# Exposure to Forced Swim Stress Alters Local Circuit Activity and Plasticity in the Dentate Gyrus of the Hippocampus

Authors: Orli Yarom; Mouna Maroun; Gal Richter-Levin
Journal: Neural Plasticity
 (2008)

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## Abstract

Studies have shown that, depending on its severity and context, stress can affect neural plasticity. Most related studies focused on synaptic plasticity and long-term potentiation (LTP) of principle cells. However, evidence suggests  that following high-frequency stimulation, which induces LTP in principal cells, modifications also take place at the level of complex interactions with interneurons within the dentate gyrus, that is, at the local circuit level. So far, the possible effects of stress on local circuit activity and plasticity were not  studied. Therefore, we set out to examine the possible alterations in local circuit activity and plasticity following exposure to stress. Local circuit activity and plasticity were measured by using frequency dependant inhibition (FDI) and commissural modulation protocols following exposure to a 15 minute-forced swim trial. Exposure to stress did not alter FDI. The application of theta-burst stimulation (TBS) reduced FDI in both control and stressed rats, but this type of plasticity was greater in stressed rats. Commissural-induced inhibition was significantly higher in stressed rats both before and after applying theta-burst stimulation. These findings indicate that the exposure to acute stress affects aspects of local circuit activity and plasticity in the dentate gyrus. It is possible that these alterations underlie some of the behavioral consequences of the stress experience.


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## Body

## 1. Introduction

Stress is defined as any condition that seriously disrupts
physiological and psychological homeostasis ranging from anxiety to
posttraumatic stress disorder [1], and affects cognitive functions both in
animal models and in humans [2–4]. The hippocampus is of special significance
in this respect because it has been shown to play a major role in regulating
stress [5, 6], and to be involved in some aspects of learning and memory [7–13].At present, long-term potentiation (LTP) of synaptic
transmission in the hippocampus is the most studied neurophysiological model
for learning and memory processes in the mammalian nervous system. LTP, like
behavior, appears to be affected by stress. Depending on the type of stress and
the procedures used, stress has been shown to have different effects on
different measures of synaptic plasticity. There is a general agreement that
LTP in area CA1 of the hippocampus is impaired following stress [4, 14–18]. Some studies have also shown that stress impairs LTP in the dentate
gyrus (DG) of the hippocampus [16, 19, 20], while others reported intact LTP
in the DG following stress [14, 21]. Thus, DG LTP is considered to be less
sensitive to stress compared to LTP in CA1 [22].Although LTP is a widely accepted model of learning and
memory, debates continue over its validity, and controversial results regarding
its behavioral correlates are reported (for review, see [23]). A different
level of processing that is likely to be relevant to memory formation is local
circuit activity. When examining this level of processing, the focus is on
interactions between local, mostly inhibitory GABAergic neurons and pyramidal
or granular principle cells in the hippocampus and cortex [24, 25].  This is
in contrast to the focus on LTP of input excitatory synapses onto principle
cells, which is responsible for transmitting information from one region to
another. Inhibitory interneurons exert a powerful control over local circuit
activity through feedforward and feedback inhibition. Modification of local
circuits can affect the computational properties of the region, and therefore
affect its involvement in behavior.In the current study, local circuit activity and plasticity
were measured by using frequency-dependent inhibition (FDI) and commissural
modulation protocols, following exposure to behavioral stress.FDI is suggested to reflect GABA-mediated inhibition by
perforant path- (PP-) activated interneurons onto granule cells [26].
Increasing stimulus frequency from 0.1 Hz to 1.0 Hz results in the reduction of
the population spike (PS) of the field potential response to stimulation of the
PP [27]. Our lab has previously shown that FDI in the DG is NMDA-dependent
[28], GABA-mediated, and that delivering theta-burst stimulation (TBS) to the
PP of the hippocampus induced a lasting reduction in FDI [18].The DG commissural pathway is activated by stimulating the
contralateral DG at different intervals prior to PP stimulation. Stimulation of
the commissural pathway induces a biphasic, inhibitory/excitatory effect on
granule cell responsiveness to PP stimulation. The inhibitory phase is a
result of activation of feedforward inhibition [29].Although the effect of behavioral stress on induction of
hippocampal LTP has been studied extensively, to our knowledge no research has
established the relationship between stress and local circuit activity and
plasticity. The current study addresses this issue in order to further explore
the potential relevance of local circuit activity to learning and memory. Our
aim in this study was to characterize local circuit activity and plasticity in
the DG of the hippocampus following exposure to behavioral stress.


## 2. Methods

### 2.1. Subjects

Adult, male Sprague Dawley rats, weighing 240–330 g, from Harlan
(Jerusalem, Israel) maintained five per cage on a 12-hour light/dark cycle
with water and laboratory rodent chowad libitum.


### 2.2. Corticosterone Radioimmunoassay

Trunk blood was collected following decapitation and
samples were centrifuged at 3000 r.m.p. for 20 minutes at 4ºC. Serum was stored
at −80ºC. Corticosterone was measured using a radioimmunoassay kit
(Coat-A-Count, Diagnostic Products Corporation, Los Angeles, Calif, USA).


### 2.3. Electrophysiology

Rats were anaesthetized (6% chloral hydrate in 100 mL
saline; 0.5 mL/100 g. IP) and prepared for acute stimulation of the
perforant path and for recording of field potentials in the dentate gyrus as
described before [29].Rats were placed in a head holder in a
stereotaxic frame and small holes were drilled in the skull to allow the
insertion of electrodes in the brain. A recording microelectrode (glass, tip
diameter 2–5μm filled with 2 M NaCl, resistance 1–4 MΩ) was placed in the dentate gyrus (coordinates: 4 mm posterior to bregma, 2.5 mm lateral to midline).
A bipolar 125 μm stimulating electrode was implanted in the ipsilateral
angular bundle to stimulate the perforant path (coordinates: 8 mm posterior to bregma, 4 mm lateral to midline). The
depth of the electrodes was adjusted to maximize the size of the evoked
positive-going excitatory postsynaptic potential (EPSP) recorded in the hilus
of the dentate gyrus.Evoked responses were digitized
(10 kHz) and analyzed using the Cambridge Electronic Design 1401+ and its
Spike2 software. Offline measurements were made of the amplitude of the PS and
the slope of the EPSP using averages of 5 successive responses to a given stimulation
intensity applied at 0.1 Hz. Test stimuli (monopolar pulses, 100-microsecond duration,
intensity adjusted to yield a PS of 30–50% of the maximal pretetanus value)
were delivered at 0.1 Hz. After positioning the electrodes, the rat was left
for 20 minutes before commencing the experiment. During recording the rats were
maintained at 37 ± 1°C with a homeothermic blanket system (Harvard).


### 2.4. Long-Term Potentiation

LTP was induced by a TBS (3 sets of 10 trains,
each consisting of 10 pulses at 100 Hz. Intertrain interval: 200 milliseconds,
and the interval between each set: 1 minute, trains are delivered at 2x test
stimulus intensity).LTP was measured as the difference in EPSP
slope before and 60 minutes after TBS.
We defined LTP as an increase of the least 20% in the EPSP slope of the
evoked potentials 60 minutes after application of TBS.


### 2.5. Local Circuit Activity

Frequency-dependent inhibition
To determine FDI, 10
pulses were delivered at 0.1 Hz followed by 10 pulses at 1.0 Hz, as described
before [18]. This pattern was repeated twice. The pulses given were at test
stimulus intensity. The PS or EPSP slope of the 10 responses at   0.1 Hz were averaged and compared to the 10
responses at 1.0 Hz in each set. The results of the two sets were averaged.
Inhibition is expressed as an FDI index which was assessed by dividing the
averaged response at 1.0 Hz by the averaged response at 0.1 Hz.Commissural-induced modulation
The DG commissural
pathway was activated by stimulating the contralateral DG at different
intervals prior to PP stimulation (15, 30, 80, and 150 milliseconds) as
described before [30].  Stimulation of
the commissural pathway induces a biphasic inhibitory/excitatory effect on
granule cell responsiveness to PP stimulation [31, 32].
Commissural-induced modulation is expressed by commissural
index which was evaluated as the ratio of the size of the response to PP stimulation
after commissural stimulation divided by that of the response to PP stimulation
with no priming stimulation.
In order to measure TBS-induced alterations on frequency-dependent
inhibition or on commissural-induced inhibition and facilitation, the
stimulation intensity following the induction of LTP was adjusted to yield a PS
size comparable to pre-TBS level.


### 2.6. Behavior

#### 2.6.1. Elevated Plus Maze

Apparatus
The maze employed is a four-armed
black opaque Plexiglas platform, elevated 50 cm above ground. Two opposite arms are
enclosed by 40 cm high Plexiglas walls on both sides and on the outer edges of the platform, that
is, “closed,” while the two remaining opposite arms are “open,” and are
surrounded only by a 1 cm high Plexiglas rim, which serves as a tactile guide to animals in the open
areas. Individual rats were placed
in the central platform, faced towards different arms in randomized order.Procedure
Each
five-minute session was recorded using an overhead video camera connected to a
monitor/recorder in an adjacent observation room. Animals were placed in the
central platform and allowed to explore for five minutes. Animals were then
subjected to the acute swim stress procedure, and then allowed to rest for 90
minutes. After the resting period animals were placed once again in the maze
for a five-minute poststress test. Time spent in the open arms was
measured. Animals were
scored as being in an open or closed arm only when all four paws passed over the
open/closed dividing line.


### 2.7. Induction of Behavioral Stress

Apparatus
The water container used for the forced
swim procedure consists of a circular container of water (50 cm in diameter with a rim 75 cm high). Water depth was 50 cm and temperature was maintained at 23 ± 1°C.Acute Swim Stress (ASS) procedure
Rats were
subjected to ASS as previously described [33]. Individual rats were subjected
to a single 15-minute swim session in the water container. After this single
swim session, rats were allowed to dry in a resting cage for 30 minutes, and
then anesthetized and taken to electrophysiology.


## 2.1. Subjects

Adult, male Sprague Dawley rats, weighing 240–330 g, from Harlan
(Jerusalem, Israel) maintained five per cage on a 12-hour light/dark cycle
with water and laboratory rodent chowad libitum.


## 2.2. Corticosterone Radioimmunoassay

Trunk blood was collected following decapitation and
samples were centrifuged at 3000 r.m.p. for 20 minutes at 4ºC. Serum was stored
at −80ºC. Corticosterone was measured using a radioimmunoassay kit
(Coat-A-Count, Diagnostic Products Corporation, Los Angeles, Calif, USA).


## 2.3. Electrophysiology

Rats were anaesthetized (6% chloral hydrate in 100 mL
saline; 0.5 mL/100 g. IP) and prepared for acute stimulation of the
perforant path and for recording of field potentials in the dentate gyrus as
described before [29].Rats were placed in a head holder in a
stereotaxic frame and small holes were drilled in the skull to allow the
insertion of electrodes in the brain. A recording microelectrode (glass, tip
diameter 2–5μm filled with 2 M NaCl, resistance 1–4 MΩ) was placed in the dentate gyrus (coordinates: 4 mm posterior to bregma, 2.5 mm lateral to midline).
A bipolar 125 μm stimulating electrode was implanted in the ipsilateral
angular bundle to stimulate the perforant path (coordinates: 8 mm posterior to bregma, 4 mm lateral to midline). The
depth of the electrodes was adjusted to maximize the size of the evoked
positive-going excitatory postsynaptic potential (EPSP) recorded in the hilus
of the dentate gyrus.Evoked responses were digitized
(10 kHz) and analyzed using the Cambridge Electronic Design 1401+ and its
Spike2 software. Offline measurements were made of the amplitude of the PS and
the slope of the EPSP using averages of 5 successive responses to a given stimulation
intensity applied at 0.1 Hz. Test stimuli (monopolar pulses, 100-microsecond duration,
intensity adjusted to yield a PS of 30–50% of the maximal pretetanus value)
were delivered at 0.1 Hz. After positioning the electrodes, the rat was left
for 20 minutes before commencing the experiment. During recording the rats were
maintained at 37 ± 1°C with a homeothermic blanket system (Harvard).


## 2.4. Long-Term Potentiation

LTP was induced by a TBS (3 sets of 10 trains,
each consisting of 10 pulses at 100 Hz. Intertrain interval: 200 milliseconds,
and the interval between each set: 1 minute, trains are delivered at 2x test
stimulus intensity).LTP was measured as the difference in EPSP
slope before and 60 minutes after TBS.
We defined LTP as an increase of the least 20% in the EPSP slope of the
evoked potentials 60 minutes after application of TBS.


## 2.5. Local Circuit Activity

Frequency-dependent inhibition
To determine FDI, 10
pulses were delivered at 0.1 Hz followed by 10 pulses at 1.0 Hz, as described
before [18]. This pattern was repeated twice. The pulses given were at test
stimulus intensity. The PS or EPSP slope of the 10 responses at   0.1 Hz were averaged and compared to the 10
responses at 1.0 Hz in each set. The results of the two sets were averaged.
Inhibition is expressed as an FDI index which was assessed by dividing the
averaged response at 1.0 Hz by the averaged response at 0.1 Hz.Commissural-induced modulation
The DG commissural
pathway was activated by stimulating the contralateral DG at different
intervals prior to PP stimulation (15, 30, 80, and 150 milliseconds) as
described before [30].  Stimulation of
the commissural pathway induces a biphasic inhibitory/excitatory effect on
granule cell responsiveness to PP stimulation [31, 32].
Commissural-induced modulation is expressed by commissural
index which was evaluated as the ratio of the size of the response to PP stimulation
after commissural stimulation divided by that of the response to PP stimulation
with no priming stimulation.
In order to measure TBS-induced alterations on frequency-dependent
inhibition or on commissural-induced inhibition and facilitation, the
stimulation intensity following the induction of LTP was adjusted to yield a PS
size comparable to pre-TBS level.


## 2.6. Behavior

### 2.6.1. Elevated Plus Maze

Apparatus
The maze employed is a four-armed
black opaque Plexiglas platform, elevated 50 cm above ground. Two opposite arms are
enclosed by 40 cm high Plexiglas walls on both sides and on the outer edges of the platform, that
is, “closed,” while the two remaining opposite arms are “open,” and are
surrounded only by a 1 cm high Plexiglas rim, which serves as a tactile guide to animals in the open
areas. Individual rats were placed
in the central platform, faced towards different arms in randomized order.Procedure
Each
five-minute session was recorded using an overhead video camera connected to a
monitor/recorder in an adjacent observation room. Animals were placed in the
central platform and allowed to explore for five minutes. Animals were then
subjected to the acute swim stress procedure, and then allowed to rest for 90
minutes. After the resting period animals were placed once again in the maze
for a five-minute poststress test. Time spent in the open arms was
measured. Animals were
scored as being in an open or closed arm only when all four paws passed over the
open/closed dividing line.


## 2.6.1. Elevated Plus Maze

Apparatus
The maze employed is a four-armed
black opaque Plexiglas platform, elevated 50 cm above ground. Two opposite arms are
enclosed by 40 cm high Plexiglas walls on both sides and on the outer edges of the platform, that
is, “closed,” while the two remaining opposite arms are “open,” and are
surrounded only by a 1 cm high Plexiglas rim, which serves as a tactile guide to animals in the open
areas. Individual rats were placed
in the central platform, faced towards different arms in randomized order.Procedure
Each
five-minute session was recorded using an overhead video camera connected to a
monitor/recorder in an adjacent observation room. Animals were placed in the
central platform and allowed to explore for five minutes. Animals were then
subjected to the acute swim stress procedure, and then allowed to rest for 90
minutes. After the resting period animals were placed once again in the maze
for a five-minute poststress test. Time spent in the open arms was
measured. Animals were
scored as being in an open or closed arm only when all four paws passed over the
open/closed dividing line.


## 2.7. Induction of Behavioral Stress

Apparatus
The water container used for the forced
swim procedure consists of a circular container of water (50 cm in diameter with a rim 75 cm high). Water depth was 50 cm and temperature was maintained at 23 ± 1°C.Acute Swim Stress (ASS) procedure
Rats were
subjected to ASS as previously described [33]. Individual rats were subjected
to a single 15-minute swim session in the water container. After this single
swim session, rats were allowed to dry in a resting cage for 30 minutes, and
then anesthetized and taken to electrophysiology.


## 3. Results

### 3.1. Acute Swim Stress Induces Elevated Levels of Serum
Corticosterone and Increased Levels of Anxiety

Elevated plus maze
In order to validate that
animals subjected to the acute swim stress procedure were indeed behaviorally
affected, we used the elevated plus maze test [34].
A paired samplet-test revealed that in the poststress
session, animals spent less time in the open arms (see Figure
1(a), t(7) = 4.25, P < .005).(a)Time spent in open arms of the elevated
plus maze before and after exposure to stress. Following
exposure to acute swim stress, rats spent less
time in the open arms of the maze in comparison to the time spent in the open
arms prior to the stress procedure (n = 8, P < .005), indicating
increased anxiety. (b) Levels of corticosterone in control and
stress rats. Acute swim stress induced elevation of serum corticosterone
in the stressed rats (n = 6) compared to control rats (n = 8, P <  .005).
(a)(b)Levels of corticosterone
Levels of serum
corticosterone were measured for control and stressed rats. An independentt-test
revealed that stress was associated with a significant increase in
corticosterone levels (see Figure 1(b), t(13) = 4.29, P < .005)  (see Figure 1(b)).


### 3.2. Acute Swim Stress does not Affect Baseline Responses in
PP-DG Pathway

The stimulation intensity used to elicit a baseline
response was not different between the control and stress groups (t(25) = 0.93, n.s.).
There was no significant difference between control and stressed rats in the
amplitude of the baseline PS (t(25) = 1.44, n.s.))  and fEPSP slope (t(25) = 0.26, n.s.).


### 3.3. The Effect of Behavioral Stress on Frequency-Dependent
Inhibition

Upon altering the frequency of stimulation from 0.1 Hz to
1.0 Hz, a marked reduction of the PS was observed in both control and stressed
rats as  indicated  by the FDI index (see Figure2(a)). This
reduction was apparent prior to TBS application and 60 following it (see Figure
2(b), F(1,14) = 101.16, P < .05). No significant differences in FDI index were
found between the two groups on both tests (F(1,14) = 0.18, n.s.)(a) Left:
representative field potential response of dentate gyrus granule cells to stimulating
of the PP at 0.1 Hz.  Right:
representative field potential response of dentate gyrus granule cells to
stimulation of the PP at 1.0 Hz. Time unit: second. (b)FDI before and after TBS application. The application of TBS significantly reduced FDI in both
control (n = 8) and stressed  (n = 8) rats (P < .05).
(a)(b)Although no main effect was observed for group under the
different conditions, the analysis of variance indicated a significant group X
test interaction (F(1,14) = 6.53,P < .05). The initial pre-TBS FDI
index of stressed rats was lower than that of controls whereas the post-TBS FDI
index was higher.Altering stimulation frequency from 0.1 Hz to 1.0 Hz resulted
in a slight but significant reduction in the EPSP slope in both groups. In
control rats, stimulation at 1.0 Hz reduced the slope of the EPSP to0.85±0.05 of its size during stimulation at 0.1 Hz (t(7) = 11.49, P < . 05).
In stressed rats, stimulation at 1.0 Hz reduced the slope of the EPSP to 0.88±0.02 of its size during stimulation at 0.1 Hz [t(7) = 3.66, P < 0.05].
There was no significant difference in EPSP FDI between the two groups and, in
contrast to the PS, the application of TBS did not affect EPSP FDI in either
group.


### 3.4. The Effects of Behavioral Stress on Commissural Modulation

In both control and stressed rats, the response to the PP
stimulation was inhibited by priming stimulation of the commissural path at
interpulse intervals of 15 milliseconds and 30 milliseconds (see Figure3(a)).
When using the 15-millisecond interpulse interval, exposure to stress induced
significantly higher inhibition, as expressed by the small commissural index of
stressed rats compared to controls. This increase in inhibition was apparent
before and after TBS application (F(1,12) = 64.36, P < .005). The
application of TBS did not cause any significant changes in commissural-induced
inhibition in either group (F(1,12) = 0.126, n.s). (See Figures 3(b) and 3(c).)(a)The effects of commissural priming on
responses of the DG to PP stimulation. Top: representative field potential responses of  dentate gyrus cells to stimulating the PP
without commissural priming (left), and to stimulating the PP with priming
stimulation to the contralateral DG at 15 milliseconds (right) in control rats. Bottom: representative field potential responses of  dentate gyrus cells to stimulating the PP
without commissural priming (left), and to stimulating the PP with priming
stimulation to the contralateral DG at 15 milliseconds (right) in stressed
rats. Time unit: second. (b) Commissural-induced modulation prior to TBS
application. Commissural-induced inhibition was significantly higher in
stressed rats (n = 6) at 15-millisecond and 30-millsecond interpulse intervals in
comparison with control rats (n = 6) (P < .05). (c) Commissural-induced modulation following
TBS application. Commissural-induced inhibition was significantly higher in
stressed animals at 15-millisecond and 30-millisecond intervals n = 6 (P <
.05).
(a)(b)(c)When priming stimulation of the commissural path at interpulse
interval of 30 milliseconds, stressed rats exhibited significantly higher
commissural-induced inhibition than control rats before and after TBS
application (F(1,12) = 90.42,P < .001), as seen in the 15-millisecond interpulse
interval. Interestingly, both groups have shown a significant decrease in commissural-induced
inhibition following TBS application (F(1,12) = 13.68, P < .005).
(See Figures
3(b) and 3(c).)When using the 80-millisecond and 150-millisecond interpulse
intervals, the response to the PP stimulation was facilitated in both control
and stressed rats, as expressed by  the
large commissural indexes. In both groups, commissural-induced facilitation was
significantly increased after TBS application (80 milliseconds: F(1,12) = 17.55,P <
.005; 150 milliseconds: F(1,12) = 7.87, P < .05). No differences were
found between the two groups before or after TBS application (80 milliseconds:
F(1,12) = 0.68, n.s; 150 milliseconds: F(1,12) = 0.057, n.s). (See Figures 3(b)
and 3(c).)


### 3.5. The Effect of the Stressor on LTP Induction in the
Dentate Gyrus

Sixty minutes following the application of TBS, there was a
clear potentiation of the slope of the EPSP in control rats (38% ± 5.9, t(13) =
6.38,P < .001) and stressed rats (35% ±  9.3, t(10) = 3.54, P < .05), compared
to pre-TBS levels.  The application of
TBS also resulted in an increase in PS amplitude in both groups (control: 128%
± 32, t(13) = 4.02, P < .005);
stress: 127% ±  38, t(10) = 4.6, P <
.05).No significant difference in LTP was found between the two
groups (EPSP-LTP: t(23) = 0.33; PS-LTP: t(23) = 0.93, n.s)] (see Figure4).Figure 4
TBS-induced potentiation in control and stress
rats. Both control (n = 14) and stress (n = 11) rats have shown an
increase in the slope of the EPSP. The magnitude of the EPSP potentiation was
not significantly different between the two groups.


## 3.1. Acute Swim Stress Induces Elevated Levels of Serum
Corticosterone and Increased Levels of Anxiety

Elevated plus maze
In order to validate that
animals subjected to the acute swim stress procedure were indeed behaviorally
affected, we used the elevated plus maze test [34].
A paired samplet-test revealed that in the poststress
session, animals spent less time in the open arms (see Figure
1(a), t(7) = 4.25, P < .005).(a)Time spent in open arms of the elevated
plus maze before and after exposure to stress. Following
exposure to acute swim stress, rats spent less
time in the open arms of the maze in comparison to the time spent in the open
arms prior to the stress procedure (n = 8, P < .005), indicating
increased anxiety. (b) Levels of corticosterone in control and
stress rats. Acute swim stress induced elevation of serum corticosterone
in the stressed rats (n = 6) compared to control rats (n = 8, P <  .005).
(a)(b)Levels of corticosterone
Levels of serum
corticosterone were measured for control and stressed rats. An independentt-test
revealed that stress was associated with a significant increase in
corticosterone levels (see Figure 1(b), t(13) = 4.29, P < .005)  (see Figure 1(b)).


## 3.2. Acute Swim Stress does not Affect Baseline Responses in
PP-DG Pathway

The stimulation intensity used to elicit a baseline
response was not different between the control and stress groups (t(25) = 0.93, n.s.).
There was no significant difference between control and stressed rats in the
amplitude of the baseline PS (t(25) = 1.44, n.s.))  and fEPSP slope (t(25) = 0.26, n.s.).


## 3.3. The Effect of Behavioral Stress on Frequency-Dependent
Inhibition

Upon altering the frequency of stimulation from 0.1 Hz to
1.0 Hz, a marked reduction of the PS was observed in both control and stressed
rats as  indicated  by the FDI index (see Figure2(a)). This
reduction was apparent prior to TBS application and 60 following it (see Figure
2(b), F(1,14) = 101.16, P < .05). No significant differences in FDI index were
found between the two groups on both tests (F(1,14) = 0.18, n.s.)(a) Left:
representative field potential response of dentate gyrus granule cells to stimulating
of the PP at 0.1 Hz.  Right:
representative field potential response of dentate gyrus granule cells to
stimulation of the PP at 1.0 Hz. Time unit: second. (b)FDI before and after TBS application. The application of TBS significantly reduced FDI in both
control (n = 8) and stressed  (n = 8) rats (P < .05).
(a)(b)Although no main effect was observed for group under the
different conditions, the analysis of variance indicated a significant group X
test interaction (F(1,14) = 6.53,P < .05). The initial pre-TBS FDI
index of stressed rats was lower than that of controls whereas the post-TBS FDI
index was higher.Altering stimulation frequency from 0.1 Hz to 1.0 Hz resulted
in a slight but significant reduction in the EPSP slope in both groups. In
control rats, stimulation at 1.0 Hz reduced the slope of the EPSP to0.85±0.05 of its size during stimulation at 0.1 Hz (t(7) = 11.49, P < . 05).
In stressed rats, stimulation at 1.0 Hz reduced the slope of the EPSP to 0.88±0.02 of its size during stimulation at 0.1 Hz [t(7) = 3.66, P < 0.05].
There was no significant difference in EPSP FDI between the two groups and, in
contrast to the PS, the application of TBS did not affect EPSP FDI in either
group.


## 3.4. The Effects of Behavioral Stress on Commissural Modulation

In both control and stressed rats, the response to the PP
stimulation was inhibited by priming stimulation of the commissural path at
interpulse intervals of 15 milliseconds and 30 milliseconds (see Figure3(a)).
When using the 15-millisecond interpulse interval, exposure to stress induced
significantly higher inhibition, as expressed by the small commissural index of
stressed rats compared to controls. This increase in inhibition was apparent
before and after TBS application (F(1,12) = 64.36, P < .005). The
application of TBS did not cause any significant changes in commissural-induced
inhibition in either group (F(1,12) = 0.126, n.s). (See Figures 3(b) and 3(c).)(a)The effects of commissural priming on
responses of the DG to PP stimulation. Top: representative field potential responses of  dentate gyrus cells to stimulating the PP
without commissural priming (left), and to stimulating the PP with priming
stimulation to the contralateral DG at 15 milliseconds (right) in control rats. Bottom: representative field potential responses of  dentate gyrus cells to stimulating the PP
without commissural priming (left), and to stimulating the PP with priming
stimulation to the contralateral DG at 15 milliseconds (right) in stressed
rats. Time unit: second. (b) Commissural-induced modulation prior to TBS
application. Commissural-induced inhibition was significantly higher in
stressed rats (n = 6) at 15-millisecond and 30-millsecond interpulse intervals in
comparison with control rats (n = 6) (P < .05). (c) Commissural-induced modulation following
TBS application. Commissural-induced inhibition was significantly higher in
stressed animals at 15-millisecond and 30-millisecond intervals n = 6 (P <
.05).
(a)(b)(c)When priming stimulation of the commissural path at interpulse
interval of 30 milliseconds, stressed rats exhibited significantly higher
commissural-induced inhibition than control rats before and after TBS
application (F(1,12) = 90.42,P < .001), as seen in the 15-millisecond interpulse
interval. Interestingly, both groups have shown a significant decrease in commissural-induced
inhibition following TBS application (F(1,12) = 13.68, P < .005).
(See Figures
3(b) and 3(c).)When using the 80-millisecond and 150-millisecond interpulse
intervals, the response to the PP stimulation was facilitated in both control
and stressed rats, as expressed by  the
large commissural indexes. In both groups, commissural-induced facilitation was
significantly increased after TBS application (80 milliseconds: F(1,12) = 17.55,P <
.005; 150 milliseconds: F(1,12) = 7.87, P < .05). No differences were
found between the two groups before or after TBS application (80 milliseconds:
F(1,12) = 0.68, n.s; 150 milliseconds: F(1,12) = 0.057, n.s). (See Figures 3(b)
and 3(c).)


## 3.5. The Effect of the Stressor on LTP Induction in the
Dentate Gyrus

Sixty minutes following the application of TBS, there was a
clear potentiation of the slope of the EPSP in control rats (38% ± 5.9, t(13) =
6.38,P < .001) and stressed rats (35% ±  9.3, t(10) = 3.54, P < .05), compared
to pre-TBS levels.  The application of
TBS also resulted in an increase in PS amplitude in both groups (control: 128%
± 32, t(13) = 4.02, P < .005);
stress: 127% ±  38, t(10) = 4.6, P <
.05).No significant difference in LTP was found between the two
groups (EPSP-LTP: t(23) = 0.33; PS-LTP: t(23) = 0.93, n.s)] (see Figure4).Figure 4
TBS-induced potentiation in control and stress
rats. Both control (n = 14) and stress (n = 11) rats have shown an
increase in the slope of the EPSP. The magnitude of the EPSP potentiation was
not significantly different between the two groups.


## 4. Discussion

In the present study, we have examined the effects of behavioral
stress on local circuit activity and plasticity in the DG. We report that when
using FDI [26], this form of local circuit activity was reduced following the
application of TBS in both control and stressed rats. However, this reduction
in FDI plasticity was greater in the stressed rats compared to controls.When using commissural-induced modulation, inhibition was
significantly higher in stressed rats both before and after TBS at 15- and 30-millisecond
intervals.FDI results from direct afferent excitation of inhibitory
interneurons or of other cells that excite inhibitory cells [26, 35], and thus
provides a simple method for measuring local circuit activity mediated by
dentate interneurons. In the present study, an attempt was made to find out
whether behavioral stress would have an effect on FDI activity and plasticity.
Our results reveal that both control and stressed rats have shown a decrease in
FDI following TBS, and that differences between pre- and posttetanic levels of
FDI were not significantly different between the two groups. When examining the
differences between pre- and post-TBS levels of FDI within each group, a
greater difference was observed in the stressed group, which may suggest that
undergoing behavioral stress has caused an increase in FDI plasticity.This increased plasticity is somewhat surprising since
exposure to stress is typically shown to suppress plasticity and to impair
learning [36–40]. It should be noted though that most studies
related to stress effects on plasticity focused mainly on LTP and on CA1 area
of the hippocampus. The effects of stress on LTP in other brain regions are
less consistent [14, 21, 41].
Furthermore, even in CA1, while stress may suppress LTP, it is reported
to enhance other forms of plasticity, such as LTD [17, 42].The effects of stress on commissural-induced modulation
were also examined. Stressed rats have shown a marked increase in inhibition
both before and after the application of TBS, indicating a lasting modification
in this form of local circuit activity. This result is in agreement with other
studies that have shown that stress levels of corticosterone produce a profound
and long-lasting inhibitory influence on hippocampal cell activity [43–47]. Although stress has caused a reduction in local circuit activity in
this case, no alteration in commissural local circuit plasticity was observed
after TBS application.The fact that FDI activity was not affected by the stress
while commissural-induced modulation was significantly affected suggests that
stress may affect only a subpopulation of inhibitory interneurons. Indeed, it
has been previously suggested that interneurons in the hippocampus may be
divided into those showing no activity-dependent plasticity and those that do.
It was further suggested that activity-dependnet plasticity of this sort may
contribute to mood and anxiety disorders [48].The fact that an increase in inhibitory activity was
observed in the hippocampus may imply that
deficits in cognitive functioning and flexibility might take place, as has been
suggested before for aged rats [18]. This does not necessarily mean that rats
that have undergone stress would show impaired learning in a specific task, but
it is possible that they would be less adaptive if required to shift to a new
coping strategy during task acquisition. For example, stress has been shown to
cause impairment in reversal learning and induced perseveratory behavior in the
Morris water maze without having a significant effect on task acquisition [49].In the present study, dentate gyrus LTP was not affected by
the stress employed. As indicated above, the effects of stress on dentate gyrus
LTP may depend on the exact nature of the stress experience [14, 21, 41].
Interestingly, although stress seemed to have caused alterations in local
circuit plasticity (as observed in the FDI findings) and in local circuit
activity (as observed by the commissural modulation findings), it had no effect
on LTP induction. This further supports the notion that local circuit activity
and plasticity is independent of synaptic plasticity such as LTP [18, 28].The study of the effects of stress on GABAergic
neurotransmission is of special interest because it has been suggested that
GABA plays a role in the pathophysiology of mood and anxiety disorders [50–54]. The results presented here may further support the potential role
of GABA-impaired modulation of neural activity and plasticity in stress-related
disorders.Our results suggest that stressful experience may lead to
alterations in local circuit activity and plasticity. Understanding the
alterations that take place in these local interneurons may contribute to a
better understanding of their involvement in memory formation and regulation
under normal and psychopathological conditions.



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*Source: 194097-2008-02-14.xml*