1. A brief introduction to MR physics

2. Hydrogen/Proton: A spinning top   = J
Protons, electrons and neutrons each possess a spin or angular momentum (J) of ½, which can be positive or negative. Hydrogen has one proton in its nucleic, i.e. one unpaired spin. It spins around its own axis. Each nucleic has a specific gyromagnetic ratio (gamma), which gives it a specific dipole moment (µ).
 = J

3. With and without the presence of an external magnetic field (B0) in the z direction
Randomly oriented hydrogen
atoms (spins) in absence of an
external magnetic field
The protons aligns with the external field: Low field: 0.47 T, Clinical fields: B0 = 1.5 – 3.0 T
Up or down: 2I + 1, I = ½ gives 2 allowable energy levels.
M = sum of individual dipole moments that points upwards (lower energy state, more dipole moments parallel with the external field)
The nuclei presses about an axis that is collinear with B0, which is due to the angular momentum that the nuclei posses. The combination of nuclear magnetic moment and angular moment cause the nuclei to precess about B0 in much the same way as a spinning top (snurrebass). The resonance frequency, omega0, which is the speed of the precessing proton is dependent on the gyromagnetic ratio and the field strength. At a clinical field of 1.5 T, the resonance frequency of protons is MHz, and at for example 0.47 T the resonance frequency of protons is 20 MHz.
Together they create a macroscopic
magnetic M0 vector parallel with B0
(and z)

4. Radio Frequency (RF) coils in the xy-plane (B1)
A 900 pulse flips M0 into
the xy-plane
M0 aligns with the z-axis B1
The magnetization vector, M0, can be manipulated with radiofrequency signals to produce a measurable signal in the xy-plane. We wish to measure the relaxation times of different proton containing molecules in tissue. The relaxation times, T1 and T2, are the time it takes for the magnetization vector to go/relax back to equilibrium after being manipulated by a 90 degree RF pulse, by giving up their acquired energy to their surroundings.
RF on

5. An oscillating FID signal in the xy-plane (readout plane)
When the protons aligns with the external field again they give up their energy to their surroundings. This signal is registered by a coil (like an antenna):
RF off:
The spins
relax back
to their
equilibrium
state (along z)
When the RF pulse is turned off the spins will relax back to their equilibrium state. A receiver coil can be placed in the y-plane and read-out the signal from the relaxation processes. This signal is called the free induction decay (FID) signal, and is the MR signal. The signal is very different for water and lipids, i.e. their relaxation times differ the most. Other tissue structures have relaxation times in between these two extremes.

6. The time it takes for M0 to realign with B0 after the influence of
T1 relaxation:
The time it takes for M0 to realign
with B0 after the influence of
an RF pulse. Dependent of the
exchange with the media. T1 is
short for fat and long for water
T2 relaxation:
The time it takes before the spins
(protons) reach equilibrium after
spin-spin exchange and thereafter
dephasing in the xy-plane:
T2  T1
The two relaxation patterns happens simultaneously, but can be measured separately: T1 and T2, where T2 is almost shorter or equal to T1.
Since the protons have different surroundings they react differently on the influence of external signals (fields and/or RF pulses). Protons in water, for example, are not good shielded from the external magnetic field because of their neighboring oxygen atoms. However, protons in lipids are good shielded because of their neighboring carbon atoms. Water molecules diffuse or vibrate quickly in tissue, while lipid-protons vibrate more slowly. These properties are the reason why water and lipid act so differently on the influence of RF pulses. Water is greatly influenced by the external field and keeps the energy which it absorbs for a long time, i.e long relaxation times, while lipid does not absorb a lot of energy and thereby give away the energy equally quickly, i.e. short relaxation times.

7. The MR signal Intrinsic parameters: T1, T2, n(H)
Instrument parameters: TR, TE
Pulse sequences: vary TR and TE
Contrast agents: manipulate T1 and T2
To create an image other electromagnetic fields called imaging gradients are used in addition to the RF pulses. Different types of images can be created by using different pulse sequences or programs, consisting of different instrument parameters, i.e. TR and TE, that can be varied. In addition, the physiologic parameters, T1, T2 and n(H), are intrinsic and not possible to change directly. However, T1 and T2 can be manipulated indirectly by the use of contrast agents.
In this thesis the parameters T1 and T2 have been measured in order to study contrast mechanisms, physiology and water movements in tissue.

8. Contrast Agents (CA)
Even though MR has high specificity, CA are often used to better visualize specific structures and pathology
Indirect effect on hydrogen spins. The relaxation times (T1 and T2) are reduced, thus signal changes
Either T1 or T2 CA’s, where T1 CA’s increase the signal and gives “positive” contrast, whereas T2 CA’s reduces the signal and gives “negative” contrast in the image

9. Different CA’s (exogenous)
Paramagnetic
Positive contrast: n(H) increases, T1 reduces and T2 increases
Much stronger T1 than T2 reducing effect, which in total gives an improved T1 contrast by the addition of paramagnetic agents
T1-weighted images
Some paramagnetic ions: Gd3+, Mn2+
Mn2+ is strongly paramagnetic and responsible for MRI contrast enhancement.

10. Electron spin: Non-paired electrons
Mn2+
A magnetic moment analogous to the magnetic moment of protons
The magnetic moment of an electron ~600 times stronger than the magnetic moment of protons, and will therefore have an influence on them
Paramagnetiske ioner har uparrede elektroner, dvs elektronene har et magnetisk moment analogt til kjernens magnetiske moment. Forskjellen er at elektronenes magnetiske moment er mye sterkere enn kjernens og vil dermed påvirke kjernens magnetiske moment. De forandrer kjernens magnetiske miljø. T1 og T2 reduseres.
Change the magnetic environment of the protons, where T1 and T2 will be reduced 1H
Paramagnetic
ion

11. Efficacy in a homogenous solution
Relaxivity r1 and r2:
The measured effectiveness of an CA to reduce T1 and T2
Given in (s mM)-1
A simple linear formula:
T1 -1with CA = T1-1without CA + r1 • [CA]
In a homogenous compartment

12. T1 relaxation – shorter and shorter T1 (CA)
stronger and stronger signal in a T1 weighted image where the CA is present

13. *Schering,**Amersham Health/GE Healthcare
Different types
Free metal ions are toxic in large quantities, that is why they are bound to a chelate. This “shielding” influence the effectiveness of the CA, and safety is competing with efficacy
Few side effects; does not result in severe allergic reactions as with X-ray CA’s
Extracellular (infarction, pathology)
Gd-DTPA (Magnevist*)
Liver CA (pathology)
MnDPDP (Teslascan**)
Intravascular/E.C. (few protons < 5% of all the tissue protons). Large complexes which are trapped in the blood pool
Gd-DTPA-BMA (Omniscan**)
Small molecules
Vil ikke ha frie metallioner i kroppen, fordi de binder seg til ulike strukturer og blir værende lenge.
Av og til vanlig med fett suppresjon ved bruk av Gd-agens, fordi fett og Gd-vev kan bli isointenst (like rask relaksasjontid, eller like kort T1)
*Schering,**Amersham Health/GE Healthcare

15. Water Exchange Theory

16. H O Intrinsic values t P T 2 ic : Extracellular Intracellular ec - 1
Her ser dere en enkel skisse av en celle med et intracellulært og et ekstracellulært rom (ikke tatt med andre kompartememter i cellen samt blodårer).
Vann diffunderer altså fritt inne i de ulike kompartementene og mellom dem.
Parameteren tau brukes her om levetiden til vann, hvor ic betegner det intracellulære kompartementet og ec det ekstracellulære.
Vi sier at vannutvekslingshastigheten (i Hz) over membranen mellom de ulike kompartementene er lik summen av vannutvekslingen fra de ulike kompartementene. Levetiden er i så måte en fysiologisk konstant som er avhengig av størrelsen til cellen og permeabiliteten til membranen (tau-1 = PS/V).

17. A relaxivity in each ”homogenous” water compartment
Intracellular relaxivity: r1ic

18. Monoexponential decay Inversion Recovery pulse sequence:
Mz(t) = M0 ( 1 - 2e-t/T1 )
Dvs et monoexponentielt signal. Lineær plott gir en graf med stigningstallet –1/T1 (=R1): ln(M0 – Mz) = (ln2+lnM0) – t/T1.

19. FAST EXCHANGE T1
-1
Dersom vi bruker MR som en teknikk til å studere cellene med, er oppløsningen tidsavhengig, dvs at det vi ser er avhengig av tiden prosessene tar på en MR-skala.
Relaksasjonstidene T1 og T2 til protonene er avhengige av miljøet rundt protonene, dvs at vi kan manipulere disse ved å endre protonene sitt miljø. T1 er fra like langsom til mye langsommere enn T2 (i vann er de like). Alle vev har sine egne intrinsikke (iboende/sanne) T1 og T2’er. Siden T2 er et sammensatt og lite homogent signal, skal jeg kun forholde meg til T1 nå. (mer utfyllende om T2 i avhandlingen?)
Selv om ingen av disse prosessene kan foregå alene, går det an å måle dem hver for seg. Vi måler T1 med f.eks en inversion recovery eller saturation recovery pulssekvens. I realiteten har vi altså en intrinsikk intracellulær T1ic og en ekstracellulær T1ec, med andeler på hhv pic og pec.
Vi sier at vannutvekslingen er rask (på en MR-skala) dersom den er mye større enn forskjellen i relaksasjonsraten mellom de to kompartementene vi studerer. (Klikke inn Fast exchange og likning)
Resultatet blir at vi bare klarer å måle én relaksasjonstid fra hele cellen (KLIKK!), hvor relaksasjonsraten er mye større enn summen av de kompartementelle relaksasjonsratene.
For hjerteceller har vi funnet en T1 tid på 1000 ms i litteraturen (og det har vi også selv målt på lab’en).
|R1ic – R1ec| kalles ”shutter speed” av Springer.

20. The apparent biexponential T1’s are sums
of the intrinsic relaxation rates and
the water exchange in each compartment: 1 1 1 = + T’ T  1 ic 1 ic ic 1 1 1
Men; nå er T1-tidene ikke de intrinsikke T1’tidene til cellen, men snarere en sum av den intrinsikke T1-tiden og levetiden til vannet i de enkelte kompartementene. = + T’ T’  1 ec 1 ec ec

21. T’1ic p’ic T’1ec p’ec SLOW EXCHANGE: Apparent values
Dersom vannutvekslinghastigheten er mye mindre enn differansen av de kompartementelle relaksasjonshastighetene, får vi det vi kaller langsom vannutveksling.
Resultatet blir dermed at vi detekterer to relaksasjonstider istedenfor én. I dette forenklete systemet antar vi at den ene T1-tiden kommer fra det intracellulære rommet og det andre fra det ekstracellulære.
Molekylær imaging (lavfelts?).

22. Biexponential signal:
Mz = p’ic ( 1 - 2e-t/T1’ic ) + p’ec ( 1 - 2e-t/T1’ec ))
Når vi gjøre studier med kontrastmidler som holder seg i enten det ekstracellulære eller det intracellulære rom, så kan vi ved hjelp av et dose-respons studiet, prøve å korrigere for denne feilen i T1’ene.
V(myocytter) = /- 7.0 pl in rats (n = 21), konfokal mikroskop (Satoh H., Delbridge L.M., Blatter L.A. and Bers D.M. (1996) Surface:volume relationship in cardiac myocytes studied with confocal microscopy and membrane capacitance measurements: species-dependence and developmental effects. Biophysical Journal 70, )
V(myocytter) = /- 7.3 pl (mean +/- SD) in rabbits (n = 28), /- 9.0 pl in ferrets (n = 23)
V(gjærcelle) = 65 fL (dvs pl)
1/ = (PA)/V

23. Intermediate exchange
Dersom vi har fast exchange eller utartet slow exchange er det lett å finne T1’ene – de er distinkte, men dersom vi har en situasjon som er intermediær, dvs i overgangsfasen mellom slow til fast, så blir det litt mer triksete.

24. 2 SX equation for R1ic
Da kan vi bruke en model som heter 2SX modellen (two-site water exchange) til å korrigere for vannutvekslingen som forkludrer T1’ene våre. (dette er for å finne den reelle R1-2, men vi har også tilsvarende likning for R1-1 og en tredje for å beregne de sanne andelene. Her er kontrastmiddelet i det ekstracellulære rommet)
(Dersom resultatet ikke gir noen endring i T1’ene, så kan det skyldes at levetiden så lang at dette leddet blir neglisjerbart i likningen -> slow exchange)

25. 2 SX equation for R1ec

26. 2 SX equation for pic and pec

27. Calculating relaxivities, lifetimes of water and population fractions

28. EKSTRASTOFF

29. SOLOMON-BLOEMBERGEN dipole-dipole term
c (s)
temp
nonviscous
liquids
viscous
Solids R2 R1
1/T1M ,
1/T2M
SOLOMON-BLOEMBERGEN dipole-dipole term
20 MHz
60 MHz
100 MHz
10-11
10-7

30. Extracellular Intravascular E.C. Contrast Agent Plasma Interstitium
1H20ic
1H20p
ic-1 +
Plasma
1H20*CAp
CAp
m-1
Extracellular
Vessel wall
Interstitium
I vev er det litt mer komplisert enn i den enkle figuren som jeg har skissert. Her er et litt vanskelig bilde av blodårer (plasma og blodceller), interstitium og celler med cellelegemer. CA kommer jo med blodet i første omgang. OG vi har ulike kontrastmidler som holder seg i blodbanen og i det ekstracelluære matrix.
1H20ic
 ic-1
m-1
Intracellular
1H20*CAec
CAec + 1H20ec
ec-1

31. Springer's group at the State University of New York, has published a series of Papers [64-66, 114] where they have used a set of modified Block equations that accounts for water exchange pioneered by McConnell (1958) [76]. For longitudinal magnetization (Mz), two sets of equations, corresponding to each compartment (ic and ec) were deduced.
where Ck = 1/\tau_k and k = ic/ec. The Equations in~\ref{eq:BlochMod} differ from the usual Bloch equations (Equation~\ref{eq:Bloch1} -~\ref{eq:Bloch3}) by the addition of two terms to the right-hand side, \pm C_{ic} (M_z)_{ic} and \pm C_{ec} (M_z)_{ec}. In the first equation - C_{ic} (M_z)_{ic} represents the rate at which (Mz)_{ic} decreases due to the transfer of Mz magnetization out of the ic space, and C_{ec} (M_z)_{ec} measures the rate at which (Mz)_{ic} increases due to the transfer of M_z magnetization into the ic space from the ec space.
The longitudinal two-component relaxation equation can be given as…
Fast- and slow water exchange are borderline situations. The in-between situation is also common:

32. The exact solutions for the apparent $T_\mathit{1}$ values based on the equations of Woessner [113], were first presented by Leigh (1971) [70] and McLauglin and Leigh (1973) [77] in the following form:
Similar presentations are found in Hazlewood [44], and in Winkler and Mitchel [112]:

33. The physiological parameters $\tau_{ic}$ and $p_{ic}$ are related to $\tau_{ec}$ and $p_{ec}$ by equilibrium mass balance:
The apparent population fractions can be expressed as [112]:
The 2SX water exchange model by Springer \textit{et al.} is only presented by the use of an extracellular contrast agent (GdDTPA$^{2-}$). With manganese compounds, the contrast agent (Mn$^{2+}$-ions) resides intracellular. In Figure~\ref{fig:icCA} all the different compartments in cardiac tissue are named and the water exchanges with an intracellular agent are shown.
It has also been universally assumed that the linear relationship between $R_1$ and [$CA$] in homogenous solutions applies to biological tissue. However, the hyperfine mechanism dominates the $CA$ increase of $R_1$, and requires molecular contact between $CA$ and water. The theory behind these contrast mechanisms will be presented in Section~\ref{sec:SB}.

34. Conclusions
Multi-component analyses of T1 and T2 revealed two compartments with different chemical environments in the heart cells detectable with 0.47 T MR
Mn2+ ions act as pure competitors to Ca2+ ion influx and, indirectly, as monitors of Ca2+ channel activity. However, Mn2+ uptake may occur at high extracellular Mn2+ concentrations (0-30 µM) without reducing bulk Ca2+ influx and contractile function.
Cell entry and retention of Mn2+ is inhibited early during ischemia, but is resumed in viable cells on reperfusion. Changing activities of Ca2+ channels and, possibly also, of mitochondria contribute to this phenomenon. This forms the basis for use of Mn2+ releasers as MRI markers of viability.
Recovery of longitudinal magnetization (T1 relaxation), measured by relaxometry in excised biopsies, is shortened in Mn2+ enriched hearts. T1 analysis reveals two components: one rapid (T1-1) and one slow (T1-2), representing water protons in different compartments. The T1-1 may be derived from the intracellular space or from mitochondria.
Water exchanges between tissue compartments are of central importance in MRI, and an intracellular, preferably intramitochondrial, contrast agent like Mn2+ opens for new possibilities for estimations of these phenomena
Diffusion experiments can be performed.
Relaxographic imaging (micro imaging).
Participations in clinical trials.

35. Main findings Two compartment model more suitable for T1 analysis
Close correlation between tissue Mn content and T1relaxation
Slow water exchange in excised cardiac tissue
Remarkably high relaxivity of intracellular Mn2+ ions
The main findings of my study is that:
-Mn2+ entry is dependent on Ca2+ channel activity
-That a two compartment model is suitable for T1 estimation, where a slow water exchange over the cell membrane is assumed.
-There is a close correlation between tissue Mn content and the relaxation parameters
-I will show you some figures from my results
-The potential of Mn-compounds as CA in cardiac MRI
-Physiological understanding of Mn2+-uptake and retention in the cardiomyocytes
-Can we resolve different cell compartments using contrast agents and low field MR?
-Where is the retained Mn2+ localized?

36. Physiology and Mn content