Membranes,Membranes, waterwater andand diffusion:diffusion: PotentialPotential forfor brainbrain imagingimaging

Denis Le Bihan NeuroSpin Director, CEA-Saclay, France Kyoto University, Japan Institut de France, Academy of Sciences

DLB March08 Functional (PET, fMRI): A Water Story Coupling between blood flow, metabolism and activation (Roy & Sherrington, 1894) Iron Do we need better? 15* BOLD fMRI H20 PET

Hemoglobin

Rest

Activation Òblood flow Seiji Òblood flow & oxygenation Òquantity of radioactive water Ogawa Ômagnetized water relaxation in activated tissues in/near vessels

DLB March08 Radioactive water Magnetized water BOLD fMRI: Hemodynamic events…

Coupling between CBF, metabolism and activation (Roy & Sherrington, 1894) – Rationale & mechanism? metabolism (glucose) vs neurogenic? Æ activation of glial cells by glutamate Æ arachidonic acid –> vasodilation/vasoconstriction (Gordon et al. Nature. 456, 745, 2008)

Æ pO2? large increase in blood flow but small increase in O2 consumption – fMRI BOLD not always coupled to neuronal activity or metabolism! Anticipatory signals (Sirotin et al. Nature 457, 475, 2009), pathology, drugs: indomethacin, antihistamin,…

5.9%

4.9%

3.9% AVM embolization 2.9% LehericyBiochemistryet al. Neurology, 2002 1.9% 0.9%

-0.1% Time (s) -4 -2 0 2 4 6 8 101214161820 4.7T monkey Logothetis et al. 2001. Nature Æ Functional hyperhemia & local field potentials Æ Other more direct mechanisms? " Small induced electric axonal currents (≈ EEG/MEG) : Indirect reflect of neuronal activation :" NoNeurotransmission: separation of Ca2+ inflowclustersas seen sharingwith MnMRI Biophysics vascularization :"IntrinsicStructurallimitation events inducedin spatial/temporal in cortical cells resolution Æ Water diffusion MRI [Darquie et al. PNAS 2001] DLB March08 IRM de IRMDIFFUSION

Le Bihan & Breton, CRASS, 1985 Echelle macroscopique Eau (noyaux d’hydrogène) (millimètres) FreeStroke water: D ≈ 3 10 -9 m²/s Brain @ 37°Cconnectivity Functional MRI ? 1/2 ≈ 17µm (Td = 50ms) Echelle microscopique D (micromètres) 1/2 b 17

1/2 50 Td

DLB March08 IRM de Diffusion ... Effet directionnel (anisotropie)…

Mangin, Poupon et al. MICCAI 1998, NI 2000

DLB March08… vers la connectivité cérébrale Du mouvement brownien à la connectivité cérébrale: L’imagerie du Tenseur de diffusion (Basser & Le Bihan, 1992-94)

BrainVISA software Cointepas et al. http://brainvisa.info

DLB March08 Diffusion-sensitized MRI: tissue events

1 - Cell swelling Ù Decreased water diffusion 9 Change in water diffusion coefficient with osmolality High osmolality Æ cell shrinking, increased diffusion Low osmolality Æ cell swelling, decreased diffusion 9 Decreases water diffusion have been reported in - - Extra-physiological activation: status epilepticus induced by bicuculline and cortical electroshocks (Zhong), spreading depression (Sotak, Moseley) 2 - Physiological neuronal activation Ù cell swelling NIRS data, Grinwald et al.

CA1 Hippocampal region Extracellular field potential

OPTICAL IMAGING Infra-Red transmittance

Tasaki, 1988 black: dendritic region gray: somatic region Neuron (Takagi et al. 2002) DLB March08 s Vasculature Optical imaging Diffusion-sensitized fMRI @ 3T

Diffusion fMRI

2.20%

31.80

1.70% 31.60

) 31.40

1.20% aw r

BOLD fMRI y ( it 31.20 s n e

0.70% t 31.00 al in n g I 30.80 0.20% S

30.60 -0.30% 0 5 10 15 20 25 30.40 Time (s) 0 50 100 150 Early response (

DLBLe March08Bihan et al. PNAS 103, 8263-8268, 2006 (Satoru Kohno et al. JCBFM in press) Water diffusion model in the brain: Back to Einstein…

R 2 − 1 4 Dt d P (R,td ) = e 4πDt d Free diffusion = Gaussian distribution of displacements

1/2 For MRI: S=exp (-b D)

human brain cortex = 2 Dt Clark CA & Le Bihan D, MRM, 2002

d

A Restricted ln (S) ≠ – b D

Permeable Barriers B Hindered

Displacement profil (q-space) : Gaussian 1 + Gaussian 2

BiexponentialDLB March08 behavior: diffusion is not ‘free’ Æ 2 water pools? The water molecule 3.2 Å

'Water is H2O, hydrogen two parts, oxygen one, but there is also a third thing, that makes it water and nobody knows what it is.‘ D H Lawrence (1885-1930)

Hydrogen bond

0.9572 Å 105.5° (liquid)

Water tetramer

R = 2.976 (+0.000, -0.030) Å, α = 6 ± 20°, β = 57 ± 10°

Æ Liquid water: tetrahedral H-bond network with local defects

Proton mobility (diffusion) decreases with increase degree of

DLBstructuring March08 (lower density of local defects) Marx et al. Nature 1999 Brain water diffusion model: structured water Visual cortex 5 2 KINDS OF WATER IN CELLS: SLOW + FAST DIFFUSION POOLS ffast= 65.9% 4.5 fslow= 34.1% -3 9 WaterWater- diffusionmembr ina brainne interface:is mainly hindered by cell membranes Dfast = 1.39 10 mm²/s 4

l) -4 a n Dslow = 3.25 10 mm²/s Long-range electrostatic trapping ig s ( ln 3.5 9 Diffusion behavior is well described by a biexponential model -Protein « trapping » effects: structured water 3 9 Slow-Extendand fastto adherdiffusionent water pools layers do notoutcoincide to 180nmwith the intra- and 2.5 0 1000 2000 3000 4000 extracellular(Xu & Yeung, vScienceolumes… 1998, Vogler, 1998) b value ln (S) ≠ – b D -≈ 30% of cell water (Pissis, 1987) 9ƒ but“Slow” variations diffusionin slow/fast pool: structureddiffusion fracti waterons “bound” correlate towell membraneswith those of the EC/IC volumes (cell size) ) (stroke,ADC changes osmotic withchallenges, membrane statussurfaceepilepticus, interaction spreading depression…) (cell size, anisotropy, cellularity,…)

Intra-cellular Cell swelling: ADC, fslow compartment - Extra-cellular compartment

Extra Intra ‘Fast’ ‘Slow’ Cellular Cellular Diffusion Diffusion (20%) (80%) Pool Pool =Fast? =Slow?

DLB March08 2 KINDS OF WATER IN CELLS: SLOW + FAST DIFFUSION POOLS Slow diffusion pool :membrane interactions Le Bihan Phys. Med. Biol. 52 R57-R90, 2007

Acute stroke Fast pool: Bulk water and Cell swelling proteins-bound water Slow pool: Membrane-bound water (w/cytoskeleton ± 40 nm) Cell anisotropy Y Rest Activation

Intra-cellular Z compartment Membranes: ADC, fslow Extra-cellular compartment

Dxx Dyy Dzz

Cell proliferation (cancer) Dslow 0.14 0.13 0.23 0.17

Dfast 1.12 1.13 1.39 1.21

‘Fast’ ‘Slow’ fslow 0.49 0.41 0.11 0.33 Diffusio Diffusion n Pool Pool Human brain Clark & Le Bihan, MRM, 2002 Ross B. J. Natl. Cancer Inst, 92, 2029-2036, 2000

Day 0 Day 2

Day 4 Day 8 r=.78

DayDLB 16 March08 Day 20 WaterBOLD dfMRI:iffusion Hemodynamic fMRI: Cell membrane events… events … Fast coupling between ion fluxes, membrane depolarization and water structure Coupling between CBF, metabolism and activation (≈ PET-H20, PET-FDG, NIRS) S(t)={DRF,HRF} ⊗ paradigm 0.016 –Mechanism: fluxes of ions AND water

2. 20% 0.0142.20% Baseline •Swelling of (soma, dendrites) and glial cells 0.0121.70% 0.01 DRF 1.20% •Membrane surface scales with square of cell size 0.008 HRF 0.0060.70% DRF: Tmax=1.65, alpha=0.5 Swelling •Phase transition ( order) of water near polarized 0.004 HRF: Tmax=6, alpha=5.2 0.20% 0.002 membranes AND attached polymer-gel matrix 0 5 10 15 20 25 seconds -00.30% Stress and Fold Localization in Thin Elastic Membranes 0 2 4 6 8 Ti10m11e (13s) 15 17 19 21 23 25 27 29 30 Pocivavsek et al. Science 230;912-916, 2008

Diffusion and Hemodynamic Response Functions extracted from the MRI signal time courses Æ Why and how do cell swell? A hypothesis… (Aso et al. HBM06) - Action potentials Æ Phase transition - Require cytoskeleton integrity (Matsumoto) - Cytoskeleton undergoes physical changes (Sato) -CA1Cal cHippocampalium is critical (regionTasaki, 1988) - Transient cellular expansion (swelling) and water influx (Tasaki) Extracellular field potential - Transient liberation of heat (23µ°C Tasaki)

Induction of hemodynamic responseOPTICAL(Bold)(= IMAGICold)??NG Infra-Red transmittance Tasaki, 1988 Æ Water seemsblack: dendriticto play region a central role in gray: somatic region Cytoskeleton density gradient Hodgkin & Huxley modelcell revisited: activation physiology from Tsukita & Matsumoto, 1986 Unique featureNot of a cfasttion enough?potential initiation in neoco(Takagirtical neureonst al. 2002) NaundorfDLB(see March08 NaundorfG.H. et Pollack al. at Na al.ture Nature 440;1060, 440:1060,Action 2006 2006) potential, cell swelling, heat transfer 2. 2% Water: the molecule 1. 7% BOLD Diffusion of the mind ? 1. 2% 0. 7% Le Bihan D. The ‘wet mind’ 0. 2% Phys. Med. Biol. 52:557-90, 2007 0 5 10 1 5 20 2 5 -0 .3 % Ti m e ( s ) 15* ¾Active role of water inH cell20 dePETpolarization, actionBOLD potenfMRItial? DIFFUSION fMRI -more water influx in cells than expected from ion transferts ¾Role of blood water in heat transfert (increase in CBF)? -activation --> heat release -increase of CBF and metabolism (energy demand) more to restore baseline conditions after activation, ‘housekeeping’ ¾Role of cell structure changes (swelling) in information flow -neurons & glial cells as (reversible) piezo-electric transducers -faster thanRest neurotransmission for local, intracotical networks

Activation Òblood flow Òblood flow & oxygenation Òcell size and membrane surface Òquantity of radioactive water Ômagnetized water relaxation Ôdiffusion of water near in activated tissues in/near vessels membranes

DLB March08 Radioactive water Magnetized water Plain brain water