Received: 23 September 2020 | Revised: 7 December 2020 | Accepted: 4 January 2021

DOI: 10.1111/jfbc.13622

FULL ARTICLE

Resveratrol attenuates methamphetamine-induced memory

impairment via inhibition of oxidative stress and apoptosis in
mice

Qing Zeng1 | Qi Xiong1 | Mei Zhou1 | Xiang Tian1 | Kai Yue1 | Yi Li2 | Xiji Shu1 |
Qin Ru1

1
Wuhan Institutes of Biomedical Sciences,
School of Medicine, Jianghan University, Abstract
Wuhan, China Methamphetamine (METH) abuse produces serious neurotoxicity to the central
2
Wuhan Mental Health Center, Wuhan,
nervous system along with long-term cognitive dysfunction. Resveratrol, a natural
China
polyphenol, has broad application prospects in the treatment of neurodegenerative
Correspondence
diseases. Therefore, this study was conducted to investigate whether resveratrol
Qin Ru, Wuhan Institutes of Biomedical
Sciences, School of Medicine, Jianghan might alleviate METH-induced memory deficits in vivo. We found that multiple ex-
University, No. 8 Sanjiaohu Road, Wuhan,
posures to METH significantly impaired cognitive functions and caused long-lasting
Hubei 430056, China.
Email: ruq.whibs@aliyun.com memory deficits (p < .05). Pretreatment of resveratrol (10 or 100 mg/kg) remark-
ably attenuated METH-induced memory impairment in mice (p < .05). Bioinformatics
Funding information
China Scholarship Council, Grant/Award analysis results showed that resveratrol might alleviate memory deficits by inhibit-
Number: 201908420112
ing METH-induced oxidative damage and apoptosis. Molecular docking showed that
resveratrol had hydrogen bonding interactions with Kelch-like ECH associated pro-
tein 1 (Keap1), a repressor protein of the classic antioxidant Keap1-Nrf2 pathway.
Further results validated oxidative stress parameters, apoptosis, and expression of
Keap1 were significantly increased, while the translocation and activation of nuclear
factor erythroid 2-related factor 2 (Nrf2) into the nucleus and expression of its down-
stream proteins were greatly decreased in the hippocampus after METH exposure
(p < .05). These changes caused by METH could be prevented by resveratrol (p < .05).
Therefore, these findings suggested that the prevention of resveratrol on memory
dysfunction induced by METH was possibly related to the activation of the Keap1-
Nrf2 pathway and reduction of apoptosis. Supplementation of resveratrol could be a
potential treatment for preventing the neurotoxicity of METH in the future.
Practical applications
As one of the worst commonly abused psychostimulants, methamphetamine (METH)
addiction produces serious complications including cognitive impairment and mem-
ory deficits. Resveratrol is a natural polyphenol that has important nutritional supple-
ments and protective effects in the treatment of many neurodegenerative diseases.

Abbreviations: ANOVA, analysis of variance; DA, dopamine; GCSc-γ, glutamyl-cysteine synthetase-γ; GSH, glutathione; HO-1, heme oxygenase-1; Keap1, Kelch-like ECH associated
protein 1; MDA, methane dicarboxylic aldehyde; METH, methamphetamine; NOR, novel object recognition; Nrf2, nuclear factor erythroid 2-related factor 2; Res, resveratrol; ROS,
reactive oxygen species; Sal, saline; SOD, superoxide dismutase; TAC, total antioxidant capacity; Veh, vehicle; γ-H2AX, phosphorylated H2AX.
Xiji Shu and Qin Ru are equally contributed to this article.

J Food Biochem. 2021;45:e13622. wileyonlinelibrary.com/journal/jfbc © 2021 Wiley Periodicals LLC. | 1 of 15

https://doi.org/10.1111/jfbc.13622
2 of 15 | ZENG et al.

In this study, the results of bioinformatics prediction and experimental validation

showed that resveratrol might effectively prevent memory impairment via the inter-
action with Keap1, activation of the Keap1-Nrf2 pathway, and inhibition of DNA dam-
age and apoptotic responses post METH exposure. Therefore, these findings provide
new ideas and insights into the application of resveratrol in the treatment of nervous
system damage caused by METH.

KEYWORDS

apoptosis, Keap1-Nrf2 pathway, memory deficits, methamphetamine, resveratrol

1 | I NTRO D U C TI O N brain and exert powerful neuroprotective functions (Bastianetto

As one of the worst commonly abused psychostimulants, metham-
phetamine (METH) addiction produces serious complications sys-
temically affecting multiple organs, especially the central nervous
system (Reichel et al., 2014). Multiple METH exposures can result in
neurodegenerative changes in the hippocampus and frontal cortex,
which are all related to long-term cognitive dysfunction and memory
deficits in humans and animal models (Avila et al., 2018; Casaletto
et al., 2015; Weber et al., 2012). Long-term abuse of METH reduced
addicts' awareness of their memory acquisition and recall defi-
cits, and the overestimation of memory further exacerbated their
executive dysfunction and reduced cognitive reserve (Casaletto
et al., 2015). Neurocognitive deficits did not just occur in people who
were currently abusing METH, however, it has also been found in
those who have stopped taking for a long time (Cherner et al., 2010;
Silva et al., 2014). Using magnetic resonance imaging and computa-
tional image analyses, Thompson et al. had shown that METH abuse
caused severe gray-matter deficits in the cerebral cortex that con-
tributed to impaired memory performance (Thompson et al., 2004),
indicating that the neuronal damage is likely to participate in the
memory injury caused by METH.
Memory impairment caused by various factors such as aging and
lipopolysaccharide could all be alleviated by antioxidants such as
naringin and vitamin E (Baghcheghi et al., 2018; Khajevand-Khazaei
et al., 2018). For instance, vitamin E could improve post-traumatic
stress disorder, cisplatin, or waterpipe smoking-induced memory im-
pairment possibly through affecting alterations in oxidative stress
biomarkers including superoxide dismutase (SOD), glutathione (GSH),
and methane dicarboxylic aldehyde (MDA) (Ahmed et al., 2020;
Alzoubi et al., 2019; Hosseinzadeh et al., 2020). However, less re-
search emphasis has been focused on whether antioxidants have
protective effects against METH-induced memory deficits.
Resveratrol (trans-3,5,4’-trihydroxystilbene, Res) is a stilbe-
ne-type natural polyphenol mainly found in dietary plants including
raspberries, mulberries, and the skin of grapes (Yeung et al., 2019).
Resveratrol has important nutritional supplements and chemopre-
ventive effects in the treatment of many diseases (Feng et al., 2019;
Rauf et al., 2017; Wahl et al., 2018). Although poorly water-soluble,
resveratrol shows high membrane permeability and can enter the
et al., 2015; Chen et al., 2017; Rigon et al., 2019). For instance, res-
veratrol had a significant inhibitory effect on cognitive impairment in
aged mice after surgery and Alzheimer's disease rats (Lin et al., 2018;
Wang et al., 2018). Moreover, resveratrol could protect dopaminer-
gic neurons from METH-induced neuronal cytotoxicity in vitro(Kan-
thasamy et al., 2011; Sun et al., 2015). However, few reports were
focusing on the protective effect of resveratrol on cognitive impair-
ment caused by METH in vivo. Based on these supportive results,
this study was conducted to investigate the potential beneficial ef-
fect of resveratrol against the memory deficits caused by METH. To
this end, novel object recognition (NOR) task, a classic experiment
for detecting cognitive function was used to test the memory capac-
ity of mice. Furthermore, the expression levels of oxidative stress
biomarkers including SOD and MDA as well as apoptosis induction
were evaluated, bioinformatics analysis and comet electrophoresis
were also used to analyze potential mechanisms of the neuroprotec-
tive effect of resveratrol.
2 | M E TH O DS A N D M ATE R I A L S
2.1 | Animals
C57BL/6 mice (18–22 g, male) were purchased from Beijing vital
river laboratory animal technology Co., Ltd. Every five mice were
placed in one cage, and water and food were freely available. Mice
were housed in a 12-hr light-dark cycle and the temperature of
the environment was controlled at 23 ± 20°C. All animal experi-
ment procedures were performed under the “Regulations on the
management of laboratory animals” issued by the National Science
and Technology Commission of China and approved by the Ethics
Committee of Jianghan University.
2.2 | Reagents
METH (98%) was obtained from the Hubei Public Security Bureau
and was dissolved in saline (Sal) to a stock concentration of 10 mg/
ml before injections. Resveratrol (Res, CAS Number 501-36-0,
ZENG et al. | 3 of 15

F I G U R E 1 Memory deficits induced by chronic administration of METH. (a) Experimental procedure. METH was intraperitoneally
injected for 10 consecutive days following the schedule in Table 1. Novel object recognition (NOR) experiment was carried out on days 17
and days 32. After NOR testing, the locomotor activity experiment was carried out as described. (b) Pattern diagrams of NOR training and
testing. (c and e) Discrimination index of NOR testing. Compared with saline-treated mice, the discrimination index of METH-treated mice
was significantly reduced on days 17 (c) and days 32 (e). (d and f) The total distance of movements in the locomotor activity test. The total
distance of movements among the different groups exhibited no significant differences on Day 17 (d) and Day 32 (f). Data were presented as
means ± SEM. n = 10 for per group, **p < .01 compared to saline group
4 of 15 | ZENG et al.

F I G U R E 2 Resveratrol ameliorated memory deficits induced by METH. (a) Experimental procedure. METH was intraperitoneally injected
for 10 consecutive days following the schedule in Table 1. Resveratrol (10 or 100 mg/kg) was oral administrated 1 hr before METH injection.
The habituation session of the NOR experiment was carried out on days 16. Twenty-four hours after the habituation, the training and
testing sessions of the NOR experiment were performed. The locomotor activity experiment was carried out after the NOR testing session.
After that, the comet assay and western blotting experiments were performed. (b) Discrimination index to explore the effect of resveratrol
on METH-induced memory deficits. Compared with saline-treated mice, the discrimination index of METH-treated mice was significantly
reduced. Resveratrol (10 or 100 mg/kg) pretreatment significantly ameliorated METH-induced reduction of the discrimination index. (c)
The total distance of movements in the locomotor activity test. The total distance of movements among the different groups exhibited no
significant differences. (d) Representative graphs of trajectories during the NOR testing. The green circle represented the location of the
new object. The yellow circle represented the location of the familiar object. Meanwhile, the different colored dots represented the starting
and ending points of the animal. Data were presented as means ± SEM. n = 10 for per group, **p < .01 compared to the Veh/Sal group,
##
p < .01 compared to the Veh/METH group

≥99%) was purchased from Sigma Chemical Corp (St Louis, MO, kit, MDA assay kit, and GSH assay kit were procured from Nanjing
USA) and was dissolved in 0.5% sodium carboxymethylcellulose Jiancheng Bioengineering Research Institute (Nanjing, China).
(vehicle, Veh). Total antioxidant capacity (TAC) assay kit, SOD assay Antibodies against heme oxygenase-1 (HO-1, abs131494), glutamic
ZENG et al.
TA B L E 1 Dosing schedule in the chronic METH exposure group
Day 1 Day 2 Day 3 Day 4 Day 5
(mg/kg) (mg/kg) (mg/kg) (mg/kg) (mg/kg)
T1 0.5 1 1.5 2 2.5
T1 + 2 hr 0.5 1 1.5 2 2.5
T1 + 4 hr 0.5 1 1.5 2 2.5
T1 + 6 hr 0.5 1 1.5 2 2.5
Note: Mice in chronic METH exposure group were intraperitoneally injected with METH four times at an interval of 2 hr for 10 consecutive days.
The saline control group (vehicle group) mice received a similar volume of physiological 0.9% saline via i.p. injections according to the same schedule.
T1 = the time of day for the first injection. Use the same time for each day (i.e., always give the first injection at 9:30 a.m.)
cysteine synthetase-γ (GCSc-γ, abs138070), Bax (abs130057), Bcl-2
(abs131701), cleaved caspase-3 (abs132005), Lamin B (abs131244)
and phosphorylated H2AX (γ-H2AX, abs130814) were purchased
from Absin Bioscience Co., Ltd (Shanghai, China). Antibody against
GAPDH (BM3876), Kelch-like ECH associated protein 1 (Keap1,
PB0813), nuclear factor erythroid 2-related factor 2 (Nrf2) and
horseradish peroxides (HRP) conjugated goat anti-rabbit antibody
(BA1050) and HRP conjugated goat anti-mouse antibody (BA1056),
protein extraction buffer, protease, and phosphatase inhibitors
were obtained from Wuhan Boster Biological Technology Co., Ltd
(Wuhan, China). The nuclear and cytoplasmic protein extraction kit
was from Beyotime Company (Haimen, China).
2.3 | Animals treatment
Specifically, animal experiments were divided into two parts, and ex-
perimental procedures were detailed in Figures 1a and 2a.
Experiment 1: Mice were divided randomly into the saline group
and the METH group. Mice in the METH group were injected in-
traperitoneally with METH following the schedule in Table 1 (Li
et al., 2017). Mice in the saline group received injections with an
equivalent volume of saline according to the same schedule. Multiple
incremental METH exposure patterns were designed to mimic the
clinical situation of METH abuse, which usually includes gradual in-
creases in drug intake. Behavioral tests were performed on Day 17
and Day 32 and were conducted in separate animal groups.
Experiment 2: Mice were divided randomly into Veh/Sal group,
Veh/METH group, Res-L/METH group, and Res-H/METH group.
METH was administrated following the schedule in Table 1. One
hour before the first daily injection of METH, 10 mg/kg (Res-L) or
100 mg/kg (Res-H) resveratrol was intragastrically administrated.
The concentrations of resveratrol were chosen according to refer-
ences (Corpas et al., 2019; Wang et al., 2018). Behavioral experi-
ments, comet assay, and western blot were performed as follows.
2.4 | Novel object recognition task
The NOR task was carried out according to the literature with
minor modifications (Maroso et al., 2016). Briefly, the experimental
| 5 of 15
Day 6 Day 7 Day 8 Day 9 Day 10
(mg/kg) (mg/kg) (mg/kg) (mg/kg) (mg/kg)
3 3.5 4 4.5 5
3 3.5 4 4.5 5
3 3.5 4 4.5 5
3 3.5 4 4.5 5
device consisted of a 40 × 40 × 40 cm Plexiglas open field box and
a video tracking system XR-XX117 (Shanghai Xinruan Information
Technology Co.Ltd, China). The NOR task procedure included three
phases: habituation, training, and testing. During the habituation
phase, mice can explore freely in the box for 10 min without an ob-
ject to adapt to the environment. After 24 hr, two objects were fixed
symmetrically on the bottom of the box, and mice were allowed to
explore two objects freely. The training phase lasted for 5 min. Then
mice were returned to their home cages immediately. A novel object
was used to replace one of the familiar objects 4 hr later, and mice
were returned to the box and freely explored. The testing phase also
lasted for 5 min. When the mouse's head entered a range of less than
or equal to 2 cm around the object, the mouse was thought to be
exploring the object. Time spent exploring the novel (N) and familiar
(F) object was recorded respectively. This formula (N − F/N + F) was
used to calculate the discrimination index during the testing phase.
2.5 | Locomotor activity testing
The experimental device of the locomotor activity test included
a 40 × 40 × 35cm plexiglass chamber and tracking software
(YH2018RLA16, Wuhan Yihong Technology Co., Ltd.). After the
NOR testing phase, mice were tested for locomotor activity accord-
ing to the experimental arrangement shown in Figures 1a and 2a. To
detect the effects of METH and resveratrol on the locomotor per-
formance of mice, the total distance and movement speed of mice in
10 min were recorded.
2.6 | Prediction of drug targets for resveratrol and
collection of gene targets for METH-induced
memory impairment
The potential protein targets of resveratrol, related genes of METH,
and related genes of memory impairment were collected from the
GeneCards database (https://www.genec​ards.org/). Then, the pro-
tein targets of resveratrol were mapped with related genes of METH
and related genes of memory impairment on the Bioinformatics &
Evolutionary Genomics website (http://bioin​forma​tics.psb.ugent.be/
webtools/Venn/). The gene ontology (GO) analyses were conducted
6 of 15 | ZENG et al.
using the functional annotation tool of DAVID Bioinformatics
Resources 6.7 (http://david.abcc.ncifc​r f.gov/). Terms with thresh-
olds of Count ≥ 10 and p values ≤ .05 were chosen in functional
annotation clustering.
2.7 | Molecular docking of resveratrol with Keap1
The Keap1 amino acid sequence was downloaded from the PDB pro-
tein structure database (http://www.rcsb.org/, PDB ID: 6LRZ). This
is a co-crystal structure of Keap1 in complex with dimethyl fumarate.
The atoms of dimethyl fumarate were discarded using Pymol 1.4.
Furthermore, AutoDock v.4.2.6 was used to add the hydrogens, to
calculate charges, and to merge the non-polar hydrogens of protein
structure(Zulkipli et al., 2020). Next, the protein file was saved in
PDBQT format. The three-dimensional (3D) structure of resveratrol
was downloaded from the PubChem structure database (https://
pubch​em.ncbi.nlm.nih.gov/, Compound CID: 445,154). The SDF for-
mat of resveratrol was converted to PDB format using OpenBabel
2.4.1(Kumar et al., 2020). AutoDock v.4.2.6 was used to convert the
PDB format of resveratrol to PDBQT format and was used for mo-
lecular docking of resveratrol with Keap1 to understand the inter-
action between them. In AutoDock software, the grid dimensions
were set at 129 Å × 129 Å × 129 Å to cover the entire binding site
of Keap1. Besides, Pymol 1.4 was used to verify the hydrogen bonds
between resveratrol with Keap1.
2.8 | Determination of antioxidant capacity and
lipid peroxidation
Mice were sacrificed by cervical dislocation under anesthesia after
the indicated treatment. Blood samples were collected and centri-
fuged at 3,000 rpm for 10 min at 4°C, and the activities of SOD or
contents of GSH in serum were measured. The detection method
was performed according to the manufacturer's instructions. MDA
concentration, which was usually used to measure the degree of lipid
peroxidation, was measured using the MDA detection kit according
to the manufacturer's instructions. The TAC of serum from different
groups of mice was also determined.
2.9 | Comet assay
The comet assay is an effective method to determine DNA damages
at the single-cell level(Freese et al., 2018). In brief, the hippocam-
pus was immediately taken out to obtain a single-cell suspension.
The isolated cells were counted in a Cell counter to determine cell
concentration and survival by trypan blue exclusion assay. Then
low melting point agarose (0.75%) was mixed with cells, and cells
were further seeded on microscope slides precoated with a layer of
1% normal melting point agarose. The slides were placed in a cell
lysis solution at 4°C for 1.5 hr and electrophoresed for 20 min to
make DNA to disentangle. Finally, slides were washed and stained
with SuperStain (Cwbio company, China) for 20 min avoiding light.
Pictures were analyzed with fluorescence microscopy (Olympus,
Japan). Comet assay software project 12.2 software was used to
analyze the tail length and percentage of tail DNA.
2.10 | Western blotting analysis
Hippocampal were isolated, lysed, and centrifuged for 15 min (4°C)
at 12,000 g. The nuclear Nrf2 protein was obtained through a nu-
clear protein extraction kit. After the detection of concentration,
the supernatants were mixed with loading buffer and denatured for
5 min. Protein samples were separated using a gel electrophoresis
system and transferred to the polyethylene difluoride membranes.
After blocking for 1 hr in 5% nonfat milk, membranes were incu-
bated with the primary antibody and then with the HRP secondary
antibody. The following primary antibodies were used: anti-Keap1
(1:500), anti-Nrf2 (1:500), anti-HO-1 (1:1,000), anti-Bax (1:1,000),
anti-GCSc-γ (1:2,000), anti-Bcl-2 (1:1,000), anti-cleaved caspase-3
(1:1,000), anti-Lamin B (1:1,000), anti-γ-H2AX (1:1,000), and anti-
GAPDH (1:1,000). Enhanced chemiluminescence (Thermo Fisher,
USA) was used to observe the bands using a chemiluminescence de-
tector (Gene Corporation, Hong Kong). The intensity of each band
was determined quantitatively by Image J and calibrated by the cor-
responding internal reference protein, and the results were shown as
normalized for the Veh/Sal group.
2.11 | Statistical analysis
Data were expressed as mean ± standard error (SEM). All results
were analyzed using SPSS 23.0 software. The independent-sample
t test was used to assess the differences between the groups in
Figure 1. Other data used one-way ANOVA and Tukey's HSD post
hoc test. p less than .05 was considered statistically significant.
3 | R E S U LT S
3.1 | Multiple METH exposures induced memory
deficits
The NOR task was carried out to examine whether the multiple ex-
posures of METH caused long-term memory deficits in mice. Animal
treatment and the time points of the behavioral test were shown in
Figure 1a and Table 1, and the procedure of the NOR task was shown
in Figure 1b. Figure 1c demonstrated that mice in the saline group
showed exploratory preferences for the novel subject during the
testing phase. The discrimination index in the METH group was sig-
nificantly lower on Day 17 (Figure 1c, p < .01) than that of the saline
group, and this phenomenon lasted until Day 32 (Figure 1e, p < .01),
suggesting multiple METH exposures could induce long-term memory
ZENG et al.
F I G U R E 3 (a) The potential protein targets analysis of resveratrol on memory deficits induced by METH. (b) The 14 most significance of
gene ontology enrichment analysis of therapy target genes of resveratrol on memory deficits induced by METH. c. Molecular docking results
of resveratrol and Keap1
impairment. At the same time, the locomotor performance was similar
in both groups, and there was no difference between saline and METH
groups on Day 17 (Figure 1d, p > .05) and Day 32 (Figure 1f p > .05) in
total distance. The lack of alteration in the total distance in locomotor
activity testing indicated that mice in the METH group did not display
major motor deficits. These data suggested that multiple METH expo-
sures could result in long-time (22 days after the last METH injection)
memory deficits without affecting locomotor activities.
3.2 | Resveratrol ameliorated memory deficits
induced by METH
To study the preventive effect of resveratrol, different doses of
resveratrol were orally administrated 1 hr before the first METH
injection every day. None of the experimental groups showed
any changes in toxic behavior or mortality, and no adverse events
were observed. Figure 2a showed the arrangement of subsequent
| 7 of 15
experiments. The discrimination index of mice in the METH group
was significantly reduced in comparison with the saline group
(Figure 2b and d, p < .01). Pretreatment with resveratrol (10 or
100 mg/kg) significantly inhibited the decrease of the discrimination
index induced by METH (p < .01). Besides, there was no significant
difference among the different groups in terms of the total distance
in locomotor activity testing (Figure 2c, p > .05), indicating that the
locomotor activity of mice was not affected by METH or resveratrol.
These results showed that resveratrol exerted a remarkable neuro-
protective effect on memory impairment caused by METH without
affecting locomotor activities.
3.3 | Potential mechanisms identification of
resveratrol on memory deficits induced by METH
To investigate the potential mechanism by which resveratrol allevi-
ated METH-induced learning and memory impairment, the targets
8 of 15 | ZENG et al.

F I G U R E 4 Effect of resveratrol on the oxidative enzyme system after METH exposure. After the behavior test, mice were sacrificed
and the serums were collected for measurements of the levels of SOD (a), TAC (b), GSH (c), and MDA (d) using corresponding commercial
detection kits. Data were presented as means ± SEM. n = 10 for per group, *p < .05, **p < .01 compared to the Veh/Sal group, #p < .05,
##
p < .01 compared to the Veh/METH group

of resveratrol and related targets for METH and memory impair-
ment were searched on the GeneCards database. Six hundred and
ninety-one targets of resveratrol, 8,397 memory impairment-related
proteins, and 422 METH-related proteins were collected from the
GeneCards database. Then, these protein targets were mapped on
the Bioinformatics & Evolutionary Genomics website. As a result,
110 targets of resveratrol were associated with METH-induced
memory impairment (Figure 3a). To identify the biological character-
istics of putative targets of resveratrol with METH-induced memory
impairment in detail, the GO enrichment analyses of involved tar-
gets were conducted via the functional annotation tool of DAVID
Bioinformatics Resources 6.7. There were 827 terms of biological
process (BP) which met the requirements of Count ≥ 10 and p val-
ues ≤ .05. The top 14 significantly enriched terms (Count ≥ 60 and p
values ≤ 1 × 10–30) in BP were shown in Figure 3b, which indicated
that resveratrol may regulate neuron function via apoptosis process,
programmed cell death, response to oxygen-containing compound to
exert its therapeutic effects on METH-induced memory impairment.
The Keap1-Nrf2 pathway is the major regulator of cytopro-
tective responses to oxidative stresses, thereby is responsible for
cell defense against apoptosis in nervous system disorders like
cognitive impairment (Baird & Yamamoto, 2020; Sharma et al., 2020).
Molecular docking is a computational approach to identify possible
binding modes of the selected compound against its biological target
(Pingaew et al., 2018). Therefore, Autodock software was used to do
molecular docking between resveratrol and Keap1 and analyze the
possible interaction between them. The Molecular Docking results
showed that resveratrol and Keap1 had a good affinity, and its bind-
ing energy is −5.42 kcal/mol, and Leu365, Leu 557, and Ile559 were
critical amino acids responsible for the formation of hydrogen bonds
(Figure 3c). These results indicated that Keap1 might be the poten-
tial target of resveratrol, and resveratrol may improve antioxidant
capacity by influencing the conformation of Keap1 for alleviating
METH-induced memory impairment.
3.4 | Effect of resveratrol on the oxidative system
after METH exposures
The contents of MDA and GSH, SOD activities, and total antioxi-
dant capacity in serum were examined to verify the possible ef-
fect of resveratrol in METH-induced oxidative stress. As shown
ZENG et al.
in Figure 4, METH significantly reduced SOD activities (Figure 4a,
p < .05), GSH content (Figure 4c, p < .01) and total antioxidant ca-
pacity (TAC, Figure 4b, p < .01) in comparison with the saline group.
Furthermore, SOD activities, GSH levels, and total antioxidant ca-
pacity were greatly enhanced in the serum after 100 mg/kg resvera-
trol pretreatment (p < .05). Additionally, compared with the saline
group, the serum MDA content in the METH group was significantly
higher (Figure 4d, p < .01) compared with the control group. In con-
trast, pretreatment of resveratrol (100 mg/kg) greatly lowered the
content of MDA (p < .05). These data showed that pretreatment of
resveratrol may exert a neuroprotective effect against memory im-
pairment induced by METH via reducing oxidative stress.
3.5 | Resveratrol prevented METH-induced
abnormal expression of proteins in the Keap1-
Nrf2 pathway
Nrf2 is an essential redox-regulated transcription factor that can
regulate the transcription of cytoprotective genes, such as HO-1,
GCSc-γ, and SOD during oxidative stress in the brain, Nrf2 stores in
the cytoplasm as an inactive form via binding to Keap1, the confor-
mational change of Keap1 could facilitate the dissociation of Nrf2
from Nrf2-Keap1 complex, which allows Nrf2 to translocate into the
nucleus (He et al., 2020; Taguchi et al., 2020). So the effect of res-
veratrol on the expressions of related proteins in the Keap1-Nrf2
pathway was shown in Figure 5. METH significantly upregulated the
expression of Keap1 (p < .05), inactivated the nuclear translocation
of Nrf2, and downregulated expressions of its downstream protein
HO-1, and GCSc-γ (p < .05) in the hippocampus. Furthermore, the
pretreatment of resveratrol (10 or 100 mg/kg) effectively activated
the nuclear translocation of Nrf2 and increased protein expressions
of HO-1, and GCSc-γ post METH exposures, and decreased the ex-
pression of Keap1 (p < .05). These results proved that resveratrol
may, at least in part, enhance the antioxidant capacities of the hip-
pocampus by activating the Keap1-Nrf2 signaling pathway to allevi-
ate METH-induced memory impairment.
3.6 | Resveratrol prevented METH-induced
apoptosis in hippocampus
To evaluate the effect of METH-induced apoptosis and the interven-
tion effect of resveratrol, Western blotting was used to detect ex-
pressions of Bax, Bcl-2, and cleaved caspase-3 proteins. Compared
to the control group, multiple METH exposures significantly en-
hanced protein expressions of Bax and cleaved caspase-3 (p < .01,
Figure 5a and d), and remarkably reduced the level of Bcl-2 in the
hippocampus (p < .05). Moreover, pretreatment with resveratrol (10
or 100 mg/kg) could significantly increase the level of Bcl-2 and de-
crease the protein level of cleaved caspase-3 and Bax (p < .05). The
result suggests that resveratrol could prevent METH-induced dam-
age through the inhibition of apoptosis.
| 9 of 15
3.7 | Resveratrol reduced DNA break after
METH exposures
DNA break was further assessed for the hippocampus cells using
alkaline comet assay as a marker of brain oxidative apoptosis caused
by METH. The results in Figure 6a showed that DNA damages were
all increased remarkably compared to the control group post METH
treatment. Figure 6b and c demonstrated the percentage of tail DNA
and tail length were all increased remarkably after METH treatment
(tail DNA: p < .01; tail length: p < .01). Interestingly, DNA damage
was observably decreased in resveratrol (10 or 100 mg/kg) pre-
treatment groups. Comet experiment results showed that after the
supplement of resveratrol, both the percentage of tail DNA and tail
length decreased significantly (p < .01).
Immunoblotting of γ-H2AX, a marker of DNA strand break,
was carried out to further evaluate the protection of resveratrol
on DNA damage. METH significantly increased the protein expres-
sion of γ-H2AX in the hippocampus compared with the saline group
(Figure 5a and e, p < .05). In contrast, pretreatment with resvera-
trol (10 or 100 mg/kg) could significantly reduce the protein level of
γ-H2AX (p < .05). Therefore, we speculated that resveratrol could
significantly decrease the DNA break-induced by METH in the
hippocampus.
4 | D I S CU S S I O N
A growing number of studies have shown that METH abuse can lead
to cognitive impairment and memory deficits (Hajheidari et al., 2017;
Zuloaga et al., 2016), but the exact mechanism is still unclear. The
present study showed that multiple METH exposures impaired
memory function and caused long-lasting memory deficits without
affecting spontaneous locomotion. Meanwhile, pretreatment of an-
tioxidant resveratrol showed therapeutic effects on memory deficits
after repeated METH exposures in mice. Results of bioinformatics
analysis showed that resveratrol might alleviate METH-induced
memory deficits by inhibiting oxidative damage and apoptosis.
Molecular docking showed that resveratrol had hydrogen bonding
interactions with Keap1, the repressor protein of the Keap1-Nrf2
pathway. Further results validated oxidative stress, DNA break, and
apoptosis were significantly increased, while expressions of related
proteins in the Keap1-Nrf2 pathway were greatly decreased in the
hippocampus of mice after METH exposures. Furthermore, resvera-
trol pretreatment could reverse these changes caused by METH,
confirming the importance of oxidative stress and apoptosis in the
mechanism of cognitive impairment after METH exposure.
Previous studies have shown that METH addicts scored lower on
cognitive tests than healthy controls and exhibited overall memory
impairment (Ballard et al., 2015; Zhong et al., 2016), and memory defi-
cits could persist for several months or even years after abstinence
in some abusers (Iudicello et al., 2010; Zhong et al., 2016). Consistent
with these clinical studies, our results proved that repeated adminis-
tration of METH caused significant memory deficits after 7 days and
10 of 15 | ZENG et al.

22 days post-injection, indicating a long-lasting memory impairment et al., 2019; Wang et al., 2019; Zhang et al., 2019). Resveratrol has
behavior. Resveratrol is an effective natural product that could in- been reported to be protective against METH damage in cultured
hibit the generation of free radicals, reduce oxidative stress-induced cells in vitro, however, its effect on METH damage in vivo is also
neuronal damage, and ameliorate cognitive impairment (Gocmez worth further study. Interestingly, we found that resveratrol could
ZENG et al. | 11 of 15

F I G U R E 5 Resveratrol prevented METH-induced changes in oxidative stress-, DNA damage-, and apoptosis-related proteins in the
hippocampus. (a) Representative bands of Keap1, Nrf2, HO-1, GCSc-γ, Bax, Bcl-2, cleaved caspase-3, and γ-H2AX. (b) Nrf2 protein
expression in the nucleus of the hippocampus area was significantly decreased and resveratrol (10 or 100 mg/kg) pretreatment could
greatly increase the expression of nucleus Nrf2 protein expression. (c) Pretreatment with resveratrol (10 or 100 mg/kg) prevented METH-
induced decrease of HO-1 and GCSc-γ protein expression and increase of Keap1 expression in the hippocampus. (d) Bax protein expression
was significantly increased while Bcl-2 protein expression was significantly decreased after METH treatment. Resveratrol (10 or 100 mg/
kg) pretreatment could remarkably reverse these changes of protein expression induced by METH. (e) Pretreatment with resveratrol (10
or 100 mg/kg) prevented METH-induced increase of cleaved caspase-3 and γ-H2AX protein expression in the hippocampus. Data were
presented as means ± SEM. n = 3 for per group, *p < .05, **p < .01 compared to the Veh/Sal group, #p < .05, ##p < .01 compared to the Veh/
METH group

remarkably improve the ability of mice to discriminate novel object
in the NOR task, and the lack of alteration in the total distance in-
dicated that resveratrol enhances memory function without affect-
ing their locomotor activity. These findings proved that resveratrol
could be a potential candidate for treating METH-induced nerve in-
jury, however, whether the mechanism was related to its antioxidant
effect still need further experiments.
Bioinformatics analysis may offer a direction for the mechanistic
study of resveratrol. In the current study, we used the GeneCards
database to clarify the potential mechanism by which resveratrol
alleviated METH caused memory impairment. From the integrated
drug target prediction and GO analysis, resveratrol may exert its
neuroprotective effect on memory deficits induced by METH via
the apoptosis process, response to oxygen-containing compound,
which were characterized as the important mechanism of neurode-
generative progression. METH can enter dopaminergic neurons via
dopamine (DA) transporter, increase intracellular DA level, promote
the auto-oxidation of DA, and form various toxic substances such

F I G U R E 6 Resveratrol reduced DNA damage after METH exposure. (a) DNA damages were measured by comet assay, all the images
were taken at 200X magnification. The corresponding statistical analysis was shown in (b) and (c). Compared to the Veh/Sal group, the
percentage of tail DNA and tail length were all increased remarkably after METH treatment (all p < .01). Moreover, METH-induced DNA
damage was remarkably decreased in resveratrol (10 or 100 mg/kg) pretreatment groups. Data were presented as means ± SEM. n = 12 for
per group, **p < .01 compared to the Veh/Sal group, ##p < .01 compared to the Veh/METH group
12 of 15 | ZENG et al.
as reactive oxygen species, leading to oxidative stress (Krasnova &
Cadet, 2009). To further validate the postulation, serum GSH, SOD,
TAC, and MDA which may be responsible for impaired memory
were observed. Compared with the healthy control, MDA levels of
METH abusers were significantly increased and SOD activities were
decreased (Huang et al., 2013). Of importance, our results proved
that METH-induced memory impairment behavior in vivo was as-
sociated with an increased level of MDA and decline of GSH, SOD,
TAC in serum. Additionally, resveratrol pretreatment increased the
antioxidant capacity of mice, which was shown by the increase of
GSH, SOD, TAC, and decreased the lipid peroxidation of mice, which
was manifested by the decrease in MDA levels. The result of the
animal serum antioxidant capacity test verified the speculation of
bioinformatics analysis, indicating that reducing oxidative stress may
be involved in the protective effect of resveratrol on METH-induced
memory deficits.
The Keap1-Nrf2 pathway is the major regulator of cytoprotec-
tive responses to exogenous and endogenous stresses caused by
free radicals including reactive oxygen species (ROS) and electro-
philes. The key signaling proteins within the pathway are the Nrf2
that binds together with small Maf proteins to the antioxidant re-
sponse element (ARE) in the transcriptional regulatory regions of
FIGURE 7 Proposed molecular mechanism describing the protective effect of resveratrol on inhibiting METH-induced memory deficits
cytoprotective genes, such as HO-1, and GCSc-γ (He et al., 2020;
Taguchi et al., 2020), and Keap1, a repressor protein that binds to
Nrf2 and promotes its degradation by the ubiquitin-proteasome
pathway (Kansanen et al., 2013). Nrf2 stores in the cytoplasm as
an inactive form via binding to Keap1 and the sensitive amino acid
residues on Keap1 could combine with some stimulus, which will
cause the change of the conformation and lose the ability to bind to
Nrf2, then, Nrf2 can be activated after entering into the nucleus and
regulate the transcription of target genes (He et al., 2020; Taguchi
et al., 2020). Several studies documented that Nrf2-knockout mice
exhibit greater METH-initiated loss of dopaminergic terminals and
postnatal neurodevelopmental deficits (Ramkissoon & Wells, 2013,
2015). Therefore, the computational modeling approach was applied
to understand the molecular basis of the biological function of res-
veratrol. Our results indicated that the interaction energy between
resveratrol and Keap1 was −5.42 kcal/mol, and it formed three hy-
drogen-bonded interactions with Leu365, Leu 557, and Ile559 of
Keap1. Computational modeling through molecular docking based
binding study of interactions between resveratrol and Keap1 re-
vealed the potential role of resveratrol as an antagonist of Keap1.
Results of the immunoblotting analysis proved that multiple METH
exposures significantly increased the level of Keap1 and reduced
ZENG et al.
the levels of nucleus Nrf2 and its downstream antioxidant proteins
(HO-1 and GCSc-γ) in the hippocampus, and pretreatment of resver-
atrol could reverse the effects of METH. These results indicated that
resveratrol might bind to Keap1, release its inhibitory effect on Nrf2,
and promote the entry of Nrf2 into the nucleus and transcription of
downstream genes.
Previous studies have shown that oxidative stress caused DNA
damage response, contributing to the mechanism of memory impair-
ment, behavioral disorders, and other neurodegenerative diseases
(Coppede & Migliore, 2015; Dabrowska & Wiczkowski, 2017). As
a target molecule for free radicals, DNA was also oxidized by free
radicals caused by METH, resulting in DNA breaks and apoptosis
(Ru et al., 2019). Accordingly, we postulated that the increase of free
radicals caused by METH may cause severe damage to DNA and
lead to memory impairment. H2AX belongs to the histone family,
and its serine at position 139 is rapidly phosphorylated after DNA
double-strand breaks, and the appearance γ-H2AX (phosphorylated
H2AX) is concurrent with the initiation of DNA fragmentation (Singh
et al., 2015). Interestingly, we found that the percentage of tail DNA,
tail DNA length, and the expression of γ-H2AX were greatly elevated
after METH treatment. Resveratrol treatment significantly attenu-
ated the increase of tail DNA percentage and tail DNA length and
reduced the upregulation of γ-H2AX in the hippocampus compared
to that of the METH group. Therefore, these data proved that res-
veratrol could be a potential bioactive molecule with the capability
to protect the central nervous system from oxidative DNA damage.
It is now widely believed that neuronal apoptosis is a common
feature of the brain in patients with neurodegenerative diseases. The
previous report has shown that METH could significantly decrease
the expression of Bcl-2, increase the level of cleaved caspase-3 in the
prefrontal cortex, and lead to cognitive deficits (Long et al., 2017).
In agreement with this finding, we found that the expressions of
pro-apoptotic factor Bax and cleaved caspase-3 were significantly
increased while the expression of Bcl-2 (anti-apoptotic factor) was
significantly inhibited after repeated METH exposure. Moreover,
pretreatment with resveratrol significantly attenuated the downreg-
ulation of Bcl-2 in the hippocampus and the upregulation of Bax and
cleaved caspase-3. Therefore, we proposed the following putative
molecular cascade to explain the memory dysfunction induced by
METH. Multiple METH exposures induced the production of free
radicals, resulting in oxidative stress, inhibition of the Keap1-Nrf2
pathway, and an increase in DNA damage. This led to cell apopto-
sis that was responsible for memory deficits. Resveratrol pretreat-
ment significantly reduced free radical damage to cells, activated the
Keap1-Nrf2 pathway, decreased oxidative DNA break and apoptosis
in the hippocampus, and improved memory deficits caused by multi-
ple METH exposures (Figure 7).
5 | CO N C LU S I O N
In the present study, we demonstrated that METH exposures in
dose-escalation mode caused long-lasting memory deficits, and
| 13 of 15
resveratrol pretreatment significantly attenuated memory impair-
ment induced by METH. A new strategy of a combination of com-
putational prediction and experimental validation was adopted to
identify the underlying pharmacological mechanism. We demon-
strated that the supplement of resveratrol might effectively prevent
memory impairment via the interaction with Keap1, activation of the
Keap1-Nrf2 pathway, and inhibition of DNA damage and apoptotic
responses post METH exposure. To our knowledge, there were few
studies on the neuroprotective effects of resveratrol on METH-
induced nervous system injury. However, a minor pitfall of this study
is that only the NOR test was used. It will be important to use other
methods to consolidate these findings in our follow-up study. We
believed the potential therapeutic effects of resveratrol on METH-
induced nervous system injury may benefit from this study and fur-
ther studies, and these findings will provide the impetus for further
application of resveratrol in the nervous system.
AU T H O R C O N T R I B U T I O N S
Qing Zeng: Data curation; Investigation; Project administration;
Writing-original draft. Qi Xiong: Investigation; Writing-review & ed-
iting. Mei Zhou: Investigation; Software. Xiang Tian: Investigation;
Software. Kai Yue: Investigation. Yi Li: Investigation; Supervision. Xiji
Shu: Project administration; Supervision. Q Ru: Conceptualization;
Formal analysis; Funding acquisition; Investigation; Supervision;
Writing-original draft; Writing-review & editing.
DATA AVA I L A B I L I T Y S TAT E M E N T
The data that support the findings of this study are available on re-
quest from the corresponding author. The data are not publicly avail-
able due to privacy or ethical restrictions.
ORCID
Qin Ru https://orcid.org/0000-0002-2169-7471
REFERENCES
Ahmed, M., Alzoubi, K. H., & Khabour, O. F. (2020). Vitamin E prevents
the cognitive impairments in post-traumatic stress disorder rat
model: Behavioral and molecular study. Psychopharmacology (Berl),
237(2), 599–607. https://doi.org/10.1007/s0021​3-019-05395​-w
Alzoubi, K. H., Halboup, A. M., Alomari, M. A., & Khabour, O. F. (2019).
The neuroprotective effect of vitamin E on waterpipe tobacco
smoking-induced memory impairment: The antioxidative role. Life
Sciences, 222, 46–52. https://doi.org/10.1016/j.lfs.2019.02.050
Avila, J. A., Zanca, R. M., Shor, D., Paleologos, N., Alliger, A. A., Figueiredo-
Pereira, M. E., & Serrano, P. A. (2018). Chronic voluntary oral metham-
phetamine induces deficits in spatial learning and hippocampal protein
kinase Mzeta with enhanced astrogliosis and cyclooxygenase-2 levels.
Heliyon, 4(2), e00509. https://doi.org/10.1016/j.heliy​on.2018.e00509
Baghcheghi, Y., Beheshti, F., Shafei, M. N., Salmani, H., Sadeghnia, H. R.,
Soukhtanloo, M., Anaeigoudari, A., & Hosseini, M. (2018). The ef-
fects of vitamin E on brain derived neurotrophic factor, tissues oxida-
tive damage and learning and memory of juvenile hypothyroid rats.
Metabolic Brain Disease, 33(3), 713–724. https://doi.org/10.1007/
s1101​1-017-0176-0
Baird, L., & Yamamoto, M. (2020). The molecular mechanisms regulat-
ing the KEAP1-NRF2 pathway. Molecular and Cellular Biology, 40(13).
https://doi.org/10.1128/MCB.00099​-20
14 of 15 | ZENG et al.
Ballard, M. E., Weafer, J., Gallo, D. A., & de Wit, H. (2015). Effects of
acute methamphetamine on emotional memory formation in hu-
mans: Encoding vs consolidation. PLoS One, 10(2), e0117062. https://
doi.org/10.1371/journ​al.pone.0117062
Bastianetto, S., Menard, C., & Quirion, R. (2015). Neuroprotective action
of resveratrol. Biochimica Et Biophysica Acta, 1852(6), 1195–1201.
https://doi.org/10.1016/j.bbadis.2014.09.011
Casaletto, K. B., Obermeit, L., Morgan, E. E., Weber, E., Franklin, D. R.,
Grant, I., & Woods, S. P. (2015). Depression and executive dysfunc-
tion contribute to a metamemory deficit among individuals with
methamphetamine use disorders. Addictive Behaviors, 40, 45–50.
https://doi.org/10.1016/j.addbeh.2014.08.007
Chen, W. J., Du, J. K., Hu, X., Yu, Q., Li, D. X., Wang, C. N., Zhu, X. Y., &
Liu, Y. J. (2017). Protective effects of resveratrol on mitochondrial
function in the hippocampus improves inflammation-induced de-
pressive-like behavior. Physiology & Behavior, 182, 54–61. https://doi.
org/10.1016/j.physb​eh.2017.09.024
Cherner, M., Suarez, P., Casey, C., Deiss, R., Letendre, S., Marcotte,
T., Vaida, F., Atkinson, J. H., Grant, I., & Heaton, R. K. (2010).
Methamphetamine use parameters do not predict neuropsycholog-
ical impairment in currently abstinent dependent adults. Drug and
Alcohol Dependence, 106(2–3), 154–163. https://doi.org/10.1016/j.
druga​lcdep.2009.08.010
Coppede, F., & Migliore, L. (2015). DNA damage in neurodegenerative
diseases. Mutation Research, 776, 84–97. https://doi.org/10.1016/j.
mrfmmm.2014.11.010
Corpas, R., Grinan-Ferre, C., Rodriguez-Farre, E., Pallas, M., & Sanfeliu,
C. (2019). Resveratrol induces brain resilience against Alzheimer
neurodegeneration through proteostasis enhancement. Molecular
Neurobiology, 56(2), 1502–1516. https://doi.org/10.1007/s1203​
5-018-1157-y
Dabrowska, N., & Wiczkowski, A. (2017). Analytics of oxidative stress
markers in the early diagnosis of oxygen DNA damage. Advances in
Clinical and Experimental Medicine: Official Organ Wroclaw Medical
University, 26(1), 155–166. https://doi.org/10.17219/​acem/43272
Feng, L., Yasmeen, R., Schoene, N. W., Lei, K. Y., & Wang, T. T. Y. (2019).
Resveratrol differentially modulates immune responses in human
THP-1 monocytes and macrophages. Nutrition Research, 72, 57–69.
https://doi.org/10.1016/j.nutres.2019.10.003
Freese, L., Almeida, F. B., Heidrich, N., Hansen, A. W., Steffens, L.,
Steinmetz, A., Moura, D. J., Gomez, R., & Barros, H. M. T. (2018).
Environmental enrichment reduces cocaine neurotoxicity during
cocaine-conditioned place preference in male rats. Pharmacology,
Biochemistry and Behavior, 169, 10–15. https://doi.org/10.1016/j.
pbb.2018.04.001
Gocmez, S. S., Sahin, T. D., Yazir, Y., Duruksu, G., Eraldemir, F. C., Polat, S.,
& Utkan, T. (2019). Resveratrol prevents cognitive deficits by atten-
uating oxidative damage and inflammation in rat model of streptozo-
tocin diabetes induced vascular dementia. Physiology & Behavior, 201,
198–207. https://doi.org/10.1016/j.physb​eh.2018.12.012
Hajheidari, S., Miladi-Gorji, H., & Bigdeli, I. (2017). Environmental en-
richment prevents methamphetamine-induced spatial memory defi-
cits and obsessive-compulsive behavior in rats. Iranian Journal of
Psychiatry, 12(1), 8–14.
He, Y., Jiang, K., & Zhao, X. (2020). Taraxasterol protects hippocampal
neurons from oxygen-glucose deprivation-induced injury through
activation of Nrf2 signalling pathway. Artificial Cells, Nanomedicine,
and Biotechnology, 48(1), 252–258. https://doi.org/10.1080/21691​
401.2019.1699831
Hosseinzadeh, M., Alizadeh, A., Heydari, P., Kafami, M., Hosseini, M.,
Beheshti, F., Marefati, N., & Ghanbarabadi, M. (2020). Effect of vi-
tamin E on cisplatin-induced memory impairment in male rats. Acta
Neuropsychiatrica, 1–6. https://doi.org/10.1017/neu.2020.34
Huang, M. C., Lin, S. K., Chen, C. H., Pan, C. H., Lee, C. H., & Liu, H. C. (2013).
Oxidative stress status in recently abstinent methamphetamine
abusers. Psychiatry and Clinical Neurosciences, 67(2), 92–100. https://
doi.org/10.1111/pcn.12025
Iudicello, J. E., Woods, S. P., Vigil, O., Scott, J. C., Cherner, M., Heaton, R.
K., Atkinson, J. H., Grant, I., & the HIV Neurobehavioral Research Ce.
(2010). Longer term improvement in neurocognitive functioning and
affective distress among methamphetamine users who achieve sta-
ble abstinence. Journal of Clinical and Experimental Neuropsychology,
32(7), 704–718. https://doi.org/10.1080/13803​39090​3512637
Kansanen, E., Kuosmanen, S. M., Leinonen, H., & Levonen, A. L. (2013).
The Keap1-Nrf2 pathway: Mechanisms of activation and dysregu-
lation in cancer. Redox Biology, 1, 45–49. https://doi.org/10.1016/j.
redox.2012.10.001
Kanthasamy, K., Gordon, R., Jin, H., Anantharam, V., Ali, S., Kanthasamy,
A. G., & Kanthasamy, A. (2011). Neuroprotective effect of res-
veratrol against methamphetamine-induced dopaminergic apop-
totic cell death in a cell culture model of neurotoxicity. Current
Neuropharmacology, 9(1), 49–53. https://doi.org/10.2174/15701​
59117​95­017353
Khajevand-Khazaei, M. R., Ziaee, P., Motevalizadeh, S. A., Rohani, M.,
Afshin-Majd, S., Baluchnejadmojarad, T., & Roghani, M. (2018).
Naringenin ameliorates learning and memory impairment following
systemic lipopolysaccharide challenge in the rat. European Journal
of Pharmacology, 826, 114–122. https://doi.org/10.1016/j.ejphar.
2018.03.001
Krasnova, I. N., & Cadet, J. L. (2009). Methamphetamine toxicity and
messengers of death. Brain Research Reviews, 60(2), 379–407. https://
doi.org/10.1016/j.brain​resrev.2009.03.002
Kumar, P., Kumar, M., Wisdom, K. S., Pathakota, G. B., Nayak, S. K., Reang,
D., Nagpure, N. S., & Sharma, R. (2020). Characterization, docking and
molecular dynamics simulation of gonadotropin-inhibitory hormone
receptor (GnIHR2) in Labeo Catla. Cellular Physiology and Biochemistry,
54(5), 825–841. https://doi.org/10.33594/​0 0000​0272
Li, B., Chen, R., Chen, L., Qiu, P., Ai, X., Huang, E., Huang, W., Chen, C.,
Liu, C., Lin, Z., Xie, W. B., & Wang, H. (2017). Effects of DDIT4 in
methamphetamine-induced autophagy and apoptosis in dopaminer-
gic neurons. Molecular Neurobiology, 54(3), 1642–1660. https://doi.
org/10.1007/s1203​5-015-9637-9
Lin, Y. T., Wu, Y. C., Sun, G. C., Ho, C. Y., Wong, T. Y., Lin, C. H., Chen,
H. H., Yeh, T. C., Li, C. J., Tseng, C. J., & Cheng, P. W. (2018). Effect
of resveratrol on reactive oxygen species-induced cognitive impair-
ment in rats with angiotensin II-induced early Alzheimer's disease
(dagger). Journal of Clinical Medicine, 7(10). https://doi.org/10.3390/
jcm71​0 0329
Long, J. D., Liu, Y., Jiao, D. L., Wang, Y. J., Zan, G. Y., Ju, Y. Y., Zhao, M., &
Liu, J. G. (2017). The neuroprotective effect of memantine on meth-
amphetamine-induced cognitive deficits. Behavioral Brain Research,
323, 133–140. https://doi.org/10.1016/j.bbr.2017.01.042
Maroso, M., Szabo, G. G., Kim, H. K., Alexander, A., Bui, A. D., Lee, S.
H., Lutz, B., & Soltesz, I. (2016). Cannabinoid control of learning and
memory through HCN channels. Neuron, 89(5), 1059–1073. https://
doi.org/10.1016/j.neuron.2016.01.023
Pingaew, R., Mandi, P., Prachayasittikul, V., Prachayasittikul, S.,
Ruchirawat, S., & Prachayasittikul, V. (2018). Synthesis, molecular
docking, and QSAR study of sulfonamide-based indoles as aromatase
inhibitors. European Journal of Medicinal Chemistry, 143, 1604–1615.
https://doi.org/10.1016/j.ejmech.2017.10.057
Ramkissoon, A., & Wells, P. G. (2013). Developmental role of nuclear fac-
tor E2-related factor 2 in mitigating methamphetamine fetal toxic-
ity and postnatal neurodevelopmental deficits. Free Radical Biology
and Medicine, 65, 620–631. https://doi.org/10.1016/j.freer​adbio​
med.2013.07.043
Ramkissoon, A., & Wells, P. G. (2015). Methamphetamine oxidative
stress, neurotoxicity, and functional deficits are modulated by nu-
clear factor-E2-related factor 2. Free Radical Biology and Medicine, 89,
358–368. https://doi.org/10.1016/j.freer​adbio​med.2015.07.157
ZENG et al.
Rauf, A., Imran, M., Suleria, H. A. R., Ahmad, B., Peters, D. G., & Mubarak,
M. S. (2017). A comprehensive review of the health perspectives
of resveratrol. Food & Function, 8(12), 4284–4305. https://doi.
org/10.1039/C7FO0​1300K
Reichel, C. M., Gilstrap, M. G., Ramsey, L. A., & See, R. E. (2014). Modafinil
restores methamphetamine induced object-in-place memory deficits
in rats independent of glutamate N-methyl-D-aspartate receptor
expression. Drug and Alcohol Dependence, 134, 115–122. https://doi.
org/10.1016/j.druga​lcdep.2013.09.018
Rigon, R. B., Fachinetti, N., Severino, P., Durazzo, A., Lucarini, M.,
Atanasov, A. G., Mamouni, S. E., Chorilli, M., Santini, A., & Souto,
E. B. (2019). Quantification of trans-resveratrol-loaded solid
lipid nanoparticles by a validated reverse-phase HPLC photodi-
ode array. Applied Sciences, 9(22), 4961. https://doi.org/10.3390/
app92​24961
Ru, Q., Xiong, Q., Tian, X., Chen, L., Zhou, M., Li, Y., & Li, C. (2019). Tea
polyphenols attenuate methamphetamine-induced neuronal dam-
age in PC12 cells by alleviating oxidative stress and promoting DNA
repair. Frontiers in Physiology, 10, 1450. https://doi.org/10.3389/
fphys.2019.01450
Sharma, V., Kaur, A., & Singh, T. G. (2020). Counteracting role of nu-
clear factor erythroid 2-related factor 2 pathway in Alzheimer's
disease. Biomedicine & Pharmacotherapy, 129, 110373. https://doi.
org/10.1016/j.biopha.2020.110373
Silva, C. D., Neves, A. F., Dias, A. I., Freitas, H. J., Mendes, S. M., Pita,
I., Viana, S. D., de Oliveira, P. A., Cunha, R. A., Fontes Ribeiro, C.
A., Prediger, R. D., & Pereira, F. C. (2014). A single neurotoxic dose
of methamphetamine induces a long-lasting depressive-like be-
haviour in mice. Neurotoxicity Research, 25(3), 295–304. https://doi.
org/10.1007/s1264​0 -013-9423-2
Singh, I., Ozturk, N., Cordero, J., Mehta, A., Hasan, D., Cosentino, C.,
Sebastian, C., Kruger, M., Looso, M., Carraro, G., Bellusci, S., Seeger,
W., Braun, T., Mostoslavsky, R., & Barreto, G. (2015). High mobil-
ity group protein-mediated transcription requires DNA damage
marker gamma-H2AX. Cell Research, 25(7), 837–850. https://doi.
org/10.1038/cr.2015.67
Sun, D., Yue, Q., Guo, W., Li, T., Zhang, J., Li, G., Liu, Z., & Sun, J. (2015).
Neuroprotection of resveratrol against neurotoxicity induced by
methamphetamine in mouse mesencephalic dopaminergic neu-
rons. BioFactors, 41(4), 252–260. https://doi.org/10.1002/biof.
1221
Taguchi, K., Izumi, Y., Takada-Takatori, Y., Akaike, A., & Kume, T. (2020).
Protective effect of 2',3'-Dihydroxy-4',6'-dimethoxychalcone
on glutamate-induced neurotoxicity in primary cortical cultures.
Biological &/and Pharmaceutical Bulletin, 43(1), 184–187. https://doi.
org/10.1248/bpb.b19-00718
Thompson, P. M., Hayashi, K. M., Simon, S. L., Geaga, J. A., Hong, M.
S., Sui, Y., Lee, J. Y., Toga, A. W., Ling, W., & London, E. D. (2004).
Structural abnormalities in the brains of human subjects who use
methamphetamine. Journal of Neuroscience, 24(26), 6028–6036.
https://doi.org/10.1523/JNEUR​OSCI.0713-04.2004
| 15 of 15
Wahl, D., Bernier, M., Simpson, S. J., de Cabo, R., & le Couteur, D. G.
(2018). Future directions of resveratrol research. Nutrition and
Healthy Aging, 4(4), 287–290. https://doi.org/10.3233/NHA-170035
Wang, B., Ge, S., Xiong, W., & Xue, Z. (2018). Effects of resveratrol pre-
treatment on endoplasmic reticulum stress and cognitive function
after surgery in aged mice. BMC Anesthesiology, 18(1), 141. https://
doi.org/10.1186/s1287​1-018-0606-5
Wang, N., He, J., Pan, C., Wang, J., Ma, M., Shi, X., & Xu, Z. (2019).
Resveratrol activates autophagy via the AKT/mTOR signaling path-
way to improve cognitive dysfunction in rats with chronic cere-
bral hypoperfusion. Frontiers in Neuroscience, 13, 859. https://doi.
org/10.3389/fnins.2019.00859
Weber, E., Blackstone, K., Iudicello, J. E., Morgan, E. E., Grant, I., Moore,
D. J., & Woods, S. P. (2012). Neurocognitive deficits are associated
with unemployment in chronic methamphetamine users. Drug and
Alcohol Dependence, 125(1–2), 146–153. https://doi.org/10.1016/j.
druga​lcdep.2012.04.002
Yeung, A. W. K., Aggarwal, B. B., Erdoganorhan, I., Horbańczuk, O. K.,
Nikolay, T. T., & Atanasov, A. G. (2019). Resveratrol, a popular dietary
supplement for human and animal health: Quantitative research lit-
erature analysis—A review. Animal Science Papers and Reports, 37(2),
103–118.
Zhang, Y., Li, Y., Wang, Y., Wang, G., Mao, L., Zhang, D., & Wang, J. (2019).
Effects of resveratrol on learning and memory in rats with vascular
dementia. Molecular Medicine Reports, 20(5), 4587–4593. https://doi.
org/10.3892/mmr.2019.10723
Zhong, N., Jiang, H., Du, J., Zhao, Y., Sun, H., Xu, D., Li, C., Zhuang,
W., Li, X., Hashimoto, K., & Zhao, M. (2016). The cognitive impair-
ments and psychological wellbeing of methamphetamine depen-
dent patients compared with health controls. Progress in Neuro-
Psychopharmacology and Biological Psychiatry, 69, 31–37. https://doi.
org/10.1016/j.pnpbp.2016.04.005
Zulkipli, N. N., Zakaria, R., Long, I., Abdullah, S. F., Muhammad, E. F.,
Wahab, H. A., & Sasongko, T. H. (2020). In silico analyses and cyto-
toxicity study of asiaticoside and asiatic acid from Malaysian plant
as potential mTOR inhibitors. Molecules, 25(17), 3991. https://doi.
org/10.3390/molec​ules2​5173991
Zuloaga, D. G., Wang, J., Weber, S., Mark, G. P., Murphy, S. J., & Raber,
J. (2016). Chronic methamphetamine exposure prior to middle cere-
bral artery occlusion increases infarct volume and worsens cognitive
injury in Male mice. Metabolic Brain Disease, 31(4), 975–981. https://
doi.org/10.1007/s1101​1-016-9808-z
How to cite this article: Zeng Q, Xiong Q, Zhou M, et al.
Resveratrol attenuates methamphetamine-induced memory
impairment via inhibition of oxidative stress and apoptosis in
mice. J Food Biochem. 2021;45:e13622. https://doi.
org/10.1111/jfbc.13622