Thymol reverses depression‐like behaviour and upregulates hippocampal BDNF levels in chronic corticosterone‐induced depression model in female mice

Abstract

Objectives:
Based on this, the central therapeutic effects of thymol were verified in the neurotrophic pathway.

Methods:
Female swiss mice were divided into four groups: control, corticosterone (Cort), thymol (Cort + thymol) and fluvoxamine (Cort + Flu). The administration of corticosterone was used to induce depressive symptoms for 23 days. After the treatment, the animals were exposed the behavioural tests, such as forced swimming test, tail suspension test, sucrose preference test, light/dark test, social interaction test, Y-maze test, plus-maze test and hole-board test. The hippocampus was also removed, and BDNF was measured by ELISA and Western blot.

Key findings:
As a result, thymol and fluvoxamine were able to reverse the depressive symptoms, as well as to improve the anxious frame. The anhedonic and short-term memory was restored with the treatment. In the neurochemical tests, both thymol and fluvoxamine restored BDNF levels, improving the depressive condition.

Conclusions:
This work opens up new investigations aiming at the use of this molecule as a therapeutic alternative for treating depression disorders.

Full text

Thymol reverses depression-like behaviour and upregulates
hippocampal BDNF levels in chronic corticosterone-induced
depression model in female mice
Victor Celso Cavalcanti Capibaribe, Auriana Serra Vasconcelos Mallmann, Iardja St
efane Lopes,
Iris Cristina Maia Oliveira, Natalia Ferreira de Oliveira , Raquell de Castro Chaves,
Mariana Lima Fernandes, Mariana Albuquerque de Araujo, Daniel Moreira Alves da Silva,
Jos
e Tiago Valentim, Adriano Jos
e Maia Chaves Filho, Danielle Silveira Mac^
edo , Silv^
ania Maria
Mendes de Vasconcelos , Alyne Mara Rodrigues de Carvalho and
Francisca Cl
ea Florenc
ßo de Sousa
Drug Research and Development Center—NPDM, Federal University of Cear
a, Fortaleza, Brazil
Keywords
BDNF; corticosterone; depression; thymol
Correspondence
Dr. Francisca Cl
ea Florenc
ßo de Sousa, Drug
Research and Development Center—NPDM,
Federal University of Cear
a, Rua Coronel
Nunes de Melo, 1000 Rodolfo Te
ofilo,
Fortaleza –CE, 60430-275, Brazil.
E-mail: cleaflorenco@yahoo.com.br
Received July 3, 2019
Accepted August 22, 2019
doi: 10.1111/jphp.13162
Abstract
Objectives Based on this, the central therapeutic effects of thymol were verified
in the neurotrophic pathway.
Methods Female swiss mice were divided into four groups: control, corticos-
terone (Cort), thymol (Cort +thymol) and fluvoxamine (Cort +Flu). The
administration of corticosterone was used to induce depressive symptoms for 23
days. After the treatment, the animals were exposed the behavioural tests, such as
forced swimming test, tail suspension test, sucrose preference test, light/dark test,
social interaction test, Y-maze test, plus-maze test and hole-board test. The hip-
pocampus was also removed, and BDNF was measured by ELISA and Western
blot.
Key findings As a result, thymol and fluvoxamine were able to reverse the
depressive symptoms, as well as to improve the anxious frame. The anhedonic
and short-term memory was restored with the treatment. In the neurochemical
tests, both thymol and fluvoxamine restored BDNF levels, improving the depres-
sive condition.
Conclusions This work opens up new investigations aiming at the use of this
molecule as a therapeutic alternative for treating depression disorders.
Introduction
Depression affects more than 300 million people worldwide
in all age groups and has a higher frequency among other
psychiatric disorders. It is one of the leading causes of dis-
ability around the world, which can induce suffering,
reduce productivity at work and school, and negatively
influence family relationships. Approximately 800 000 peo-
ple die of suicide each year due to the depressive disorders,
and it causes considerable damage to the economy and
health in terms of the functional impairment and the costs
related to the treatment.
[1–3]
When associated with other
pathologies such as cardiovascular, endocrine and pul-
monary diseases, it increases mortality levels, besides the
already high risk of suicide.
[4,5]
It is more prevalent in
female sex, being two to three times more frequent in
women than in men.
[3,6]
The main focus of the development of pharmacological
therapy for depression, for decades, has been based on the
theory of monoaminergic dysregulation.
[1,7]
Another
hypothesis recently studied is the deficiency in the neu-
rotrophic pathway, where the low levels of several neu-
rotrophins, especially brain-derived neurotrophic factor
(BDNF), have been associated with depressive disorder.
[8]
The interest in evaluating the therapeutic effects of natu-
ral products, mainly plant-derived ones, in mental disor-
ders such as depression has been a topic of research in the
last decade.
[9–11]
Within the large variety of plants and nat-
ural products, St. John’s wort is a promising example for
the depressive disorder.
[11–14]
©2019 Royal Pharmaceutical Society, Journal of Pharmacy and Pharmacology,71 (2019), pp. 1774–17831774
Research Paper
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Thymol, a natural monoterpenoid phenol, is a com-
mon compound found in approximately 81 species of
plants, including Thymus species.
[15,16]
Several pharmaco-
logical activities of thymol have been described, such as
fungicide,
[17]
antibiotic
[16]
and anti-inflammatory.
[18]
More recently, the antidepressant-like effect of thymol
administration was also demonstrated and related to its
action in immune-mediated pathways.
[19]
In this scenar-
io, our study aims to investigate the antidepressant-like
effects of thymol in the neuroendocrine model of
depression induced by chronic administration of corti-
costerone in female mice, focusing on its effects on hip-
pocampal BDNF levels.
Materials and Methods
Animals
Adult female Swiss mice (21–25 g, age: 8–10 weeks) were
chosen based on previous studies
[20,21]
and randomly dis-
tributed in four groups of 8–12 animals. All drugs adminis-
trations were performed between 10–12 a.m. The animals
were housed at a temperature of 22–25°C, with no food or
water deprivation, in a 12-h light/dark cycle. All procedures
in this study were performed and approved according to
the UFC’s ethics committee (protocol-61/2016).
Drugs
Thymol (Sigma
â
, St Louis, MO, USA) was dissolved in dis-
tilled water containing 0.1% dimethyl sulfoxide (DMSO)
and 0.1% Tween
â
80 at a final dose of 50 mg/kg, by oral
gavage (po).
[22]
Corticosterone (Sigma
â
) was dissolved in
saline containing 0.1% DMSO, and 0.1% Tween
â
80 was
administered at a dose of 20 mg/kg subcutaneously (sc).
Fluvoxamine (Abott
â
, New Jersey, USA), in a 50 mg/kg
dose was diluted in distilled water.
Experimental procedure
The control group (vehicle) was treated with saline (sc) for
23 consecutive days, and during the last 9 days, they also
received distilled water (po) containing 0.1% DMSO and
0.1% of Tween
â
80. The corticosterone group (Cort)
received corticosterone (sc) for 23 consecutive days, and
during the last 9 days, the mice also received distilled water
(po) containing 0.1% DMSO and 0.1% of Tween
â
80. The
thymol group (Cort +thymol) and fluvoxamine group
(Cort +Flu) were treated with corticosterone (sc) for 23
consecutive days and additionally received thymol or flu-
voxamine (po) during the last 9 days, respectively. At the
end of the treatment, after the last drug administration, the
animals were submitted to behavioural tests to evaluate
depressive, anxious and cognitive deficit behaviour, such as
open field test (OFT), tail suspension test (TST), forced
swimming test (FST), sucrose preference test (SPT), light/
dark test (LDT), hole-board test (HBT), plus-maze test
(PMT), social interaction test (SIT) and Y-maze test
(YMT). Animals were euthanized, and the hippocampus
was removed for BDNF analysis (Figure 1).
Behavioural tests
Open field test (OPT)
The test was performed in an acrylic apparatus with transpar-
ent walls and black bottom, and with markings on the bottom
dividing the total base area of the apparatus into nine squares.
Each mouse was placed in the middle square, and the number
of squares crossed was counted during a 5-min test.
[23]
Tail suspension test (TST)
Each mouse was suspended by the tail to a height approxi-
mately 58 cm from the surface and fixed by an adhesive
Figure 1 Experimental protocol. Cort: corticosterone; sc: subcutaneous; Flu: fluvoxamine; BDNF: brain-derived neurotrophic factor. [Colour figure
can be viewed at wileyonlinelibrary.com]
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Victor Celso Cavalcanti Capibaribe et al. Thymol reverses depression-like behavior
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tape placed approximately 1 cm from the tip of the tail.
The immobility time was recorded for 6 min.
[24]
Forced swimming test (FST)
The animals were placed in a cylinder with approximately
30 cm high, containing water, and the duration of immo-
bility in its entirety was recorded for a period of 5 min.
[25]
Sucrose preference test (SPT)
On the 21st and 22nd days, mice were habituated to two
bottles, each day for 18 h, of sucrose solution (2%, w/v) or
tap water, respectively. On the 23rd day, animals were
exposed to two bottles, one bottle contained 2% (w/v)
sucrose solution and the other one water. The consumption
preference was assessed and recorded.
[26]
Light/Dark test (LDT)
Each animal was placed in a test box divided into two areas,
one dark and one light. The two areas were connected by
an entrance measuring 7.5 cm wide and 7.5 cm high, and
the animal could move freely between them. The mouse
was placed in the light area, far away from the dark area,
and the time spent by each animal in the light area was
measured.
[27–29]
Hole-board test (HBT)
The apparatus used was a perforated board containing 16
holes spaced uniformly. The parameter used for analysis
was the number of times the animal inserted the head into
the holes (head-dips) during the 5-min period.
[30]
Plus-Maze test (PMT)
The apparatus consists of two open arms and two walled
arms, forming a raised cross, elevated 45 cm from the
ground. All arms are connected by a central platform. Each
animal was placed on the central platform with the head
facing one of the walled arms, and its behaviour was
observed for a period of 5 min. The number of entries and
the time spent in both open and closed arms were evalu-
ated; however, for the statistical analysis, the data were
related to the open arms.
[31]
Social interaction test (SIT)
Each animal was placed individually in an acrylic test box
divided into three equal space chambers. The test box had
an entrance between the chambers, with the animal having
free access to the three chambers. The animal being
analysed was placed in the middle chamber. An iron cage
containing a mouse was placed in the right chamber, called
the social chamber (SCham), and a similar cage, but with-
out any animal, was placed in the left chamber, called the
non-social chamber (NSCham). For 5 min, the mouse
placed in the central chamber was analysed and the time
spent in the three chambers was recorded.
[32,33]
Y-maze test (YMT)
Each mouse was placed in one of the identical arms, pro-
viding it the opportunity to explore and move freely
throughout the maze. The number of spontaneous alterna-
tions of each animal in the apparatus was recorded, alterna-
tion being defined as the entries into the three arms
consecutively. The percentage of alternations was calculated
by the ratio of correct alternations to number of possible
alternations multiplied by 100.
[34–37]
BDNF assessment
ELISA test
The animals were sacrificed, and the encephalic hippocam-
pal portion of the animals was collected. The BDNF levels
in the brain were assessed using the anti-BDNF sandwich
enzyme immunoassay ELISA kit (Chemicon
â
,S
~
ao Paulo,
Brazil) as per the manufacturer’s instructions.
Western blotting
Hippocampi were homogenized in RIPA lysis buffer with
protease inhibitor. Protein concentrations were determined
by the Lowry method according to the manufacturer’s pro-
tocol. SDS polyacrylamide gel electrophoresis (10%) was
performed using 50 lg of protein, transferred onto PVDF
membrane, blocked with BSA 5% and incubated with rab-
bit anti-BDNF IgG primary antibody (1:2000; Alomone
Labs
â
, Jerusalem, Israel) and horseradish peroxidase-con-
jugated goat anti-rabbit IgG secondary antibody (1:1000;
Thermo Scientific
â
, Waltham, MA, USA). Signal was
detected using the ECL system (Bio-Rad
â
, California, CA,
USA) according to the manufacturer’s instructions, and the
bands were captured with a CCD camera using the Chemi-
Doc system (Bio-Rad). Densitometry quantification of
bands was performed with ImageLab Bio-Rad (Bio-Rad)
software.
Statistical analyses
Statistical data analysis were performed using GraphPad
Prism 7.0a (GraphPad Software, San Diego, CA, USA) soft-
ware. Shapiro–Wilk normality was performed and chosen
©2019 Royal Pharmaceutical Society, Journal of Pharmacy and Pharmacology,71 (2019), pp. 1774–17831776
Thymol reverses depression-like behavior Victor Celso Cavalcanti Capibaribe et al.
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one-way ANOVA and Tukey (post hoc) or Kruskal–Wallis
and Dunn’s (post hoc), according to parametric or non-
parametric data, respectively. Results were expressed as the
mean standard error of the mean (SEM) and considered
significant at P<0.05.
Results
Behavioural tests
Open field test
In the OFT, the locomotor activity was measured by the
number of square crossings, and no significant change was
observed after chronic corticosterone (20 mg/kg), thymol
(50 mg/kg) or fluvoxamine (50 mg/kg) administration
(Figure 2).
Tail suspension test
In the tail suspension test (Figure 3), Cort-treated animals
showed an increase in immobility time compared to con-
trol group (P<0.0001) while treatment with the drug test
thymol significantly reversed the immobility time
(P<0.0001; F(3, 36) =47.32).
Forced swimming test
Figure 4 shows that corticosterone administration dur-
ing 23 days can increase significantly the immobility
time in the forced swimming test (P<0.0001). Treated
thymol group (50 mg/kg) had a shorter immobility time
(P<0.0001; F(3, 36) =58.18) compared to Cort
group.
Sucrose preference test
In the sucrose preference test, the Cort group showed an
considerable decrease in sweet solution consumption when
compared to the vehicle group (P=0.0075). The groups
treated with thymol (P=0.0075) and fluvoxamine
(P=0.0002) during 9 days significantly reversed the lack
of interest in sucrose solution [F(3, 36) =4.552]. The
reversion of the described anhedonic state is shown in
Figure 5.
Light/Dark test
In the light/dark test (Figure 6), results showed no signifi-
cant alteration compared to groups related to time spent in
the light chamber parameter (F(3, 36) =1.207).
Figure 2 Number of squares crossed in the open field test. Results
are expressed as the mean SEM (n= 10 per group). Statistical anal-
ysis was performed by Kruskal–Wallis, followed by Dunn’s post hoc
test. Significant values: *P<0.05 vs Cort group.
Figure 3 Immobility time analysis in the tail suspension test. Results
are expressed as the mean SEM (n= 12 per group). Statistical anal-
ysis was performed by one-way ANOVA, followed by Tukey’s post
hoc test. Significant values: *P<0.0001 vs control and **P<0.0001
vs Cort group.
Figure 4 Immobility time analysis of the forced swimming test. Sta-
tistical analysis was performed by one-way ANOVA, followed by
Tukey’s post hoc test. Results are expressed as the mean SEM
(n=9–10 per group). Significant values: *P<0.0001 vs control group
**P<0.0001 vs Cort group.
©2019 Royal Pharmaceutical Society, Journal of Pharmacy and Pharmacology,71 (2019), pp. 1774–1783 1777
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Hole-board test
In the hole-board test, the exploratory activity of the ani-
mals was evaluated and corticosterone treatment signifi-
cantly decreased the number of head-dips (P=0.0102)
compared to control group while thymol administration
was able to reverse this parameter (P<0.0001; F(3,
30) =11.78) (Figure 7).
Plus-Maze test
In the plus-maze test, the administration of corticosterone
for 23 days caused a significant reduction in the percentage
of entries in open arms (P=0.0019) and the thymol
(P=0.0003) and fluvoxamine (P=0.0045) administration
reversed this parameter. Cort group animals also decreased
the time spent in open arms (P=0.0002; F(3,
36) =8.763) compared to control group. Thymol
(P=0.0182)- and Flu (P=0.0010)-treated groups showed
an significant increasing in time spent in open arms when
compared to Cort group, as shown in Table 1.
Social interaction test
The group treated with corticosterone alone (Cort), when
compared to the control group, presented lower social
interaction (P=0.0025); F(3, 36) =10.62), as shown in
Figure 8. The group treated with corticosterone followed by
treatment with thymol (P<0.0001) showed greater social
interaction when compared to the Cort group treated.
Y-maze test
In the Y-maze test, used to evaluate work memory, chronic
treatment of corticosterone decreased the number of
Figure 5 Percentage of sucrose consumption in the sucrose prefer-
ence test. Results are expressed as the mean SEM (n= 9 per
group). Statistical analysis was performed by one-way ANOVA, fol-
lowed by Tukey’s post hoc test. Significant values: *P<0.01 vs con-
trol group and **P<0.01, ***P<0.001 vs Cort group.
Figure 6 Time spent in light the animals in the light/dark test.
Results are expressed as the mean SEM (n= 7 per group). Statisti-
cal analysis was performed by one-way ANOVA, followed by Tukey’s
post hoc test.
Figure 7 Number of the head-dips in the hole-board test. Results
are expressed as the mean SEM (n=8–10 per group). Statistical
analysis was performed by one-way ANOVA, followed by Tukey’s post
hoc test. Significant values: *P<0.05 vs control group. **P<0.0001
and Cort group.
Table 1 Percentage of the total entries and the total time spent in
open arms analysis of animals in the elevated plus-maze
Group
% of entries in opens
arms
% time spent in open
arms
Control 30.00 2.140 26.96 1.877
Cort 11.11 2.196*3.0.20 2.222**
Cort + Thymol 30.00 8.015***** 19.69 2.644***
Cort + Flu 31.25 1.857**** 25.00 1.755*****
Cort, corticosterone; Flu, fluvoxamine; Sc, subcutaneous. Results are
expressed as the mean SEM (n=11–12 per group). Statistical anal-
ysis was performed by Kruskal–Wallis, followed by Dunn’s post hoc
test for the % of entries in the open arms and one-way ANOVA, fol-
lowed by Tukey’s post hoc test in the % time spent in the open arms.
Significant values: *P<0.01; **P<0.0001 vs control group and
***P<0.05; ****P<0.01 and *****P<0.001 vs Cort group.
©2019 Royal Pharmaceutical Society, Journal of Pharmacy and Pharmacology,71 (2019), pp. 1774–17831778
Thymol reverses depression-like behavior Victor Celso Cavalcanti Capibaribe et al.
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correct alternations compared to control group
(P=0.0042) and the treatment with thymol was able to
increase this parameter (P=0.0023; F(3, 32) =8.269) as
shown in Figure 9.
BDNF assessment
Total level of BDNF was measured using ELISA (sandwich),
while the level of the mature form (mature BDNF) was
determined using Western blotting. In ELISA assay, treat-
ment with corticosterone or tested drugs showed no alter-
ations in BDNF levels, as shown in Figure 10A.
Interestingly, in Western blot analysis (Figure 10B), only
thymol treatment significantly increased the expression of
mature BDNF compared to Cort group (P=0.0395).
Discussion
The present study shown that thymol reverses depression-
like behaviour and upregulates hippocampal BDNF levels
in chronic corticosterone-induced depression model in
female mice. These results are important in the current sce-
nario where depression, unfortunately, is still an ongoing
condition with a chronic and recurrent course
[38]
and often
associated with significant physical and mental health
impairment in the patients
[39]
and that existing medicines
still do not solve completely.
For a long time, the biological underpinnings of depres-
sive disorders have been supported by the monoaminergic
hypothesis of depression, which does not explain why 30%
of antidepressants are refractory.
[40]
In this way, other theo-
ries have been proposed, including those that focus on the
participation of the neurotrophic factors such as BDNF.
The antidepressant-like effects of thymol were positively
demonstrated after its oral administration, as shown by
decrease observed in the immobility time in forced swim-
ming and tail suspension tests. These tests are based on the
premise that these present stressful situations activate the
trigger of fight/flight behavior.
[11,24,41]
So, we can consider
that immobility is directly related to despair and depres-
sion-like behaviour, wherein thymol proved to be effective
in the reversal of this parameter.
However, the forced swimming and tail suspension tests
can produce false-positive results if the drug tested is a gen-
eral stimulant. In this case, this hypothesis can be ruled out
using the open field test, described first by Hall,
[42]
that
consists of evaluating a series of parameters, including
hyperactivity,
[43]
exploratory behaviour
[44]
and locomotor
activity.
[45,46]
The absence of interference in the pattern of
square crossings observed in the present study allowed us
to conclude that corticosterone, as well as fluvoxamine and
thymol, did not interfere in the locomotor activity of the
animals.
Another parameter that was evaluated was anhedonia.
This term was first defined as the inability to feel pleasure
and was first described by Ribot.
[47]
Currently, DSM-V, in
addition to defining anhedonia as the reduced capacity to
have pleasure resulting from positive stimuli or degradation
of the memory of a previously experienced pleasure, reports
that loss of interest or pleasure is one of the crucial symp-
toms for the diagnosis of depression.
[48–50]
Therefore, thy-
mol presented significant result when compared to the
corticosterone group, in reference to one of the main symp-
toms presented in the major depressive disorder. This result
indicates thymol as a possible therapeutic tool for this chal-
lenging trait of depression.
The complexity of depressive disorder is also manifested
by the fact that it coexists with other psychiatric conditions
such as anxiety, leading the patient to manifest autonomic
Figure 8 Percentage of the social preference in the social interaction
test. Data are expressed as the mean SEM (n=10–12 per group).
Statistical analysis was performed by one-way ANOVA, followed by
Tukey’s post hoc test. Significant values: *P<0.001 vs control group,
**P<0.001 and ***P<0.0001 vs Cort group.
Figure 9 Percentage of the number of correct alternations obtained
in the Y-maze results. Results are expressed as the mean SEM
(n= Statistical analysis was performed by one-way ANOVA, followed
by Tukey’s post hoc test. Significant values: *P<0.01 vs control
group and **P<0.01 and ***P<0.001 vs Cort group.
©2019 Royal Pharmaceutical Society, Journal of Pharmacy and Pharmacology,71 (2019), pp. 1774–1783 1779
Victor Celso Cavalcanti Capibaribe et al. Thymol reverses depression-like behavior
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symptoms. The coexistence of another psychiatric disorder
with depression has a major impact on the course of the
disease, resulting in delayed recovery, increased risk of
relapse, disability and suicide attempts.
[51,52]
Several studies
have identified social anxiety disorder as a significant risk
factor for the future onset of depression in all ages.
[53,54]
Thus, it is relevant to assess whether thymol shows any
improvement in anxiety symptoms.
The plus-maze test is considered the most commonly
accepted and validated test for the search for new anxiolytic
agents. It is based on the fact that the animals have aversion
to open and elevated places, thus restricting most of their
activities in closed arms.
[55]
The present study showed that
thymol administration was able to induce anxiolytic effects
in the plus-maze test.
In order to confirm the anxiolytic activity of thymol, it
was decided to use the hole-board test which also shows
that exploratory behaviour is gradually inhibited by anxi-
ety. This model is also used to induce anxiety in experimen-
tal animals, and it has been shown that anxiolytic drugs
increase the number of head-dips.
[56]
Our results showed
that thymol significantly increased the number of head-
dips, also indicating a possible anxiolytic effect.
In addition, thymol demonstrated an increase in the resi-
dence time of animals in the light division in the light/dark
test. Such, the test demonstrates that anxiolytic drugs
increase the latency for entry into the dark division.
[57,58]
Given the tests performed, we can see that thymol has
potential antidepressant accompanied by an ability to
reduce symptoms associated with depression such as
anxiety.
Gotlib and Joormann,
[59]
in their review, suggest that
people’s thoughts, conclusions, attitudes and interpreta-
tions, as well as how they attend to and recall events may
increase the risk of developing depressive episodes.
Segrin
[55]
claims that the combination of deficiency in
social skills and negative life events are corroborating
factors for the onset of depressive symptoms. More
recently, Bolsoni-Silva and Loureiro
[56]
concluded that a
good repertoire of social skills helps an individual to
overcome stressful events, since the individual’s capacity
to identify the quantity of social support needed to
transpose this situation is affected by their social interac-
tions. The authors also report that deficits in social skills
may be associated, or still be associated with depressive
symptoms.
In this way, the ability of an antidepressant drug to pre-
sent a significant response in social interactions is very pos-
itive. The reference drug fluvoxamine, a selective serotonin
reuptake inhibitor (SSRI), has clinical action against several
aspects of anxiety disorder such as panic disorders, obses-
sive–compulsive disorder and social anxiety disorders.
[54]
In our study, thymol, similar to fluvoxamine, increased
social interaction. This finding is important since some
patients continue suffering from residual social and func-
tional impairment, even after antidepressant treatment.
[60]
Studies suggest the relation between depression and defi-
cits in executive functioning, working memory and atten-
tion.
[61,62]
This can be observed in patients with major
depression that often manifest difficulties in concentration
and problem solving, consistent with DSM-V criteria for
diagnosing a major depressive episode.
[63]
Figure 10 (a) Evaluation of the quantity of BDNF picograms per gram of tissue—ELISA methodology. Results are expressed as the mean SEM
(n=4–6 per group). Statistical analysis was performed by Kruskal–Wallis, followed by Dunn’s post hoc test. (b) Evaluation the quantity of BDNF
pictograms per gram of tissue—Western blot methodology. Results are expressed as the mean SEM (n=3–4 per group). Significant values:
*P<0.05 vs Cort group.
©2019 Royal Pharmaceutical Society, Journal of Pharmacy and Pharmacology,71 (2019), pp. 1774–17831780
Thymol reverses depression-like behavior Victor Celso Cavalcanti Capibaribe et al.
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The Y-maze test is widely used to analyse exploratory
behaviour, learning and memory.
[57–59]
According to
Yan,
[60]
the test can measure the functioning of the imme-
diate spatial memory, which is classified as a type of short-
term memory. Some studies have shown that the memory
deficits observed in the Y-maze test may be related to low
BDNF levels, resulting in impairment of cognition along
with the symptoms of depression.
[64]
The brain-derived neurotrophic factor (BDNF) is a
member of the neurotrophin family, which is responsible
for the regulation of neuronal differentiation and synaptic
plasticity, two complex processes that directly influence
memory and learning throughout life.
[61,62]
The ability of a
synapse to change in strength (synaptic plasticity) is con-
sidered the cellular basis of most cognitive processes,
including memory formation and is extensively study
through CA1–3 synapses in the hippocampus.
[64]
Neuronal activity, at the CA1–CA3 junction, induces
presynaptic release of BDNF, and the subsequent interac-
tion of BDNF mature form, with the postsynaptic neu-
rotrophic tyrosine kinase receptor B (TrkB), is required for
the long-term potentiation (LTP) and synapse consolida-
tion.
[65]
For this reason, BDNF and its receptor TrkB have
come under investigation as potential targets for treating
cognitive symptoms.
[63,64]
Unfortunately, antidepressant drug treatment has also been
associated with poor performance on tasks evaluating working
memory and verbal learning and memory
[66]
specially in
elderly.
[67]
Thus, thymol administration showed an increase
in mature BDNF in stressed mice hippocampus, unlike flu-
voxamine, exhibiting a differential effect on cognition.
Most of the cognitive theories propose that alterations in
cognition lead to improvement of other symptoms of
depression, including sustained negative affect and anhedo-
nia. For Gotlib and Joormann,
[59]
the antidepressant effect
of thymol and its ability to improve the short-term mem-
ory may be related to neurotrophins regulation.
These results, therefore, indicates for the first time, as far
as we know, the thymol antidepressant-like effect, and it
was followed by an upregulation of BDNF, opening per-
spective of as a possible therapeutic tool for this challenging
trait of depression in the future.
Conclusion
The present study confirmed the marked antidepressant-
like effect of thymol, as demonstrated here initially by its
ability to reverse chronic corticosterone-associated beha-
viour and decrease in BDNF levels. The field of neurogenic
pathway is still new, however, very promising, and thymol
seems to exert a positive effect on BDNF expression and
maturation in the hippocampus. The present work opens
the door to further research, mainly related to thymol,
envisioning future elucidation of this drug’s mechanisms of
action and its use as a therapeutic antidepressant alterna-
tive.
Declarations
Conflict of interest
The Author(s) declare(s) that they have no conflicts of
interest to disclose.
Acknowledgements
This study was financed in part by the Coordenac
ß
~
ao de
Aperfeic
ßoamento de Pessoal de N
ıvel Superior—Brasil
(CAPES)—Finance Code 001; by Conselho Nacional de
Desenvolvimento Cient
ıfico e Tecnol
ogico (CNPq-Process
numbers: no 12/2017, no 306746/2013-1, no 446120/2014-
6 and no 407567/2013-5) and Fundac
ß
~
ao Cearense de Apoio

a Pesquisa (FUNCAP-Cear
a-Brazil).
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