%

Na+-dependent dopamine co-transporter (DAT)

CocaineAmphetamine

Most

Some

LDCV

Na+-dependent norepinepherine co-transporter (DAT)

Amphetamine

Leaking

Packagingintovesicles

Thecatecholamines(dopamineandepinepherine)aretransportedintovesiclesbyanATP-dependentprocesslinkedtoaprotonpump.ProtonsarepumpedintothevesiclesbyavesicularATPase(V-ATPase).Theprotonsthenexchangeforthepositively-chargedcatecholamineviathetransporterVMAT(vesicularmonoaminetransporter).

COMTandMAO

Parkinson’s disease

Inactivation is dependent on SAM,

vitamin B12 and folate

Regulation

• Tyrosinehydroxylase– Shortterm:

• Inhibitionbyfreecytosoliccatecholamines,whichcompetewithBH4bindingtoenzyme

• Activationbydepolarization,whichactivatesseveralproteinkinasesincludingPKC,PKA,Ca2-calmodulin–dependentkinasesthatphosphorylatetyrosinehydroxylase.ThismakestheenzymebindmoretightlyBH4and,consequently,lesssensitivetoendproductinhibition.

– Long-term(plusdopamineb-hyroxylase)Prolongedsympatheticneuronalactivityincreasestheamountsoftyrosinehydroxylaseanddopamine–hydroxylasemRNAs.

TRYPTOPHAN-DERIVEDNEUROTRANSMITTERS

Serotoninandmelatonin

requiresBH4

Serotonin

5-hydroxyindoleacetic acid

urine

MAO

Antidepressants , called selective serotonin re-uptake inhibitors (SSRIs) like Prozac® inhibit the reuptake process resulting in prolonged serotonin presence in the synaptic cleft.

VMAT

Melatonin

Serotoninissynthesizedinthepinealglandandservesasaprecursorforthesynthesisofmelatonin,whichisaneurohormoneinvolvedinregulating:

sleeppatternsSeasonalandcircadian(daily)rythymsDark-lightcycle

VMAT vesicles

GLUTAMATEANDASPARTATE

Glutamateandaspartate

NonessentialaminoacidsDonotcrossBBB

mustbesynthesizedinneuronsMainsyntheticcompartments

neuronsglialcells

Bothareexcitatoryneurotransmitters.

Synthesisofglutamate

Sources:1. Glycolysis® Krebscycle®dehydrogenationofa-ketoglutarate

2. Glutamine(deamination)3. Aspartate(transamination)Removal

excitatoryaminoacidcarrier-1(EAAC1)glutamatetransporter-1(GLT-1)andglutamate—aspartatetransporter(GLAST)

GABA

glutaminaseGlutaminesynthetase

transaminase 1

2

3

a-KG

GluDehydro

Sourcesofglutamate(supplementary)

Aspartate

Avesicularuptakemechanismforaspartatehasnotyetbeendemonstrated,somewhatweakeningthecaseforconsideringaspartatetobeaneurotransmitterPrecursor:oxaloacetate(transamination)

Glycine

AmajorinhibitoryneurotransmitterItissynthesizedfromserinebyserinehydroxymethyltransferasethrough3-phosphoglycerateRemoval:high-affinitytransporter

GABA

GABAispresentinhighconcentrations(millimolar)inmanybrainregions.

Theseconcentrationsareabout1,000timeshigherthanconcentrationsoftheclassicalmonoamineneurotransmittersinthesameregions.

TheGABAshuntisaclosed-loopprocesswiththedualpurposeofproducingandconservingthesupplyofGABA.

GABAshunt

Gln ® Glubyglutaminase.Glu® GABAbyglutamatedecarboxylase(GAD),whichrequirespyridoxalphosphate(vitaminB6).GABAisstoredinvesiclesuntilreleased.GABAiseither

takenupintopresynapticterminalandrepackagedgoesintotheGABAShuntwhereitistakenupintothegliaandconvertedtoGlu.

GluisconvertedintoGln,whichistransportedintotheneighboringnerveterminalstosynthesizeGlu.

Diet

MembranePL

Synthesisofacetylcholine

Choline+acetylcoenzyme-AbycholineacetyltransferaseincytoplasmTransportedintoandstoredinvesicles.Removal:hydrolysisbyacetylcholinesterase

Histamine

itdoesnotpenetratetheblood-brainbarrierand,hence,mustbesynthesizedinthebrain.Histamineisinactivatedbytwoenzymes—histaminemethyltransferaseanddiamineoxidase(histaminase).

Pyridoxal phosphate

Astrocytes(MAO) Neuron

XNo recycling

Histamine is packaged into vesicles by VMAT

Nitricoxide(NO)

Glutamateisreleased(1)andactsonNMDAreceptorslocatedonthepost-synapticneuron(2)Ca2+entersthepostsynapticneuronactivatingNOS(3),whichformsNOfromsarginine(4).NOstimulatesguanylatecyclaseformingcGMP(5),whichresultsinaphysiologicalresponse(6)NOcandiffuseout:a)tothepresynapticterminal(retrogrademessenger)(7)prolongingeffectandb)intoadjacentneurons(8)andglialcells(9)stimulatingguanylatecyclase. Half-life: 2-4 seconds

NO is inhibited by hemoglobin and other heme proteins which bind it tightly

IsNOaneurotransmitter?

Yes,but:ItisnotstoredinvesiclesItisnotreleasedbycalcium-dependentexocytosis(itdiffuses)Itsinactivationispassive(thereisnoactiveprocessthatterminatesitsaction)

ItdecaysspontaneouslyItdoesnotinteractwithreceptorsontargetcells

Itssphereofactiondependsontheextenttowhichitdiffuses,anditsactionisnotconfinedtotheconventionalpresynaptic-postsynapticdirection.

NOactsasaretrogrademessengerandregulatesthefunctionofaxonterminalspresynaptictotheneuroninwhichitissynthesized.

NOsynthase

IsoformI(nNOSorcNOS)Neuronsandepithelialcellsactivatedbytheinfluxofextracellularcalcium

isoformII(iNOS)Macrophagesandsmoothmusclecellsinducedbycytokines

andisoformIII(eNOS)Endothelialcellsliningbloodvesselsactivatedbytheinfluxofextracellularcalcium

AllthreeisoformsrequireBH2asacofactorandnicotinamideadeninedinucleotidephosphate(NADPH)asacoenzyme.

NotethedifferencesbetweenneuropeptidesandneurotransmittersActivity(slowvs.fast),response(slowvs.fast),anddurationofaction(longvs.short)Receptortargets(multiplevs.single)Geneexpression(yesvs.no)Synthesis,transport,andpackagingConcentrationforaction(lowvs.high)Speedofrelease(slowvs.fast)Concentrationof[Ca+2]forreleaseSiteofsynthesis,modification,actionFate

Neuropeptidesvs.Neurotransmitters

Neuropeptides Neurotransmitters

Size Largermolecules(3to36aminoacids)

Smallermoleculesofdifferentprecursors

Re-uptake Not–re-takenupagain Possible re-uptake

Post-release modification Yesby extracellularpeptidases NomodificationsStorage Large densecorevesicles SmallsynapsevesiclesLocation Anywhereintheneuron Inthepresynapticaxon

terminalSecretions Co-releasedAction Slow-acting Fast-actingSynthesis Ribosomes, ER,Golgibodies,etc. Cytoplasm ofpresynapticendEfficiencyofsignaling Moreefficient LessefficientConcentrations Lower Higher

DiffusionatReleaseSite Diffuseaway Donotdiffusefar