11Drug TargeT InsIghTs 2014:8

Open Access: Full open access to
this and thousands of other papers at
http://www.la-press.com.

Drug Target
Insights

Introduction
Several lines of evidence have suggested that impaired cog-
nition is an element of depression and that antidepressant
therapy may improve cognitive function.1 Classic antidepres-
sants with a central monoaminergic function are associated
with a delayed therapeutic response and side effects, thereby
limiting their usage. Although the pathology of depression
and the mechanism of action of antidepressant drugs are

well known, the adequacy of current antidepressant targets is
questionable.

Stress is an important risk factor in the development
of depression,2 and stress-induced memory impairments are
commonly reported.3 Therefore, the use of an antidepressant
to ameliorate stress-induced cognitive deficits is of therapeutic
relevance. Drugs that elevate the mood of depressed patients
are associated with synaptic effects, that is, the ability to

The Antidepressant Agomelatine Improves Memory Deterioration and
Upregulates CREB and BDNF Gene Expression Levels in Unpredictable
Chronic Mild Stress (UCMS)-Exposed Mice

esen gumuslu1, Oguz Mutlu2, Deniz sunnetci1, guner ulak2, Ipek K. Celikyurt2,
naci Cine1, Furuzan akar2, Hakan Savlı1 and Faruk erden2
1Department of Medical Genetics, Kocaeli University Medical Faculty, Kocaeli, Turkey. 2Pharmacology, Kocaeli University Medical Faculty,
Kocaeli, Turkey.

Abstr Act: Agomelatine, a novel antidepressant with established clinical efficacy, acts as an agonist of melatonergic MT1 and MT2 receptors and as
an antagonist of 5-HT2C receptors. The present study was undertaken to investigate whether chronic treatment with agomelatine would block unpredict-
able chronic mild stress (UCMS)-induced cognitive deterioration in mice in passive avoidance (PA), modified elevated plus maze (mEPM), novel object
recognition (NOR), and Morris water maze (MWM) tests. Moreover, the effects of stress and agomelatine on brain-derived neurotrophic factor (BDNF)
and cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) messenger ribonucleic acid (mRNA) levels in the hippocam-
pus was also determined using quantitative real-time polymerase chain reaction (RT-PCR). Male inbred BALB/c mice were treated with agomelatine
(10 mg/kg, i.p.), melatonin (10 mg/kg), or vehicle daily for five weeks. The results of this study revealed that UCMS-exposed animals exhibited memory
deterioration in the PA, mEPM, NOR, and MWM tests. The chronic administration of melatonin had a positive effect in the PA and +mEPM tests,
whereas agomelatine had a partial effect. Both agomelatine and melatonin blocked stress-induced impairment in visual memory in the NOR test and
reversed spatial learning and memory impairment in the stressed group in the MWM test. Quantitative RT-PCR revealed that CREB and BDNF gene
expression levels were downregulated in UCMS-exposed mice, and these alterations were reversed by chronic agomelatine or melatonin treatment. Thus,
agomelatine plays an important role in blocking stress-induced hippocampal memory deterioration and activates molecular mechanisms of memory storage
in response to a learning experience.

Key words: agomelatine, melatonin, depression, memory, BDNF, CREB

CITATIoN: gumuslu et al. The antidepressant agomelatine Improves Memory Deterioration and upregulates CreB and BDnF gene expression Levels in unpredictable
Chronic Mild stress (uCMs)-exposed Mice. Drug Target Insights 2014:8 11–21 doi: 10.4137/DTI.s13870.

RECEIvED: December 11, 2013. RESUBMITTED: January 19, 2014. ACCEpTED FoR pUBLICATIoN: February 6, 2014.

ACADEMIC EDIToR: Prithviraj Bose, editor in Chief

TYpE: Original research

FUNDING: author(s) disclose no funding sources.

CoMpETING INTERESTS: Author(s) disclose no potential conflicts of interest.

CopYRIGhT: © the authors, publisher and licensee Libertas academica Limited. This is an open-access article distributed under the terms of the Creative Commons
CC-BY-nC 3.0 License.

CoRRESpoNDENCE: oguzmutlu80@hotmail.com

http://www.la-press.com
http://www.la-press.com
http://dx.doi.org/10.4137/DTI.S13870
mailto:oguzmutlu80@hotmail.com

Gumuslu et al

12 Drug TargeT InsIghTs 2014:8

increase the extracellular levels of serotonin, noradrenaline,
and dopamine in the brain.4 The monoamine hypothesis of
depression fails to explain all the effects of antidepressants5
and receptor activation as a consequence of the elevation in
synaptic monoamines, which represent the primary molecular
results of antidepressants.

One of the more recent novel antidepressant mechanisms
identified is the control of circadian rhythms. Melatonin, the
major hormone of the pineal gland, is an endogenous agonist
of MT1/MT2 receptors and is involved in the regulation of
the sleep–wake cycle. The recognition that circadian rhythm
desynchronization also plays a key role in mood disorders has
led to the development of agomelatine. Agomelatine, the first
melatonergic antidepressant, is an agonist of the melatonergic
MT1 and MT2 receptors

6 and an antagonist of the serotonergic
5-HT2C receptors,

7 and it mimics the actions of melatonin in
the synchronization of circadian rhythm patterns in rodents.8
The antidepressant-like activity of agomelatine is attributed to
the synergy between these sets of receptors, which are impor-
tant components of the circadian timing system. Agomelatine
has demonstrated antidepressant-like activity in several ani-
mal models of depression9 and in a transgenic mouse model
of depression.10 Clinically, it has demonstrated efficacy in
major depressive disorders in several trials.11 Agomelatine is
an effective treatment for depression because it resynchronizes
circadian rhythms12 that are disturbed in depression. Indeed,
it has been speculated for a considerable amount of time that
the disorganization of internal circadian rhythms plays a criti-
cal role in the development of major depression.13 Agomela-
tine has a favorable clinical profile of antidepressant properties
and fewer side effects than traditional antidepressants.11

Unpredictable chronic mild stress (UCMS) is an impor-
tant behavioral model that resembles human depression.14
The hippocampus and its connections within limbic–cortical
networks may play a crucial role in the pathogenesis of major
depression. Acute stress and chronic stress disturb hippocam-
pal-dependent memory and prevent the formation of long-
term potentialization, which plays a role in the formation of
synaptic plasticity and memory.

The expression of genes implicated in neuronal plasticity,
such as brain-derived neurotrophic factor (BDNF) and cyclic
adenosine monophosphate (cAMP) response element bind-
ing protein (CREB), have been shown to be downregulated
in stressed mice.15 BDNF is a neurotrophin that modulates
neuronal plasticity, which is frequently associated with anti-
depressant treatment.16 Neurotrophin expression is activity-
dependent17 and may be regulated by the light and dark cycle
in rats18 and in humans.19 The regulation of the neurotrophin
BDNF, whose gene and protein expression and function may
be defective in mood disorders,20 has been extensively inves-
tigated in recent years as one of the mechanisms of antide-
pressants. The modulation of BDNF represents a key element
in long-term adaptive changes induced by antidepressant
drugs. Moreover, such molecular analyses were preceded by

behavioral assays to investigate the antidepressant activity of
agomelatine via the forced swimming test21 and to evaluate
the effect of agomelatine on recognition memory in the novel
object recognition (NOR) task.22 Chronic administration of
agomelatine leads to the upregulation of BDNF-LTP (long-
term potentialization)-related genes and reverses depression-
like symptoms. CREB is a core component of the molecular
switch that converts short-term memory to long-term mem-
ory. Recent studies have established the role of CREB in
learning and memory in mammals in addition to providing
insight into the molecular mechanisms of CREB regulation
and function. The involvement of CREB and the upstream
signaling pathways leading to its activation in learning-
associated plasticity makes them attractive targets for drugs
aimed at improving memory function in both diseased and
healthy individuals.23

As stress plays an important role in the development of
depression, we aimed to investigate whether chronic treat-
ment with agomelatine, a novel antidepressant that has a
unique receptor profile as a MT1/MT2 melatonergic agonist

6
and 5-HT2C receptor antagonist,

7 would block UCMS-
induced cognitive deterioration in mice in the passive avoid-
ance (PA), modified elevated plus maze (mEPM), NOR, and
Morris water maze (MWM) tests. As the genes involved in
neurite remodeling are among the primary targets of regu-
lation by chronic stress, the effects of stress and the chronic
administration of agomelatine on BDNF and CREB messen-
ger ribonucleic acid (mRNA) expression in the hippocampus
of stressed mice were also determined using quantitative real-
time polymerase chain reaction (RT-PCR).

Methods
Animals. Male, inbred BALB/cByJ mice (MAM

TUBİTAK, Gebze, Kocaeli, Turkey), seven to eight weeks
old at their arrival to the laboratory, were used in this study.
The animals (four to five per cage) were kept in the laboratory
at 21 ± 1.5 °C with 60% relative humidity under a 12 hour
light/dark cycle (lights on at 8:00 p.m.) for two weeks before
experimentation. The animals were assigned to one of two
treatment groups, the non-stressed group (controls) and mice
subjected to the UCMS procedure. Non-stressed mice were
housed in groups (eight mice per cage) during the experiment,
whereas mice in the stressed group were housed individually
in cages (length: 268 mm, width: 135 mm, height: 81 mm)
from the start of the chronic stress until the end of the study.
All animals received food and water ad libitum. A group-
housed control group was preferred to an individual-housing
condition because social isolation is highly stressful for mice
and should thus per se contribute to the effects of chronic
stress.24,25 All procedures described in this paper were con-
ducted in accordance with the European Community Council
directive for the Ethical Treatment of Animals (86/609/EEC)
and with the ethical approval of the Kocaeli University Eth-
ics Committee (Number: AEK 9/3 2010, Kocaeli, Turkey).

http://www.la-press.com

Agomelatin improves memory in stress exposed mice

13Drug TargeT InsIghTs 2014:8

All animals were naive to the experimental apparatus, and
different animals were used for each test.

experimental groups and drug administration. Mela-
tonin was purchased from Merck Chemical Company (Merck,
Hohenbrunn, Germany), and agomelatine was purchased
from Wuhan Sunrise Technology Development Company
Limited (Wuhan, China). Both were dissolved in saline sup-
plemented with 10% DMSO. All drugs were freshly prepared
and administered in a volume of 0.1 ml per 10 g body weight.
The control groups received the same volume of vehicle. At
the end of two weeks of drug-free UCMS, the mice were
assigned to six experimental groups (n = 15 per group) in a
semi-randomized manner such that the initial coat state and
body weights were equivalent in all of the groups. Melatonin
(10 mg/kg), agomelatine (10 mg/kg), or vehicle was admin-
istered intraperitoneally (i.p.) each day at 17:00 hours (two
hours before the lights were turned off ) for five weeks to both
stressed and non-stressed animals.

On the final day of injections (day 35), agomelatine-,
melatonin-, or vehicle-treated mice (n = 7 per group) were
sacrificed without behavioral testing to examine the effects of
the drugs on the gene expression levels of BDNF and CREB.
The remaining animals (n = 8 per group) underwent training
in the elevated plus maze, PA, and MWM tests. All behav-
ioral testing and tissue and blood sampling were conducted
two to five hours following the final injection, between 19:00
and 22:00 hours.

UcMs procedure. The UCMS regimen used in this
study was based on the procedure originally designed by Will-
ner et al.26 and adapted to mice.27 This stress model consists of
repeated mild physical and psychological stressors. Mice were
subjected to different types of stressors in a chronic, inevita-
ble, and unpredictable way several times a day for seven weeks.
Stressors were administered in a pseudo-random manner and
could occur at any time of night or day. In this respect, the
stressor sequence was changed every week to make the stress
procedure unpredictable. In all of the experiments, the first
two drug-free weeks of UCMS were followed by five weeks
of UCMS application during which the mice were treated
with drug or vehicle. For further details on the procedure, see
Yalcin et al.28

PA test. Animals were trained in a one-trial, step-
through PA apparatus to evaluate memory based on contex-
tual fear conditioning and instrumental learning.29 A decrease
in retention latency indicates an impairment in memory in the
PA task. The apparatus consisted of a box with an illuminated
part (L 7 × 12.5 × h 14 cm) and a dark part (L 24 × 12.5 × h
14 cm), both equipped with a grid floor composed of steel bars
(0.3 cm diameter) spaced 0.9 cm apart. The inhibitory avoid-
ance task consisted of two trials. On the first day of training, the
mice were individually placed into the light compartment and
allowed to explore the boxes. The intercompartment door was
opened after a 60 second acclimation period. In the acquisition
trial, each mouse was placed in the illuminated compartment,

which was lit by a bright bulb (2000 lux). The animals received
drugs prior to acquisition training. If the mouse stepped into
the dark compartment (2/3 of the tail in the dark compart-
ment), the door was closed by the experimenter, and an ines-
capable foot shock (0.25 mA/1 second) was delivered through
the grid floor of the dark compartment. A cutoff time of five
minutes was selected. The time taken to enter the dark com-
partment (training latency) was recorded. Immediately after
the shock, the mouse was returned to the home cage. The
retention trial started 24 hours after the end of the acquisition
trial. Each mouse was placed in the illuminated compartment
as in the training trial. The door was opened after a 30 second
acclimation period. The step-through latency in the retention
trial (with a maximum 300 seconds cutoff time) was used as
the index of retention of the learned experience. A shock was
not applied during the retention trial.

mePM test. Cognitive behavior was evaluated using the
mEPM learning task, which measures spatial long-term mem-
ory.30 The maze was made of wood and consisted of two open
arms (29 × 5 cm) surrounded by a short (1 cm) Plexiglas edge
to avoid falls and two enclosed arms (29 × 5 × 15 cm) arranged
such that the two open arms were opposite to each other. The
arms were connected by a central platform (5 × 5 cm). The
maze was elevated 40 cm above the floor. The principle of
this experiment is based upon the aversion of rodents to open
spaces and heights. The animals prefer the enclosed, protected
areas of the maze.

The procedure was performed as described previously.30,31
During the acquisition session (day 1), each mouse was gently
placed at the distal end of an open arm facing away from the
central platform. The time required for the mice to move from
the open arm to either of the enclosed arms (transfer latency)
was recorded. Training (repeated exposure of animals to the
open arms) shortened this parameter, possibly as a conse-
quence of learning acquisition and retention. If the mouse did
not enter the enclosed arm within 90 seconds, it was excluded
from further experimentation. Animal entry into the enclosed
arm required the animal to cross an imaginary line separating
the enclosed arm from the central space with all four legs.
After entering the enclosed arm, mice were allowed to move
freely in the maze in both the open and enclosed arms for
10 seconds. Mice were then returned to their home cage. The
retention session occurred 24 hours after the acquisition ses-
sion (on day 2). Mice were placed in the open arm, and the
transfer latency was recorded again. Experiments were con-
ducted between 10:00 and 14:00 hours in a dimly lit, semi-
soundproof room under natural light.

Nor. We used a NOR test protocol based on that of
Ennaceur and Delacour22 with slight modifications. The appa-
ratus consisted of a circular open field (40 cm diameter and
30 cm height) made of PVC with a black-and-white striped
cardboard pattern (30 × 20 cm) nailed to one of the walls and a
Plexiglas floor. A light bulb above the central section provided
constant illumination of approximately 100 lux. The NOR

http://www.la-press.com

Gumuslu et al

14 Drug TargeT InsIghTs 2014:8

task procedure consisted of the following three components:
habituation, training, and retention. Each mouse was indi-
vidually habituated to the apparatus for five minutes in the
absence of objects (habituation trial). The mouse was placed in
the apparatus for the training trial and two identical objects
(moon or butterfly) were placed in a symmetrical position
10 cm above the side wall 30 minutes after the habituation
trial. The order of objects used for each subject per trial was
determined randomly. The total time spent exploring the two
objects was recorded by the experimenter over five minutes.
Exploration of an object was defined as directing the nose
toward the object and/or touching it with the nose. After a
predetermined retention interval of one hour, the mouse was
placed back into the apparatus for the retention trial; how-
ever, during this trial, two dissimilar objects were presented,
a familiar one and a new one. The object not used in the train-
ing trial was used as the novel object in the retention trial. The
animals were allowed to explore freely for five minutes and the
time spent exploring each object was recorded. If recognition
memory was intact, the mouse would be expected to spend
more time exploring the novel object.30 The ratio index (RI)
was calculated as the time spent exploring the new object (N)
divided by the total time exploring both objects (N + R) mul-
tiplied by 100. A higher RI was considered to reflect greater
memory retention.

MwM. The MWM was a circular pool (90 cm diam-
eter and 30 cm height) filled with water (22ºC) to a depth
of 14 cm and rendered opaque by the addition of small black
balls. The pool was located in a dimly lit, soundproof test room
with a various visual cues, including a white and black poster
on the wall, a halogen lamp, a camera, and the experimenter.
The maze was divided into four quadrants, and three equally
spaced points served as starting positions around the edge of
the pool. The order of the release positions varied systemati-
cally throughout the experiment. A circular escape platform
(6 cm diameter and 12 cm high) was located in one quadrant
1 cm above the water surface during the familiarization session
and 1 cm below the water surface during the other sessions.

Video tracking was conducted with a video camera
focused on the full diameter of the pool. The navigation
parameters were analyzed using the Ethovision 3.1 video
analysis system (Noldus, The Netherlands). The mice were
trained in the MWM five times daily (familiarization session,
S1, S2, S3, S4).

One familiarization and four acquisition sessions were
performed using the MWM. During the familiarization ses-
sion and acquisition phase of the experiment, each mouse
was subjected to three trials. The delay between the trials was
60 seconds, and a one-day interval was used between each ses-
sion. For each trial, the mouse was taken from the home cage
and placed into the water maze at one of three randomly deter-
mined locations with its head facing the center of the water
maze. After the mouse found and climbed onto the platform,
the trial was stopped, and the escape latency was recorded. If

the mouse did not climb onto the platform in 60 seconds, the
trial was stopped, and the experimenter guided the mouse to
the platform; an escape latency of 60 seconds was recorded.

A “probe trial” was used to assess the spatial memory
retention of the location of the hidden platform 24 hours after
the last acquisition session. During this trial, the platform was
removed from the maze, and the mouse was allowed to search
the pool for 60 seconds. The percent of time spent in each
quadrant was recorded.

tissue sampling, rNA isolation, and quantitative
rt-Pcr. One day after the final stress session, the mice
were decapitated by cervical dislocation. The left and right
hippocampi were surgically removed and stored in liquid
nitrogen. Total RNA was isolated with the RNeasy Mini Kit
extraction procedure (Qiagen, Valencia, CA, USA). Briefly,
tissues were homogenized in RLT lysis buffer containing
β-mercaptoethanol using a Thermo Savant FastPrep FP120
Homogenizer. Sample homogenates were applied to RNeasy
Mini Spin columns (Qiagen) and processed according to the
manufacturer’s instructions. An on-column DNase digestion
was performed to remove any residual genomic DNA con-
tamination. RNA samples were eluted in RNase-free water,
and the concentration was measured spectrophotometrically
using the NanoDrop ND-1000 Spectrophotometer (Nano-
Drop ND-1000; NanoDrop Technologies, Wilmington,
DE). Subsequently, cDNA was synthesized using a RevertAid
First Strand cDNA synthesis kit (Fermentas Inc., Maryland,
USA). Quantitative RT-PCR was performed according to the
methods described in previous studies.32,33 Standard curves
were obtained via serial dilutions of the beta-globulin gene.
Primers specific to the genes under investigation (Table 1)
were obtained from Integrated DNA Technologies (Iowa,
USA) and IONTEK Inc. (Merter, Istanbul, Turkey). The
gene expression values obtained were normalized using the
BACT housekeeping gene. Gene expression levels were cal-
culated with the REST (Relative Expression Software Tool)
program. Changes in the CREB and BDNF gene expression
levels were calculated in the stress-exposed and non-stressed
(n = 7/each group) animal groups. The effects of agomela-
tine and melatonin (n = 7/each group) on CREB and BDNF
expression levels were also evaluated in the stress-exposed
group.

statistics
One-way analysis of variance (ANOVA) and the post-hoc
Tukey’s test were used to analyze the mEPM, MWM, and
NOR tests. To evaluate the differences among drug treat-
ment groups during the first and second transfer latencies in
the PA test, the Kruskal–Wallis non-parametric test was used
followed by Dunn’s post-hoc test. Data are expressed as the
mean values ± SEM. P , 0.05 was accepted as statistically
significant. Statistical evaluation of BDNF and CREB gene
expressions was performed with the REST (Relative Expres-
sion Software Tool) program.

http://www.la-press.com

Agomelatin improves memory in stress exposed mice

15Drug TargeT InsIghTs 2014:8

results
effects of drugs on learning and memor y in the PA

test. There was no significant difference in first day latency
among the groups (H = 6.98, P . 0.05, Figure 1A). The
second day latency (retention latency) significantly differed
between the groups (H = 17.82, P = 0.003). Stress signifi-
cantly shortened the retention latency compared to the non-
stressed control group (P , 0.05). Melatonin prolonged

the retention latency in stressed animals (P , 0.05), while
agomelatine had a partial but statistically insignificant
effect (Fig. 1B).

effects of drugs on learning and memory in the mePM
test. After chronic injection of melatonin (10 mg/kg) or ago-
melatine (10 mg/kg) for five weeks, there was no signifi-
cant difference in first day latency (TL1) among the groups
[F(5,35) = 2.22; P = 0.07, Figure 2A]. TL2 (latency on the
second day) was significantly different when all groups were
compared [F(5,35) = 3.38; P = 0.01, Figure 2B]. TL2 sig-
nificantly increased in the stressed control group compared
to the non-stressed control group (P , 0.05), and this effect
was significantly reversed by melatonin (P , 0.05), while ago-
melatine had a partial effect but failed to reach to a statistically
significant value (Fig. 2B).

effects of drugs on visual memory in the Nor test.
A significant difference was observed between the groups
[F(5,41) = 9.80; P , 0.001] when the effects of melatonin or
agomelatine were evaluated during the retention trial of the
NOR test. The RI between the stressed control and non-
stressed control mice were significantly different (P , 0.001).
Both melatonin and agomelatine significantly increased the

Table 1. Primary sequences of genetic studies.

GENE pRIMARY SEqUENCE

Beta2 microglobulin (F) 5’ Tga CTT TgT CaC agC CCa
aga Ta 3’
(r) 5’ aaT CCa aaT gCg gCa TCT TC 3’

BaCT (F) 5’ agC CaT gTa CgT agC CaT CCa 3’
(r) 5’ TCT CCg gag TCC aTC aCa aTg3’

CreB (F) 5’ agC Tgg CCT gTC CCa CTg CT 3’
(r) 5’ aCC aTT CTg aaC aCa aag Cag
CCa3’

BDnF (F) 5’ gCC Caa Cga aga aaa CCa Taa 3’
(r) 5’ gga ggC TCC aaa ggC aCT T 3’

PA test

Drugs

F
ir

s
t

d
a
y
l

a
te

n
c
y

PA test

Drugs

R
e

te
n

ti
o

n
l

a
te

n
c

Figure 1. effects of melatonin (10 mg/kg) or agomelatine (10 mg/kg) on (A) first day latency and (B) retention latency in the passive avoidance test in mice.
Notes: The data are expressed as the mean ± seM values. *P , 0.05 vs. non-stressed control group. #P , 0.05 vs. stressed control group.

http://www.la-press.com

Gumuslu et al

16 Drug TargeT InsIghTs 2014:8

mEPM test

Drugs

T
L

-1
nC

mEPM test

Drugs

T
L

-2

Figure 2. effects of melatonin (10 mg/kg) or agomelatine (10 mg/kg) on (A) transfer latency on the first day (B) transfer latency on the second day.
Notes: The data are expressed as mean ± seM values. *P , 0.05 vs. non-stressed control group. #P , 0.05 vs. stressed control group.

NOR test

Drugs

R
a

ti
o

i
n

d
e

Figure 3. effect of melatonin (10 mg/kg) or agomelatine (10 mg/kg) on the rI in the novel object recognition test.
Notes: The data are expressed as mean ± seM values. *P , 0.001 vs. non-stressed control group. #P , 0.001 vs. stressed control group.

RI compared to stress-exposed control mice (P , 0.001)
(Fig. 3).

effects of drugs on learning and memory in the MwM
test. There was a significant difference in escape latency in all
sessions during the evaluation of drug groups [F(5,41) = 8.22,

P , 0.001; F(5,41) = 10.77, P , 0.001; F(5,41) = 8.93,
P , 0.001; F(5,41) = 12.40, P , 0.001; F(5,41) = 10.32,
P , 0.001, respectively; Figure 4a]. Stress significantly
increased the escape latency during all sessions (P , 0.001)
in control animals, whereas both melatonin and agomelatine

http://www.la-press.com

Agomelatin improves memory in stress exposed mice

17Drug TargeT InsIghTs 2014:8

MWM test

0
20
40
60
80

100
120
140
160

Sessions

MWM test

15
20

Drugs

MWM test

0
5

10
15
20
25
30
35
40

Drugs

MWM test

Drugs

E
s

c
a

p
e

l
a

te
n

c
y

(
s

)
T

im
e
s

p
e

n
t

in
t

a
rg

e
t

q
u

a
d

ra
n

t
in

p
ro

b
e

t
ri

a
l

(s
)

M
e
a
n

d
is

ta
n

c
e
t

o
p

la
tf

o
rm

i
n

p
ro

b
e
t

ri
a
l

(c
m

)
S

p
e
e
d

(
c
m

/s
)

Fam ses 1st ses 2nd ses 3rd ses 4th ses

Figure 4. effects of melatonin (10 mg/kg) or agomelatine (10 mg/kg) on (A) escape latency, (B) the time spent in escape platform quadrant, (C) mean
distance to platform, and (D) swim speed in the probe trial (60 seconds) of the MWM test. The data are expressed as the mean ± seM values. *P , 0.001
vs. non-stressed control group. #P , 0.05, ##P , 0.01, ###P , 0.001 vs. stressed control group.

significantly shortened the escape latency in the familiariza-
tion session (P , 0.05 and P , 0.01; respectively) and in the
other sessions (P , 0.001) in stressed animals (Fig. 4A).

A significant difference was observed among all drug
groups in the time spent in the target quadrant in probe
trial of MWM test [F(5,41) = 6.27; P = 0.0003; Figure 4B].
Stressed control animals significantly decreased the time

spent in the escape platform quadrant (P , 0.001) compared
to non-stressed animals, and both melatonin (P , 0.05) and
agomelatine reversed this effect; agomelatine had a higher
impact than melatonin (P , 0.01; Figure 4B).

The mean distance traveled by the mice to the platform
in the probe trial of the MWM test was significantly dif-
ferent between the drug groups [F(5,41) = 3.98; P = 0.005;

http://www.la-press.com

Gumuslu et al

18 Drug TargeT InsIghTs 2014:8

Figure 4C]. Stress significantly increased the mean distance
traveled to the platform (P , 0.01) compared to the non-
stressed control group. Melatonin (P , 0.05) and agomelatine
significantly reversed this effect; agomelatine had a higher
impact (P , 0.01; Figure 4C).

Each treatment group did not significantly differ in
swimming speed [F(5,41) = 1.32; P = 0.27; Figure 4D] in the
probe trial of the MWM test.

effects of drugs on creb and bdNF gene expression.
In evaluating plasticity-related genes, we measured mRNA
levels in the hippocampus of mice subjected to UCMS/drug
treatment using quantitative RT-PCR. Decreased BDNF and
CREB expression might indicate both stress and cognitive
impairment. To determine whether downregulation of gene
expression can be prevented by antidepressant treatment,
melatonin or agomelatine was administered for 35 days to
mice subjected to UCMS.

Our results demonstrated that mRNA levels of the neu-
rotrophin family member BDNF, which has been studied in
relation to the stress response, were reduced in the hippocam-
pus of the mice subjected to UCMS. We also measured hip-
pocampal mRNA expression levels for CREB, a transcription
factor that regulates BDNF in response to drug treatment.
UCMS caused a reduction in CREB mRNA levels in stressed
animals. Melatonin or agomelatine treatment significantly
reversed UCMS-induced downregulation of CREB and
BDNF gene expression. Our results are in agreement with
data from similar studies.34,35 The gene expression observed in
each group is shown in Table 2.

discussion
Stress is a known risk factor in the development of many
neuropsychiatric disorders, including depression.2 More-
over, because stress-induced memory impairment is com-
monly reported in stress-related psychopathologies,3 there is
therapeutic relevance of the use of antidepressant treatments
to prevent stress-induced cognitive deficits.1

Both chronic mild stress and learned helplessness sig-
nificantly diminish the cognitive performance of mice in the

MWM test, and animals treated with antidepressants exhibit
significantly enhanced cognitive performance.35 The chronic
administration of agomelatine produced antidepressant-like
effects in the chronic mild stress model of depression36 and
improved learned helplessness in a model of depression-induced
avoidance learning deficits. In our study, stress-induced a sig-
nificant deterioration of memory in the PA, mEPM, NOR,
and MWM tests, and agomelatine ameliorated these effects
in NOR and MWM tests while it had a partial effect in the
PA and mEPM tests.

Agomelatine, a novel antidepressant with established
clinical efficacy,6 is an agonist of the melatonin MT1 and
MT2 receptors

37 and has a potent antagonistic activity on
serotonergic 5-HT2C receptors.

7 The affinity of agomelatine
for melatonin receptors is comparable with that of mela-
tonin.38 Melatonin produced in the pineal gland during peri-
ods of darkness plays a key role in the regulation of circadian
rhythms.8 It has a short half-life and is extensively metabo-
lized, leading to poor bioavailability. Moreover, the antide-
pressant-like activity of agomelatine in the rat CMS model
of depression is independent of the time of drug administra-
tion, while melatonin has no antidepressant-like activity after
administration in the morning.9 The search for metabolically
stable analogs with new and innovative properties resulted in
the discovery of agomelatine.39

Agomelatine-induced molecular changes may play a
role in its antidepressant and pro-cognitive effects and may
be attributed to synergy between the 5-HT2C antagonist and
melatonergic agonist properties of the drug.6 Agomelatine
increased the release of noradrenaline and dopamine in accor-
dance with its 5-HT2C antagonist properties.

7 These may in
turn be associated with the functional output of β-adrenergic
D1 and D2 receptors and activation or inhibition of the cAMP-
PKA pathway, which is postulated to modulate microtubule
dynamics40 and synaptic plasticity.41 Numerous findings
indicate that reduced function of 5-HT2C receptors may be
involved in the mechanism by which antidepressants allevi-
ate depression.42 The 5-HT2C receptor is involved in circadian
rhythm resynchronization,43 and 5-HT2C receptor antagonists
prevent the inhibitory effects of light on melatonin synthesis.

In mammals, specific M1 and M2 receptors are located
mainly in suprachiasmatic nuclei in the central nervous sys-
tem and in some peripheral sites. Moreover, the MT1 and
MT2 agonistic properties of agomelatine might also play a
role because both receptors modulate several signaling path-
ways such as PKA and protein kinase C (PKC), which have
been implicated in synaptic plasticity regulation.40 Circadian
rhythms are disturbed in depressed patients,44 and the UCMS
procedure causes a generalized disorganization of circadian
rhythms, which is suggested to play an important role in the
pathophysiology of depression, among other biochemical,
physiological, and behavioral impairments.45 Agomelatine can
resynchronize experimentally disturbed circadian rhythms,36
an effect that is independent of the time of administration.36

Table 2. gene expression levels in uCMs- exposed mice and effects
of drugs on gene expressions in uCMs- exposed mice. Melatonin
(10 mg/kg) or agomelatine (10 mg/kg) was given intraperitoneally
for 35 days to mice subjected to unpredictable chronic mild stress
(uCMs) (n = 7/each group). all of the treatments begun after 2 weeks
of stress regimen and were administered during 5 weeks.

GRoUpS CREB BDNF

uCMs + vehicle 2,408 ↓ 1,208 ↓

uCMs + melatonin 6,409 ↑ 1,164 ↑

uCMs + agomelatine 2,088 ↑ 1,060 ↑

Notes: ↓, Decrease in expression. ↑, Increase in expression.
Abbreviations: CreB, cyclic adenosine monophosphate (caMP) response
element binding protein; BDnF, brain-derived neurotrophic factor.

http://www.la-press.com

Agomelatin improves memory in stress exposed mice

19Drug TargeT InsIghTs 2014:8

Basic and clinical studies provide evidence for the neu-
rotrophic hypothesis of depression and antidepressant activ-
ity.20 The neurotrophin family member BDNF is involved in
neuronal differentiation and survival as well as the synaptic
plasticity associated with learning and memory.46 The cAMP
signaling pathway (in particular, the downstream effector
CREB) has also been shown to play an important role in neu-
ronal and synaptic plasticity.47 Therefore, decreased expression
of BDNF or CREB could contribute to the atrophy of the
hippocampus in response to stress, and the upregulation of
BDNF and CREB could contribute to the action of antide-
pressant therapy.48 It has been postulated that chronic stress
caused a downregulation of hippocampal BDNF or CREB
levels and that this reduction could be upregulated through
antidepressant therapy.48

Moreover, Molteni et al.49 demonstrated that acute
agomelatine treatment modulates the expression of BDNF
through a functional interaction between melatonergic MT1/
MT2 and serotonergic 5-HT2C receptors, supporting the con-
cept that intracellular events can be regulated via the synergis-
tic activity of different neuromodulatory systems. Our results
are consistent with this hypothesis and demonstrate that ago-
melatine treatment improves UCMS-induced memory deteri-
oration and upregulates hippocampal CREB and BDNF gene
expression levels.

BDNF belongs to the neurotrophic factor family and
plays a crucial role in the development, regeneration, sur-
vival, and maintenance of neuronal function in the cen-
tral nervous system.16 These neurotrophins are abundantly
expressed in the hippocampus,50 where they are important
modulators of spatial learning51 and activity-dependent
synaptic plasticity, such as long-term potentiation.52 More-
over, BDNF plays an important role in the formation, reten-
tion, and recall of spatial memory, and decreases in BDNF
expression result in the impairment of spatial learning and
memory.53 The results of clinical studies have shown that
depressive patients exhibit diminished plasma BDNF lev-
els and that antidepressant treatment increases plasma
BDNF levels.54 Interestingly, the expression of BDNF is
also influenced by light and dark cycles in rats55 as well as
in humans.19 It is speculated that acute agomelatine treat-
ment can upregulate the expression of BDNF mRNA levels
in the prefrontal cortex through the functional interaction
between melatonergic MT1/MT2 and serotonergic 5-HT2C
receptors,56 thus preventing the circadian downregulation
of the neurotrophin.

Soumier et al.57 postulated that agomelatine produced
major transcriptional changes in the hippocampus, where sig-
nificant upregulation of BDNF was observed. Moreover, the
levels of BDNF protein were elevated by agomelatine in both
the hippocampus and the prefrontal cortex.58 These findings
support the hypothesis that alteration of hippocampal BDNF
expression is correlated with antidepressant response in the
hippocampus.16

Chronic agomelatine treatment decreased BDNF expres-
sion in the amygdale and this effect might be related to a nega-
tive feedback mechanism in response to the high magnitude
of neuronal remodeling or to a distinct and yet unknown
neurochemical event.57 Chronic agomelatine has neurogenic
effects in the hippocampus in rats57 and a reversed depression-
induced decrease in neurogenesis.59 Agomelatine exposure
increases neurite outgrowth of granule cells in hippocampal
primary cell culture and accelerates the maturation of newly
formed granule cells in rats.57 The results of our study revealed
that chronic agomelatine increased BDNF expression in the
hippocampus, and our findings are consistent with those of
recent studies.57,60 These findings provide new information
regarding the molecular mechanisms that contribute to the
chronic effects of the new antidepressant agomelatine on brain
function. The ability of agomelatine to modulate the expres-
sion of these neuroplastic molecules, which follow a circadian
rhythm, may contribute to its antidepressant action.

BDNF is involved in the etiology of mood disorders,
and it is thought to participate in the structural remodeling
associated with antidepressant therapy.58 As neurotrophin
expression is activity-dependent17 and may be regulated by the
light and dark cycle in rats18 and humans,19 it may be inferred
that its transcription can be modulated by acute agomelatine
administration and may represent a downstream target of its
synaptic effects. On this basis, we investigated BDNF mRNA
levels in the rat hippocampus.

The cAMP-CREB signal transduction cascade is known
to be responsible for the sustained alterations that occur in
cellular and behavioral models of learning and memory.61
Chronic but not acute antidepressant administration, includ-
ing norepinephrine and selective serotonin reuptake inhibitors,
increases CREB expression, phosphorylation, and function in
limbic brain structures including the hippocampus and cere-
bral cortex.62

Agomelatine has a favorable clinical safety and tolerabil-
ity profile of antidepressant properties with few side effects
and no withdrawal syndrome.11 In contrast to SSRIs,63 it has
few sexual side effects and there are no reports of impotence,
ejaculation difficulties, or decreased libido. The novel mode of
action of agomelatine (selective binding profile, not inducing
serotonin release or increasing extracellular serotonin levels
and no effect on 5-HT1 A receptors)7 might be responsible for
its favorable safety profile.

Overall, the present study suggests that chronic admin-
istration of the novel antidepressant agomelatine, with its
distinct mechanism of action based on synergy between the
melatonergic and 5-HT2C pathways and the advantages of a
favorable clinical safety/tolerability profile, improves memory
deterioration and upregulates CREB and BDNF gene expres-
sion levels in UCMS-exposed mice. Thus, agomelatine appears
to play an important role in blocking stress-induced hippocam-
pal memory deterioration and activates the molecular mecha-
nisms of memory storage in response to a learning experience.

http://www.la-press.com

Gumuslu et al

20 Drug TargeT InsIghTs 2014:8

Author contributions
EG, OM, DS, and GU conceived and designed the experi-
ments. EG, OM, DS, IKC, and NC analyzed the data. EG,
OM, GU, and IKC wrote the first draft of the manuscript.
NC, FA, HS, and FE contributed to the writing of the manu-
script. EG, OM, DS, GU, IKC, NC, FA, HS, and FE agree
with manuscript results and conclusions. EG, OM, GU, FA,
HS, and FE jointly developed the structure and arguments
for the paper. FA, HS, and FE made critical revisions and
approved final version. All authors reviewed and approved the
final manuscript.

DISCLoSURES AND EThICS
As a requirement of publication the authors have provided signed confirmation of their
compliance with ethical and legal obligations including but not limited to compliance
with ICMJe authorship and competing interests guidelines, that the article is neither
under consideration for publication nor published elsewhere, of their compliance with
legal and ethical guidelines concerning human and animal research participants (if
applicable), and that permission has been obtained for reproduction of any copy-
righted material. This article was subject to blind, independent, expert peer review.
The reviewers reported no competing interests.

reFereNces
1. Yau JL, Noble J, Hibberd C, et al. Chronic treatment with the antidepressant

amitriptyline prevents impairments in water maze learning in aging rats. J Neu-
rosci. 2002;22:1436–42.

2. Bremner JD, Vermetten E. Stress and development: behavioral and biological
consequences. Dev Psychopathol. 2001;13:473–89.

3. de Quervain DJ, Roozendaal B, Nitsch RM, McGaugh JL, Hock C. Acute
cortisone administration impairs retrieval of long-term declarative memory in
humans. Nat Neurosci. 2000;3:313–4.

4. Hirschfeld RM. History and evolution of the monoamine hypothesis of depres-
sion. J Clin Psychiatry. 2000;61(6):4–6.

5. Castren E. Is mood chemistry? Nat Rev Neurosci. 2005;6:241–6.
6. Audinot V, Mailliet F, Lahaye-Brasseur C, et al. New selective ligands of human

cloned melatonin MT1 and MT2 receptors. Naunyn Schmiedebergs Arch Pharma-
col. 2003;367:553–61.

7. Millan MJ, Gobert A, Lejeune F, et al. The novel melatonin agonist agomela-
tine (S20098) is an antagonist at 5-hydroxytryptamine2C receptors, blockade
of which enhances the activity of frontocortical dopaminergic and adrenergic
pathways. J Pharmacol Exp Ther. 2003;306:954–64.

8. Pitrosky B, Kirsch R, Malan A, Mocaer E, Pevet P. Organization of rat circadian
rhythms during daily infusion of melatonin or S20098, a melatonin agonist. Am J
Phys. 1999;277:812–28.

9. Bertaina-Anglade V, la Rochelle CD, Boyer PA, Mocaer E. Antidepressant-like
effects of agomelatine (S 20098) in the learned helplessness model. Behav Phar-
macol. 2006;17:703–13.

10. Barden N, Shink E, Labbe M, Vacher R, Rochford J, Mocaer E. Antidepressant
action of agomelatine (S 20098) in a transgenic mouse model. Prog Neuropsychop-
harmacol Biol Psychiatry. 2005;29:908–16.

11. Kennedy SH, Emsley R. Placebo-controlled trial of agomelatine in the treatment
of major depressive disorder. Eur Neuropsychopharm. 2006;16:93–100.

12. Quera Salva MA, Vanier B, Laredo J, et al. Major depressive disorder, sleep
EEG and agomelatine: an open label study. Int J Neuropsychopharmacol.
2007;10:691–6.

13. McClung CA. Circadian genes, rhythms and the biology of mood disorders.
Pharmacol Ther. 2007;114:222–32.

14. Willner P. Validity, reliability and utility of the chronic mild stress model of depres-
sion: a 10 years review and evaluation. Psychopharmacology. 1997;134:319–29.

15. Gumuslu E, Mutlu O, Sunnetci D, et al. The effects of tianeptine, olanzapine
and fluoxetine on the cognitive behaviors of unpredictable chronic mild stress-
exposed mice. Drug Res. 2013;63:532–9.

16. Tardito D, Perez J, Tiraboschi E, Musazzi L, Racagni G, Popoli M. Signal-
ing pathways regulating gene expression, neuroplasticity, and neurotrophic
mechanisms in the action of antidepressants: a critical overview. Pharmacol Rev.
2006;58:115–34.

17. Bramham CR, Messaoudi E. BDNF function in adult synaptic plasticity: the
synaptic consolidation hypothesis. Prog Neurobiol. 2005;76:99–125.

18. Cirelli C, Tononi G. Differential expression of plasticity related genes in wak-
ing and sleep and their regulation by the noradrenergic system. J Neurosci.
2000;20:9187–94.

19. Begliuomini S, Lenzi E, Ninni F, et al. Plasma brain-derived neurotrophic factor
daily variations in men: correlation with cortisol circadian rhythm. J Endocrinol.
2008;197:429–35.

20. Calabrese F, Molteni R, Racagni G, Riva MA. Neuronal plasticity: a link between
stress and mood disorders. Psychoneuroendocrinology. 2009;34(1):208–16.

21. Porsolt RD, Le Pichon M, Jalfre M. Depression: a new animal model sensitive to
antidepressant treatments. Nature. 1977;266:730–2.

22. Ennaceur A, Delacour J. A new one-trial test for neurobiological studies of mem-
ory in rats. 1 Behavioral data. Behav Brain Res. 1988;31:47–59.

23. Barco A, Pittenger C, Kandel ER. CREB, memory enhancement and the treat-
ment of memory disorders: promises, pitfalls and prospects. Expert Opin Ther
Targets. 2013;7(1):101–14.

24. Muscat R, Willner P. Suppression of sucrose drinking by chronic mild unpredict-
able stress: a methodological analysis. Neurosci Biobehav Rev. 1992;16:507–17.

25. Arbe MD, Einstein R, Lavidis NA. Stressful animal housing conditions and
their potential effect on sympathetic neurotransmission in mice. Am J Physiol
Regul Integr Comp Physiol. 2002;282:1422–8.

26. Willner P, Muscat R, Papp M. Chronic mild stress-induced anhedonia: a realis-
tic animal model of depression. Neurosci Biobehav Rev. 1992;16:525–34.

27. Ducottet C, Belzung C. Behaviour in the elevated plus-maze predicts coping
after subchronic mild stress in mice. Physiol Behav. 2004;81:417–26.

28. Yalcin I, Aksu F, Belzung C. Effects of desipramine and tramadol in a chronic
mild stress model in mice are altered by yohimbine but not by pindolol. Eur J
Pharmacol. 2005;514:165–74.

29. Ogren SO, Johansson C, Magnusson O. Forebrain serotonergic involvement in
avoidance learning, Neurosci Lett. 1985;58:305–9.

30. Reddy DS, Kulkarni SK. Possible role of nitric oxide in the nootropic and anti-
amnesic effects of neurosteroids on aging- and dizocilpine-induced learning
impairment. Brain Res. 1998;799:215–29.

31. Hlinak Z, Krejci I. Concurrent administration of subeffective doses of scopol-
amine and MK-801 produces a short-term amnesia for the elevated plus-maze in
mice. Behav Brain Res. 1998;91:83–9.

32. Savlı H, Aalto Y, Nagy B, Knuutila S, Pakkala S. Gene expression analysis of
1,25(OH)2D3 dependent differentiation of HL-60 cells—a cDNA array study.
Br J Haematol. 2002;118(4):1065–72.

33. Savlı H, Karadenizli A, Kolaylı F, Gundes S, Ozbek U, Vahaboglu H. Expres-
sion stability of six housekeeping genes: A proposal for resistance gene quanti-
fication studies of Pseudomonas aeruginosa by real-time quantitative RT-PCR.
J Med Microbiol. 2003;52:403–8.

34. Alfonso J, Frasch AC, Flugge G. Chronic stress, depression and antidepressants:
effects on gene transcription in the hippocampus. Rev Neurosci. 2005;16:43–56.

35. Song L, Che W, Min-Wei W, Murakami Y, Matsumoto K. Impairment of the
spatial learning and memory induced by learned helplessness and chronic mild
stress. Pharmacol Biochem Behav. 2006;83:186–93.

36. Papp M, Gruca P, Boyer PA, Mocaer E. Effect of agomelatine in the chronic mild
stress model of depression in the rat. Neuropsychopharmacol. 2003;28:694–703.

37. Ying SW, Rusak B, Delagrange P, Mocaer E, Renard P, Guardiola- Lemaitre
B. Melatonin analogues as agonists and antagonists in the circadian system and
other brain areas. Eur J Pharmacol. 1996;296:33–42.

38. Bonnefond C, Martinet L, Lesieur D, Adam G, Guardiola-Lemaître B. Charac-
terization of S-20098, a new melatonin analogue. In: Touitou Y, Arendt J, Pevet
P, eds. Melatonin and the Pineal Gland. Amsterdam: Elsevier Science Publishers;
1993:123–6.

39. Guardiola-Lemaitre B. Melatoninergic receptor agonists and antago-
nists: pharmacological aspects and therapeutic perspective. Ann Pharm Fr.
2005;63:385–400.

40. Bianchi M, Hagan JJ, Heidbreder CA. Neuronal plasticity, stress and depres-
sion: involvement of the cytoskeletal microtubular system? Curr Drug Targets
CNS Neurol Disord. 2005;4:597–611.

41. Warner-Schmidt JL, Duman RS. Hippocampal neurogenesis: opposing effects
of stress and antidepressant treatment. Hippocampus. 2006;16:239–49.

42. Sanchez C, Hyttel J. Comparison of the effects of antidepressants and their
metabolites on reuptake of biogenic amines and on receptor binding. Cell Mol
Neurobiol. 1999;19:467–89.

43. Kennaway DJ, Moyer RW. Serotonin 5-HT2C agonists mimic the effect of light
pulses on circadian rhythms. Brain Res. 1998;806:257–70.

44. Wirz-Justice A. Biological rhythms in mood disorders. In: Bloom FE, Kupfer DJ,
eds. Psychopharmacology: the Fourth Generation of Progress. New York: Raven Pres;
1995:999–1017.

45. Wehr TA, Wirz-Justice A. Circadian rhythm mechanisms in affective illness
and in antidepressant drug action. Pharmacopsychiatry.1982;15:31–9.

46. Branchi I, Francia N, Alleva E. Epigenetic control of neurobehavioural plastic-
ity: The role of neurotrophins. Behav Pharmacol. 2004;15:353–62.

47. Schulman H. Protein phosphorylation in neuronal plasticity and gene expres-
sion. Curr Opin Neurobiol. 1995;5:375–81.

http://www.la-press.com

Agomelatin improves memory in stress exposed mice

21Drug TargeT InsIghTs 2014:8

48. Duman RS, Malberg J, Nakagawa S, D’Sa C. Neuronal plasticity and survival in
mood disorders. Biol Psychiatry. 2000;48:732–9.

49. Molteni R, Calabrese F, Cattaneo A, et al. Acute stress responsiveness of the
neurotrophin BDNF in the rat hippocampus is modulated by chronic treatment
with the antidepressant duloxetine. Neuropsychopharmacology. 2009;34:1523–32.

50. Ernfors P, Ibáñez CF, Ebendal T, Olson L, Persson H. Molecular cloning and
neurotrophic activities of a protein with structural similarities to nerve growth
factor: developmental and topographical expression in the brain. Proc Natl Acad
Sci U S A. 1990;87:5454–8.

51. Scaccianoce S, Del Bianco P, Caricasole A, Nicoletti F, Catalani A. Relationship
between learning, stress and hippocampal brain-derived neurotrophic factor.
Neuroscience. 2003;121:825–8.

52. Lu B. BDNF and activity-dependent synaptic modulation. Learn Mem.
2003;10:86–98.

53. Mu JS, Li WP, Yao ZB, Zhou XF. Deprivation of endogenous brain-derived
neurotrophic factor results in impairment of spatial learning and memory in
adult rats. Brain Res. 1999;835:259–65.

54. Aydemir C, Yalcin ES, Aksaray S, et al. Brain-derived neurotrophic factor
(BDNF) changes in the serum of depressed women. Prog Neuropsychopharmacol
Biol Psychiatry. 2006;30:1256–60.

55. Bova R, Micheli MR, Qualadrucci P, Zucconi GG. BDNF and trkB mRNAs
oscillate in rat brain during the light-dark cycle. Brain Res Mol Brain Res.
1998;57:321–4.

56. Molteni R, Calabrese F, Pisoni S, et al. Synergistic mechanisms in the modulation
of the neurotrophin BDNF in the rat prefrontal cortex following acute agomela-
tine administration. World J Biol Psychiatry. 2010;11:148–53.

57. Soumier A, Banasr M, Lortet S, et al. Mechanisms contributing to the phase-
dependent regulation of neurogenesis by the novel antidepressant, agomelatine,
in the adult rat hippocampus. Neuropsychopharmacol. 2009;34:2390–403.

58. Calabrese F, Molteni R, Maj PF, et al. Chronic duloxetine treatment induces spe-
cific changes in the expression of BDNF transcripts and in the subcellular local-
ization of the neurotrophin protein. Neuropsychopharmacology. 2007;32:2351–9.

59. Morley-Fletcher S, Mairesse J, Soumier A, et al. Chronic agomelatine treatment
corrects behavioral, cellular, and biochemical abnormalities induced by prenatal
stress in rats. Psychopharmacology (Berl). 2011;217(3):301–13.

60. Ladurelle N, Gabriel C, Viggiano A, Mocaër E, Baulieu EE, Bianchi M. Ago-
melatine (S20098) modulates the expression of cytoskeletal microtubular pro-
teins, synaptic markers and BDNF in the rat hippocampus, amygdala and PFC.
Psychopharmacology. 2012;221:493–509.

61. Barad M, Bourtchouladze R, Winder DG, Golan H, Kandel E. Rolipram,
a type IV specific phosphodiesterase inhibitor, facilitates the establishment of
long-lasting long-term potentiation and improves memory. Proc Natl Acad Sci
USA. 1998;95:15020–5.

62. Thome J, Sakai N, Shin K, et al. cAMP response element-mediated gene
transcription is upregulated by chronic antidepressant treatment. J Neurosci.
2000;20:4030–6.

63. Ferguson JM, Shrisvastava RK, Stahl SM, et al. Re-emergence of sexual dys-
function in patients with major depressive disorder: doubleblind comparison of
nefadozone and sertraline. J Clin Psychiatry. 2001;62:24–9.

http://www.la-press.com