Complexity and vulnerability of the GABAergic system in the

pathophysiology of the cerebral cortex

Senior collaboratorsHumberto Salgado (Hideyo Noguchi, UADY, Merida Mexico)Kuei Tseng (Rosalind Franklin University, Chicago, IL)

Laura Martinolich (Carnegie Mellon Univ., Pittspburgh, PA)Stefan Rose-John (Christian Albrecht Univ., Kiel Germany)

Students

Lu Dinh Anwesha BanerjeeAnkur PatelFrancisco Garcia-Oscos

Outline of the presentation:

1) Introduction on the GABAergic system and its role in neurologic and psychiatric disease

2) GABAergic signaling is impaired by stress through IL-6

3) The GABAergic system is severely impaired in a rat model of autism

Excitatory synapsesrelease glutamate

Inibitory synapsesrelease GABA

Shared factor among Epilepsy, Schizophrenia, and Autism

The balance between synaptic excitation and inhibition (= E/I ratio) is altered in favor of excitation

GABAergic neurons are potential target for cytotoxic damage caused by

- Genetic predisposition (fragile X chromosome, Rett syndrome, Prader-Willy, allelic variations in chromosome 14q22, and many other genetic or chromosomal variants)

- Stress (different types: metabolic, psychologic, toxicants, temperature, during different stages -prenatal, perinatal, developmental, or adult)

Data from many laboratories correlate GABAergic deficit with many neurological and psychiatric conditions like

Epilepsy Schizophrenia

Autism

(but also anxiety –including PTSD-, depression, tinnitus, and multiple other conditions)

Embryologic origin of the different IN types

The majority of murine cortical interneurons are derived from progenitor zones of the

Ganglionic Eminences (GE) in the ventral telencephalon. These zones can be divided into lateral, medial, and caudal (LGE, MGE, and CGE, respectively)

The two most studied groups of interneurons (PV and SST expressing) are derived exclusively from the MGE

It has also been reported that different regions of the MGE give rise to different subtypes, with progenitors giving rise to SST and PV being biased to dorsal versus ventral MGE, respectively

Multiple genes are responsible for the proper migration and differentiation of the different groups of cortical GABAergic INs

. All of these features are absent in PV-containing cells, but these cells express presynaptic M2 muscarinic receptors [59]. Knowing that anxiolytic effects of benzodiazepines are mediated solely by α2-subunit-containing GABAA receptors, it is important to note that synapses formed by CCK-positive basket cells on pyramidal cells operate mostly via α2-subunit-containing GABAA receptors, whereas PV-positive basket-cell synapses contain largely α1 subunits.

Major differences between parvalbumin (PV)- and cholecystokinin (CCK)-containing basket cells in their connectivity features and receptor expression patterns. Each axon terminal synapsing on the interneurons [glutamatergic (pink) or GABAergic (light blue)] corresponds to 1000 ∼synapses, reflecting true differences in the relative weight of excitatory and inhibitory inputs. CCK-containing cells express presynaptic CB1 cannabinoid receptors and postsynaptic 5-HT3 5- receptors and nicotinic α7 (or α4) ACh receptors. They receive input from serotonergic median raphe afferents and local interneuron-selective inhibitory cells expressing calretinin (CR IS cell; green).

From Freund, 2003 TINs

Variability of GABA-releasing neurons

Morphological features• Soma: shape; size; orientation; other• Dendrite: arborization polarity; branch metrics; fine structure; postsynaptic element; other• Axon: initial segment; arbor trajectory; terminal shape; branch metrics; boutons; synaptic targets;• Connections: chemical and electrical; source; location and distribution; otherMolecular features• Transcription factors• Neurotransmitters or their synthesizing enzymes• Neuropeptides

• Calcium-binding proteins (Parvalbumin, Calbindin, Calretinin)• Receptors: ionotropic; metabotropic• Structural proteins• Cell-surface markers• Ion-channels• Connexins• Transporters: plasma membrane; vesicularPhysiological features• Passive or subthreshold parameters: resting membrane potential; membrane time constants; inputresistance; oscillation and resonance; rheobase and chronaxie; rectification• Action potential (AP) measurements: amplitude; threshold; half-width; afterhyperpolarization;afterdepolarization; changes in AP waveform during train.• Dendritic backpropagation• Depolarizing plateaus• Firing pattern: oscillatory and resonant behaviour; onset response to depolarizing step; steadystate resp to dep step• Response to hyperpolarizing step: rectification; rebound• Spiking recorded extracellularly: phase relationship to oscillations; functional response specificity;cross-correlation and other dynamics• Postsynaptic responses: spontaneous and evoked; ratio of receptor subtypes; spatial and temporalsummation; short- and long-term plasticity; gap junctions

Petilla terminology: nomenclature of features of GABAergic interneurons of the cerebral cortex.Petilla Interneuron Nomenclature Group, Ascoli GA, Alonso-Nanclares L, Anderson SA, Barrionuevo G, Benavides-Piccione R, Burkhalter A, Buzsáki G, Cauli B, Defelipe J, Fairén A, Feldmeyer D, Fishell G, Fregnac Y, Freund TF, Gardner D, Gardner EP, Goldberg JH, Helmstaedter M, Hestrin S, Karube F, Kisvárday ZF, Lambolez B, Lewis DA, Marin O, Markram H, Muñoz A, Packer A, Petersen CC, Rockland KS, Rossier J, Rudy B, Somogyi P, Staiger JF, Tamas G, Thomson AM, Toledo-Rodriguez M, Wang Y, West DC, Yuste R.

Nat Neurosci 2008

A major problem even after the Petilla categorization of INs was a

Large overlapping of the families with any classification

Rudy:

Three Major NON overlapping Groups of Neurons:

(1) PV-expressing basket and chandelier neurons;

(2) SST-expressing neurons; and

(3) 5HT3a receptor-expressing neurons

account for nearly 100% of GABAergic interneurons in somatosensory cortex

Bernardo Rudy, Gordon Fishell, SooHyun Lee, Jens Hjerling-LefflerThree groups of interneurons account for nearly 100% of neocortical GABAergic neurons, 2010

Proportions of GAD-67 mRNA-expressing neurons also expressing mRNA for parvalbumin (PV), somatostatin (SST), or 5HT3aR based on the study by Lee et al. ( 2010).

Classification of neocortical GABAergic interneurons. Nearly 100% of all neocortical GABAergic neurons belong to one of three groups defined by the expression of parvalbumin (PV), somatostatin (SST), and the ionotropic serotonin receptor 5HT3a (5HT3aR

Non overlapping Distribution of three groups of GABAergic interneurons

GABAergic deficit and epilepsy

Several factors confer exquisite vulnerability to GABAergic interneurons, whether they are in the

NeocortexHippocampusStriatumOr other brain areas

1) A highly specialized genetic expression (Ca-binding proteins, peptides, specialized K+ channels), different and larger compared to protein expressed in glutamatergic cells may be responsible for GENETIC VULNERABILITY

2) Their small size, which makes them prone to metabolic damage or and physico-chemical challenge like the following can confer VULNERABILITY TO STRESS

- osmotic trauma- hypoxia- glucose deprivation- free radical (ROS-reactive oxygen species) oxydation- physical trauma

MECP2 MECP2 mutations explain most cases of Rett syndrome. These patients often display seizures. Amir et al. [147]; Buyse et al. [148]

GABRG2

Mutations in the gamma2 subunit of the GABAAR cause childhood absence epilepsy ± febrile seizure Wallace et al. [174]; Kananura et al. [183]

Truncation of GABRG2 causes generalised epilepsy with febrile seizure (GEFS) Harkin et al. [184]

GABRA1

Mutations in the alpha1 subunit of the GABAAR cause juvenile myoclonic epilepsy Cossette et al. [185]

Mutations in the alpha1 subunit of the GABAAR can also cause childhood absence epilepsy Maljevic et al. [186]

Scn1a↓ sodium currents are specific to GABAergic interneurons in Scn1a+/− and Scn1a−/− Yu et al. [199]

Selective loss of Scn1a in interneurons recapitulates seizure disorder Martin et al. [200]

Arx

Role in neuronal proliferation and migration Fricourt et al. [201, 202]Specific requirement of Arx for interneuron migration Friocourt and Parnavelas [203]; Poirier et al. [204]

Arx(GCG)10+7 mice display seizures including spasms and ↓ no. of CB and NPY interneurons Price et al. [206]

Cacna1aInterneuron selective ablation of Cacna1a leads to multiple types of generalised seizures incl. absences

Rossignol et al. [63] (abstract)

Cacnb4 Thalamic tonic GABAA currents enhance rebound bursting of TC cells in Cacnb4lh/lh Cope et al. [217]

Dlx1/2

Dlx1−/−Dlx2−/− mice die perinatally and display a failure of IN migration to cortex and olfactory bulb Anderson et al. [23, 218]; Bulfone et al. [219]

Dlx1−/−Dlx2+/− abnormal laminar distribution of IN and simplified morphology Cobos et al. [220]

Dlx1−/− morphological defect and postnatal loss of SST+/CR+ interneurons: spontaneous seizures Cobos et al. [221]

Nkx2.1

Nkx2.1−/− die perinatally. Nkx2.1 is required for MGE interneuron generation. Sussel et al. [222]

Interneuron specific removal of Nkx2.1 results in misspecification of MGE cells into CGE cells, and seizures

Butt et al. [22]

Sox6

Conditional loss of Sox6 in interneurons results in misplaced/ectopic and immature basket cells (loss PV)

Batista-Brito et al. [28]; Azim et al. [223]

Conditional loss of Sox6 in interneurons results in a severe epileptic encephalopathy

From Elena Rossignol, Neur Plast, 2012

genes causing epilepsy in humans and interneuron dysfunctions

GABAergic impairment in schizophrenia and psychosis

in schizophrenia the specific ability of fast-spiking interneurons to control and synchronize disparate cortical circuits is disrupted and that this disruption may underlie many of the schizophrenia symptoms.

The high vulnerability of corticolimbic fast-spiking interneurons to genetic predispositions and to early environmental insults--including excitotoxicity and oxidative stress--might help to explain their significant contribution to the development of schizophrenia

GAD67

↓ GAD67 in prefrontal cortexVolk et al. [109]Akbarian et al. [110]

Preserved # number of PV cells, cortexWoo et al. [111]Hashimoto et al. [112]

↓ GAD67 level in PV cells, cortex Hashimoto et al. [112]

Association with polymorphisms in GAD67 promoter Addington et al. [113]

Chandelier Decrease in chandelier cells cartridges (GAT1+) in prefrontal cortexWoo et al. [114]Volk et al. [115]

SST ↓ levels of SST in microarray analysis and ↓ number of SST cells, prefrontal cortex Hashimoto et al. [116]

NPY/CCK ↓ levels of NPY and CCK in microarray analysis Hashimoto et al. [116]

Gamma

Gamma oscillations are triggered by working memory tasks + selective attentionTallon-Baudry et al. [79]

Howard et al. [81]

Decreased power of cortical gamma oscillations and phase locking to memory taskSpencer et al. [82, 84]

Cho et al. [85]

Erb4 Selective interneuron loss of Erb4: “schizophrenia-like behaviors” Wen et al. [126]

Erb4/Nrg1 Erb4 in PV cells is required for Nrg1-dependant regulation of LTP (hippocampus) Chen et al. [127]

NR1 Selective loss of the NMDAr NR1 subunit in PV cells: decreased excitatory input to PV cells results in “schizophrenia-like behaviors” and ↓ expression of PV and GAD67 Belforte et al. [128]

BDNF BDNF regulates activity-dependant maturation of PV cells Bdnf−/− and Trkb−/− : ↓ synaptic GAD67 and GABA and behavioral anomalies

Huang et al. [129]Cotrufo et al. [130]

Hashimoto et al. [131]

PSA/NCAM Activity-mediated expression of PSA regulates PV cells maturation and visual plasticity Di Cristo et al. [132]

GammaGamma oscillations are triggered by stimulating PV cells: enhanced performance

Cardin et al. [77]

Sohal et al. [83]

Gamma oscillations depend on PV cells-mediated fast-synaptic inhibition Bartos et al. [72]

GABAergic system deficits in schizophrenia

A combination of genetic disposition, maternal stress, infection or malnutrition, and obstetric complications may lead to

-NMDAR hypofunction of cortical interneurons, especially parvalbumin (PV)-containing fast-spiking neurons, during development,

- pathophysiological phenotypes (cortical disinhibition, impaired oscillatory activity, dopaminergic dysregulation, and oxidative stress)

factors that may precipitate the emergence of major schizophrenia-like symptoms after adolescence.

GABAergic neuronal dysfunction hypothesis of schizophrenia

The relationship between

- DA and glutamate (hyperfunction)

And

- GABA and NMDA receptor (hypofunction)

is matter of debate and controversy

NMDAR deletion in cortical PV neurons would down-regulate GABA synthesis and release, which not only results in cortical disinhibition but also impairs the synchronized activity of principal neurons.

Cellular mechanisms for psychosis and schizophrenia

This may reduce the cortical output to VTA and thereby increases dopamine activity in the nucleus accumbens. modified from Lewis and Gonzalez-Burgos (2006).

Behavioral phenotype observed Onset of symptom Effect of social

isolation

Nest building impairment After 12 week old Exacerbated

Mating deficit After 12 week old

Impaired saccharine preference test N/D Exacerbated

Anxiety-like behaviors After adolescence Exacerbated

A transgenic mouse where NMDA receptors where not expressed selectively GABAergic IN displays the following alterations:

From Nakazawa et al., Neuropharmacology, 2012

How does stress lead to epileptic or psychotic episodes?

Neurotransmitters and other endogenous substances modulate brain activity during periods

of great energetic demand (physiological stress), by

1) Enhancing intrinsic (voltage dependent) neuronal excitability, mainly by blocking K currents

2) Temporarily altering synaptic function with sophisticated mechanisms, in order to co-ordinate behavioral responses in multiple brain areas, inducing behavioral states like:

-alert (high-levels of neuromodulators) vs. non alerted (low- levels)

and acting selectively on excitatory and inhibitory synapses

Many other substances are released after stress

Effect of pro-inflammatory cytokines

A large number of clinical as well as basic research studies in the last decade suggest that stress increases the levels of pro-inflammatory cytokines which affect the CNS

Immune system cells interact through small peptides that synthesize, release, and target other immune cells

cytokine network

Membrane receptors associated with interleukins (members of the cytokine)

Besides systemic immune cells, alsomicroglia, astrocytes and neurons produce interleukins in the CNS

Cytokines diffuse from and to system and CNS

The BBB limits diffusion from and to the brain.

Yet, specific transport systems allow most cytokines relatively large systemic-CNS diffusion

Epilepsy, ASD, schizophrenia and interleukin 6 (IL-6)

Experimental data show an association between high levels of IL-6 and the occurrence of several conditions,in particular:

- Epilepsy: high levels of IL-6 are present in the cephalo rachidian liquid of epileptic patients

- increased de la IL-6 has been shown to induce psychotogenic (schizophrenia) alterations in animal models

- an increased IL-6 in pregnant mother has been proposed to be a risk factor for offspring for diseases of the autistic spectrum (ASD)

How can IL-6 affectneuronal excitability?

Exemple: efects of TNF- a (another proinflammatory interleukin).

TNF-a enhances excitability acting directly at the synaptic level, increasing the excitatory – inhibitory ratio (E/I)

Can IL-6 increase cell excitability through a process similar to TNF-a?

Our work demonstrate that IL-6 impairs GABAergic, but not glutamatergic signals

time (min)

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in gabazine

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IL-6 decreases selectively GABAergic but not glutamatergic synaptic responses

Garcia-Oscos et al., Biol. Psych. 2012

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Garcia-Oscos et al., Biol. Psych. 2012

GABAAR trafficking

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Dependence on:

PI3K

Golgi-membrane protein redistribution

Membrane anchoring

Vesicle motility

The effect of IL-6 depends on protein trafficking

Garcia-Oscos et al., Biol. Psych. 2012

Does this phenomenon occur after actual systemic stress?

Injection of LPS(lipopolysaccharide)From gram-negative bacteriaActivation of TLR4

Electric footshock

Experimental induction of IL-6 with:

IL-6

IL-6 specificity of the effect of stress on synaptic transmission

IL-6 trans-signaling and its inhibition

Intraventricular injections of the IL-6 antagonist soluble gp130 prevent the effect of stress

AIP

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Garcia-Oscos et al., Biol. Psych. 2012

Conclusions:

IL-6 has is an important mediator of neuronal excitability after stress

IL-6 pathway antagonism has a therapeutic potential in the treatment of neurologic and psychiatric conditions

Diverse causes converge to increase IL-6 in pregnant mothers as risk factor for mental disease

We speculate that similar processes in uterus can be critical for the development of mental disease, opening a potential for pre- and peri-natal prevention, intervention, and treatment of interleukin 6 -related mental disease

Study of the GABAergic system in an autism model

Autistic spectrum disorders (ASD) are defined by

1) Impairment of communication2) Impairment of the social conduct 3) Repetitive behavior and limited interests

Other ASD common symptoms include anxiety, hyperactivity, sensory hypersensitivity, and general altered emotion

A currently favored theory on autism is that ASDs are due to an unbalance between inhibition and excitation in favor of the latter, whether because of too much excitability, o because of too little inhibition

GAD65/67 ↓ levels of GAD65/67 in cortex Fatemi et al. [143]

15q11-13Maternal duplications in 15q11-13 in nonsyndromic autism Baker et al. [145]including GABRA5, GABRG3, GABRB3 (GABAAR subunits) Hogart et al. [146]

Mice

Fmr1 Fmr1 k/o: behavioral anomalies improve with glutamatergic antagonists Dolen et al. [151]Bear et al. [152, 153]

Neuroligins/neurexins

NRL1/2 expression in nonneuronal cells trigger synapse formation in presynaptic cells Scheiffele et al. [154]

NL-1 overexpression in hippocampal neurons promotes assembly of excitatory and inhibitory synapses and knock-down results in loss of inhibitory > excitatory synapses

Chih [155]

Presynaptic β-neurexin induces GABA and glutamate synapse differentiation in postcell Graf et al. [156]

NRL1,3,4 localise at glutamatergic synapses, NRL2 at both excitatory and inhibitory Graf et al. [156]

MecP2Binds methylated CPG islands and exerts epigenetic control of UBE3A and GABR3 Samaco et al. [157]Interneuron selective loss of MecP2 recapitulates the Rett-like behavioral aN in mice Chao et al. [158]

uPAR, HGF, MET

uPAR−/− displays 50% loss of IN in cortex and seizure susceptibility Powell et al. [11]

uPAR is required for the processing of HGF (an interneuron motogen), Powell et al. [159]

HGF, through its receptor MET, can rescue the phenotype of uPAR−/− mice Bae et al. [160]Interneuron selective MET ablation: ↓ PV cortex, ↑ striatal PV cells, disrupts reversal learning Martins et al. [161]

GABA signaling related chromosomal or genetic alterations in autism

The distribution of parvalbumin positive cell is altered in VPA rats

While PV+ cells have a relatively uniform distribution in the neocortex, VPA animals alternate areas of the brain with normal distribution with others shown PV+ cell clusters and void areas

Unpublished data

GABAergic transmission is severely altered in VPA rats

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Unpublished data

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Miniature currents (mIPSC) have lower frequency and slower decay time in VPA rats

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Col 7 vs Col 8 Col 9 vs Col 10 Col 11 vs Col 12 Col 13 vs Col 14 Col 15 vs Col 16 Col 17 vs Col 18 Col 19 vs Col 20 Col 21 vs Col 22 Col 23 vs Col 24 Col 25 vs Col 26 Col 27 vs Col 28 Col 29 vs Col 30

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Pharmacologic differences in GABAergic currents

VPA synaptic currents loose their sensitivity to zolpidem (modulator of intrasynaptic subunity alpha1-4 –not effective on extrasynaptic alpha 5subunits, but not topero no al gaboxadol which modulates the extrasynaptic delta subunit Unpublished data

β and α2 adrenoceptors are present in PV+ interneuronas

(Salgado et al., Synapse, 2011)

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Impaired adrenergic modulation in VPA rats

Unpublished data

Conclusions

GABAergic signaling is severely impaired in the VPA model of autism, suggesting that behavioral or pharmacological therapies aimed at enhancing GABAergic function in a developing ASD may be beneficial

Colaboradores senior

• Humberto Salgado (Hideyo Noguchi, UADY, Merida Mexico)• Kuei Tseng (Rosalind Franklin University, Chicago, IL)• Emily Tansey (University of Ohio, Columbus OH)• Chris McBain (NICHD, NIH, Bethesda MD)• Saobo Lei (University of North Dakota, Grand Forks ND)• Patrick Kanold (Univ. Maryland College Park, MD)• Jorge Flores-Hernandez (BUAP, Puebla MX)• Juan Carlos Pineda (UADY, Mérida MX)• Rodrigo Paz (Hospital Horwitz Barak, Santiago, Chile)

Estudiantes

Justin Nichols (Baylor, Houston)Timothy Bellay (Brown, NH)Lu Dinh (Fort Worth, TX)Anwesha Banerjee (UTD)Ankur Patel (UTSW)Laura Martinolich (Univ. Pittsburgh)Francisco Garcia (UNT, Denton)Clay Brown (UTD)Ruben Wolff (Politechn. Univ. Zurich)Dayra Lorenzo (Politechn. Univ. Zurich)Erica Sherry (UTD)

NE modulates GABA

Acute effect of NE

GABAA Glu AMPA

Layer I ↓ pre & post (α1)

↓post (α1)

Layer II/III ↑pre (α2, β) ↓post (α1)

This work, together with previous work from ours and other laboratories, show an important difference between muscarinic and adrenergic modulation of synaptic response

Ac. effect of Ach(oxotremorine)

GABAA Glu AMPA

Layer I ↓ pre (M1, M2) ↓pre (M1-M3?)

Layer II/III ↓ pre (M1, M2) ↓pre (M1-M3?)

Conclusions

Altogether, these data suggest that endogenous factors (neuromodulators) and exogenous factors (stress and associated molecules) can act differentially or specifically on GABAergic interneurons

GABAergic system is impaired in an ASD model (VPA)

Registros dobles demuestran la componente presinaptica de la modulacion adrenergica de los eIPSC de L2/3

Efecto presinaptico de la NE sobre las eIPSC de L2/3

Neurona piramidal(capa 2/3)

Neurona GABAergica(capa 1)

(Salgado et al., Synapse, 2011)

el bloqueo de la PLC inhibe el decremento de LI eIPSC pero no el aumento de los LII/III eIPSC

una componente postsinaptica de la modulacion por NE es evidente por el decremento de la señal inducida por aplicacion de muscimol (agonista GABAA)

(Salgado et al., Synapse, 2011)

pulse number

0 1 2 3 4 5 6IP

SC

n /

IP

SC

1

0.5

1.0

pulse number

0 2 4 6 8 10

IPS

Cn

/ I

PS

C1

0.5

1.0

pulse number

0 5 10 15 20

IPS

Cn

/ I

PS

C1

0.5

1.0

1.5

A B

C D

E F

50

pA

100 ms

50

pA

100 ms

50

pA

100 ms

control

NE

control

NE

control

NE

**

**

*

**

5 Hz

10 Hz

20 Hz

layer I stimulation

pulse number

0 1 2 3 4 5 6

IPS

Cn

/ I

PS

C1

0.5

1.0

pulse number

0 2 4 6 8 10 12

IPS

Cn

/ I

PS

C1

0.5

1.0

pulse number

0 5 10 15 20 25

IPS

Cn

/ I

PS

C1

0.0

0.5

1.0

10

0 p

A

100 ms

10

0 p

A

100 ms

10

0 p

A

100 ms

A B

C D

E F

5 Hz

10 Hz

20 Hz

layer II / III stimulation

controlNE

control

NE

control

NE

*

(Salgado et al., Hear. Res., 2011)

La respuesta GABAergica de capa 1 o de capa 2/3 es opuesta a altas frecuencias

Interpretacion

La interpretacion de estos resultados es dificil por via de la falta de conocimiento de los roles especificos de las inhibicion de capa I y capa II/III

Hay que considerar que

1) las fibras GABAergica de L I (long-distance or Martinotti cells) llevan informacion sobre el contexto (informacion global, no tonotopica, de motivacion o emotiva, Malmierca 2003),

2) las fibras GABAergicas de L II/III son activadas por circuitos locales, y llevan informacion sensorial especifica (dependente de frecuencia) sincronizando los circuitos locales con oscilaciones gamma:

La NE podria contribuir a la integracion de la señal sensorial (auditiva) aumentando la eficacia de las oscilaciones gamma (mediadas por interneuronas de PV),

bajando el umbral para la activacion de las celulas piramidales en respuesta a la activacion del sistema limbico (bajando la inibicion en capa 1)

time (min)0 20 40 60

eIP

SC

s A

mp

litu

de

(pA

)

-200

-150

-100

-50

0 IL-6

100ms

50

pA

***

eIP

SC

s A

mp

litu

de

(pA

)

0

100

200

300

con

tro

l

IL-6

PP

R

0.0

0.4

0.8

1.2

IL-6

con

tro

l

n.s.

1

2

1

2

time (min)

0 20 40 60 80 100 120

eEP

SC

s A

mp

litu

de

(pA

)

-25

-20

-15

-10

-5

IL-6

2

2

1

1

50ms

10 p

A

eEP

SC

s A

mp

litu

de

(p

A)

0

10

20

30

40

50

n.s.

n.s.

con

tro

l

IL-6 co

ntr

ol

IL-6PP

R

0.0

0.4

0.8

1.2

2D Graph 1

E F G

A B C D

H

CV

0.0

0.1

0.2

0.3

con

tro

l

IL-6

n.s.

CV

0.0

0.1

0.2

0.3

n.s.

IL-6

con

tro

l

IL-6 decreases selectively GABAergic but not glutamatergic synaptic responses