Title: Effects of Whisker-trimming on GABAA receptors in SI Cortex
Or in adult barrel cortex
CHAPTER 1
INTRODUCTION
Sensory deprivation has been a productive approach to investigate the effects of
environmental stimuli on adult and developing brain. Whereas lack of normal excitatory inputs
leaves some cortical neurotransmitter systems unaffected (Goodman et al., 1993; Schlaggar et al,
1993), it can lead to down regulation of the gamma-aminobutyric acid (GABA)-ergic system. For
instance, the cortex of monkeys, cats, and rats shows particular changes in GABAergic
components after different types of sensory or visual deprivation. In the rat somatosensory cortex,
deprivation of whisker input results in decreases in GABAergic circuitry elements, such as the
number and proportion of GABA-immunoreactive synaptic contacts in layer IV (Micheva and
Beaulieu, 1995), glutamic acid decarboxylase (GAD) levels (Akhtar and Land, 1991), and
muscimol binding to GABAA receptors (Fuchs and Salazar, 1998).
Alterations in specific GABA receptors subunits might help understand GABAergic
function. This is the first investigation where whisker deprivation regulates specific GABAA
receptor subunits.
The Whisker to Barrel System of the Rat
The rodent somatosensory whisker to barrel pathway has several features that make it a
desirable system to study sensory deprivation. For instance, there is a 1:1 topographic relationship
1
between each of the whiskers in the rat’s face and a group of neurons that constitute a ‘barrel’ in
layer VI of Somatosensory Cortex (SI) (Woolsey and Van der Loos, 1970): each barrel responds
primarily to one principal whisker. This feature enabled the discovery of cytoarchitectonic
(Woolsey and Van der Loos, 1970; Welker and Woolsey, 1974; Van der Loos and Woolsey, 1973)
and physiological (Welker, 1971, 1976; Simons, 1978; Simons and Woolsey, 1979) effects of
whisker stimulation and/or deprivation, and to investigate the experience-dependent maintenance
of synapses (Micheva and Beaulieu, 1996), neurotransmitters (Micheva and Beaulieu, 1995), and
neurotransmitter receptors (Fuchs, 1995). Moreover, the SI cortex of rats is accessible to surgical
procedures, allowing for a variety of chemical, physiological and mechanical preparations and
manipulations, such as measurements of changes of GABA receptor subunits after topical cortical
blockade of NMDA receptors (Penschuck et al., 1999).
Sensory Deprivation and GABA A Receptors
Effects of sensory deprivation on GABAergic cortical circuitry have been widely studied.
Pioneer studies on the adult monkey’s visual system showed that depriving visual input from one
eye results in decreases of both GABA and its synthesizing enzyme GAD in the deprived cortical
neurons (Hendry and Jones, 1986). In the SI cortex of adult rodents, similar effects of deprivation
have been observed. First of all, GAD is reduced in deprived barrels after trimming whiskers for 6
weeks beginning in the adult, but not beginning in the neonate (Akhtar and Land, 1991).
Physiological studies showed that simply trimming rat whiskers leads to signs of disinhibition in
the deprived barrel neurons, such as higher spontaneous activity, and a decreased selectivity to
respond to specific angles of whisker deflection (Simons and Land, 1987). These physiological
changes remain even after allowing neonatally deprived rats to regrow their whiskers for several
2
weeks, indicating the dramatic, long-lasting effect of neonatal deprivation. What is the chemical
basis of these physiological changes? Since GABA is the main inhibitory neurotransmitter in
cortex, it was important to consider GABA and its receptors as suitable candidates responsible for
these physiological changes. Blocking GABAA receptors with the antagonist bicuculline results in
signs of cortical disinhibition (Kyriazi et al, 1996) that are similar to those from deprived barrel
cortex. Furthermore, binding of the GABA agonist [3H]muscimol, which selectively binds to
GABAA receptors, is reduced in the deprived barrels (Fuchs and Salazar, 1998). This effect was
observed in both neonatally and adult deprived rats, and was still present even after allowing the
rats to grow their whiskers for ten additional weeks after the trimming period. These overall
decreases after deprivation were suggested as a down-regulating mechanism that compensates for
the reduced sensory input (Fuchs and Salazar, 1998). Recent studies showed that whisker trimming
for 2 months, starting at birth, reduced the numerical density of both intracortical and
thalamocortical symmetrical synapses, upon inhibitory neurons in barrel neuropil (Sadaka et al.,
2003).
GABA inhibitory effects in cortex depend on the types of GABA receptors involved.
Inhibition through the GABAA receptor, a chloride channel itself, is relatively fast. These receptors,
when activated, increase the chloride conductance of the membrane, affecting transmitter release in
the presynaptic neuron, and producing hyperpolarization or shunting inhibition of the postsynaptic
neuron (MacDonald and Olsen, 1994; Xi and Akasu, 1996). Through the metabotropic GABAB
receptor inhibition is slower and involves changes in potassium and calcium permeability at
presynaptic and postsynaptic synapses (Misgeld et al., 1995; Howe et al., 1987; Deisz and Prince,
1989; Deisz et al., 1997).
3
GABA Receptor Subunits and Sensory Deprivation
GABAA receptors subunits comprise a family of at least 17 subunits (Davies et al., 1997).
Each subunit is expressed in a particular laminar pattern in SI and visual cortex (V1). For instance,
in SI and V1, the α1 subunit, which is present in the majority of the GABAA receptors, is densest in
layers III-IV (Fritschy et al., 1994). This laminar-specific expression might represent distinct
functional areas which can be differentially affected by sensory deprivation paradigms. For
instance, while monocular intravitreal injection of TTX in monkey induces a reduction in α1, β2,
and γ2 subunit mRNAs subunits levels in deprived visual cortex, it leaves levels of α2, α4, and β1
unchanged (Huntsman et al., 1994; Jones, 1997). In focal cortical malformations induced by
neonatal freeze lesions of SI, α1 and α5-GABAA subunits decrease in rat SI (Redecker, 2000).
Furthermore, electrolytic lesion of thalamus in the newborn decreases α1 in layers III-IV, but
increases α2, α3, and α5 in the same SI layers (Paysan, 1997).
When whiskers are trimmed during a critical period of early postnatal development,
stimulation of the regrown whiskers causes a degraded tuning of layer II/III receptive fields in the
corresponding deprived column (Lendvai et al., 2000; Stern et al., 2001). Similarly, plucking
whiskers from birth results in weaker responses of neurons in layers II/III and IV of the related
barrel column (Fox, 1992, 1994), but causes stronger responses from neighboring barrel columns
(Simons and Land, 1987; Fox, 1992, 1994). Similar deprivation effects are recorded in layer II/III
and IV barrel neurons after adult deprivation in rats (Glazewski and Fox, 1996; Wallace and Fox,
1999). However, it has not been determined whether there are decreases in GABAergic
components of layers II/IIII that could account for the modified excitation in these layers.
GABAA receptors decrease in layer IV of deprived barrel neurons (Fuchs and Salazar,
4
1998). These effects might not only be restricted to this layer, but might be present in layers II and
III of the same deprived whisker barrel columns. Moreover, changes in GABAA receptor binding,
as assessed by autoradiography, might be paralleled by changes in GABAA receptor subunits. A
favorable candidate α1-GABAA subunit, since it is the one that predominates in layers II/III of
barrel columns (
), and is constituent of most GABAA receptors (
). In the
present study, to determine whether sensory deprivation affects GABAA receptors in layers II/III
and IV, quantitative autoradiographic tests were performed on adult whisker trimmed rats. To
determine whether the same deprivation paradigm affects the α1-GABAA receptor subunits,
immunocytochemical methods were used.
5
CHAPTER II
MATERIALS AND METHODS
Subjects
The subjects were Long-Evans hooded rats (Simonsen, Gilroy, CA), maintained on a
12:12-hour light: dark cycle, with food and water available ad libitum. Whiskers from either the
middle row C or the other rows ABDE were trimmed for six weeks.
Whisker Deprivation
Six-week-old rats had whiskers trimmed every other day for 6 weeks, ensuring that their
vibrissae were kept shorter than 1 mm. All experimental rats had the mystacial vibrissae in either
row C, rows ABDE, or all rows clipped on one or both sides of the face. During the procedure,
no anesthetic was used, and rats were hand-held with gentle restraint. These protocols were
approved by the Institutional Animal Care and Use Committee at the University of North Texas.
Histology
Unperfused rats were killed at the end of their deprivation schedules by decapitation. For
tangential sections the deprived barrel region was dissected out from the brain, and was flattened
and frozen at -44˚C with the heat dissipater of a cryostat (2800 Frigocut N, Reichert-Jung,
6
Cambridge Instruments, Deerfield, IL). Cryostat sections 16 µm thick were cut at -20˚ C
tangentially to the pial surface, and were thaw-mounted onto gelatin subbed microscope slides.
For coronal sections the whole brain was frozen in -40˚C isopentane for 5 min, transferred to 80˚C isopentane, and cut and mounted as above. Sections were air dried for 0.5 to 3 h and then
stored desiccated at -80˚C until their use for either immunochemistry or receptor
autoradiography. After receptor autoradiography, some sections were Nissl stained, to examine
whether cellular and immunological observations can be paralleled. Other sections were stained
for cytochrome oxidase (CYO) activity (Wong-Riley, 1979) to localize barrels and to analyze
any similarities between CYO activity and receptor binding.
Immunocytochemistry
Subunit-specific antisera were used to visualize GABA subunits. The α1-GABAA receptor
subunit antisera were raised in rabbit against synthetic peptides derived from mouse subunit cDNA
(generously provided by J.M. Fritschy).
Prior to the immunological procedure, slides with rat sections were transferred to a 10˚C
refrigerator for 5-10 min. Then they were dried in a slide warmer for 45 sec, immersed horizontally
for 20 min in 0.5% paraformaldehyde in 0.15 M phosphate buffer (pH 7.4) (Fristchy and Mohler,
1995) for 20 min, and rinsed for 10 min in ice-cold 0.5 M Tris-saline buffer (TBS), pH 7.6. This
and all of the following steps were performed with gentle agitation using a rocker table. Slides
were then washed in preincubation solution consisting of TBS containing 10% normal goat serum
and 0.05% Triton X-100.
Sections were and incubated for two days and two nights at 4˚C in primary antibody
solution (1:100,000) diluted in TBS buffer containing 10% normal goat serum (NGS). Sections
7
were then washed three times for 1.5 hr in TBS and incubated for 30 min at room temperature in
TBS containing 2% NGS and goat-raised anti-rabbit biotinylated secondary antibodies (Jackson
Immunoresearch Laboratories Inc., West Grove, PA) diluted 1:300 in TBS pH 7.4. After 1 hr
additional washing in TBS, sections were transferred to the avidin-peroxidase solution (Vectastatin
Elite kit; Vector Laboratories, Burlingame, CA) for 30 min, and then washed 30 min in TBS.
Sections were then stained using two equal volumes of 0.1% diaminobenzidine hydrochloride
(DAB) in 50 mM TBS. To one of the volumes of substrate solution 0.02% hydrogen peroxide was
added to the sections to start the reaction, and sections were vigorously agitated for 1-2.5 min until
the desired color was obtained. The reaction was stopped by transfer into ice-cold TBS. After three
more 10 min washes in TBS, sections were dried in a slide-warmer, dehydrated with ascending
series of ethanol, cleared in xylenes and coverslipped using DPX.
Ligand binding
GABAA receptors were assessed using [3H]muscimol (Mower et al., 1986; Schwark et al.,
1994). These same sections were previously used to examine changes in layer IV after deprivation
(Fuchs and Salazar, 1998). Sections were preincubated 20 minutes at 4oC in 0.31 M Tris-citrate
buffer, pH 7.1, and then were incubated for 40 minutes at 4oC in the buffer containing 10 nM
[3H]muscimol (12-20 Ci/mmol; DuPont NEN, Boston, MA). Sections then were rinsed twice for
30 seconds in cold buffer, dipped briefly in dH20, and dried in a stream of air. Nonspecific binding
was assessed in the presence of 1 mM GABA.
Autoradiography
8
The brain sections and tritium standards (Microscale, Amersham) were exposed
simultaneously in the same cassette to tritium-sensitive Hyperfilm (Amersham). Following 2-4
month exposure periods, the film was developed with Kodak D-19 and processed according to the
manufacturer’s instructions.
Data analysis: Immunostained and Nissl stained sections
For semiquantitative image analysis, sections were digitized using a video-based
computerized image analysis system (MCID, Imaging Research, St. Catherine, Ont., Canada).
Measures of relative optical density were obtained from all the tangential α1-GABAAimmunostained sections, from cortical layer I (whenever possible) to layer IV. Samples were taken
within a computer-generated circle over each barrel column, allowing for comparisons between
deprived and intact rows.
Measures from the readily visible barrels in layer IV were taken first. Readings from upper
layers were then obtained. Patterns in the positions of radially oriented blood vessels were used to
confirm the exact location of each barrel column. Determination of boundaries between layers
II/III and IV was aid with a light microscope, taking into consideration cellular differences
between granular layer IV and supragranular layer III, and that septa are more conspicuous in layer
IV than in layer III.
Within each section, the mean ratio of densities in deprived/nondeprived row was
determined, using rows B, C, and D only. The average ratio for each subject was then calculated
from the section means. Then the average ratio for all subjects was obtained. To test the null
hypothesis that this ratio was not different from 1, Student’s t-test (two-tailed) was used.
Experimental groups were compared with one another by using analysis of variance with post hoc
9
t-tests. The significance level was 0.05. Percent decrease within each section was calculated as the
mean for deprived barrels minus the mean for nondeprived barrels, divided by the mean for
nondeprived barrels. Standard error measurements (S.E.M.) were calculated first for each subject.
For display, images were contrast-enhanced in pseudocolor.
Data analysis: Autoradiographs
[3H]Muscimol was quantitatively analyzed using a video-based computerized image
analysis system (MCID, Imaging Research, St. Catharines, Ont., Canada). Tritium standards were
used to calibrate autoradiographic densities. Ligand binding densities were automatically
calibrated by using a best-fit equation based on the plastic tritium standards, which had been
calibrated in nCi/mg tissue wet weight based on standards made from rat brain tissue (Fuchs and
Schwark, 1993). The analysis of data was done by the same procedures as the immunostained and
Nissl stained sections.
10
CHAPTER III
RESULTS
Effects of Whisker Deprivation on α1-GABAA receptor subunit
Coronal sections from control subjects depict the normal laminar distribution for α1GABAA receptor subunit immunostaining, in which layer IV appears the darkest (Fig. 1). Adult
rats with whiskers trimmed for 6 weeks showed in tangential sections a clear decrease in optical
density immunopositive staining for α1-GABAA receptor subunit in the deprived barrel columns.
Deprived columns showed less staining than the adjacent non-deprived ones. This decrease was
larger and readily detected in layers II/III of the deprived barrel columns 6% ± 0.6 (P<0.005). The
effect in layer IV was smaller, but still significant (-3.3% ± 0.9, P<0.005, Fig. 3, Table 1). Control
subjects showed no difference between C and adjacent rows (
11
).
Nissl Staining
Identification of barrel columns boundaries was aided by the differential staining and cell
density between barrel septa and hollow. Figure 4( with coronal and tangential sections). Trimming
whiskers for 6 weeks in adult rats resulted in an overall decrease in layers II, III, and IV of -8.7%
± 0.9 (mean ± S.E.M., P<0.005) as compared to the adjacent non deprived columns. This
decrease was largest in supragranular layers II and III (-11.1% ± 1.5, P<0.005), and smaller in
layer IV (-5.6% ± 0.7, P<0.005), where the difference was less obvious. (fig 4, Table 1). In
control subjects visual inspection rendered no difference between C and adjacent rows.
GABAA receptor autoradiography
[3H]Muscimol binding showed an overall reduction of -11.0% ± 0.9 in the deprived
columns as compared to the adjacent intact ones (mean ± S.E.M., P<0.001). The decrease was
similar for layers II/III (-11.4% ± 0.9, P<0.001), and IV (-10.2% ± 0.9, P<0.005). Comparison
between these two percentages from supragranular and granular layers was non significant.
Control subjects showed no difference between C and adjacent rows.
12
Cortical Layers
[3H] Muscimol
Binding
II, III, and IV
II/III
IV
Difference (II/III) - IV
-11.0 ± 0.9***
-11.4 ± 0.9***
-10.2 ± 0.9***
n.s.
α1-GABAA receptor
subunit
Immunoreactivity
-4.9 ± 0.6***
-6.0 ± 0.6***
-3.3 ± 0.9***
*
Nissl
-8.7 ± 0.9**
-11.1 ± 1.5***
-5.6 ± 0.7***
**
Table 1. Percentage decrease in the deprived barrel rows relative to intact rows (mean ± S.E.M.)
*P<0.05;**P<0.005; ***P<0.001; n.s., not significant.
13
Effects of Whisker Trimming on Layers II,III, and IV of SI
14
% decrease on deprived vs intact columns
12
10
8
6
4
2
0
α1-GABAA receptor subunit
[3H] Muscimol Binding
Nissl
Fig 1. Effects of whisker trimming on α1-GABAA receptor subunit immunoreactivity,
[3H]Muscimol binding, and Nissl staining on deprived barrel columns. After 6 weeks of whisker
trimming in adult rats, deprived rows containing layers II, III, and IIV showed significant
decreases in all three markers (**p<0.005) compared to adjacent undeprived rows.
Each value represents the mean ± S.E.M.
14
14
Percent decrease in deprived barrel layers
12
10
8
6
4
2
0
II/III**
IV*
α1-GABAA receptor subunit
II/III**
IV*
[3H] Muscimol Binding
II/III*
IV*
Nissl
Fig X. Effects of whisker trimming on α1-GABAA receptor subunit immunoreactivity,
[3H]muscimol binding, and Nissl staining on deprived barrel columns of layers II/III, and IV of
SI of rats. The effects were larger in supragranular deprived layers II/III than in deprived layer
IV for all paradigms. For α1-GABAA receptor subunit immunoreactivity the decrease in layers
II/III was 6% ± 0.6, P<0.001, and decrease in layer IV was 3.3% ± 0.9, P<0.001. For
[3H]muscimol binding the decrease in layers II/III was 11.4% ± 0.9, P<0.001, whereas in layer
IV it was 10.2% ± 0.9, P<0.001. For Nissl staining the decrease in layers II/III was 11.1% ± 1.5,
P<0.001, and in layer IV it was 5.6% ± 0.7, P<0.001. The difference in percent deprivation effect
in supragranular versus granular layer was significant, both for α1-GABAA receptor subunit
immunoreactivity (P<0.005) and for Nissl staining (p<0.05). Each value represents the mean ±
S.E.M.
15
CHAPTER IV
DISCUSSION
This is the first study indicating a decrease in GABAA receptors and α1-GABAA receptor
subunit in layers II/III and IV of SI after adult whisker trimming in rats. This suggests that GABA
receptors might play an important role not only in the long-lasting disinhibition of deprived barrel
columns (Simons and Land, 1987), but also in the specific signs of adult-induced plasticity
observed in layers II/III (Glazewski and Fox, 1996; Fox, 2002).
This cortical disinhibition seems to be compensatory, perhaps serving to amplify the
diminished input from sensory stimuli to the brain brought about by whisker deprivation (Hendry
et al., 1990; Siucinska and Kossut, 1996). Also, it might arise within the cortex (Simons and Land,
1994), since deprivation effects can be avoided by blocking postsynaptic activity in cortex through
the placement of a slow-release polymer containing muscimol, a GABA agonist, over the barrel
field (Wallace et al., 2001). Moreover, similar effects of whisker trimming can be induced through
microiontophoresis of the GABA antagonist bicuculline (Kiriazi et al., 1996).
The present results show that GABAA receptor binding decreases not only in layer IV
(Fuchs and Salazar, 1998), but also in the deprived layers II/III of the same column. While layer IV
presents a high degree of functional independence from supragranular layers (Goldreich, et al.,
1999; Huang et al., 1998), intracortical activity is required for potentiation and depression of
sensory responses in barrel cortex (Wallace et al., 2001). Thus, a reduction in GABAA receptors in
layers II/III could significantly contribute to observed experience-dependent plasticity (Lendvai et
al., 2000; Stern et al., 2001; Fox, 1992, 1994; Simons and Land, 1987; Fox, 1992, 1994; Glazewski
16
and Fox, 1996; Wallace and Fox, 1999; Fuchs and Salazar, 1998). More importantly, because of
the predominant nature of horizontal connections in layers II/III, a decrease in GABAA receptors
might allow excitation from adjacent non-deprived columns to spread in these layers and thus
modify axonal excitability/connections
This GABAA receptor downregulation after adult deprivation might be correlated to cell
number reduction, diminished connectivity, shrinkage of the barrel column, or a combination of
these factors. Decreases in GABA-containing neurons and inhibitory synaptic contacts targeting
dendritic spines have been identified in layer IV after neonatal whisker trimming (Micheva and
Beaulieu, 1995b, 1997; Sadaka et al., 2003). Nissl staining also identifies decreases in intracortical
projections after neonatal deprivation (Keller and Carlson, 1999). Consistent with these reports,
our results show that adult trimming of rows C or ABDE also leads to decreases in Nissl staining
in all layers of deprived SI. It is possible that this decrease at least in part might reflect shrinkage of
deprived columns or expansion of adjacent, undeprived ones. In fact, injections of fluorescent
dyes reveal increases in barrel size and axonal connectivity in adjacent-to-deprived layers I-IV
(Kossut and Juliano, 1999). Barrel size analysis after unilateral deprivation from birth, shows either
no change (Sadaka et al., 2003), or paradoxically, an increase after cortical blockage of activity
using the NMDA-activity-blocker MK-801 in cortical implants (Penschuck et al., 1999). However,
due to involvement of activity-dependent competition between adjacent barrels, unilateral
deprivation leads to a different set of functional and structural conditions than deprivation of one
whisker or row only from one side produces (Glazewski and Fox, 1996; Wallace et al., 2001).
Within a column, structural and functional modifications in layers II/III and IV are closely
interrelated. Layer IV neurons normally make powerful monosynaptic contact with layers II/III
within the correspondent column (Feldman, 2000; Lubke et al., 2000). Whisker deprivation during
17
development does not affect axonal length within a column (Bender et al., 2001). However, it
causes exuberant growth of axonal branches ascending from layer IV to II/III, extending beyond
their correspondent barrel column, and results in changes in layers II/III receptive fields (Stern et
al., 2001). Furthermore, deprivation decreases unitary EPSP amplitudes within layer II/III neurons,
and thus reduces their connectivity within the deprived column (Petersen et al., 2004). These
changes reflect a decrease in synaptic density in layer III neurons, the likely site of termination of
cortico-cortical inputs (Kossut, 1998).
Decreases in α1-GABAA receptor subunit, as shown in this experiment through
immunocytochemistry, have also been found in cortex after other deprivation paradigms. These
reductions might be correlated to decreases in number and/or or size of inhibitory contacts, but it
might also reflect a specific decrease of synthesis of receptors, or to changes in subunit
composition (Lech, 2001). Electrolytic lesion of thalamus in the newborn decreases α1-subunit in
layer III-IV whisker barrels, and increases α2, α3, and α5 in the same areas (Paysan et al., 1997).
Similarly, focal cortical malformations induced by neonatal freeze lesions of SI shows a decrease
in α1 and α5-GABAA subunits (Redecker, 2000). In contrast, blocking cortical NMDA activity
with MK-801, using a cortical implant, increases barrel size, without affecting α1-subunit laminar
distribution or immunostaining intensity (Penschuck, et al., 1998). This increase is paradoxical,
since NMDA was expected to reduced overall cortical activity. As mentioned above, trimming
adjacent whiskers introduces competition for input among neighboring barrels, enhancing
deprivation effects, such as in chessboard pattern whisker deprivation (Glazewski and Fox, 1996;
Wallace et al., 2001). In this sense, comparisons between adjacent deprived-undeprived barrels
could resemble effects of monocular deprivation in the visual system of primates. In primary visual
cortex, unilateral injection of tetradotoxin to one eye of adult monkey for 8-21 days reduces the
18
levels of α1, β2, and γ2 subunit mRNA in the deprived ocular dominance columns. This reduction
is fairly specific, since mRNA for other subunits (α2, α4, and β1) remains unchanged (Hendry et
al., 1994; Huntsman et al., 1994; Jones, 1997).
While the majority of studies have applied sensory deprivation during developmental
critical periods, some reports have shown effects of adult sensory deprivation (Glazewski and Fox,
1996; Akhtar and Land, 1991; Fuchs and Salazar, 1998). Adult plasticity differs from
developmental plasticity in fundamental ways. Thalamocortical synapses in barrel cortex can be
functionally identified as early as postnatal day 3, but the adult number of synapses per neuron is
achieved only until postnatal day 21 (Micheva and Beaulieu, 1996). Thus, deprivation during
development can sometimes result in irreversible cortical modifications (Lendvai et al., 2000),
linked to changes in collateralization and arborization of thalamocortical fibers (Antonini and
Stryker, 1993). In the adult, experience-regulated plasticity may be a more readily reversible
event, which may rely on more subtle and specific changes in synaptic strength than on synapse or
axonal formation and elimination (Trachtenberg et al., 2000; Rausell and Jones, 1995; Huntley,
1996). Whisker deprivation changes the excitability of neurons in cortex in a layer-specific
manner. Layer IV becomes more sensitive to whisker deprivation around postnatal day 7 (Fox,
1992), while layers II/III reach mature receptive field organization around postnatal day 14 (Stern
et al., 2000). Plasticity in II-IV remains in the adult (Huang et al., 1998; Goldreich et al., 1999;
Armstrong-James et al., 1994; Glazewski and Fox, 1996; Lendvai et al., 2000, Sheperd et al.,
2003). For instance, [14C] 2-deoxyglucose labeling shows that normal metabolic activity decreases
in layers II/III and IV of barrel columns corresponding to plucked whiskers if the deprivation
period begins in young adult rats. In rats deprived at older age, decreases in metabolic activity are
less reduced and only measurable in layers II/III, and not in layer IV (Kossut, 1998; Skibinska et
19
al., 2000). Moreover, plastic changes in responses from adult cortex after peripheral manipulations
appear as early as 24 hrs later in layers II/III (Diamond et al., 1994). These observation that this
induced changes occur in layer II/III without changes in layer IV after sensory manipulation,
suggests that supragranular layers are indeed a site of experience-dependent plasticity in SI
(Glazewski and Fox, 1996). However the changes in layers II/III might occur at the level of
synapses from layer IV to II/III neurons, since whisker deprivation induces depression at these
sites (Allen et al., 2003; Sheperd et al., 2003).
This study provides a structural substrate which might help explain the high degree of
functional plasticity observed in layers II-IV of SI. In particular, down-regulation of GABAA
receptors and α1-GABAA receptors subunit might contribute to the notable neurophysiological
changes induced by adult whisker trimming in layers II/III deprived barrel cortex.
20
REFERENCES
-
Akhtar, N.D. and Land, P.W., Activity-dependent regulation of glutamic acid decarboxylase
in the rat barrel cortex: Effects of neonatal versus adult sensory deprivation. J. Comp.
Neurobiol., 307 (1991) 200-213.
-
Ambardekar, A. V., Ilinksy, I.A., Froestl, W., Bowery, N.G., and Kultas-Ilinsky, K.
Distribution and properties of GABAB antagonist [3H] CGP 62349 binding in the rhesus
monkey thalamus and basal ganglia and the influence of lesions in the reticular thalamic
nucleus. Neuroscience, 93-4 (19) 1339-1347.
-
Armstrong-James M, Fox K. Spatiotemporal convergence and divergence in the rat S1 ‘barrel’
cortex. J. Comp. Neurol., 263 (1987) 265-281. BUSCAR
-
Armstrong-James, M., Diamond, M.E., Ebner, F.F. An innocuous bias in whisker use in adult
rats modifies receptive fields of barrel cortex neurons. J Neurosc., 14 (1994) 69786991.BUSCAR
-
Antonini, A., Stryker, M. Rapid remodeling of axonal arbors in the visual cortex. Science, 260
(1993) 1819-1821. BUSCAR
-
Bender, K.J., Rangell, Juliana, and Ferldman, D.E. Development of columnar topography in
the excitatory layer 4 to layer 2/3 projection in rat barrel cortex. J. Neurosc. 23-25 (2003)
8759-8770.
-
Bowery, N.G., Hill, D.R., Hudson, A.L., Doble, A., Middlemass, D.N., Shaw, J., and Turnbull,
M. (-) Baclofen decreases neurotransmitter release in the mammalian CNS by an action at a
novel GABA receptor. Nature, 283 (1980) 92-94.
21
-
Bowery, N.G., Hudson, A.L., and Price, G.W. GABAA and GABAB receptor site distribution in
rat central nervous system. Neuroscience. 20 (1987) 365-383.
-
Chmielowska, J, Carvel, G.E., and Simons, D.J. Spatial organization of thalamocortical and
corticothalamic projection systems in the rat SmI barrel cortex. J.Comp. Neurol. 285 (1989)
325-338.
-
Chu, D.C.M., Albin, R.L., Young, A.B., and Penney, J.B. Distribution and kinetics of GABAB
binding sites in rat central nervous system: a quantitative autoradiographic study.
Neuroscience. 34-2 (1990) 341-357.
-
Davies, P.A, Hanna, M.C, Hales, T.G, Kirknesss, E.F. (1997) Nature (London) 385, 820-823
-
Deisz, R.A. and Prince, D.A. Frequency-dependent depression of inhibition in guinea-pig
neocortex in vitro by GABAB receptor feed-back on GABA release. J. Physiol. Lond. 424
(1989) 513-541.
-
Deisz, R.A. Presynaptic and Postsynaptic GABAB receptors of neocortical neurons of the rat in
vitro: differences in pharmacology and ionic mechanisms. Synapse. 25 (1997) 62-72.
-
Deisz, R.A. GABAB receptor-mediated effects in human and rat neocortical neurones in vitro.
Neuropharmacology. 38 (1999) 1755-1766).
-
Diamond, M.E., Huang, W., Ebner, F.F. Laminar comparison of somatosensory cortical
plasticity. Science 265: 1885-1888. BUSCAR
-
Feldman, D.E., Timing-based LTP and LTD at vertical inputs to layer II/III pyramidal cells in
rat barrel cortex. Neuron. 27 (2000) 45-56
-
Fox, K. A critical period for experience-dependent synaptic plasticity in rat barrel cortex. J.
Neurosc. 12 (1992) 1826-1838.
-
BUSCAR
Fox, K. The cortical component of experience-dependent synaptic plasticity in the rat barrel
22
cortex. J. Neurosc. 14 (1994) 7665-7679. BUSCAR
-
Fox, K. Anatomical pathways and molecular mechanisms for plasticity in the barrel cortex.
Neurosci. 111-4 (2002) 799-814.
-
Fritschy, J.M., Paysan, J., Enna, A., Mohler, H. (1994) J. Neurosci. 143, 5302-5324.
-
Fritschy, J.M, Mohler, H. J. Comp. Neurol., 359 (1995) 154-194
-
Fuchs, J.L. and Salazar, E. Effects of whisker trimming on GABAA receptor binding in the
barrel cortex of developing and adult rats. J. Comp. Neurol., 395 (1998) 209-216.
-
Glazewski, S., Fox., Time course of experience-dependent synaptic potentiation and depression
in barrel cortex of adolescent rats. J. Neurophysiol., 75 (1996) 1714-1729. BUSCAR
-
Goodman, C.S, Shatz, C.J. Neuron 10. suppl., (1993) 77-78.
-
Huntley, G.W., Correlation between patterns of horizontal connectivity and the extent of shortterm representational plasticity in rat motor cortex. Cereb. Cortex, 7 (1996) 143-156.
-
Hendry, S.H.C. and Jones, E.G. Reduction in number of immunostained GABAergic neurons
in deprived-eye dominance columns of monkey area 17, Nature, 320 (1986) 750-753.
-
Hendry, S.H.C., Huntsman, M.M., Vinuela, A. Mohler, H. de Blas, A.L., Jones, E.G. GABAA
receptor subunit immunoreactivity in primate visual cortex: distribution in macaques and
humans and regulation by visual input in adulthood. J. Neurosci. 14 (1994) 2383-2401.
-
Howe, J.R., Sutor, B., and Zieglgansberger, W. Baclofen reduces post-synaptic potentials of rat
cortical neurons by an action other than its hyperpolarizing action. J. Physiol. Lond. 384
(1987) 539-570.
-
Huang
-
Huntsman, M.M., Isackson, P.J., and Jones, E.G. Lamina-specific expression and activitydependent regulation of seven GABAA receptor subunit mRNAs in monkey visual cortex. J.
23
Neurosc. 14-4 (1994):2236-59.
-
Jones, E.G. Area and lamina-specific expression of GABAA receptors subunit mRNAs in
monkey cerebral cortex. Can. J. Physiol. Pharmacol. 75-5 (1997) 452-469.
-
Jones, ’95 and 98
-
Keller, A., Carlson, G.C. Neonatal whisker clipping alters intracortical, but not thalamocortical
projections, in rat barrel cortex. J. of Comp. Neurol., 412 (1999) 83-94.
-
Kiriazy, H.T., Carvell, G.E., Brumberg, J.C., and Simons, D.J. Quantitative Effects of GABA
and Bicuculline Methiodide on Receptive Field properties of neurons in real and simulated
whisker barrels. Journal of Neurophysiology. 75-2 (1996) 547-560
-
Kossut, M., Hand, P.J., Greenberg, J., Hand, C.L. Single vibrissa cortical column in SI cortex
of rat and its alterations in neonatal and adult vibrissa-deafferented animals: a quantitative
2DG study. J Neurophysiol.60 (1988) 829-852.
-
Kossut, M. Plasticity of the barrel cortex neurons. Prog. Neurobiol. 39 (1992) 389-422.
-
Kossut, M., Experience-dependent changes in function and anatomy of adult barrel cortex.
Exp. Brain Res. 123 (1998) 110-116.
-
Kossut M., and Juliano, S.L., Anatomical correlates of representational map reorganization
induced by partial vibrissectomy in the barrel cortex of adult mice. Neuroscience. 92-3 (1999)
807-817.
-
Lech, Monica, 2001 BUSCAR BUSCAR BUSCAR
-
Lendvai B., Stern, E., Svoboda, K., Experience-dependent plasticity of dendritic spines in the
developing rat barrel cortex in vivo. Nature 404 (2000) 876-881.BuUSCAR
-
Lebedev, M.A., Mirabella, G., Erchova, I., Diamond, M.E. Experience-dependent plasticity of
rat barrel cortex: redistribution of activity across barrel columns. Cerebral Cortex. 10-1 (2000)
24
23-31.
-
Lubke, J., Egger, V., Sakmann, B., Feldmeyer, D. Columnar organization of dendrites and
axons of single and synaptically coupled excitatory spiny neurons in layer 4 of the rat barrel
cortex. J. Neurosc. 20 (2000) 5300-5311.
-
MacDonald, R.L, Olsen R.W.GABAA receptor channels. Annual Review of Neuroscience.17
(1994) 569-602.
-
Micheva K.D., and Beaulieu, C. Neonatal sensory deprivation induces selective changes in the
quantitative distribution of GABA-immunoreactive neurons in the rat barrel field cortex. J.
Comp. Neurol. 361-4 (1995) 574-84.
-
Micheva, K.D., and Beaulieu, C. An anatomical substrate for experience-dependent plasticity
of the rat barrel field cortex. Proc. Natl. Acad. Sci. USA. 92 (1995) 11834-11838.
-
Micheva, K.D., Beaulieu, C. Development and plasticity of the inhibitory neocortical circuitry
with an emphasis on the rodent barrel field cortex: a review. Can J. Physiol Pharmacol. 75-5
(1997) 470-478. BUSCAR BUSCAR
-
Maravall, M., Koh, I.Y., Lindquist, W.B., Svoboda, K., Experience-dependent changes in basal
dendritic branching of layer 2/3 pyramidal neurons during a critical period for developmental
plasticity in rat barrel cortex. Cerebral Cortex, advances March 28 (2004) 1-10
-
Micheva, K.D. and Beaulieu, C. An anatomical substrate for experience-dependent plasticity
of the rat barrel field cortex. Proc Natl Acad Sci. 92-25 (1995) 11834-8.
-
Micheva K.D., and Beaulieu, C. Postnatal development of GABA neurons in the rat
somatosensory barrel cortex: a quantitative study. Eur. J. Neurosci. 7-3 (1995) 419-30
-
Micheva, K.D. and Beaulieu, C. Quantitative aspects of synaptogenesis in the rat barrel field
cortex with special reference to GABA circuitry. ’ J. Comp. Neurol., 373-3 (1996) 340-354.
25
-
Micheva, K.D. and Beaulieu, C. Development and plasticity of the inhibitory neocortical
circuitry with an emphasis on the rodent barrel field cortex: a review. Can J Physiol
Pharmacol. 75-5 (1997) 470-8.
-
Misgeld, U., Bijak, M., and Jarolimek, W. A physiological role for GABAB receptors and the
effects of baclofen in the mammalian central nervous system.. Prog. Neurobiol., 46(4) (1995)
423-62
-
Muñoz, A., DeFelipe, J., and Jones, E.G. Patterns of GABA (B)R1a,b receptor gene
expression in monkey and human visual cortex. Cerebral Cortex, 11-2 (2001) 104-113.
-
Payssan, J., Kossel, A., Bolz,,,, J., and Fritschy, J. Area-specific regulation of γ-aminobutyric
acid type A receptor subtypes by thalamic afferents in developing rat neocortex. Proc. Natl.
Acad. Sci. USA. 94 (1997) 6995-7000.
-
Penschuck, S., Giorgetta, O., and Fritschy, J.M. Neuronal activity influences the growth of
barrels in developing rat primary somatosensory cortex without affecting the expression pattern
of four major GABAA receptor  subunits. Brain Research. 112 (1999) 117-127.
-
Petersen, C.C.H., Brecht, M., Hahn, T.T.G., Sakmann, B. Synaptic changes in Layer 2/3
underlying map plasticity of developing barrel cortex. Science, 304 (2004) 739-742.
-
Rausell, E., Jones, E. Extent of intracortical arborization of thalamocortical axons as a
determinant of representational plasticity in monkey somatic sensory cortex. J. Neurosci., 15
(1995) 4270-4288.
-
Redecker, C., Luhmann, H.J., Hagemann, G., Fritschy, J., and Witte, W. Differential
Downregulation of GABAA receptor subunits in widespread brain regions in the freeze-lesion
model of focal cortical malformations. J. Neuroscience. 20-13 (2000) 5045-5053.
-
Rema, V., Armstrong-James, M., Ebner, F.F. Experience-dependent plasticity is impaired in
26
adult rat barrel cortex after whiskers are unused in early postnatal life. J. Neurosc. 23-1 (2003)
358-366.
-
Sadaka, Y., Weinfeld, E., Lev, D.L., and White, E. Changes in mouse barrel synapses
consequent to sensory deprivation from birth. J. Comp. Neurol,, 457 (2003) 75-86.
-
Schlaggar, B.L., Fox, K., O’Leary, D.D.M. Postsynaptic control of plasticity in developing
somatosensory cortex. Nature (London) 364 (1993) 623-626.
-
Sheperd, G.M., Pologruto, T.A., Svodoba, K. Circuit analysis of experience-dependent
plasticity in the developing rat barrel cortex. Neuron, 38-2 (2003) 277-289.
-
Skibinska, A., Glazewski, S., Fox, K. Age-dependent response of the mouse barrel cortex to
sensory deprivation: a 2-deoxyglucose study. Exp. Brain Res. 132 (2000) 132-134.
-
Simons, D.J. Response properties of vibrissa units in the rat SI somatosensory neocortex. J.
Neurophysiol., 41 (1978) 798-820.
-
Simons, D.J. and Land, P.W. Early experience of tactile stimulation influences organization of
somatic sensory cortex. Nature, 326 (1987) 694-697.
-
Simons, D.J. and Woolsey, T.A., Functional organization of mouse barrel cortex. Brain
Research, 165 (1979) 327-332.
-
Stern, E.A., Maravall, M., and Svoboda, K. Rapid development and plasticity of layer 2/3
maps in rat barrel cortex in vivo. Neuron, 31-2 (2001) 305-315.
-
Siucinska, E., Kossut, M. Short-lasting classical conditioning induces reversible changes of
representational maps of vibrissae in mouse SI cortex: a 2DG study. Cereb. Cortex 6 (1996)
506-513. BUSCAR
-
Trachtenberg, J.T., Chen., B.E., Knott. G.W., Feng, G., Sanes., J.R., Welker, E., Svodoba, K.,
Long-term in vivo imaging of experience-dependent synaptic plasticity in adult cortex. Nature
27
420 (2002) 788-794. BUSCAR
-
Van der Loos, H. and Woolsey, T.A. Somatosensory cortex: structural alterations following
early injury to sense. Science, 179-71 (1973) 395-398.
-
Wallace, H.S., Glazewski, S., Fox, K. The role of cortical activity in experience-dependent
potentiation and depression of sensory responses in rat barrel cortex. J Neurosc. 21 (2001)
3881-3894.BUSCAR
-
Welker, C. and Woolsey, T.A., Structure of layer IV in the somatosensory neocortex of the rat:
description and comparison with the mouse. J. Comp. Neurol., 158 (1974) 437-453
-
Welker, C., Microelectrode delineation of fine grain somatotopic organization of (SmI)
cerebral neocortex in albino rat. Brain Research 26-2 (1971) 259-275.
-
Welker, C. Receptive fields of barrels in the somatosensory neocortex of the rat. J. Comp.
Neurol., 166-2 (1976) 173-189.
-
Woolsey, T.A. and Van der Loos, H. The structural organization of layer IV in the
somatosensory region (SI) of mouse cerebral cortex, Brain Research, 17 (1970) 205-242.
-
Xi, Z X; Akasu, T. Presynaptic GABAA receptors in vertebrate synapses. The Kurume
Medical Journal, 43-2 (1996) 115-12
28