University of Southern California Dissertations and Theses

DHM reduces cognitive impairment induced by social isolation and ethanol consumption

(USC Thesis Other)

DHM reduces cognitive impairment induced by social isolation and ethanol consumption

PDF

Download

Open document

Flip pages

Download a page range

Download transcript

Copy asset link

Request this asset

Transcript (if available)

Content

DHM Reduces Cognitive Impairment Induced by Social Isolation and Ethanol
Consumption

by

Zidan Yuan

A Thesis Presented to the
FACULTY OF THE USC ALFRED E. MANN SCHOOL OF PHARMACY AND
PHARMACEUTICAL SCIENCES
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
MASTER OF SCIENCE
(CLINICAL AND EXPERIMENTAL THERAPEUTICS)

May 2023

2023 Zidan Yuan

ii

Acknowledgements

Time is cracking on. I will be graduating from the beautiful USC School of Pharmacy
soon to complete my master's degree. The memories of the past two years rushed into
my mind.
First of all, I want to express my sincere gratitude to my supervisor, Dr. Jing Liang. I
spent two enjoyable years in this great lab, learning many experimental skills and
research theories. With her help, I successfully completed my thesis. I am extremely
grateful to lab member Saki Watanabe for leading me in my experimental research
and giving me a lot of support and help. I also like to thank Dr. Liana Asatryan and
Dr. Stan Louie for reviewing my thesis carefully and providing me with useful
suggestion in writing thesis. I wish to thank all the lovely people I met at USC.
I would like to thank my parents. They have selflessly provided me with emotional
and financial support. Their words of advice and whispers are the loudest trumpets in
the fog. I also want to thank my good friends. Every day studying and preparing for
exams together, every chat, every trip, are the memories I cherish the most.
Finally, I would like to thank myself. I would like to thank myself for setting high
standards for myself from the time I entered school and trying to do it from the
beginning to the end; I would like to thank myself for the days when I was confused
about the future, but I was able to make the right decision. I will continue to work
hard to become a better person in the future.
Happy graduation!

iii

Table of Contents
Acknowledgements ................................................................................................................... ii
List of Figures .......................................................................................................................... iv
Abstract ..................................................................................................................................... v
Chapter 1 Background ............................................................................................................... 1
1.1 Anxiety ............................................................................................................................ 1
1.1.1 Introduction .............................................................................................................. 1
1.1.2 Current drugs for anxiety disorders .......................................................................... 2
1.2 Alcohol use disorders (AUD) .......................................................................................... 6
1.2.1 Introduction .............................................................................................................. 6
1.2.2 AUD and GABA ...................................................................................................... 7
1.3 GABA A receptors ............................................................................................................ 9
1.4 Therapeutic role of DHM .............................................................................................. 11
1.5 Alzheimer's disease and DHM ...................................................................................... 13
1.6 Goals and specific aims ................................................................................................. 15
2.1 Materials ........................................................................................................................ 16
2.2 Methods ......................................................................................................................... 18
2.2.1 Behavior Test .......................................................................................................... 18
Novel object recognition (NOR) ............................................................................. 18
Novel context recognition test (NCR) ..................................................................... 19
Elevated plus maze (EPM) ...................................................................................... 21
2.2.2 Protein extraction and Western blot analysis.......................................................... 22
2.2.3 Statistical analysis .................................................................................................. 22
Chapter 3 Results ..................................................................................................................... 24
3.1 DHM reverted cognition and memory decline caused by Social isolation, EtOH
treatment .............................................................................................................................. 24
3.2 Western Blot Analysis ................................................................................................... 26
3.3 DHM reduces alcohol preference in mice ..................................................................... 27
Discussion ............................................................................................................................... 29
Study limitations ...................................................................................................................... 33
References ............................................................................................................................... 34

iv

List of Figures
Figure 1. The scheme of a GABA-A receptor ............................................................. 10
Figure 2. Plant origin and chemical structure of DHM ............................................... 11
Figure 3. Experimental timeline .................................................................................. 17
Figure 4. Schematic diagram of the experimental design (NOR) ................................ 18
Figure 5. Schematic diagram of the experimental design (NCR) ................................ 19
Figure 6. Novel object recognition tests. ..................................................................... 24
Figure 7. Novel context recognition test ...................................................................... 25
Figure 8. Representative blots for Phosphorylated Tau protein ................................... 26
Figure 9. Relative values of phosphorylated Tau protein expression were quantified 26
Figure 10. Preference for water/alcohol in mice at six weeks ..................................... 27
Figure 11. Body weight changes in mice over six weeks ............................................ 28

v

Abstract

Social isolation is a stress-responsive stimulus that is associated with the etiology of
anxiety-related disorders. Chronic stress is a common precursor to cognitive and
memory impairment associated with Alzheimer's disease (AD). When coupled with
psychosomatic disease and chronic ethanol (EtOH) intake, this rate of
progression/decline can be greatly increased. Interestingly, since the COVID-19
pandemic, studies have demonstrated an upward trend in social isolation, psychological
distress, and alcohol sales. We found that γ-aminobutyric acid A receptors (GABAARs)
play a similar regulatory role in anxiety disorders, Alzheimer's disease, and alcohol use
disorders. As a positive allosteric modulator of GABAARs, dihydromyricetin (DHM)
has emerged as a highly promising therapeutic for development because of its ability to
reduce social isolation (SI)-induced anxiety and EtOH-related impairments.

The study presented in this thesis aims to explore the effects of social isolation, long-
term EtOH consumption, and DHM treatment on cognition in mice. We established an
SI-induced anxiety + EtOH drinking model consisting of 8-week-old male C57BL/6
mice. Behaviorally, DHM-treated mice exhibited improved cognitive performance and
reduced anxiety levels. Pathologically, DHM significantly reduced the expression of
phosphorylated tau protein in the mouse brain. DHM was shown to potentially reverse
or prevent the cognitive and memory decline induced by social isolation and long-term
EtOH consumption, exert anxiolytic effects and have possible therapeutic effects on

vi

AD, consistent with our expectations. The findings in this study further suggests the
need for exploring the intrinsic mechanism of action of DHM.

1

Chapter 1 Background
1.1 Anxiety
1.1.1 Introduction
Anxiety, a psychiatric disorder in which the patient feels distinctly fearful and anxious
about the future, usually leads to tachycardia, sleepless nights, nervousness, and
irritability, as well as shortness of breath, frequent urination, and extreme weakness.
Complications such as depression may also present in anxiety[1]. Currently,
approximately 12% of the global population has an anxiety disorder, with women
having twice the prevalence of men [2]. Anxiety disorders include several types of
conditions: generalized anxiety disorder, social anxiety disorder (social phobia), object-
specific phobia, and separation anxiety disorder. Of these, social anxiety disorder
accounts for about 10% of all types of anxiety. Anxiety disorders are long-term, harmful
illnesses that occur at any time during a person's life, causing extreme personal suffering
and a huge economic and social burden [3].

Treatment for anxiety disorders includes the use of anti-anxiety medications such as
benzodiazepines and psychotherapy such as exposure therapy, cognitive therapy and
cognitive behavioral therapy (CBT) [4]. Although the specific mechanisms behind
various types of anxiety disorders have not been fully elucidated, the role of the GABA
system in the mechanism is widely recognized. Most anxiolytic drugs also target the

2

GABA system for modulation, including benzodiazepines, which are allosteric
modulators of GABAARs, and gabapentin, which targets GABA metabolism to increase
brain GABA levels. However, their clinical use is limited by poor selectivity for GABA
subtypes and unavoidable side effects such as sedation and cognitive impairment. In
the last two decades, no anxiolytic drug with a novel mechanism has been approved
and marketed for the treatment of anxiety disorders. In previous studies, our team has
demonstrated that DHM is a positive modulator of GABAARs at the BZ site without
tolerance or addiction potential, suggesting that it has the potential to modulate anxiety-
like behaviors through GABA receptors [5].

1.1.2 Current drugs for anxiety disorders

Selective serotonin reuptake inhibitors (SSRIs) and serotonin-noradrenaline
reuptake inhibitors (SNRIs)
SSRIs and SNRIs belong to the monoamine class of drugs. Because of their excellent
benefit/risk balance, they are now widely used to treat major depressive disorder and
related psychiatric disorders, including anxiety disorders.
SSRIs include citalopram, escitalopram, fluoxetine, and fluvoxamine. The mechanism
of action of SSRIs is to block neuronal reuptake (reuptake) of serotonin and increase
the level of serotonin (5-HT) in the brain, which would allow more serotonin for
neurotransmission. Although SSRIs act selectively on the 5-HT system, they are not
specific for various 5-HT receptor isoforms. They act on 5-HT1 receptors to exert

3

therapeutic effects, but also affect 5-HT2 receptors and 5-HT3 receptors, causing side
effects such as anxiety, nausea, and headache. So, SSRIs both relieve anxiety on the
one hand and cause anxiety on the other [6].
SNRIs include desvenlafaxine, duloxetine, levomilnacipran, and milnacipran. These
drugs block the reuptake of the neurotransmitters serotonin and norepinephrine (NA)
in the brain. SNRIs act on two receptors to ensure its effectiveness and are rather
insensitive to other receptors limiting its adverse effects [7].
Both SSRIs and SNRIs take 2 to 4 weeks after dosing for noticeable effects and have a
wide therapeutic window [4]. Based on the need for long-term use and non-immediate
relief, many patients do not achieve remission. Moreover, they are easily tolerated, and
withdrawal reactions may occur after cessation of use.

Tricyclic antidepressants (TCA)
Tricyclic antidepressants include promethazine, clomipramine, and amitriptyline.
TCAs exert their therapeutic effects by blocking the reuptake of NA and 5-HT by
norepinephrine (NA)-ergic and 5-hydroxytryptamine (5-HT)-ergic nerve endings and
increasing the concentration of monoamine transmitters in the synaptic gap. This class
of drugs was once the drug of first choice, but its clinical use was limited by frequent
and severe side effects such as cardiovascular toxicity, which led to a narrow safety
range [8]. They are now generally used after SSRIs and SNRIs.

4

Benzodiazepines
Benzodiazepines (BZs) are widely used anxiolytics, with about 55-94% of patients with
anxiety disorders in the United States receiving treatment with this class of drugs,
including diazepam, clozapine, and others. Unlike the first two classes of drugs,
benzodiazepines are not used to treat depression, but are used as sedative-hypnotics and
anxiolytics [9]. Its target of action is the inhibitory neurotransmitter GABA, which
causes central depression by inducing enhanced opening of GABA receptor-coupled
chloride channels, inhibiting postsynaptic potentials, and reducing central firing of
certain important neurons such as norepinephrine. Therefore, drug efficacy is correlated
with the level of GABA in the body. However, the inhibitory effect on the central
nervous system also leads to sequelae such as fatigue and impaired cognitive function.
Importantly, the long-term use may lead to addiction of these drugs. Therefore, BZs are
generally not used as a first-line therapeutic agents [10].

Other drugs
In addition to these major classes of anxiolytics, there are also some anxiolytics such
as buspirone and monoamine oxidase (MAOI) inhibitors. As the first antidepressants to
be developed, MAOIs were effective, but have usually been replaced by safer
antidepressants with fewer side effects. Its use often requires dietary restrictions and
avoidance of interactions with other medications, and may pose a risk of suicide.
Buspirone has been shown to be used to treat generalized anxiety [11], but its clinical
use is limited. Overall, due to receptor selectivity and the specific structure of the brain,

5

anti-anxiety medications, despite their variety, are still not well suited for long-term
widespread use in the treatment of this disorder. In clinical practice, the use of
medications is adjusted according to the patient's age, the specific type of anxiety
disorder, and the clinical response.

6

1.2 Alcohol use disorders (AUD)
1.2.1 Introduction
Alcohol use disorder (AUD) is one of the most prevalent mental disorders worldwide
[12], characterized by compulsive heavy drinking and loss of control over alcohol
intake behaviors, and with a tendency to become more severe and escalate over time,
even leading to considerable disability [13] and impairment such as cirrhosis of the liver
[14]. According to the World Health Organization, more than 3 million people died from
harmful use of alcohol in 2016, accounting for one-twentieth of all deaths. Males are
the main affected population with a prevalence rate approximately five times higher
than that of females [15]. In addition, socioeconomic status also influences the
epidemiology of AUD; with those of low socioeconomic status generally being at
greater risk than those of high socioeconomic status [16].

During the COVID-19 epidemic, studies have demonstrated that increased financial
hardship, psychological distress from social isolation, uncertainty about the future, and
inadequate and redistributed human resources for health may lead to increased alcohol
intake and trigger further exacerbation of liver disease [17]. Despite the enormous
emotional and economic burden that this disorder has imposed on individuals, families,
and society, the treatment for this disorder remains unsatisfactory. In clinical practice,
reducing alcohol intake is the most important aim of treatment [18], and studies have
shown that this action reduces subsequent disease and mortality.

7

Current FDA-approved drugs for AUD include naltrexone, acamprosate, and disulfiram,
and extended-release injectable naltrexone. Benzodiazepines are generally not
recommended for AUD because of their addictive nature and lack of safety and other
side effects [19]. While naltrexone is therapeutic [20], it has side effects such as
withdrawal symptoms, nausea, irritability, and fatigue, as well as impaired thinking and
anxiety. Acamprosate is more effective in promoting alcohol abstinence and less
effective in treating AUD [21]. Compliance with disulfiram is poor [22], and
acetaldehyde buildup can cause many unpleasant side effects. Thus, more useful AUD
drugs with fewer side effects is needed.

1.2.2 AUD and GABA
Studies have confirmed that EtOH acts at many neurotransmitter systems after crossing
the blood-brain barrier, including GABA, acetylcholine, and glutamate. GABA, the
primary inhibitory neurotransmitter in the brain, may be the source of alcohol's benefits
[23], bringing about sedative and tranquilizing effects, which rebound after withdrawal
[24]. While there are many different receptors for GABA, scientists are gradually
discovering that ethanol acts by mediating GABAA receptors and that GABAA subtypes
respond differently to different concentrations [25] and different times of exposure to
alcohol. This difference is due to the fact that EtOH acts on certain isoforms of
GABAARs, thus inducing alterations in the assembly of GABAA subunits and leading
to functional differences in GABAARs [26].

8

In past studies, we demonstrated that a single, acute drink of EtOH (5-10 mmol/L)
increases the exposure to GABAARs and acts as a sedative, and this exposure is
accompanied by changes in the levels of α4, β3 and δ isoforms [27]. Per NIAAA
guidelines, acute social drinking means less than four drinks a day and less than 14
drinks a week for men, and less than three drinks a day and less than seven drinks a
week for women. Animal studies have shown that this level of acute alcohol
consumption leads to increases in GABAAR α4/δ isoforms in the hippocampus of
mice. As rat models of chronic intermittent EtOH (CIE) with chronic and repeated
EtOH consumption exhibit increased anxiety and impaired memory, we proposed that
chronic EtOH exposure causes alteration in GABAARs plasticity, with a decrease in
α1-containing receptors and an increase in α4-containing receptors [28]. All these
findings suggest that changes in GABAAR subunit composition are a potential
mechanism for EtOH behavior.

9

1.3 GABAA receptors
The GABA receptor, which was first discovered in humans in the 1970s, is an ionotropic
receptor and is a class of ligand-gated ion channel [29]. The endogenous ligand for this
channel is a neurotransmitter known as GABA.
GABA is an inhibitory neurotransmitter formed by the removal of the carboxyl group
at the alpha position from the precursor glutamate by the action of glutamic acid
decarboxylase. Scientific studies have found that most psychiatric disorders including
anxiety disorders, depression, epilepsy, and Parkinson's are associated with reduced
levels of GABA in the brain [30].
GABA receptors have three subtypes, namely GABAA receptors, GABAB receptors,
and GABAC receptors. Among them, GABAA receptors are pentagonal heterogeneous
polypeptide oligomers composed of five subunits embedded in the dual lipid-like
protoplasmic membrane of neuronal cells, which in turn possess multiple isoforms.
GABA acts by activating GABAA receptors on neuronal cell membranes, opening
chloride channels, leading to an intracellular flow of chloride ions, generating inhibitory
postsynaptic potentials, and suppressing neuronal excitation [31].
Usually, many GABAARs contain two α subunits, two β subunits and one γ subunit.
Each subunit has several isoforms and consists of a family of 19 related isoforms. Six
alpha (alpha1-6), three beta (beta1-3), three gamma (gamma1-3), three ρ (rho1-3) and
one δ (delta), ε (epsilon), π (pi) and θ (theta) [32][33], which lead to the production of
a large number of receptor isoforms, constitute an extremely complex pharmacological
and physiological functions. The pharmacological interactions between BZDs and

10

GABA(A) receptors occur at different sites, between α and γ subunits and between α
and β subunits, respectively (Figure 1) [34].

Figure 1. The scheme of a GABAA receptor [34].

As such, understanding the relationship between GABAA receptor defects and
partitioning, as well as CNS disorders, will have significant implications for the
discovery of disease pathogenesis as well as drug development.

11

1.4 Therapeutic role of DHM
Dihydromyricetin[(2R,3R)-3,5,7-trihydroxy-2-(3,4,5-trihydroxyphenyl)-2,3-
dihydrochromen-4-one (DHM)] is a flavonoid compound of herbal medicines which
extracted from Ratan tea and Hovenia dulcis.

Figure 2.Plant origin and chemical structure of DHM.

DHM is currently used primarily as an herbal supplement for headaches and
hangovers, helping people cope with the acute and long-term effects of alcohol, as
well as having hepatoprotective properties. Our team has confirmed in previous
studies that DHM works primarily by reducing EtOH concentration and increasing the
amount and efficiency of alcohol dehydrogenase and acetaldehyde dehydrogenase,
making it easier for the body to metabolize and then eliminate. However, a more
importantly, DHM has been shown to be a positive allosteric modulator (PAM)
capable of increasing GABAA receptor (GABAAR) activity. These properties can be
used to manage acute alcoholism and alcohol withdrawal symptoms (anxiety) [5].
In addition, we have previously demonstrated that DHM can reduce anxiety in social

12

isolation (SI)-induced anxiety mice by restoring GABAAR function, gephyrin
expression, and adenosine triphosphate (ATP) levels [35]. These beneficial effects
demonstrate the therapeutic potential of DHM, which warrants further more detailed
investigation of the molecular mechanism for DHM action.

13

1.5 Alzheimer's disease and DHM
Alzheimer's disease (AD) is the leading cause of dementia, a progressive
neurodegenerative disease that affects millions of people worldwide and imposes a
huge socioeconomic burden on all societies. The prevalence of AD doubles every 5
years after age 60, from 1% of those aged 60 to 64 to 40% of those aged 85 and older
[36]. It is more prevalent in women than in men. Alzheimer’s disease is characterized
by the deposition of β-amyloid peptide (Aβ) in the brain and neurogenic fiber tangles
of hyperphosphorylated tau protein [37]. Biomarkers such as β-amyloid peptide
(Aβ42), total tau protein, and the amount of phosphorylated tau protein can reflect the
pathology of AD and help determine the condition [38]. Studies have shown that these
markers can be used to detect the condition of AD patients over time and as markers
of treatment efficacy [39].
Currently, donepezil, galantamine, Exelon and aducanumab are approved by the FDA
for the treatment of AD. In July 2021, FDA accepted Eisai’s Biologics License
Application (BLA) for aducanumab under the accelerated approval pathway and
granted Priority Review, which is an anti-Aβ protofibril antibody, it has shown
clinical demonstration of positive outcomes and is expected to be the first
symptomatic AD drug [40].

In the team's past work, we use transgenic mouse model of AD demonstrated that
while GABAAR γ2 subunit expression was not significantly altered, GABA-mediated
mIPSC frequency, amplitude, total charge transfer was decreased and tonic currents

14

were reduced. This represents an impaired transmission function of the GABA
system. Also, DHM can reduce Aβ peptide and improve memory function in mice
[40]. Therefore, out of the effect of DHM on GABAAR plasticity and the identified
improvement in cognitive memory, we considered its possibility as an AD therapy.

15

1.6 Goals and specific aims
To further understand the pharmacological mechanisms of DHM as an anxiolytic and
anti- EtOH, we developed a social isolation-induced anxiety + EtOH drinking model
in C57BL/6 mice that induces anxiety and fosters alcohol dependence by reducing
social interactions as a chronic stressor. This model would have a significant effect on
rodent behavior and brain structure to prove our conjecture.
Thesis goal: The goal of my thesis was to examine the effects of DHM on anxiety,
alcohol-preferring behaviors, and cognitive impairment caused by social isolation and
alcohol intake. In addition, we set to explore the role of DHM when both anxiety and
alcohol were present.

Specific aims:
(1) To test whether the EtOH intake of mice changes over time after DHM treatment.
(2) To test whether the behavior of mice, including memory and cognition, changed
after DHM treatment.
(3) Exploring possible associations between anxiety, alcohol, and Alzheimer's disease.

16

Chapter 2 Experimental Procedures
2.1 Materials
All animal experiments were performed according to the protocols approved by the
University of California (UCLA) and University of Southern California (USC)
Institutional Animal Care and Use Committee (IACUC), and all methods were carried
out in accordance with relevant guidelines, regulations, and recommendations,
including the ARRIVE guidelines. Eight-week-old male C57BL/6 mice (Charles River
Laboratories, Hollister, CA) were utilized in social isolation-induced anxiety and EtOH
drinking model. Mice were either group-housed or singly-housed in the vivarium under
a 12 h light/dark cycle with direct bedding and free access to food and water. Three
mice were allocated in each cage for group-housed group. Social isolation mice were
singly housed in opaque cages with reduced bedding, no environmental stimuli, and
given free access to food, water and 10% EtOH. After a two-week acclimation period,
sucrose (vehicle) or DHM (5 mg/kg) were orally administered daily for 4 weeks. Tissue
biochemical analyses were conducted at the University of Southern California (USC).

Groups were randomly separated as follows for a total of 6 weeks before sacrifice:
a. Group-housed mice without any drug administration for 2 weeks, and then given
daily sucrose agar as a vehicle for an additional 4 weeks (G2 + Veh4). Provide only
water as a water source.
b. Isolated mice without any drug administration for 2 weeks, and then given daily

17

sucrose agar as a vehicle for an additional 4 weeks for a total isolation period of 6 weeks
(Iso2 + Veh4). Provide both water and alcohol as water sources.
c. Isolated mice without any drug administration for 2 weeks, and then given daily
DHM for an additional 4 weeks for a total isolation period of 6 weeks (Iso2 + D4).
Provide both water and alcohol as water sources.

Figure 3.Experimental timeline

DHM (HPLC purified ≥98%, Master Herbs Inc., Pomona, CA) 2 mg/kg was prepared
in a 3% agar cube containing 5% sucrose. Mice were given as treatment by cutting the
agar into 0.5 × 0.5 × 0.5 cm cubes. Mice were given the sucrose agar and DHM cubes
orally once daily for four weeks during the dark period of the 12-hour light/dark cycle
with minimal disturbance to the mice. Ensure that each mouse receives and only one
sucrose-DHM agar cube and observe that the mice consume it completely to ensure
complete consumption of the corresponding treatment.

18

2.2 Methods
2.2.1 Behavior Test
Novel object recognition (NOR)
We first performed an observational study within one week of completing DHM
training to identify associations between factors and recognition. Novel object
recognition (NOR) is the cognition-related behavioral test implemented in this study. It
is widely used to test the neurobiology of non-spatial memory in rodents [42][43], but
the underlying mechanisms are not yet clear. Normally, when animals are exposed to
both the familiar object and the new object, they will prefer to approach and explore
the new rather than the familiar, as shown by the time spent exploring with their noses.

On the first day, mice were habituated in an empty open field for 5 minutes. On day
two, mice were subjected to a 5-min exploration period in the same field with two
identical objects placed equidistantly. On day three, mice were exposed to the same
field in the presence of the familiar and novel object to test long-term recognition
memory. The time spent exploring objects was recorded, and the cognitive level was
Figure 4. Schematic diagram of the experimental design
Novel object recognition (NOR).

19

measured by calculating the object recognition index (ORI%), where ORI%=(tn-
tf)/(tn+tf), and tf and tn represent the time spent exploring familiar objects and new
objects, respectively. Since mice are nocturnal and their state is influenced by their
surroundings, we chose to conduct behavioral tests under red illumination during their
active time (after 12:00 noon daily) to ensure that the disturbance of circadian rhythm
cycles and lights is minimized and to reduce the stress of mice performing test activities
in a new environment.

Novel context recognition test (NCR)
In addition to the object itself, the environment also influences the animal's choice [44].
Therefore, NCR was designed. The conditional association between environment and
novelty acceptance effects was explored by testing the preferences generated by mice
for object-environment pairings [45].

Figure 5.Schematic diagram of the experimental design (NCR).

The specific method for days 1-3 is habituation (once/day). In this experiment, we

20

used two containers with similar areas but different environments as experimental
apparatus. A is a rectangular container and B is a cylindrical container. The container
was open at the top and was equipped with a video camera to record the behavior of
the animals. Each animal was placed in Context A without toys for 5 minutes and then
returned to the cage for 30 minutes. The animals were then placed in Environment B
without toys for 5 minutes. Day 4 was a familiarization session. Two sets of
differently shaped toys were used as familiarization objects, called T1 and T2. The
two toys of T1 were placed at specific locations in Context A, generally at three
equidistant points on the diagonal. Each animal was placed in Context A and allowed
to explore T1 for 5 minutes; the animal then returned to its cage for 30 minutes. two
toys of T2 were placed at specific locations in Context B. Each animal was allowed to
explore T2 in Context B for 5 minutes. Day 5 (24 hours after the previous day's
training): A final experiment was conducted to determine each animal's memory
retention of familiar objects. A toy in Container A was swapped with a toy in
Container B, and each animal was allowed to explore the object in Context A or
Context B for 3 minutes. Testing and video analysis. We recorded the time spent by
mice exploring familiar objects (e.g., T1) and exchanged objects (e.g., T2), and we
considered exploration as equivalent to mice touching the object with their nose or
paw. The recognition index (RI) was calculated as: index = (tn - tf/(tn + tf)), where tf
represents the time spent exploring familiar objects previously encountered in the
same environment and tn represents the time spent exploring objects in different
environments. Generally, in the NCR test, mice explored more objects that appeared

21

in different environments than those previously encountered in the same environment,
and we believe that mice developed a memory for the environment.

Elevated plus maze (EPM)
The elevated plus maze (EPM) is an important experimental method used to evaluate
anxiety responses in rodents [40]. Compared to other methods that induce anxiety
through injurious stimuli such as electrical stimulation and odor stimulation, this
method is the least harmful to the animals, while being simple to operate and the results
are visualized. The experimental principle is to place the test animal into a cross-shaped
device consisting of an open arm and a closed arm at a certain height from the ground.
The mice explore the open arm out of curiosity, but this conflict leads to anxiety due to
the height of the ground and the nature of the mice to prefer darkness. This behavior is
similar to the exploration and fearful behavior of humans for cliff hazards. We evaluated
the mice's anxiety response by recording the time they stayed in the open and closed
arms and the central area. Statistical plotting was performed using the open arm dwell
time ratio (OT%), where OT%= OT/(CT+OT) *100%. The experiments were
scheduled after NOR and tested under both dark cycle and red illumination. The
experimental procedure requires attention to create an ideal testing environment that is
as clean, odor-free, and free of sound interference as possible, taking care to place the
mice in the central area of the cross with the animals facing the open arm.

22

2.2.2 Protein extraction and Western blot analysis
Approximately 20 mg of collected mouse brain tissue was dissected with a clean tool,
lysis buffer and 1% protease inhibitor was added, and the tissue was homogenized with
Sonicator-Branson SONI. The samples are then homogenized by centrifugation at
13,000 rpm for 20 minutes and all supernatants are transferred to new tubes. Cell
extracts were quantified using the BCA Protein Assay Kit (Pierce Biotechnology,
Rockford, IL) according to the manufacturer's instructions and handbook of use.
Twenty μg of protein was separated on a 4-20% sodium dodecyl sulfate polyacrylamide
gel electrophoresis and transferred to a PVDF membrane for Western blot analysis (Bio-
Rad Laboratories, Hercules, CA). The transferred membranes were blocked with
blocking buffer containing 3% BSA in 1X Tris-buffered saline with Tween 20
(Thermofisher) for 1 hr, and then added with primary antibody (p-tau) appropriately
diluted in 1X PBST and incubated overnight at 4°C. The membranes were washed three
times with 1X PBST for 5 min each on alternate days and incubated with secondary
antibody in 1X PBST for 1 h, followed by visualization with enhanced
chemiluminescence detection reagents and Chemi-Doc (Bio-Rad) imaging equipment.
Phosphorylated tau protein was purchased from Cell Signaling Technology. All other
primary and secondary antibodies were purchased from Bio-Rad All procedures were
repeated three times to confirm the changes in protein expression.

2.2.3 Statistical analysis
All assays other than mouse behavioral tests were performed at least three times. All

23

behavioral records were observed and analyzed in a double-blind manner. One-way or
two-way analysis of variance (ANOV A) was performed, followed by Holm-Sidak
multiple comparison test with significance level set at p ≤ 0.05.

24

Chapter 3 Results
3.1 DHM reverted cognition and memory decline caused by Social
isolation, EtOH treatment
The cognitive function after DHM treatment administration were as examined using
novel object recognition test. We found that control mice spent more time exploring the
novel objects with a higher Object Recognition Index (ORI = 54.0 ± 2.2%) than SI,
EtOH-administered mice (ORI = 45.6 ± 2.3%) (P = 0.018). DHM improved ORI of SI
mice by nearly 9% (54.8 ± 1.4%) and exhibited similar behavior to the control.

Figure 6. Novel object recognition tests.
ORI = object recognition index. Bars represent mean ± SEM. One-way ANOV A
followed by multiple comparisons with Holm-Sidak method. *, P ≤ 0.05. vs Iso2 +
D4.

25

Figure 7.Novel context recognition test.

As for the NCR, compared with the G2+Veh4 control, the treatment group exhibited
increased RI (43.53± 3.8 %). According to our previous study, the socially isolated
alcohol group should show less exploratory behavior, while DHM reverses this
phenomenon. Therefore, for the abnormal results, we speculate that it is caused by
some disturbance in the behavioral test for the mice, including the scaring of mice
during cage changes.

26

3.2 Western Blot Analysis
DHM significantly reduced phospho-tau in the brain
DHM significantly reduced the concentration of hyperphosphorylated tau protein in SI,
ethanol-consuming mice compared to the control, suggesting less neurotoxicity and
more effective neurotransmission.

Figure 8. Representative blots for Phosphorylated Tau protein.

Figure 9. Relative values of phosphorylated Tau protein expression were quantified.
SI and EtOH consumption increased levels while DHM-treated significantly lowered
them.

27

3.3 DHM reduces alcohol preference in mice
For both control and treatment groups, all socially isolated mice had unrestricted
access to alcohol and water. We recorded the amount of water and alcohol (ml)
consumed by the mice over two/three days on Mondays, Wednesdays, and Fridays for
six weeks. By determining the effect of DHM on EtOH consumption, we can derive
the role of DHM on EtOH dependence. When processing the data, we chose to select
one data every other data.

Results show that social isolation causes some alcohol preference in mice, while there
is a trend of first increase and then decrease in the DHM treatment group, indicating
that DHM has a mitigating effect on alcohol addiction in mice in the beginning of the
treatment.

Figure 10.Preference for water/alcohol in mice at six weeks

28

Along with recording the drinking data, we also measured the body weight of the mice.
For the grouped mice, we randomly selected one mouse to be weighed. Overall, we
found that during the experiment, the control group gained 0.255 percent of body
weight, the SI-EtOH group gained 0.145 percent, and the DHM-treated group gained
only 0.09 percent.

Figure 11.Body weight changes in mice over six weeks.
Growth percentage: Control 0.255%, SI+EtOH 0.145%, SI+EtOH+DHM 0.09%

Cumulatively, our data show that DHM can potentially reverse or prevent cognition,
memory decline, and NFTs pathogenesis induced by social isolation and chronic EtOH
consumption.

29

Discussion
The main objective of this study was to establish the relationship between EtOH
consumption and anxiety using a mouse model to verify the anxiolytic properties and
anti-alcohol dependence of DHM. The source of anxiety was social isolation. In this
study, we found that social isolation in C57BL/6J mice increased anxiety-like behavior
in mice and led to increased EtOH consumption. Mice that were socially isolated in
behavioral tests exhibited a lack of exploration, increased anxiety, and demonstrated
impaired cognition and memory. These abnormal behavioral states are very similar to
humans and are predictable and understandable. In this state, DHM ameliorated and
reversed the development of maladaptive directions, with reduced anxiety-like
behaviors, while cognitive function was significantly better than the untreated group. It
represents DHM as a potential drug for the prevention and treatment of anxiety
disorders at the molecular and cellular level. These results are consistent with our team's
previous performance of separately designed anxiolytic effects and anti-alcohol effects.
Notably, the study used 2 mg/kg of DHM administered orally, a dose that was selected
as the minimum effective dose based on the team's previous experimental results [5].
Although this dose of DHM has been shown to improve anxiety disorders, we cannot
say that the dose is adequate for it and the dose-response relationship needs to be
validated by designing a new study. Also, phosphorylated tau (ptau) is definite
biomarker of Alzheimer's disease (AD) [38], we demonstrate here using ptau that DHM
does improve in AD, but the human ptau brain expresses six subtypes and the exact

30

mechanism of DHM needs to be further determined.

The results from the EtOH preference test demonstrated DHM shows some
improvement, such effect is not statistically significant. We therefore suspect that
there was a leak in the device during the recording and replenishment of the water
source, which led to a bias in the data. In a previous study by our team, a similar
experiment was done using rats, but with only alcohol as a variable. In the
experiment, alcohol consumption was significantly reduced in the DHM group, and
that effect was longer lasting [5]. Also, when measuring the body weight of the mice,
the grouped mice was randomly selected from one of three mice and it happens that
there were mice with abnormal body weight. Therefore, this random method
decreases the rigor of the experiment. We should use a marker to mark the tail of each
mouse for differentiation.

In a previous study, we have verified the mechanism of action of DHM by measuring
the expression of the inhibitory synaptic scaffolding protein Gephyrin in the mouse
brain. Gephyrin is a key protein that anchors, clusters, and stabilizes GABAergic
synapses [46]. So, its expression and function can affect GABAergic neurotransmission
and aggregation. Furthermore, the overall expression level of gephyrin appears to
indicate the total amount of GABAARs at a given time. So, considering the role of
GABAAR in explaining anxiety, we can consider it as a biomarker for GABA function.
We found that after 2 and 4 weeks of social isolation, gephyrin protein expression in

31

the hippocampus of mice was reduced, along with a decrease in ATP levels. After two
weeks of DHM treatment, mice showed a significant reduction in anxiety-like behavior,
along with higher gephyrin expression in the hippocampus, suggesting that DHM
restored ATP levels and maintained gephyrin expression in isolated mice [47]. These
sustained levels of gephyrin may contribute to GABAAR clustering on the neuronal
synapse, restoring GABAergic neurotransmission.
As we known, social isolation (SI) induced stress triggers a series of responses
including disruption of the hypothalamic-pituitary-adrenal axis (HPA), resulting in
decreased expression of synaptic GABAARs, resulting in impaired GABAergic
transmission. Disruption of GABAAR by microglia causes activation of microglia and
astrocytes, leading to further neuroinflammatory damage and anxiety-like behaviors.
Thus, microglia from socially isolated mice exhibited a more compact shape, but a
reduced area, as reflected in a reduced cell outline perimeter. Our current and future
research will focus on determining the benefits of DHM administration in reshaping
anxiety circuits and its potential to mitigate downstream-related cognitive decline.
However, the study on Gephyrin protein was not repeated in this project for reasons of
consumables and experimental design.

More specifically, for the GABA receptor, it is composed of many different subunits.
Previous scientists accompanied by a deeper understanding of benzodiazepines found
that different subunits mediate different functions. In order to better achieve specific
effects and avoid side effects, we tried to selectively bind to specific subunits of the

32

GABAA receptor and achieve the separation of beneficial and detrimental effects by
modulating the affinity. It has been proven that GABAA receptors containing α2 and α3
are related to anxiolysis [48], and GABA A receptors containing α5 may be related to
memory, so these three subunit receptors may be reduced in anxiety animal model.

By investigating the time-dependence of GABAA receptor composition and function in
rats with EtOH intoxication, we uncover the mechanism by which repeated EtOH
administration produces persistent GABAAR alterations that contribute to chronic
alcohol addiction. However, the mechanism by which alcohol intoxication alters the
assembly and localization of GABAAR subunits is unclear, so it is necessary to track
the changes more carefully in the content of different subunits, as well as when and
where they are assembled. In addition, we cannot determine which specific brain
regions can produce similar effects, such as the hippocampus, thalamus, etc., and there
will be some temporal differences in the effect of drugs in different regions. In future
studies, we still need to be more patient to investigate which subunit of GABA affects
the disease and what role DHM plays in it.

33

Study limitations
Because of the team's inadequate pre-planning of animal experiments, the lack of mice
in the DHM treatment group resulted in insufficient data and less significant results
than expected. Also, we did not set up treatment groups after group alcohol
administration, as well as were unable to simultaneously compare isolated mouse
samples with isolated alcohol mouse samples at the same time, so for the two variables
of social isolation and alcohol, we could not fully demonstrate the role played by
alcohol here. Also, the experimental mice used were all males.

Currently, our team is studying the effects of DHM on mice of different sexes and at
different time points, and certain results have been obtained. Despite the limitations of
the experiment, the current study provides important support for understanding the role
of DHM in improving anxiety-like behavior. In the future, we will also do more
experiments on mechanism exploration to have a clearer idea of how GABA receptors
and DHM work.

34

References
[1] Chand SP, Marwaha R. Anxiety. [Updated 2022 May 8]. In: StatPearls [Internet].
Treasure Island (FL): StatPearls Publishing; 2022 Jan-. Available from:
https://www.ncbi.nlm.nih.gov/books/NBK470361/
[2] Bandelow B, Reitt M, Rover C, Michaelis S, Gorlich Y , Wedekind D. Efficacy of
treatments for anxiety disorders: a meta-analysis. Int Clin Psychopharmacol.
2015;30(4):183–92. Epub 2015/05/02
[3] Kessler RC. The global burden of anxiety and mood disorders: putting the
European Study of the Epidemiology of Mental Disorders (ESEMeD) findings into
perspective. J Clin Psychiatry. 2007;68(Suppl. 2):10–19.
[4] Baldwin DS, Anderson IM, Nutt DJ, Allgulander C, Bandelow B, den Boer JA, et
al. Evidence-based pharmacological treatment of anxiety disorders, post-traumatic
stress disorder and obsessive-compulsive disorder: a revision of the 2005
guidelines from the British Association for Psychopharmacology. J
Psychopharmacol. 2014; 28:403–39.
[5] Shen Y , Lindemeyer AK, Gonzalez C, Shao XM, Spigelman I, Olsen RW, Liang J.
Dihydromyricetin as a novel anti-alcohol intoxication medication. J Neurosci.
2012 Jan 4;32(1):390-401. doi: 10.1523/JNEUROSCI.4639-11.2012. PMID:
22219299; PMCID: PMC3292407.
[6] Hyttel J. Pharmacological characterization of selective serotonin reuptake
inhibitors (SSRIs)[J]. International clinical psychopharmacology, 1994, 9: 19-26.
[7] Lambert O, Bourin M. SNRIs: mechanism of action and clinical features[J]. Expert
review of neurotherapeutics, 2002, 2(6): 849-858.
[8] Thanacoody HK, Thomas SH. Tricyclic antidepressant poisoning: cardiovascular
toxicity. Toxicol Rev. 2005;24(3):205–14.
[9] Starcevic V . The reappraisal of benzodiazepines in the treatment of anxiety and
related disorders. Expert Rev Neurother. 2014;14(11):1275–86.
[10] Rickels K, Schweizer E, Case WG, Greenblatt DJ. Long-term therapeutic use of
benzodiazepines. I. Effects of abrupt discontinuation [published erratum appears
in Arch Gen Psychiatry 1991; 48(1):51]. Arch Gen Psychiatry. 1990;47(10):899–
907.
[11] Chessick CA, Allen HA, Thase ME, et al. (2006) Azapirones for generalized

35

anxiety disorder. Cochrane Database Syst Rev 19: CD006115.
[12] Carvalho A F, Heilig M, Perez A, et al. Alcohol use disorders[J]. The Lancet, 2019,
394(10200): 781-792.
[13] Samokhvalov A V , Popova S, Room R, et al. Disability associated with alcohol
abuse and dependence[J]. Alcoholism: Clinical and Experimental Research, 2010,
34(11): 1871-1878.
[14] Schwarzinger M, Thiébaut S P, Baillot S, et al. Alcohol use disorders and
associated chronic disease–a national retrospective cohort study from France[J].
BMC public health, 2018, 18(1): 1-9.
[15] World Health Organization. Global status report on alcohol and health 2018[M].
World Health Organization, 2019.
[16] Collins S E. Associations between socioeconomic factors and alcohol outcomes[J].
Alcohol research: current reviews, 2016, 38(1): 83.
[17] Kim J U, Majid A, Judge R, et al. Effect of COVID-19 lockdown on alcohol
consumption in patients with pre-existing alcohol use disorder[J]. The Lancet
Gastroenterology & Hepatology, 2020, 5(10): 886-887.
[18] Rehm J, Dawson D, Frick U, et al. Burden of disease associated with alcohol use
disorders in the United States[J]. Alcoholism: Clinical and Experimental Research,
2014, 38(4): 1068-1077.
[19] Roerecke M, Gual A, Rehm J. Reduction of alcohol consumption and subsequent
mortality in alcohol use disorders: systematic review and meta-analyses[J]. The
Journal of Clinical Psychiatry, 2013, 74(12): 17165.
[20] Miller N S, Gold M S. Management of withdrawal syndromes and relapse
prevention in drug and alcohol abuse[J]. American family physician, 1998, 58(1):
139.
[21] Oncken C, Van Kirk J, Kranzler H R. Adverse effects of oral naltrexone: analysis
of data from two clinical trials[J]. Psychopharmacology, 2001, 154: 397-402.
[22] Maisel N C, Blodgett J C, Wilbourne P L, et al. Meta‐analysis of naltrexone and
acamprosate for treating alcohol use disorders: when are these medications most
helpful [J]. Addiction, 2013, 108(2): 275-293.
[23] Fuller R K, Gordis E. Does disulfiram have a role in alcoholism treatment today
[J]. Addiction (Abingdon, England), 2004, 99(1): 21-24.

36

[24] Koulentaki, M., Kouroumalis, E. GABAA receptor polymorphisms in alcohol use
disorder in the GWAS era. Psychopharmacology 235, 1845–1865 (2018).
[25] Grobin AC, Matthews DB, Devaud LL, Morrow AL (1998) The role of GABA(A)
receptors in the acute and chronic effects of ethanol. Psychopharmacology 139:2–
19
[26] Lobo IA, Harris RA (2008) GABAA receptors and alcohol. Pharmacol Biochem
Behav 90:90–94. https://doi.org/10.1016/j.pbb.2008.03.006
[27] Kang M, Spigelman I, Sapp D W, et al. Persistent reduction of GABAA receptor-
mediated inhibition in rat hippocampus after chronic intermittent ethanol
treatment[J]. Brain research, 1996, 709(2): 221-228.
[28] Liang, J., Olsen, R. Alcohol use disorders and current pharmacological therapies:
the role of GABAA receptors. Acta Pharmacol Sin 35, 981–993 (2014).
[29] Chebib M, Johnston GA. The ‘ABC’ of GABA receptors: a brief review. Clin Exp
Pharmacol Physiol. 1999; 26:937–40.
[30] Erika T G, Juan G S, BH José, et al. Novel-SubstitutedHeterocyclic GABA
Analogues. Enzymatic Activity against the GABA-AT Enzyme fromPseudomonas
fluorescens and In Silico Molecular Modeling[J]. Molecules, 2018,23(5):1128.
[31] Horenstein J, Wagner DA, Czajkowski C, Akabas MH. Protein mobility and
GABA-induced conformational changes in GABA(A) receptor pore-lining M2
segment. Nat Neurosci. 2001; 4:477–85.
[32] Barnard EA, Skolnick P, Olsen RW, Mohler H, Sieghart W, Biggio G, Braestrup C,
Bateson AN, Langer SZ (1998) International Union of Pharmacology. XV .
Subtypes of g-aminobutyric acidA receptors: classification on the basis of subunit
structure and receptor function. Pharmacol Rev 50:291–313
[33] Sieghart W, Sperk G (2002) Subunit composition, distribution and function of
GABA(A) receptor subtypes. Curr Top Med Chem 2:795–816
[34] Chen, X., van Gerven, J., Cohen, A. et al. Human pharmacology of positive
GABA-A subtype-selective receptor modulators for the treatment of anxiety. Acta
Pharmacol Sin 40, 571–582 (2019). https://doi.org/10.1038/s41401-018-0185-5
[35] Silva J, Shao A S, Shen Y , et al. Modulation of hippocampal GABAergic
neurotransmission and gephyrin levels by dihydromyricetin improves anxiety[J].
Frontiers in Pharmacology, 2020, 11: 1008.

37

[36] Cummings J L, Cole G. Alzheimer disease[J]. Jama, 2002, 287(18): 2335-2338.
[37] Sery O, Povova J, Misek I, et al. Molecular mechanisms of neuropathological
changes in Alzheimer’s disease: a review. Folia Neuropathol. 2013; 51:1–9.
[38] de Souza LC, Sarazin M, Teixeira-Junior AL, et al. Biological markers of
Alzheimer’s disease. Arq Neuropsiquiatr. 2014; 72:227–231.
[39] Buchhave P, Blennow K, Zetterberg H, et al. Longitudinal study of CSF biomarkers
in patients with Alzheimer’s disease. PLoS One. 2009; 4: e6294.
[40] Söderberg, Linda, et al. "Lecanemab, Aducanumab, and Gantenerumab—Binding
Profiles to Different Forms of Amyloid-Beta Might Explain Efficacy and Side
Effects in Clinical Trials for Alzheimer’s Disease." Neurotherapeutics (2022): 1-
12.
[41] Liang, J., Kerstin Lindemeyer, A., Shen, Y . et al. Dihydromyricetin Ameliorates
Behavioral Deficits and Reverses Neuropathology of Transgenic Mouse Models of
Alzheimer’s Disease. Neurochem Res 39, 1171–1181 (2014).
https://doi.org/10.1007/s11064-014-1304-4
[42] Goulart BK, de Lima MNM, de Farias CB, Reolon GK, Almeida VR, Quevedo J,
Kapczinski F, Schröder N, Roesler R. Ketamine impairs recognition memory
consolidation and prevents learning-induced increase in hippocampal brain-
derived neurotrophic factor levels. Neuroscience. 2010; 167:969–973. doi:
10.1016/j.neuroscience.2010.03.032.
[43] Silvers JM, Harrod SB, Mactutus CF, Booze RM. Automation of the novel object
recognition task for use in adolescent rats. J Neurosci Met. 2007; 166:99–103. doi:
10.1016/j.jneumeth.2007.06.032.
[44] Balderas, I. et al. The consolidation of object and context recognition memory
involve different regions of the temporal lobe. Learn Mem. 15, 618–624.
https://doi.org/10.1101/lm.1028008 (2008).
[45] Bevins RA, Besheer J, Palmatier MI, Jensen HC, Pickett KS, Eurek S. Novel-
object place conditioning: behavioral and dopaminergic processes in expression of
novelty reward. Behav Brain Res. 2002; 129:22–50. doi: 10.1016/S0166-
4328(01)00326-6.
[46] Choii G, Ko J. Gephyrin: a central GABAergic synapse organizer. Exp Mol Med.
2015;47: e158.
[47] Al Omran, A.J., Shao, A.S., Watanabe, S. et al. Social isolation induces

38

neuroinflammation and microglia overactivation, while dihydromyricetin prevents
and improves them. J Neuroinflammation 19, 2 (2022).
https://doi.org/10.1186/s12974-021-02368-9
[48] Rodolph U, Crestani F, Mohler H (2001) GABA(A) receptor subtypes: dissecting
their pharmacological functions. Trends Pharmacol Sci 22:188–194

Abstract (if available)

Abstract Social isolation is a stress-responsive stimulus that is associated with the etiology of anxiety-related disorders. Chronic stress is a common precursor to cognitive and memory impairment associated with Alzheimer's disease (AD). When coupled with psychosomatic disease and chronic ethanol (EtOH) intake, this rate of progression/decline can be greatly increased. Interestingly, since the COVID-19 pandemic, studies have demonstrated an upward trend in social isolation, psychological distress, and alcohol sales. We found that γ-aminobutyric acid A receptors (GABAARs) play a similar regulatory role in anxiety disorders, Alzheimer's disease, and alcohol use disorders. As a positive allosteric modulator of GABAARs, dihydromyricetin (DHM) has emerged as a highly promising therapeutic for development because of its ability to reduce social isolation (SI)-induced anxiety and EtOH-related impairments.

The study presented in this thesis aims to explore the effects of social isolation, long-term EtOH consumption, and DHM treatment on cognition in mice. We established an SI-induced anxiety + EtOH drinking model consisting of 8-week-old male C57BL/6 mice. Behaviorally, DHM-treated mice exhibited improved cognitive performance and reduced anxiety levels. Pathologically, DHM significantly reduced the expression of phosphorylated tau protein in the mouse brain. DHM was shown to potentially reverse or prevent the cognitive and memory decline induced by social isolation and long-term EtOH consumption, exert anxiolytic effects and have possible therapeutic effects on AD, consistent with our expectations. The findings in this study further suggests the need for exploring the intrinsic mechanism of action of DHM.

Linked assets

University of Southern California Dissertations and Theses

Conceptually similar

PDF

Dihydromyricetin as a potential therapeutic for Alzheimer's disease-like changes induced by social isolation

PDF

Social isolation anxiety induced synaptic failure restored by dihydromyricetin

PDF

Dihydromyricetin improves social isolation anxiety induced synaptic loss and astrogliosis in mice hippocampus

PDF

The study of DHM effects on counteracting ethanol intoxications

PDF

DHM ameliorates social isolation induced anxiety and alcohol preference

PDF

The therapeutic effects of dihydromyricetin (DHM) on anxiety disorders

PDF

Development of dihydromyricetin (DHM) as a novel therapy for alcoholic liver disease (ALD) and alcohol use disorder (AUD)

PDF

RPE secretome for the treatment of retinal degeneration in the RCS rat

PDF

Investigating sodium butyrate effects on liver in a disrupted gut microbiome model of binge-like alcohol consumption in mice

PDF

Small molecule lipoxin analogue for prevention of retinal degeneration

PDF

Neuropsychiatric complications of COVID-19 and major depression disorder with pharmacological strategies for management

PDF

Evaluating the effects of dihydromyricetin on dopamine mediated behaviors using a Parkinson's disease model

PDF

Investigating sodium butyrate as a potential treatment for alcohol liver disease through the gut-liver axis

PDF

Laser induced modulation of ocular renin-angiotensin system for treatment of dry age-related macular degeneration

PDF

Mechanisms of P2XR-mediated ethanol consumption

PDF

Alterations in renin-angiotensin system signaling in SARS-CoV2 infection

PDF

Sodium butyrate prevents antibiotic-induced increase in ethanol drinking in C57BL/6J mice by modulating neuroinflammatory response

PDF

Renin-angiotensin system modulation for the prevention and treatment of metabolic dysfunction

PDF

Investigation into the role of the intestinal microbiome on ethanol consumption behaviors

PDF

Pharmacokinetic and pharmacodynamic optimization of CFTR modulator therapy to mitigate potential drug interactions and adverse events in people with cystic fibrosis

Asset Metadata

Creator Yuan, Zidan (author)

Core Title DHM reduces cognitive impairment induced by social isolation and ethanol consumption

School School of Pharmacy

Degree Master of Science

Degree Program Clinical and Experimental Therapeutics

Degree Conferral Date 2023-05

Publication Date 04/27/2023

Defense Date 04/27/2023

Publisher University of Southern California (original), University of Southern California. Libraries (digital)

Tag Anxiety,dihydromyricetin,ethanol intake,GABA receptor,OAI-PMH Harvest,social isolation

Format theses (aat)

Language English

Contributor Electronically uploaded by the author (provenance)

Advisor Liang, Jing (committee chair), Asatryan, Liana (committee member), Louie, Stan Gee (committee member)

Creator Email yuanzidanx@gmail.com,zidanyua@usc.edu

Permanent Link (DOI) https://doi.org/10.25549/usctheses-oUC113089396

Unique identifier UC113089396

Identifier etd-YuanZidan-11735.pdf (filename)

Legacy Identifier etd-YuanZidan-11735

Document Type Thesis

Format theses (aat)

Rights Yuan, Zidan

Internet Media Type application/pdf

Type texts

Source 20230501-usctheses-batch-1033 (batch), University of Southern California (contributing entity), University of Southern California Dissertations and Theses (collection)

Access Conditions The author retains rights to his/her dissertation, thesis or other graduate work according to U.S. copyright law. Electronic access is being provided by the USC Libraries in agreement with the author, as the original true and official version of the work, but does not grant the reader permission to use the work if the desired use is covered by copyright. It is the author, as rights holder, who must provide use permission if such use is covered by copyright.

Repository Name University of Southern California Digital Library

Repository Location USC Digital Library, University of Southern California, University Park Campus MC 2810, 3434 South Grand Avenue, 2nd Floor, Los Angeles, California 90089-2810, USA

Repository Email cisadmin@lib.usc.edu