Download E I in cortical neuron balance

R. Grantyn
Dept. Sensory and Developmental Physiology,
Institute of Neurophysiology, Charité Berlin
JOHANNES-MÜLLER-INSTITUT FÜR PHYSIOLOGIE
Mainz GRK 2005
Development of synaptic inhibition
in cortical structures:
Focus on mechanisms regulating
the E/I balance of synaptic input
The group in Berlin
Christian
Henneberger
Jochen
Meier
Bhumika
Singh
Our collaborators in Madrid
Rubenstein & Merzenich* (2003):
More excitable (more weekly inhibited) cortex is, by its nature, more poorly
functionally differentiated; this type of cortex will lead to broad-ranging abnormalities in
perception, memory and cognition, and motor control. Moreover ‘noisy‘ (hyperexcitable,
poorly functionally differentiated) cortex is inherently unstable, and susceptible to epilepsy.
A short list of diseases associated with a disturbed balance between E and I:
Epilepsy
Nonsyndromic mental retardation
Autism
Rett syndrom
*Dept. Physiology and Otolaryngology, University of California at San Francisco
Does inhibition balance excitation in the cerebral cortex?
Typical compartmental model
to study synaptic actions
Synaptic conductance
as Σgtotal
Impact of synaptic
inhibition
From: Trevelyan, Watkinson (2005) Prog Biophys Mol Biol 87:109
Hippocampal cultures
An I synaptic input cancels an E input in accordance with
its time cource (τD 15-25 ms) and a space constant λ of 10 µm
From: Liu (2004) Nat Neurosci 7:373
Cerebral cortical cultures
The hypothesis of ‘homeostatic plasticity‘ of synaptic transmission in the cerbral cortex
From: Turrigiano, Nelson (2004) Nat Rev Neurosci 5:97
Main hypothesis: The number/activity of E synapses always matches
the number/activity of I synapses
Co-scaling model
E
I
Control
E/I 2:1
Enhanced E input
leads to enhanced I input
E/I remains 2:1
Our main question:
Which signals determine the ratio of E/I input on the level of
single cortical neurons developing under definded growth conditions?
Hippocampal cultures
The E/I ratio of synaptic terminals received by a given neuron
is determined on the basis of transfection-labeling and immunostaining
EGFP
EGFP
E/I ratio: typically 2:1
IPSC
τD=17.2 ms
Probability of PSC occurrence
The E/I ratio of synaptic input can be determined on the basis of
the probability of occurence of an PSC with a certain decay kinetics
E/I ratio: typically 3:1
EPSC
0.16
AMPA
0.08
GABA
0.00
0
1
log τD
2
Hippocampal neurons
Is the relationship between E and I synapses stable
throughout a population of neurons?
From: Salama-Cohen, Arevalo, Grantyn, Rodríguez-Tébar (2005) submitted
Cultured hippocampal neurons
Is the relationship between E and I synapses stable
over the length of a dendrite?
From: Liu (2004) Nature Neurosci 7:373
Postsynaptic adjustment of E synapses to altered network activity
Cerebral cortical cultures
GluR-IR GFP-IR
From: Wierenga, Ibata, Turrigiano (2005) J Neurosci 25:2895
Block of GABAARs stimulates the formation of inhibitory synapses in situ
After 2.5 h in ACSF+BMI 50 µM
After 2.5 hr in ACSF
P1 superior colliculus slices
Immediately fixed
O hr
From: Meier, Akyeli, Kirischuk, Grantyn (2003) Mol Cell Neurosci 23:600
The induction of new inhibitory synaptic terminals
requires the activation of PKC
**
***
20
0
80
- superior colliculus
- superficial layers only
- P1 WT mouse
- tangential slices
- submerged at 25 oC
- images at 100x
60
40
20
0
0h
ACSF
PKC block
PDBu
40
** *
100
VIAAT+
GAD65+ (%)
60
0h
ACSF
PKC block
PDBu
VIAAT / VF
80
**
***
******
From: Meier, Akyeli, Kirischuk, Grantyn (2003) Mol Cell Neurosci 23:600
Elimination of the PKC phosphorylation site Ser343 impairs
postsynaptic accumulation of GFP::γ2L
GABAAR γ2L contains an additional sequence of 8 amino acids,
among them the phosphorylation site Ser343
PDBu
Meier & Grantyn (2004) J Physiol 559:355
As a rule of thumb:
Conditions promoting postsynaptic GABAAR receptor accumulation also increase
the number of inhibitory terminals
Low GABAAR activity (comp. antag.)
Low level of group I mGluR activity
Block of Ca influx
Activation of Ca-dep PKC activity
Syp
VIAAT
Syp
VIAAT
GAD65
GABAAR
High GABAAR activity
(benzodiazepines)
High level of group I mGluR activity
Permanently elevated ic Ca
Depletion of Ca-dep PKC activity
Syp
VIAAT
GAD65
Block of repeatedly elevated local Ca transients promotes filopodial growth
O hr
From: Lohmann, Finski, Bonhoeffer (2005) Nat Neurosci 8:305
Does the number of newly formed synapses correlate
with dendrite number or branching?
O hr
P<0.01
Hippocampal neurons
Number of Syp+ terminals
30
20
10
0
0
10
20
30
Number of branch points
From: Singh, Henneberger, Meier, Rodríguez-Tébar, Grantyn (2005) submitted
40
Long ago, abnormal dendrite growth
has been considered to be a likely cause of mental retardation (MR)
O hr
Dendritic spine abnormalities in Golgi-stained
cerebral cortex of normal (a) and MR (b) infant
at 10 month
From: Purpura (1972) Science 186:1126
The more recent discovery that single gene defects on the X chromosome
affect the expression of rho GTPase-regulating proteins and produce
nonsyndromic MR (NS-MR) has revived the idea that some forms of MR
are primarily caused by abnormal dendrite structure
The OPHN1-defective young male displays
MR, reduced visual acuity and congenital
strabism; the mother had neither learning disabilities,
nor visual or oculomotor deficits
The cause: genomic deletion of exon 19
causing a frameshift at the site Xq12;
OPHN1 is proposed to act as inhibitor of rhoA and
might be necessary for the maintenance of spine motility,
at least during some critical steps in the development
of visuo-motor behavior
From Bergmann, ...& Ramaekers (2003) Brain 126:1537
The XL-MR genes ARHGEF6 and OPHN1 are likely to be involved in
the control of dendrite initiation an spine motility, respectively
XLMR genes
Rho family
GTPases
Effector
Cytoskeleton
Dendrite
growth
Predicted
deficits in
dendrite growth
ARHGEF6
(αPIX)
CDc42
?
Rac1
OPHN1
RhoA GTPase
Pak3
Actin filament
assembly
Dendrite initiation
ARHGEF6-/-: Reduced dendritic
branching
Actin filament
extension
Actomyosin
contraction
Dendrite elongation
Spine motility
ZZ-/-: reduced dendrite
elongation
OPHN1-/-: Increased threshold for
spine plasticity
Overexpressing or silencing NS-XLMR genes may allow one to manipulate
dendrite initiation, dendrite elongation or spine motility, and to test for changes
in synapse formation
OPHN1::DsRedE VGluT
VIAAT
15 µm
From: Meier, Grantyn, unpubl.
Overexpression of OPHN1 promotes both the formation of new primary dendrites
and
the formation of glutamatergic synapses
From: Meier, Grantyn, unpubl.
In search for genes able to alter dendrite outgrowth and E/I relationships
we also considered the group of ‘proneural‘ genes
O hr
(Ngn3?)
Ngn1/2
Neurogenin3
Mash1
From: Bertrand, Castro, Guellemot (2002) Nature Rev Neurosci 3:517
Overexpression of Ngn3 stimulates the outgrowth of new dendrites
O EGFP
hr
From: Sallama-Cohen, Arevalo, Grantyn, Rodríguez-Tébar (2005) submitted
MAP2
Overexpression of Ngn3 increases the E/I ratio
by suppressing the formation of inhibitory synapses
From: Salama-Cohen, Arevalo, Grantyn, Rodríguez-Tébar (2005) submitted
Thus, OPHN1 and Ngn3 are potentially dangerous genes...
Reverse scaling model
Control
E/I 2:1
Disinhibition of both
OPHN1 and Ngn3
E/I 8:1
If short-term+local:
Activity-dependent synapse
reorganization (Hepp-type plasticity)
If long-term+global:
States of overexcitation
(Brain pathology)
What controls the expression/activity of these genes?
Our experiments suggest a new major role of the neurotrophin NGF
From: Salama-Cohen, Arevalo, Grantyn, Rodríguez-Tébar (2005) submitted
Treatment of hippocampal neurons with nerve growth (NGF) factor strongly
promotes inhibitory synaptogenesis
(It also suppresses the outgrowth of new dendrites, not shown)
From: Salama-Cohen, Arevalo, Grantyn, Rodríguez-Tébar (2005) submitted
Tentative scheme on the role of NGF in the control of the E/I balance
in hippocampal neurons
Do all members of the neurotrophin family reduce E/I?
The I-promoting effect of NGF is new and surprizing, considering that
Brain-derived neurotrophic factor (BDNF) is well known for its epileptogenic action.
BDNF promotes LTP of glutamatergic synapses and causes a postsynaptic
depression of GABAergic inhibition...
bdnf-/-
P12
50 pA
50 ms
P15
**
eIPSC amplitude (pA)
200
160
120
80
40
0
From:
** **
+/+ -/P12
+/+ -/- -/P15 +BDNF
Henneberger, Jüttner, Rothe, Grantyn (2002) J Neurophysiol 88:595
Henneberger,...Grantyn (2005) Eur J Neurosci 21:431
How does the presence of BDNF in a postsynaptic neuron
affect the formation/stabilization of E vs. I synapses?
Control
EGFP
VGluT
tBDNF
VIAAT
From: Singh, Henneberger, Meier, Rodríguez-Tébar, Grantyn (2005) submitted
BDNF shifts the E/I balance towards excitation.
A BDNF-expressing neuron attracts a larger number of E synapses than
a neuron free of BDNF
Control is bdnf-/-
From: Singh, Henneberger, Meier, Rodríguez-Tébar, Grantyn (2005) submitted
Thus, NGF and BDNF are antagonists with respect to the E/I ratios
(and dendrite outgrowth, not shown)
The reverse scaling model
Control
E/I 1.5:1-2:1
Exogenous NGF
E/I~0.75
tBDNF, exogenous BDNF
E/I ~5:1