uppsala university pet centre per hartvig, uppsala university pet centre central dopaminergic and...

Uppsala University PET Centre

Per Hartvig, Uppsala University PET Centre

Central dopaminergic and serotonergic function studied

with positron emission tomography


´Biosynthesis of dopamine and serotonin

Precursor amino acid (tyrosine, tryptophan)

Hydroxylase (tetrahydrobiopterin)

L-dopa or 5-hydroxytryptophan

Aromatic amino acid decarboxylase(pyridoxine, vitamin B6)

Dopamine or serotonin


Effect of the the dopamine D2 antagonist OSU6162

0.006

0.007

0.008

0.009

0.01

0.011

0.012

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0.016

BASELINE OSU6162

Dopamine synthesis rate, k3, min-1 in Rhesus monkeys before and after 3 mg/kg of OSU6162.


Effect of tyrosine and R-tetrahydrobiopterin on dopamine synthesis rate and stabilization with OSU

6162

0.01

0.011

0.012

0.013

0.014

0.015

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0.017Baseline

OSU

OSU+R-BH4

OSU+R-BH4+TYR

Baseline

R-BH4

R-BH4+TYR

Figure 2. Attenuation of R-BH4-induced upregulation of striatal L-[-11C]DOPA influx by OSU6162

Tyrosine and biopterinincreases dopamine synthesis rate

The increased rate isstabilized to baselineby OSU6162


Clinical studies using OSU6162

• Parkinson disease

• Huntington chorea

• Schizophrenia

• (Alcoholism, smoking cessation)


Apomorphine effect on dopamine synthesis rate, k3

-25

-20

-15

-10

-5

0

0.1

% C

ha

ng

e i

n d

op

am

ine

sy

nth

es

is r

ate

0.1 mg/kg/h

0.5 mg/kg/h

1.0 mg/kg/h


”Tune” dependent change of dopamine synthesis rate

-20

-15

-10

-5

0

5

0.0110 0.0120 0.0130 0.0140 0.0150 0.0160

Baseline dopamine synthesis rate

%C

ha

ng

e a

fte

r a

po

mo

rp

hin

e in

fu

sio

n

Apomorphine 0.1 mg/kg induced decrease of dopa-mine change is dependent on baseline dopamine tuning in the Rhesusmonkey


Effect of L-DOPA in early and advanced Parkinsons disease

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-20

0

20

40

60

80

100

Change%

CaudateNucleus

PutamenVentral slice

Advanced PD

Early PD

PutamenDorsal slice

L-DOPA infusion saturatesdopamine synthesis in earlyParkinson´s disease

In advanced disease a loss ofpresynaptic dopamine receptorsexplains the induction of rate


Is L-DOPA an endogenous neurotransmitter ? (Miwa, Goishima 1993)

0.006

0.009

0.012

0.015

0.018

1 2

L-DOPA infusion 3 or 15 mg/kg/h induces an increase in dopamine synthesis rate.


Effect of 5R-erythro-tetrahydro-biopterin on dopamine synthesis

L-[-11C]DOPA

Baseline 6R-BH4 infusion

6R-erythro-5,5,7,8 tetra-hydrobiopterin, the endo-genous cofactor for thehydroxylases induces anincreased dopamine synthesis rate


6R-erythro-5,6,7,8-tetrahydrobiopterin

• Pharmacological effects– Release of monoamines and serotonin– Receptor effects– Enhances synthesis of monoamines– Hydroxylase AADC

– Tyrosine L-DOPA Dopamine

– Biopterin,BH4 Pyridoxin


• Clinical studies – Infantil autistic disorder – (Double blind 3 mg/kg cross over, randomized

study in children 4-8 y with PET, neurochemistry, immunology and clinical evaluation)

– Parkinson´s disease– Alzheimer´s disease

6R-erythro-5,6,7,8-tetrahydrobiopterin


The importance of radiolabelling position, 11C Dopa vs 18F-fluoro-dopa

Baseline 6R-BH4 infusion

6-fluoro-L-[-11C]DOPA

No effect of tetrahydrobiopterindue to increased synthesis of3-O-methyl dopa which is Passing to the brain giving increased background radioactivity in the reference


Multitracer protocol on dopamine function in toxicology

• Toxic Dopamine Presynaptic Postsynaptic

• reaction synthesis terminals terminals

• MPTP ()

• Manganese

• Wilson

• disease


Regulation of presynaptic dopamine function

• Supply of tyrosine and L-DOPA

• L-DOPA catalysing effect on synthesis

• Tetrahydrobiopterin effects on synthesis and release

• Presynaptic control in Parkinson disease

• Tune dependent control

• Dopamine stabilisers


Radiotracers used with PET for studies on presynaptic serotonin

• [-11C]L-tryptophan

• Carboxy - [11C]L-tryptophan

• 5-hydroxy- [-11C]L-tryptophan

• [11C] --methyl-L-tryptophan


Brain serotonin synthesis

NNH2

COOHHO

N

CHOHO

Tryptophan

NNH2

COOH

N

HO

NH2

N

HOCOOH

5-Hydroxytryptamine (5-HT)

5-Hydroxytryptophan (5-HTP)

5-Hydroxyindoleacetic acid (5-HIAA)

TH AADC

MAO AD

CH3


Positron emission tomography

11C-Tryptophan 5-hydroxy-11C-tryptophan


Brain utilization rate 5-Hydroxi (-11C)tryptophan 11C-tryptophan

SUV 0.90 1.1

Time to peak 15 min 15 min

Rate

Striatum 7.0 x 10-3 - 2.0 x 10-3

Frontal ctx 3.3 - 1.5

Temp ctx 1.2 - 0.7 ______________________________________________________________________________

11C-TRP give insignificant 11C-HT during PET, but might show some specific uptake of the tracer


Endogenous tracer substrates• Uptake into the target tissue, passing over the BBB

• Regional tissue accumulation of tracer

• Uptake into target cells

• Complex with enzymes in target cells

• Formation of active transmitter

• Uptake of active transmitter

• Release of transmitter to the synapse

• Binding to target receptors

• Metabolism of transmitter with cumulation


Biosynthesis of dopamine and serotonin

Precursor amino acid (tyrosine, tryptophan)

Hydroxylase (tetrahydrobiopterin)

L-dopa or 5-hydroxytryptophan

Aromatic amino acid decarboxylase(pyridoxine, vitamin B6)

Dopamine or serotonin


Effect of pyridoxine on the decarboxylation rate of 5-hydroxytryptophan

0

0.002

0.004

0.006

0.008

0.01

0.012

0.014

Baseline Pyridoxine

Rat

e, k

3

1

2

3

4

5

6

7


Selectivity of aromatic amino acid decarboxylase Treatment Decarboxylation rate, K3 of L-DOPA 5-hydroxitryptophan______________________________________________Pyridoxine 10 mg bolus 0 +Tetrahydrobiopterin 1 –15 mg/kg/h + 0


Effect of bolus doses of amino acid on decarboxylation rate of L-DOPA and 5-HTP

0

20

40

60

80

100

120

0 0.5 3 5 10 15 25 30

Dose of amino acid, mg/kg

k3,5

of

bas

elin

e

5-HTP

L-DOPA


Factors regulating uptake of amino acids to the brain and neurotransmitter synthesis

• Plasma amino acids• Diurnal rythm• Age• Gender• Food and drinking

– Proteins, carbohydrates and fat– Caffeine, ethanol

• Co-factors and vitamins (Pyridoxin B6, biopterin)• Drugs, SSRI


Effect of glucose infusion on uptake of 5-hydroxytryptophan derived radioactivity


Cerebral presynaptic synthesis in depression

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0 5 10 15 20 25 30 35 40 45 50

Time, minutes

No

rmal

ized

rad

ioac

tivi

ty

Healthy

Depressed


Decarboxylation rate of 5-HTP in different brain areas

Area Controls Depressed__________________________________________________Lateral frontal cortex 0.0011 min-1 0.0022Medial frontal cortex high 0.0029 0.0060 low 0.0001* 0.0042Caudate 0.0098 0.0098Putamen 0.0072 0.0081 ________________________________________________


Brain disposition of precursor amino acids

• Disease SUV Synthesis rate• Sex F > M• Depression Med pre frt ctx• Schizophrenia Do, ganglia • Tourette ganglia • OCD ganglia • ECT


Presynaptic serotonin function in social phobia(Ina Marteinsdottir et al 2001)

Method: Statistical evaluation of PET with 5-hydroxytryptophan by a pixel wise blocked analysis of variance contrasting differences between patients and controls.

Results: A a focal hyposerotonergic tonus in social phobics as compared to controls was evident in temporal cortex (periamygdala/rhinal, temporal pole and gyrus); frontal cortex, anterior cingulatae, right insula and left basal ganglia.


Regulation of aromatic amino acid decarboxylase activity for L-DOPA and 5-

hydroxytryptophan• Several mechanisms regulate amino acid transport to the brain

and presynaptic synthesis serotonin

• Synthesis of serotonin may be regulated by a similar decarboxylase enzyme but with different selectivity

• Modulating effect of enzyme co-factors e.g. tetrahydrobiopte-rin and vitamin B6 varied for the two transmitters

• Capacity limitation in transport and enzyme activity for 5HTP

• Limited capacity of amino acid transport may influence serotonin function with special impact in affective disorders


What is measured with 5-HTP and PET ?

• 11C-labelling in carboxy and -position of 5-HTP• Blockade of central decarboxylase with NSD1015• Bioanalysis of brain radioactivity using rat brain

homogenate by HPLC shows radiolabelled 5-HTP, serotonin and metabolites

• Calculated rates analysed of brain radioactivity in rat brain are similar to rates measured in monkey and man with 5-HTP and PET


Calculation of decarboxylation rate

• Brain reference region after validation of accumulation (Patlack plot, Hartvig et al 1992)

• Simulation of a brain refrence region with negligable 5HT synthesis gives rates close to measured (Blomquist et al 2001)

• Plasma as reference with metabolite correction shows regional 5HT rates in accordance with AADC activity (Hagberg et al JBFM, 2002)


• Rapid in vivo metabolism to radiolabelled products• Low plasma concentration of radioactivity • Serotonergic activity in most brain areas - no obvious

reference area in the brain • Steady state in the brain not established in 15-20 min• Limited capacity for transport over BBB and for

synthesis• Use of tracer may occur in non-serotonergic neurons

Limitations in studies with 5-hydroxy [11C]tryptophan


Theses at UUPC• Peter Bjurling Radiosynthesis• Lars Reibring- Depression• Joakim Tedroff L-Dopa in PD• Karl Johan Lindner Validation of 5 HTP• Anna Ekesbo DA degeneration, OSU• Richard Torstenson Regulation of DA• Ina Marteinsdottir SSRI responsive diseases• Pinelopi Merachtsaki Regulation of serotonin

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