The Biological Basis of the Diffusion SignalThe Biological Basis of the Diffusion Signal

Christian Beaulieu, PhDAssociate Professor and AHFMR Senior Scholar

Dept of Biomedical Engineering, University of Alberta

June 15, 2008Human Brain Mapping

Melbourne

OutlineOutlineOutline1.1. Diffusion AnisotropyDiffusion Anisotropy

2.2. Magnetic SusceptibilityMagnetic Susceptibility

3.3. NeurofibrilsNeurofibrils / Axonal Transport/ Axonal Transport

4.4. Axonal MembranesAxonal Membranes

5.5. MyelinMyelin

6.6. Interpretation of AnisotropyInterpretation of Anisotropy

1. DIFFUSION

ANISOTROPY

1. DIFFUSION 1. DIFFUSION

ANISOTROPYANISOTROPY

Tissue Water

cellular structureshinder water diffusion

Start

Finish

Pure Water

unhindered random walk

Diffusion Samples the MicrostructureDiffusion Samples the Microstructure

Isotropic Diffusion Anisotropic Diffusion

similar moleculardisplacements in all directions

greater molecular displacementalong cylinders than across

Perpendicular to TractPerpendicular to Tract Parallel to TractParallel to Tract

Slower water diffusionSlower water diffusion Faster water diffusionFaster water diffusion

White Matter

&Diffusion

White White Matter Matter

&&DiffusionDiffusion

Diffusion Ellipsoids for Different Fractional Anisotropy

Diffusion Ellipsoids for Different Diffusion Ellipsoids for Different Fractional AnisotropyFractional Anisotropy

“CSF” “CentrumSemiovale”

“CorpusCallosum”

All have same Mean Diffusivity = 0.7 x 10-3 mm2/s

Anisotropy in Neural FibresAnisotropy in Neural Anisotropy in Neural FibresFibresQuite high degree of anisotropic water diffusion Quite high degree of anisotropic water diffusion

relative to other tissuesrelative to other tissues

Diffusing molecules sample local environment Diffusing molecules sample local environment and then infer microand then infer micro--structural characteristicsstructural characteristics

Obvious that anisotropy is related to ordered Obvious that anisotropy is related to ordered arrangement of the myelinated arrangement of the myelinated fibresfibres

Water diffusion distances are ~ 8 Water diffusion distances are ~ 8 μμm in the m in the diffusion time (~40 ms) used in MRIdiffusion time (~40 ms) used in MRI

Anisotropy in Neural FibresAnisotropy in Neural Anisotropy in Neural FibresFibres

myelinaxonal membrane neurofilament

microtubule

axon

D(//)

D(⊥)

What are the relative microWhat are the relative micro--structural structural contributions to anisotropic water diffusion ?contributions to anisotropic water diffusion ?

ADC(//)

ADC(⊥)

λλ11

λλ2,32,3

Useful Model SystemsUseful Model SystemsUseful Model Systems

Anisotropy – Not Unique to Neural FibresAnisotropy Anisotropy –– Not Unique to Neural Not Unique to Neural FibresFibres

Celery vascular bundlesCelery vascular bundles

Moseley, Topics MRI, 1991Moseley, Topics MRI, 1991

Diffusion Images With Diffusion Images With Different Gradient DirectionsDifferent Gradient Directions 2. MAGNETIC

SUSCEPTIBILITY

2. MAGNETIC2. MAGNETIC

SUSCEPTIBILITYSUSCEPTIBILITY

Susceptibility? No.Susceptibility? No.Susceptibility? No.Anisotropy, measured by NMR, could artificially result Anisotropy, measured by NMR, could artificially result

from local susceptibilityfrom local susceptibility--differencedifference--induced gradientsinduced gradients

Minimize these gradients by using bipolar gradient Minimize these gradients by using bipolar gradient pulse sequence or aligning pulse sequence or aligning fibresfibres parallel to Bparallel to Boo

Susceptibility Susceptibility notnot an issue :an issue :Porcine spinal cord inPorcine spinal cord in--vitro at 4.7Tvitro at 4.7TFour garfish nerves inFour garfish nerves in--vitro at 2.35Tvitro at 2.35THuman brain white matter at 1.5THuman brain white matter at 1.5T

Trudeau et al JMR 1995Trudeau et al JMR 1995

Beaulieu et al MRM 1996Beaulieu et al MRM 1996

Clark et al JMR 1999Clark et al JMR 1999

3. NEUROFIBRILS &

FAST AXONAL

TRANSPORT

3. NEUROFIBRILS &3. NEUROFIBRILS &

FAST AXONAL FAST AXONAL

TRANSPORTTRANSPORT

Cytoskeleton Scaffolding in AxonsCytoskeleton Scaffolding in AxonsCytoskeleton Scaffolding in Axons Microtubules / Fast Axonal Transport ??Microtubules / Fast Axonal Transport ??Microtubules / Fast Axonal Transport ??

DepolymerizeDepolymerize micromicro--tubules and inhibit tubules and inhibit fast axonal transportfast axonal transport

Garfish nerves treatedGarfish nerves treatedwith with VinblastineVinblastine

Beaulieu & Allen MRM 1994aBeaulieu & Allen MRM 1994a

ADC values // and ADC values // and ⊥⊥ reduced but anisotropy preservedreduced but anisotropy preserved

0.5 μm

Neurofilaments ??Neurofilaments ??Neurofilaments ??Giant Axon of the SquidGiant Axon of the Squid

Beaulieu & Allen MRM 1994bBeaulieu & Allen MRM 1994b

Giant axon dimension much greater than Giant axon dimension much greater than root mean square displacement of water (5root mean square displacement of water (5--10 10 μμm)m)

Isotropic, Rapid Water Diffusion in AxoplasmIsotropic, Rapid Water Diffusion in Isotropic, Rapid Water Diffusion in AxoplasmAxoplasm

Beaulieu & Allen Beaulieu & Allen MRM 1994bMRM 1994b

Takahashi et al Takahashi et al PNAS 2002PNAS 2002

300 μm

40 μm

SummaryNeurofibrils / Axonal Transport

SummarySummaryNeurofibrilsNeurofibrils / Axonal Transport/ Axonal Transport

Squid diffusion results matched Monte CarloSquid diffusion results matched Monte Carlosimulations of simulations of neurofilamentaryneurofilamentary latticelattice

Microtubules and neurofilaments Microtubules and neurofilaments do notdo not play aplay asignificant role in anisotropic water diffusionsignificant role in anisotropic water diffusion

Water diffusion in pure axoplasm is rapid and is Water diffusion in pure axoplasm is rapid and is ~ 70 ~ 70 -- 80% of that in pure water80% of that in pure water

4. AXONAL

MEMBRANES

4.4. AXONAL AXONAL

MEMBRANESMEMBRANES

Water Transport is ComplexWater Transport is ComplexWater Transport is Complex

www.bioscience.orgwww.bioscience.org, Watson, Watson

AquaporinsAquaporins

Garfish NervesGarfish NervesGarfish Nerves

Beaulieu & Allen, MRM, 1994aBeaulieu & Allen, MRM, 1994a

Non-myelinated olfactory Myelinated optic (CNS)

Myelinated trigeminal (PNS)

FA 0.59 FA 0.52

FA 0.59

Anisotropy in Non-myelinated Fibres : Other Studies

Anisotropy in NonAnisotropy in Non--myelinated myelinated FibresFibres : : Other StudiesOther Studies

Rat pups white matterRat pups white matter WimbergerWimberger et al,JCAT 1995et al,JCAT 1995Prayer et al, Prayer et al, NeuroradNeurorad 19971997

JimpyJimpy mouse optic nervemouse optic nerve Ono et alOno et alBrain Res 1995Brain Res 1995

Rat Rat vagusvagus nervenerve SeoSeo et alet alMRM 1999MRM 1999

Myelin deficient rat spinal cordMyelin deficient rat spinal cord GulaniGulani et al, MRM 2001et al, MRM 2001BitonBiton et al, MRI 2006et al, MRI 2006

Brain in developing miceBrain in developing mice Mori et al Mori et al MRM 2001MRM 2001

Anisotropy in Non-myelinated Fibres : Other Studies

Anisotropy in NonAnisotropy in Non--myelinated myelinated FibresFibres : : Other StudiesOther Studies

MyelinMyelin--deficient deficient shiverershiverer micemiceSong, Song, NeuroimageNeuroimage 2002 2002 Nair, Nair, NeuroimageNeuroimage 20052005TyszkaTyszka, , NeuroimageNeuroimage 20062006

Walking leg nerve of lobsterWalking leg nerve of lobster BeaulieuBeaulieuNMR NMR BiomedBiomed 20022002

Lamprey spinal cordLamprey spinal cord Takahashi et alTakahashi et alPNAS 2002PNAS 2002

Brain in baby rabbitsBrain in baby rabbits DrobyshevskyDrobyshevsky et alet alJ J NeurosciNeurosci 20052005

Neonatal human brain Neonatal human brain inin--vivovivo HuppiHuppi et al, et al, PedPed Res 1998Res 1998Neil et al, Radiology 1998Neil et al, Radiology 1998

Lamprey Spinal CordLamprey Spinal CordLamprey Spinal Cord

Takahashi et al PNAS 2002Takahashi et al PNAS 2002

Ani

sotr

o py

Ani

sotr

o py

Axon diameterAxon diameter

Packing DensityPacking Density

increasesincreases

decreasesdecreases

** all non** all non--myelinatedmyelinated ****

Anisotropy in Pre-myelination PeriodAnisotropy in PreAnisotropy in Pre--myelinationmyelination PeriodPeriod

DTI of white matter development in DTI of white matter development in perinatalperinatal rabbitsrabbits

DrobyshevskyDrobyshevsky et al et al J J NeurosciNeurosci 20052005

MyelinationMyelinationbegan at P5began at P5

MyelinationMyelinationbegan at P11began at P11

SummaryAxonal Membranes

SummarySummaryAxonal MembranesAxonal Membranes

Anisotropy in neural Anisotropy in neural fibresfibres NOT myelin specificNOT myelin specific

Axonal membranes sufficient for significant Axonal membranes sufficient for significant anisotropy and appear to play anisotropy and appear to play primaryprimary rolerole

Nonetheless, the loss and addition of myelin in a Nonetheless, the loss and addition of myelin in a given tract could certainly affect ADC(//), given tract could certainly affect ADC(//), ADC(ADC(⊥⊥), and the degree of anisotropy), and the degree of anisotropy

5. MYELIN5. MYELIN5. MYELIN

MyelinMyelin--Deficient Spinal Cord Rat ModelDeficient Spinal Cord Rat Model

X-linked recessive Wistar ratnear-total lack of myelination in the CNS

Normal

Toluidine Blue Stain

Myelin Deficient

FA 0.73

Anisotropy decrease of ~ 10% in absence of myelin

GulaniGulani et al, MRM, 2001et al, MRM, 2001

FA 0.66

Shiverer Mice – Study 1ShivererShiverer Mice Mice –– Study 1Study 1AutosomalAutosomal mutation of myelin basic proteinmutation of myelin basic protein

Song et al Song et al NeuroimageNeuroimage 20022002

WildWild--typetype ShivererShiverer

Relative AnisotropyRelative Anisotropy

Anisotropy decreased by 16Anisotropy decreased by 16--25% in anterior 25% in anterior commissurecommissure, , cerebral peduncle, optic nerve, and optic tract cerebral peduncle, optic nerve, and optic tract

Due to increase in ADC(Due to increase in ADC(⊥) while ADC(//) stays constant ) while ADC(//) stays constant

Shiverer Mice – Study 2aShivererShiverer Mice Mice –– Study 2aStudy 2a

FA reduced by ~10% in corpus FA reduced by ~10% in corpus callosumcallosum

Nair et al Nair et al NeuroimageNeuroimage 20052005

Shiverer Mice – Study 2bShivererShiverer Mice Mice –– Study 2bStudy 2bDonorDonor--derived derived myelinationmyelination by transplanted by transplanted

precursor cells in corpus precursor cells in corpus callosumcallosum

Nair et al Nair et al NeuroimageNeuroimage 20052005

Myelin Basic Protein

T2T2--weighting did not show thisweighting did not show this

FA Map

Shiverer Mice – Study 3ShivererShiverer Mice Mice –– Study 3Study 3High resolution DTI microscopy of fixed brainsHigh resolution DTI microscopy of fixed brains

TyszkaTyszka et al et al NeuroimageNeuroimage 20062006

ShivererShivererFAFA

ControlControlFAFA

FAFADifferenceDifference

FA reduced byFA reduced by1111--19% in 19% in shiverershiverer micemice

Jimpy MiceJimpyJimpy MiceMice

HarsanHarsan et al J et al J NeuroNeuro Res 2007Res 2007

Myelin basicMyelin basicproteinprotein

ADCADC((⊥))

FAFA

P25P25

P75P75afterafter

recoveryrecovery

Transgenic Mice - DysmyelinationTransgenic Mice Transgenic Mice -- DysmyelinationDysmyelination

HarsanHarsan et al J et al J NeuroNeuro Res 2006Res 2006

Treated : P15Treated : P15 P60P60P30P30

//⊥ FA

Morphometry - Diffusion CorrelationsMorphometryMorphometry -- Diffusion CorrelationsDiffusion Correlations

Axon Axon morphometrymorphometry correlated with ADC(//), ADC(correlated with ADC(//), ADC(⊥), ), and anisotropy in normal rat cervical spinal cordand anisotropy in normal rat cervical spinal cord

Schwartz et al Schwartz et al NeuroreportNeuroreport 20052005

PositivePositive correlations with ADC(correlations with ADC(⊥))-- extraextra--cellular volume fractioncellular volume fraction-- axon spacing (most significant)axon spacing (most significant)

NegativeNegative correlations with ADC(correlations with ADC(⊥))-- axon countsaxon counts-- myelin volume fraction myelin volume fraction

What About Human Brain?What About Human Brain?What About Human Brain?PostPost--mortem DTI and histology in MS patientsmortem DTI and histology in MS patients

SchmiererSchmierer et al et al NeuroimageNeuroimage 20072007

More myelin

SummaryMyelin

SummarySummaryMyelinMyelin

Myelin has more marked effects on the mean Myelin has more marked effects on the mean diffusivity and diffusivity and eigenvalueseigenvalues than the anisotropythan the anisotropy

MyelinationMyelination can modulate degree of anisotropycan modulate degree of anisotropy

A reduction in ADC(A reduction in ADC(⊥) coincides with ) coincides with myelinationmyelinationfor a given neural for a given neural fibrefibre

Aside: small FA changes may reflect more severe pathology Aside: small FA changes may reflect more severe pathology than expectedthan expected

6. INTERPRETATION

OF ANISOTROPY

6.6. INTERPRETATIONINTERPRETATION

OF ANISOTROPYOF ANISOTROPY

How to Interpret Reductions in Anisotropy?

How to Interpret How to Interpret Reductions in Anisotropy?Reductions in Anisotropy?

Clues from Wallerian DegenerationClues from Clues from WallerianWallerian DegenerationDegenerationRat Spinal Cord Injury Rat Spinal Cord Injury

Ford et al MRM 1994Ford et al MRM 1994

PierpaoliPierpaoli et al et al NeuroimageNeuroimage 20012001Beaulieu et al MRM 1996Beaulieu et al MRM 1996

T2 T2 ADC(ADC(⊥⊥) ) ADC(//) ADC(//)

Similar findings in frog sciatic nerve and Similar findings in frog sciatic nerve and secondary secondary fibresfibres in human chronic strokein human chronic stroke

Same FA ReductionsSame FA ReductionsSame FA ReductionsNormal FANormal FA 20% FA20% FA

≠≠

FA Reductions Could Be Interpreted Differently

FA Reductions Could Be FA Reductions Could Be Interpreted DifferentlyInterpreted Differently

FAFAADC (//)ADC (//)

ADC (ADC (⊥⊥))

and/orand/or

Need to look at the eigenvaluesNeed to look at the Need to look at the eigenvalueseigenvalues

Mouse Optic Nerve IschemiaMouse Optic Nerve IschemiaMouse Optic Nerve Ischemia

Song et al Song et al NeuroimageNeuroimage 20032003

ADC (//)ADC (//) ADC (ADC (⊥⊥))

FAFA

Days post-injury

Cuprizone Treated MiceCuprizoneCuprizone Treated MiceTreated Mice

Sun et al MRM 2006Sun et al MRM 2006

λλ////drops firstdrops first

AxonalAxonaldamagedamage

λλ⊥⊥increases laterincreases later

DemyelinationDemyelination

Mouse Brain TraumaMouse Brain TraumaMouse Brain Trauma

MacDonald et al J MacDonald et al J NeurosciNeurosci 20072007

//

⊥

MacDonald et al J MacDonald et al J NeurosciNeurosci 20072007

Mouse Brain TraumaMouse Brain TraumaMouse Brain Trauma

SubacuteSubacute InjuryInjuryAcute InjuryAcute Injury

Reduced parallelReduced parallel Increased perpendicularIncreased perpendicular

Importance of AxonsImportance of AxonsImportance of Axons

Mouse Brain TraumaMouse Brain Trauma

MacDonald et al Exp MacDonald et al Exp NeurolNeurol 20072007Wu et al Wu et al NeuroimageNeuroimage 20072007

Mouse EAE ModelMouse EAE ModelOptic NerveOptic Nerve

Ani

sotr

opy

AD

C(//

)

Axolemmal Area # of Injured Axons

Human Callosotomy - FAHuman Human CallosotomyCallosotomy -- FAFA

ConchaConcha et al et al NeuroimageNeuroimage 20062006

Are these FA reductions the same?

Post-surgical transectionof corpus callosum

Human Callosotomy - EigenvaluesHuman Human CallosotomyCallosotomy -- EigenvaluesEigenvalues

ConchaConcha et al et al NeuroimageNeuroimage 20062006

FA – Eigenvalue HypothesisFA FA –– EigenvalueEigenvalue HypothesisHypothesis

FAFA

ADC (//)ADC (//)

ADC (ADC (⊥⊥))

Axonal DamageAxonal Damage

DemyelinationDemyelination

CONCLUSIONCONCLUSIONCONCLUSION

DTI – Limited ResolutionStill Probing the Microstructure

DTI – Limited ResolutionStill Probing the Microstructure

μm’s mm’s cm’s

ConclusionConclusionConclusionIntact membranes are confirmed to be the Intact membranes are confirmed to be the

primary determinant of anisotropic water primary determinant of anisotropic water diffusion in neural diffusion in neural fibresfibres

Changes in perpendicular diffusivity may be Changes in perpendicular diffusivity may be indicative of relative degree of myelin in a indicative of relative degree of myelin in a specific white matter tractspecific white matter tract

Primary role by axonal membranes Primary role by axonal membranes

Secondary role by Secondary role by myelinationmyelination (modulate)(modulate)