Transcript Document 7723956

Overview of the striatum
dorsal
•Dorsal striatum important for
motor planning and adaptive
behaviors. Main inputs arise
from motor cortex.
•Disorders involving dorsal
striatum include Parkinson’s
disease and Huntington’s
disease.
ventral
•Ventral striatum (nucleus
accumbens) is important for
reward. Main inputs arise from
amygdala and prefrontal cortex.
•Drug addiction, depression are
disorders involving the striatum.
Neurotransmission and neuromodulation:
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Adrenaline (ca. 1900-)
Acetylcholine (ca. 1900-)
Neuropeptides (1930-)
Amino Acids (GABA and Glutamate) (1950-)
Monoamines (5-HT and Dopamine) (1950-)
Endogenous opioids (1975-)
Endogenous cannabinoids (1992-)
-CB1 receptor cloned (1990)
-discovery of endogenous endocannabinoids (1992)
-modulation of synaptic function (2001)
Endocannabinoid signaling in the striatum
CB1 immunoreactivity
Egertova and Elphick, J Comp Neurology (2000)
Corticostriatal LTD
Gerdeman et al., Nat Neurosci (2002)
Anatol Kreitzer
Sheela Singla
Brad Grueter
Percy Luu
Experimental Methods
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Acute coronal brain slices – recordings performed from medium spiny
neurons (MSNs) in dorsolateral striatum of 3- to 4-week-old rats and mice
CsMeSO3 based-internal solution, 0.2 mM EGTA, QX-314 to block Na
currents
External saline: 2 mM Ca, 1 mM Mg. Picrotoxin (50 mM) present to block
GABAA-mediated currents.
Experiments performed at room temperature (RT) and 30-32 °C
EPSC (% baseline)
Tetanus induces endocannabinoid-mediated LTD
(eCB-LTD)
100
50
control (n=15)
AM251 (n=6)
0
0
10
20
30
Time (min)
40
EPSC (% baseline)
eCB-LTD requires mGluRs
100
50
LY341495 (n=5)
0
0
10
20
30
Time (min)
40
Endocannabinoids are released by activation of
postsynaptic mGluRs
EPSC (% baseline)
DHPG (100 uM)
100
50
hp:-50 (n=11)
hp: -50, AM251 (n=5)
0
0
10
20
Time (min)
30
40
EPSC (% baseline)
eCB-LTD requires D2 receptors
100
50
sulpiride (n=5)
0
0
10
20
30
Time (min)
40
sulpiride=D2 antagonist
Model of endocannabinoid release and LTD in the
striatum
1. mGluR activation, combined
with subthreshold
depolarization, leads to
endocannabinoid (eCB)
synthesis and release
2. Dopamine D2 receptor
activation enhances eCB
synthesis, while D2 inhibition
reduces eCB synthesis
3. Released eCBs bind to
presynaptic CB1 receptors and
induce presynaptic LTD
4. Neurotransmitter release is
reduced for tens of minutes or
more
(Kreitzer and Malenka, 2005)
The striatum is not homogeneous
(2 populations of medium spiny neurons)
dorsal
ventral
Basal ganglia motor circuit: direct pathway
Motor Cortex
Premotor
Cortex
Glutamate
Caudate/
Putamen
GPe
GPi/SNr
Dopamine
+
GABA
Motor
Thalamus
SNc
STN
Direct pathway MSNs express:
Dopamine D1 receptors, Muscarinic
M4 receptors, Dynorphin, Substance P
Basal ganglia motor circuit: indirect pathway
Motor Cortex
Premotor
Cortex
Glutamate
Caudate/
Putamen
GPe
GPi/SNr
GABA
Dopamine
-
SNc
Motor
Thalamus
STN
Indirect pathway MSNs express:
Dopamine D2 receptors, Enkephalin
Basal ganglia motor circuit:
Parkinson disease
Motor Cortex
Premotor
Cortex
+ Glutamate
Striatum
GPe
GPi/SNr
Dopamine
+
- GABA
-
X
SNc
Motor
Thalamus
STN
Nucleus accumbens direct and indirect pathways?
mPFC
Ventral
Subiculum
NAc
D1
BLA
D2
VP
VTA/Substantia Nigra
M4- and D2-GFP BAC-transgenic mice label direct
and indirect pathway medium spiny neurons
1
2
3
1. CPu
2. GPe
3. SNr
M4-GFP
D2-GFP
GENSAT Program (N. Heintz, M. Hatten, W. Yang)
Direct and indirect pathway synapses display
different paired-pulse ratios
1.5
200 pA
50 ms
1.0
direct (M4+)
indirect (M4-)
A2/A1
1.5
1.0
0
Ba
250
500
ISI (ms)
D2+
2.0
A2/A1
Ab
M4+
0
Bb
250
ISI (ms)
500
1.5
1.5
200 pA
50 ms
1.0
0
250
500
ISI (ms)
A2/A1
A2/A1
Aa 2.0
direct (D2-)
indirect (D2+)
1.0
0
250
ISI (ms)
500
Indirect pathway synapses have a higher
release probability
20 pA
0.2 s
0.5
(5)
4
2
(4)
0
D
Mean Amp (pA)
C 1.0
1.0
Cum. Prob.
B
Mean Freq (Hz)
Indirect
Direct
Cum. Prob.
A
0.5
I
0.0
20
(4)
(5)
D
I
10
0
0.0
0
1
2
Inter-event Interval (s)
0
50
Amplitude (pA)
Direct and indirect pathway synapses display
different paired-pulse ratios in nucleus accumbens
D2(+)
D2(-)
PPR
1.5
1.0
0.5
0
100
200
Interstim interval (msec)
7
*
6
5
4
3
2
1
0
D2(+)
D2(-)
Average mEPSC Amplitude (pA)
Average mEPSC Frequency (Hz)
Indirect (D2+) pathway synapses have a higher
release probability in nucleus accumbens?
25
20
15
10
5
0
D2(+)
D2(-)
Indirect pathway medium spiny neurons are
more excitable
B
40
Freq (Hz)
Ab
Indirect
direct
indirect
20
0
0
20 mV
100 ms
C
ISI (norm.)
Aa
Direct
2
200 400
I (pA)
direct
indirect
1
0
0
10
Spike #
Indirect (D2+) pathway medium spiny neurons
are more excitable in nucleus accumbens
Indirect
D2(+)
D2(-)
25
Frequency (Hz)
Direct
30
*
20
*
15
10
5
0
0
100
200
I (pA)
300
400
EPSC (% baseline)
Endocannabinoid-mediated LTD occurs only at
indirect pathway synapses
100
50
direct (D2 neg) (n=8)
indirect (D2 pos) (n=6)
0
0
10
20
Time (min)
30
40
150
D2(-)
100
50
0
0
10
20
30
Time (min)
40
EPSC Amplitude (% Baseline)
EPSC Amplitude (% Baseline)
Endocannabinoid-mediated(?) LTD occurs only at
indirect pathway synapses in nucleus accumbens
150
D2(+)
100
50
0
0
10
20
30
Time (min)
40
EPSC (% baseline)
Indirect pathway eCB-LTD requires dopamine
D2 receptor activation
100
50
AM251
sulpiride
5-HT, NE antagonists
0
0
20
Time (min)
40
AM251: cannabinoid CB1 receptor antagonist
sulpiride: dopamine D2 receptor antagonist
CB1 receptor agonists inhibit transmitter
release at both direct and indirect pathway
synapses
AM251
EPSC (% baseline)
WIN
100
50
direct
indirect
0
0
20
Time (min)
40
WIN55,212: cannabinoid CB1 receptor agonist
AM251: cannabinoid CB1 receptor antagonist
Postsynaptic mGluR-driven endocannabinoid
release occurs primarily at indirect pathway
synapses
EPSC (% baseline)
DHPG
100
50
direct
indirect
0
0
20
Time (min)
40
150
D2(-)
100
50
100mM RS-DHPG
0
0
10
20
30
40
Time (min)
50
60
EPSC Amplitude (% Baseline)
EPSC Amplitude (% Baseline)
Postsynaptic mGluR-driven LTD occurs primarily at
indirect pathway synapses in nucleus accumbens
150
D2(+) (-50mV)
100
50
100mM RS-DHPG (-50mV)
0
0
10
20
30
40
Time (min)
50
60
EPSC (% baseline)
Indirect pathway eCB-LTD is absent in an animal
model of Parkinson disease
100
50
indirect (n=6)
indirect, reserpine treated (n=7)
0
0
20
Time (min)
40
Endocannabinoid biosynthesis and inactivation
Phosphatidylethanolamine (PE)
N-Acyltransferase
Calcium
Phosphatidylinositol (PI)
Phospholipase Cb
Receptor activation
(mGluR or mAChR)
Calcium
N-arachidonoyl PE
NAPE-PLD
Calcium
Anandamide (AEA)
FAAH
Arachidonic acid +
(URB597)
Ethanolamine
Diacylglycerol (DAG)
DAG Lipase
2-arachidonoylglycerol (2-AG)
MGL
(URB754)
Arachidonic acid + glycerol
EPSC (% baseline)
EPSC (% baseline)
Rescue of indirect pathway LTD in a Parkinson’s
disease model (reserpine)
100
50
reserpine-treated, quinpirole (n=5)
reserpine-treated, URB754 (n=6)
100
50
reserpine, URB597
0
0
0
0
20
Time (min)
40
20
Time (min)
40
quinpirole: dopamine D2 receptor agonist
URB754: monoacylglycerol lipase (MGL) inhibitor
URB597: FAAH inhibitor
Rescue of indirect pathway LTD in a Parkinson’s
disease model (6-OHDA)
EPSC (% baseline)
EPSC (% baseline)
150
100
50
6-OHDA
6-OHDA, quinpirole
100
50
6-OHDA, URB597
0
0
0
20
Time (min)
40
0
20
Time (min)
quinpirole: dopamine D2 receptor agonist
URB597: FAAH inhibitor
40
An important role for dopamine D2 receptors and
cannabinoid CB1 receptors in movement
D2 -/- mouse
catalepsy
Locomotor activity
Baik et al. (1995), Nature
wt
D2 -/-
wt
wt
D2 -/D2-/-
CB1 -/- mouse
Zimmer et al. (1999), PNAS
Inhibition of eCB degradation enhances D2-receptormediated recovery from catalepsy in animal models
of Parkinson disease
Inhibition of eCB degradation enhances D2-receptormediated recovery of locomotor activity in animal
models of Parkinson disease
Basal ganglia motor circuit:
Parkinson’s disease treatments
Motor Cortex
Premotor
Cortex
+ Glutamate
Caudate/
Putamen
GPe
GPi/SNr
eCB biosynthesis
Dopamine
(L-DOPA)
+
- GABA
-
X
SNc
Pallidotomy
STN
Motor
Thalamus
STN Pacemaker
Conclusions
• eCB-LTD in the striatum is restricted to indirect
pathway cells
• it is synapse specific because it requires presynaptic
activity in addition to CB1 receptor activation
• eCB-LTD is prevented by dopamine depletion but can
be rescued by a D2 agonist and an inhibitor of
eCB degradation
• inhibition of eCB degradation improves motor
performace in animals models of Parkinson
disease
• D2(+) and D2(-) cells in the accumbens also have
different synaptic properties
Anatol Kreitzer
Sheela Singla
Brad Grueter
Percy Luu
Nathaniel Heintz (Rockefeller)
X. William Yang (UCLA)
Is presynaptic eCB-LTD activity-dependent?
-Spillover onto synapses that did
not participate in depolarizing the
postsynaptic cell could cause LTD.
-Synapse specificity may be
achieved if LTD depended on
presynaptic activity and not just
receptor activation.
LTD
cannabinoids
LTD?
Testing the effect of synaptic activity on LTD
pathway 1: no
synaptic stimulation
pathway 2: 0.05 Hz-1 Hz
synaptic stimulation
A CB1 agonist is sufficient to induce LTD in the
striatum, but not the cerebellum
EPSC (% baseline)
A
B
WIN
AM251
WIN
100
100
50
50
cerebellum
striatum
0
AM251
0
0
20
Time (min)
40
0
20
Time (min)
40
WIN-induced LTD is activity-dependent
10 min WIN
20 min WIN
eCB-LTD is activity-dependent
path 1, path 2
Imbalanced activity in striatal motor circuits in vivo
in a Parkinson disease model
(direct)
(indirect)
Mallet et al (2006) J Neurosci
Muscarinic M1 receptors do not regulate striatal
endocannabinoid release or eCB-LTD
EPSC (% baseline)
A
B
pirenzepine
100 Hz tetanus
100
100
50
50
hp: -70 mV
hp: -50 mV
sulpiride
sulpiride and pirenzepine
0
0
0
10
20
Time (min)
30
0
20
Time (min)
40
pirenzepine: muscarinic M1 receptor antagonist
sulpiride: dopamine D2 receptor antagonist
Dopamine-depleted mice do not have altered
paired-pulse plasticity or sensitivity to cannabinoids
1.5
WIN
PPR
EPSC (% baseline)
indirect (D2+)
indirect (D2+), dopamine-depleted
1.0
AM251
100
50
dopamine-depleted
0
0
250
ISI (ms)
500
0
20
Time (min)
40
Indirect pathway synapses have larger
NMDA/AMPA current ratios
A 3 0.6
direct
100 pA
20 ms
A2
indirect
NMDA/AMPA ratio
A1
*
0.4
0.2
0.0
direct
indirect
Inhibition of eCB degradation enhances D2-receptormediated recovery from catalepsy in animal models
of Parkinson disease
Inhibition of eCB degradation enhances D2-receptormediated recovery of locomotor activity in animal
models of Parkinson disease
Signaling through CB1 receptors
THC (partial agonist)
CB1R
heterotrimeric
G-protein
2-AG (endogenous agonist)
agonist
Anandamide (endogenous agonist)
Adapted from DiMarzo et al., Nature Reviews Drug Discovery (2004)
Endocannabinoid biosynthesis and
inactivation
Phosphatidylethanolamine (PE)
N-Acyltransferase
Calcium
Phosphatidylinositol (PI)
Phospholipase Cb
Receptor activation
(mGluR or mAChR)
Calcium
N-arachidonoyl PE
NAPE-PLD
Calcium
Anandamide (AEA)
FAAH
Arachidonic acid +
Diacylglycerol (DAG)
DAG Lipase
2-arachidonoylglycerol (2-AG)
MGL
Ethanolamine
Arachidonic acid + glycerol
Inhibition of endocannabinoid degradation enhances
D2-receptor-mediated recovery of movement in a
Parkinson disease model
*
0
+ quinpirole/URB754
400
+ quinpirole
+ quinpirole/URB754
+ quinpirole
0
+ URB754
20
800
+ URB754
*
40
Locomotor Activity
reserpine-treated
120
Distance Traveled (cm)
B
Catalepsy
reserpine-treated
Descent Latency (s)
A
D2 receptor activation enhances
endocannabinoid release in response to brief
mGluR activation
EPSC (% baseline)
DHPG
100
50
control
quinpirole
0
0
10
20
Time (min)
30
Signaling through CB1 receptors
THC (partial agonist)
heterotrimeric
G-protein
agonist
2-AG (endogenous agonist)
Anandamide (endogenous agonist)
Adapted from DiMarzo et al., Nature Reviews Drug Discovery (2004)
Endocannabinoid biosynthesis and inactivation
Phosphatidylethanolamine (PE)
N-Acyltransferase
Calcium
Phosphatidylinositol (PI)
Phospholipase Cb
Receptor activation
(mGluR or mAChR)
Calcium
N-arachidonoyl PE
NAPE-PLD
Calcium
Anandamide (AEA)
FAAH
Arachidonic acid +
(URB597)
Ethanolamine
Diacylglycerol (DAG)
DAG Lipase
2-arachidonoylglycerol (2-AG)
MGL
Arachidonic acid
(URB754)
+ glycerol
CB1 receptor distribution in rat brain
(from Pettit et al., 1998)
Endocannabinoids are not released by medium
spiny neuron depolarization
A
( ) control
D t=10 s
control
D t=10 s
EPSC (% baseline)
100
50
( )
1 sec depol (n=4)
5 sec depol (n=3)
200 pA
0
10 ms
-50
0
50
100
mGluR-mediated endocannabinoid release
requires L-type calcium channels
EPSC (% baseline)
DHPG (100 uM)
100
50
hp:-50 (n=11)
hp:-50, nitrendipine (n=8)
0
0
10
20
Time (min)
30
40
mGluR-mediated endocannabinoid release is
enhanced by subthreshold depolarization
EPSC (% baseline)
DHPG (100 uM)
100
50
hp: -70 (n=9)
hp: -50 (n=11)
0
0
10
20
Time (min)
30
40
Medium spiny neurons: up and down states
Down state: -60 to –90 mV
calcium influx mediated by Tand R-type VSCCs
Up state: -40 to –70 mV,
calcium influx mediated by Land R-type VSCCs
Wilson and Kawaguchi (1996), J Neurosci
Carter and Sabatini (2004), Neuron
Stern et al.(1998), Nature
Imbalanced activity in striatal motor circuits in vivo
in a Parkinson disease model
(direct)
(indirect)
Mallet et al (2006) J Neurosci