2013. Vol.4, No.10A, 13-18
Published Online October 2013 in SciRes (http://www.scirp.org/journal/psych) http://dx.doi.org/10.4236/psych.2013.410A003
Copyright © 2013 SciRes. 13
Buspirone Ameliorates the Morphine Withdrawal-Induced
Anxiety through Synaptic Ultrastructural Changes in
Hippocampus of Rat*
Jialin Gao1,2#, Gang Qian1#, Suyuan Luo1†, Yan Tian1, Mingsong Wu1, Zhongxiang Yao3†
1Department of Cell Biology and Genetics, Zunyi Medical College, Zunyi, China
2Department of Endocrinology and Genetic Metabolism, Yijishan Hospital of
Wannan Medical College, Wuhu, China
3Department of Physiology, The Third Military Medical University, Chongqing, China
Email: †firstname.lastname@example.org, †email@example.com
Received August 15th, 2013; revised September 12th, 2013; accepted September 29th, 2013
Copyright © 2013 Jialin Gao et al. This is an open access article distributed under the Creative Commons Attri-
bution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the
original work is properly cited.
Morphine administration causes long-lasting neural changes in the brain that underpin the behavioral ab-
normalities, and the relationship between structural changes and behavioral symptoms is obscure. In pre-
sent study, the elevated plus-maze and transmission electron microscope were applied to validate the
anxiety-like behaviors and synaptic ultrastructural changes in the hippocampi of rats among the morphine
group (morphine administration only), the buspirone group (morphine plus buspirone administration) and
the vehicle (saline treated only). As compared with the vehicle group, lower values of OE (times of en-
tering into the open arms), OE% (percentage of entries into the open arms), OT (time spent in the open
arms), OT% (percentage of time stayed in the open arms), Ns (surface density (Sv)/numerical density
(Nv)) and S (surface area) of synapses were observed in the morphine group , but significantly, behavior
higher scores of RR (rearing), HD (head-dipping), FBA (flat back approach), and higher Nv, Sv, PSD
(postsynaptic density), LPT (length of postsynaptic thickening), WCJ (widths in synaptic cleft on junc-
tions) and CCR (curvature of the cleft region) of synapses appeared in the morphine group. However, no
significant differences in values of most of those parameters above were detected between the vehicle
group and the buspirone group. These results supported that anxiety-like symptoms of rats significantly
occurred to the rats after acute morphine withdrawal, but buspirone administration could reverse these in-
dexes. It also proved that the appearance/disappearance of anxiety-related symptoms was related to the ul-
trastructural changes/reversibility of synapses in the hippocampus with morphine and buspirone admini-
strations. So, it suggested that anxiety-related symptoms were modified in rats subjected to the synaptic
ultrastructural changes in hippocampus by morphine acute withdrawal and were further rehabilitated by
buspirone administration. It is helpful to pursue the effective therapeutic methods of morphine addiction.
Keywords: Buspirone; Morphine; Anxiety; Synaptic Ultrastructure; Hippocampus
Chronic morphine administration can induce the changes of
hippocampal CA1 neurocircuitry which modulate the synaptic
plasticity through the regulation of long-term potentiation (LTP)
and long-term depression (LTD) (Salmanzadeh et al., 2003).
The synaptic plasticity is widely studied in hippocampal CA1
and CA3 fields, which is believed to be the mechanism under-
lying certain types of learning and memory (Moron et al., 2007).
It is a commonly accepted assumption that an increase in neu-
ropil volume reflects an increase in the number of synapses pre-
sent (Devoogd et al., 1985; Turner & Greenough, 1985; Black
et al., 1990; Withers et al., 1993; Klintsova et al., 2000), but, in
many regions of the brain, this has not yet to be clearly estab-
lished (Brown et al., 2002).
Buspirone, a potent anxiolytic compound in animal models
(Riblet, et al., 1986), displays reversibility of synaptic activa-
tion of pyramidal cells in the hippocampus (Trulson et al.,1986;
Mauk et al., 1988). It showed that buspirone bind selectively to
presynaptic (dorsal raphe) and postsynaptic (hippocampus, cor-
tex) 5-hydroxytryptamine 1A (5-HT1A) receptor binding sites,
and the clinical trials of anxiety and depression may be related
to the buspirone’s interactions with gepirone at pre-synaptic
and postsynaptic 5-HT1A receptor (Yocca, 1990). Buspirone
had no effect on the immediate rewarding properties of cocaine,
but the attenuation of 5-HT neurotransmission (via the autore-
ceptor agonist properties of buspirone) could reverse the nega-
tive impact of cocaine (Ettenberg & Bernardi, 2007).
So, the correlating ethological indexes of anxious rats with
synaptic ultrastructural changes of the hippocampal CA1 and
CA3 fields should enable us to evaluate the mechanism of
*The authors had no conflicts of interest to declare in relation to this article.
#The first two authors have contributed equally to this paper.
J. L. GAO ET AL.
morphine addiction and pursue the effective therapeutic meth-
ods. We hypothesized that anxiety-related symptoms were
modified in rats subjected to the synaptic ultrastructural
changes in hippocampal CA1 and CA3 fields by morphine
acute withdrawal and were further rehabilitated by buspirone
administration. Here, we should provide an evidence in which
the appearance/disappearance of anxiety-related symptoms are
related to the synaptic ultrastructural change/reversibility of the
hippocampal CA1 and CA3 fields with morphine and buspirone
Adult male Sprague Dawley rats (weighing 200 - 250 g)
were obtained from the Laboratory Animal Center, the Third
Military Medical University (Chongqing, China). Supplied with
water and food available ad libitum, rats were maintained in
groups on a 12-h light/dark cycle (lights on at 08:00 AM) and
were allowed to acclimatize to their environment for 1 week
prior to drug administration. All treatments were strictly in
accordance with National Institutes of Health Guide for the
Care and Use of Laboratory Animals.
Morphine a nd Buspiro ne Administrat ion for the
Elevated Plus-Maze Tests
Rats were randomly assigned to three groups (n = 12 each
group): the saline treated control group (saline group), the
morphine treated model group (morphine group) and the mor-
phine plus buspirone treated experimental group (buspirone
group). Morphine (morphine sulfate) was prepared in 0.9%
sterile isotonic saline. In morphine group and buspirone group,
three escalating doses of morphine (5, 10, and 15 mg/kg, s.c.,
twice per day at 12 hr intervals) were applied for 10-day inject-
tions as described before (Morinville et al., 2003). Briefly, a
dosage of 5 mg/kg morphine was applied at the primal period
of 5 days, followed by 3-day injections of 10 mg/kg, then of 15
mg/kg in the last two days. Withdrawal syndrome was precipi-
tated with abrupt termination of morphine intake by replace-
ment of daily injections of saline for 3 days, and was further
validated by observing behaviors in a separate series of ex-
periments. After acute morphine withdrawal, each anxious rat
in the buspirone group received buspirone treatment (15 mg/kg,
i.g., twice per day at 12 hr intervals) for 3 days while the anx-
ious rats in the morphine group only received saline treatment
by the same form. The rats of saline group only received saline
as the program above.
All rats were tested just once after 72 h of the last treatments
of saline or buspirone. To begin a test session, rats were indi-
vidually placed in the center of the maze, facing towards one of
the enclosed arms, and allowed 5 min of free exploration. The
elevated plus-maze apparatus consisted of a center (10 × 10 cm
square area), two opposing open arms (50 cm length, 10 cm
width and 40 cm height), and two opposing closed arms (50 cm
length, 10 cm width and 40 cm height). Each of the four arms
in the maze was connected to the center at 90˚ relative to the
adjacent arms. All sides and floor surfaces of the open and
closed arms were constructed from black Plexiglas. The floor of
the maze was covered with a thin layer of vulcanized black
rubber to facilitate inter-trail cleaning. All trials took place in a
dimly illuminated chamber and were recorded by a video cam-
era that was linked to a video monitor, and an IBM-compatible
computer positioned outside the test chamber. Behaviors of the
rats were recorded with an automated system. Positions of the
rats in the maze were continuously tracked by photo beam ar-
rays embedded along the entire length of the base of each
closed arm, the entry point to all arms, and in clear Plexiglas
tubes that extended from the distal end of each closed arm and
ran parallel to the open arms.
An auto-recorded system was applied to collect data and fur-
ther these behavior measurements were analyzed using Motor
Monitor Software (Kinder Scientific, Poway CA). Based on
computer-recorded data, judgment of anxiety-like behaviors
were validated by the following parameters for each rat: (1)
times of rat entering into the open arms (OE); (2) the percent-
age of entries into the open arms as the total entries into both
open and closed arms (OE%); (3) time spent in the open arms
(OT); (4) the percentage of time stayed in the open arms as in
both open and closed arms (OT%); (5) partial or total rising
onto hind limbs (rearing, RR); (6) exploratory movement of
head/shoulders over the sides of the maze and down toward the
floor (Head-dipping, HD); and (7) locomotion when the animal
stretched to its full length and cautiously moves forward
(Flat-back approach, FBA).
Tissue Processing for Electron Microscopy
Hippocampal samples were harvested from rats of three
separate groups (n = 6 per group). The animals were decapi-
tated after 72 h of the last injections of saline or buspirone. Ac-
cording to standard procedures (Harris & Sultan, 1995; Shep-
herd & Harris, 1998), the brains were removed, and slices of
the hippocampi were rapidly dissected free. Brains were sec-
tioned coronally at 200 µm on a vibratome; the CA1 and CA3
fields were blocked from sections containing the dorsal hippo-
campus (bregma −2.3 to −4.3 mm, mediolateral 0 - 2 mm) with
a tissue chopper into ice-cold phosphate buffered solution con-
taining (in mM) 117 NaCl, 4.7 KCl, 26 NaHCO3, 1 NaH2PO4,
2.2 CaCl2, 1.2 MgSO4, and 10 glucose, pH 7.4. The slices were
fixed rapidly in 6% glutaraldehyde and 4% paraformaldehyde
in 100 mM cacodylate buffer for 8 sec under microwave irra-
diation (Jensen & Harris, 1989), stored overnight in the fixative,
and then rinsed in buffer. Slices were bathed with 1% osmium
and 1.5% potassium ferrocyanide in 100 mM cacodylate buffer,
cooled in ice bath to 12˚C, and microwave for 2 min at 37˚C.
After several buffer rinses, they were en bloc stained using 2%
aqueous uranyl acetate while being cooled on ice and were
microwave 2 min at room temperature. Samples were dehy-
drated in an acetone series (50%, 70%, 90%, and 100%) for 40
sec each in the microwave at 37˚C. The following treatments,
involved in infiltration and embedded coffin mold, were per-
formed as described before (Shepherd & Harris, 1998). These
sections were embedded in Agar 100 resin and polymerized for
48 hr at 60˚C. The block face was trimmed to include one entire
antennat lobe in the ultrathin, and then a subset of six 70
nm-thin sections were selected in a random systematic manner
from the entire section set for analysis using TEM. Thin sec-
tions were mounted on Pioloform-coated slot grid and counter-
stained with saturated ethanolic uranyl acetate, followed by
Reynolds lead citrate, each for 5 min. The target areas, hippo-
campal CA1 and CA3 fields between strata pyramidale and
lacunosum moleculare, were examined with a Hitachi H-600
transmission electron microscope.
Copyright © 2013 SciRes.
J. L. GAO ET AL.
Copyright © 2013 SciRes. 15
Stereological Measurements of Synapses
Serial electron micrographs (12 photographs per section, from
left top to low right) were taken on a Hitachi H-600 transmis-
sion electron microscope operating at 80 kV. Images were col-
lected onto 35 mm plate film at a final magnification of 44,000
× and enlarged onto 7 × 12 inch photographic paper. Within
each photograph, the points used for analysis were determined
by placing a 9 mm2 transparent grid over a micrograph. Stereo-
logical quantification of synapses was determined using the
physical dissector method. The accurate and unbiased compare-
sons of numerical density (Nv), surface area (S), surface density
(Sv) and the average surface density of each synapse (Ns) in the
hippocampal CA1 and CA3 fields were measured as described
before (Sterio, 1984). These parameters such as the total synapse
number per µm2 (Na), the mean synapses strip length (L), the
sample thickness (T), the total grid length (Lt), and the cross
point (Ni) between grid and synapses were recorded. Accord-
ingly, the emendation coefficient (Ko) was calculated as follows:
Ko = 1 + 3T/2L. Based on these sufficient parameters, stereo-
logical analysis was respectively described as the following
formulas: Nv = Na·L−1(Ko + T), Sv = 2Ni/Lt·Ko, and Ns = Sv/Nv.
Further Measuremen t s o f S yn a p t i c Junctio n
All identifiable synaptic junctions were marked on the mi-
crographs above and used for further measurement. Asymmet-
ric synapses were identified, and postsynaptic elements were
classified as dendritic spines or dendritic shafts, by established
morphological criteria (Scheuermann, 1991). The criterion of
parallelism was waived when terminals were sectioned at an
oblique angle to the cleft specialization if the presence of a
thickening and synaptic vesicles was still obvious. The numbers
and the areas of the synaptic junctions were recorded in order to
determine postsynaptic density (PSD).
According to description before (Weibel, 1966), the mem-
branous boundaries of terminals were outlined and a curly
probe was used to determine terminal areas by permitting point
counts for area estimate. More detailed measurements of ter-
minal parameters were undertaken as described before (Dyson
& Jones, 1980). A curvature-length probe (Jones & Devon,
1978) was used to measure the length of postsynaptic thicken-
ing (LPT) and the width in synaptic cleft on junctions (WCJ) at
the cleft edge of the thickening, and to determine junction cur-
vature (curvature of the cleft region, CCR) at these positions.
LPT was measured directly along the arc of curvature. Adjacent
junctions on the same terminal separated by membrane free of
specialization were set aside.
All data obtained from behavioral measurements of plus-
maze activity, stereological detections and synaptic terminal
junction parameters were subjected to one-way analysis of
variance (ANOVA) as dictated by the experimental design. The
mean differences were compared by least significant difference
test. All analyses were performed on a completely randomized
design using the Minitab software package and a value of p <
0.05 was considered to indicate statistical significance, p < 0.01
Anxiety-Li ke B eh a vi oral Reacti ons
As shown in Table 1, very significant withdrawal symptoms
appeared in individuals of morphine group. One-way ANOVAS
between morphine group and saline group subjects revealed
notably significant carry-over effects of morphine administri-
tion on OT, OE, OT%, OE%, RR, HD, and FBA parameters.
As compared with the saline group, lower values of OE, OE%,
OT and OT% were observed in the morphine group (p < 0.01),
but significantly, higher scores of RR, HD and FBA appeared
in the morphine group (p < 0.01). However, no significant dif-
ferences in values of these parameters above were detected
between the saline group and the buspirone group (p > 0.05)
except HD (p < 0.01). These results showed that a 10-day
morphine treatment produced robust and anxiety-like behav-
ioral response patterns in the elevated plus-maze, but the bus-
pirone administration reversed them.
Quantitatively Morphological Effects of Synapses
As shown in Table 2 and Figure 1, it could be seen that sig-
nificant differences occurred in hippocampal CA1 and CA3
fields in the morphine group, mainly in the profiles of pre-
synapse and postsynapse membranes. As compared with the
saline group, higher Nv and Sv (approximately 2 folds) were
observed in the morphine group (p < 0.01 or p < 0.05) in the
hippocampal CA1 and CA3 fields; but significantly, lower
values of S and Ns were detected in the hippocampal CA1 and
CA3 fields (p < 0.01 or p < 0.05). However, no significant dif-
ferences in values of these parameters above were detected
between the saline group and the buspirone group (p > 0.05)
except S at CA1 (p < 0.01). These results showed that a 10-day
morphine administration changed the synaptic quantitatively
morphological effects in hippocampal CA1 and CA3 fields, but
the buspirone treatment reversed them.
Scores of ethological measures in three independent groups.
OT OE OT% OE% RR HD FBA
Saline group 68.11 ± 2.08 6.80 ± 1.16 0.25 ± 0.04 0.41 ± 0.06 1.40 ± 0.66 0.16 ± 0.09 2.35 ± 0.25
Morphine group 44.03 ± 3.04* 4.70 ± 0.34* 0.11 ± 0.07* 0.24 ± 0.02* 6.82 ± 1.07* 3.92 ± 0.95* 33.56 ± 1.28*
Buspirone group 65.01 ± 7.02 7.00 ± 1.60 0.25 ± 0.06 0.42 ± 0.07 1.55 ± 0.81 2.11 ± 0.23# 1.56 ± 0.07
Values represent means ± SEM (n = 12 in each group). Significant differences: *from the respective saline group and buspirone group (P < 0.01). # from the respective
saline group and morphine group (P < 0.01). OT, time spent in the open arms; OE, times of entering into the open arms; OT%, percentage of time stayed in the open arms;
OE%, percentage of entries into the open arms; RR, rearing; HD, head-dipping; FBA, flat back approach.
J. L. GAO ET AL.
Quantitatively morphological effects of synapses in hippocampal CA1 and CA3 fields.
Saline group Morphine group Buspirone group Saline group Morphine group Buspirone group
Nv(number/mm3) 0.723 ± 0.386 1.513 ± 0.817*Δ 0.743 ± 0.567 0.667 ± 0.282 1.146 ± 0.490*Δ 0.801 ± 0.361
Sv(um2/um3) 0.077 ± 0.037 0.103 ± 0.052*Δ 0.088 ± 0.060 0.044 ± 0.018 0.052 ± 0.026# 0.051 ± 0.012
S (um2) 0.174 ± 0.245 0.079 ± 0.046*▲ 0.348 ± 0.179 0.079 ± 0.059 0.059 ± 0.055# 0.062 ± 0.041
Ns(=Sv/Nv) 0.107 0.068 0.118 0.066 0.045 0.064
Values represent means ± SEM (n = 6 in each group). Significant differences: *from the respective saline group (P < 0.01); # from the respective saline group (P < 0.05);
▲ from the respective buspirone group (P < 0.01); Δ from the respective buspirone group (P < 0.05). Nv, numerical density; Sv, surface density; S, surface area; Ns, =
(a) (b) (c)
(d) (e) (f)
Electron micrographs of the hippocampal CA1 and CA3 fields, showing the morphological differences of synapses in three inde-
pendent groups (n = 6 in each group, bar = 20 nm). Thin synaptic profiles (arrows) and low values of PSD, LPT, WCJ, and CCR on
the synaptic junctions (asterisk) at CA1 (A) and CA3 (D) fields in the saline group; Thick synaptic profiles (arrows) and high val-
ues of PSD, LPT, WCJ, and CCR on the synaptic junctions (asterisk) at CA1 (B) and CA3 (E) fields in the morphine group; Thin
synaptic profiles (arrows) and low values of PSD, LPT, WCJ, and CCR on the synaptic junctions (asterisk) at CA1(C) and CA3 (F)
fields in the buspirone group. PSD, postsynaptic density; LPT, length of postsynaptic thickening; WCJ, widths in synaptic cleft on
junctions; CCR, curvature of the cleft region.
synaptic junction regions in any way reflect their functional
states, more detailed measurements of terminal synaptic pa-
rameters were undertaken in the hippocampal CA1 and CA3
fields from three independent groups above (Table 3 and Fig-
ure 1). Significantly, higher values of PSD, LPT, WCJ and
CCR from hippocampal CA1 and CA3 fields were observed in
the morphine group than that in the saline group (p < 0.01 or p
< 0.05). To some extent, buspirone administration to the acute
morphine withdrawal animals rehabilitated synaptic profile, the
scores of most of these indexes above were only slightly in-
creased in contrast with the saline group (P > 0.05), and they
still were significant lower than that in the morphine group (p <
0.01 or p < 0.05).
Concerning the ethological measurements, anxiety related
symptoms significantly occurred to the rats suffered from the
escalating doses of morphine administration while they were
exposed to the elevated plus-maze after 72 h acute morphine
withdrawal, but buspirone administration can reverse these
indexes. Morphine withdrawal increased ethological indexes
related to exploration and locomotor behavior such as end-
exploration, head-dipping, rearing, and so on. These evidences
indicated that morphine withdrawal increased the general motor
activity and decreased aversion to the open spaces of the maze
in rats. It has been reported that systemic injections of morphine
Copyright © 2013 SciRes.
J. L. GAO ET AL.
Measurements of terminal synapse junctions in hippocampal CA1 and CA3 fields.
Salineroup Morproup Buspiro group Salineroup Morproup Buspiro group ghine gne ghine gne
PSD 7.83 ± 0.80 10.86 ± 0.86#Δ 8.37 ± 1.58 7.73 ± 1.02 12.86 ± 1.15*Δ 10.31 ± 1.02
LPT 184.108.40.206.35 ± 2.30 44.93 ± 3.48*Δ 78 ± 3.26 66 ± 3.81 99 ± 7.81#▲ 41.41 ± 3.55
WCJ 2.34 ± 0.29 3.80 ± 0.30*Δ 3.07 ± 0.22 2.38 ± 0.73 3.81 ± 0.59*Δ 2.91 ± 0.15
CCR 1.21 ± 0.11 1.37 ± 0.12*Δ 1.29 ± 0.14 1.23 ± 0.18 1.39 ± 0.30*Δ 1.30 ± 0.14
ValpresentM (n = 6 iignificant di the respectiv (P < 0.01pective saline 05);
▲e resne groupom the respne groupP < 0tsynapticngth of postening;
was commenced establishment of anxiety symptoms in rats
s on which they are acting and on the
nt with the idea that synaptic potentiation on hippo-
havioral changes associated with olfactory learning and mem-
arance of anxiety-
(No. 30860373), t Development of
Guizhou Province 109) and the Key
A comparative study of the effects of morphine in the dorsal periaq-
ueductal gray and nucleus accumitted to the ele-
vated plus-maze testsearch Experimentelle
means ± SE
n each group). S
(P < 0.01); Δ fr
e saline group
.05). PSD, pos
); # from the res
density; LPT, le
group (P < 0.
synaptic thick (
WCJ, widths in synaptic cleft on junctions; CCR, curvature of the cleft region.
may produce euphoric or dysphoric effects (Ebert et al., 1998). only with shifts in the tasks being performed but also with be-
by opiate administration because of the similarities in time
course and intensities of craving and anxiety experienced dur-
ing acute opiate withdrawal (Swift & Stout, 1992). The intrap-
eritoneal injections of morphine (0.1 and 0.3 mg/kg) in rats
increased both the total entries into the arms and the percentage
of entries and time spent in the open arms of the maze
(Anseloni et al., 1999).
Opioid ligands may mediate reward or aversive process de-
pending on the structure
ceptor types with which they interact (Mucha & Herz, 1985;
Bals-Kubik et al., 1989). It is a commonly accepted assumption
that an increase in neuropil volume reflects the increase in the
number of synapses (Klintsova et al., 2000), but in many re-
gions of brain, this has yet to be clearly established (Brown et
al., 2002). It has been suggested that the increase in the motor
activity caused by opiate may be due to the activation of dopa-
minergic cells in mesolimbic system (Koob, 1992). Indeed,
morphine injections into the ventral tegmental area (VTA) can
increase the dopamine-dependent locomotion (Yuan et al.,
1992). Previous models of dysphoria-like and anxiety-like be-
haviors resulting from acute opioid dependence suggested the
dopaminergic pathway may also play a prominent role in the
transition from casual to compulsive use (Zhang & Schulteis,
In the present study, both behavioral and synaptic data are
mpus is critical for the appearance in anxiety-like behavior.
The collaterals of axons between cortex and the granule cells in
hippocampal dentate gyrus (DG) innervate the distal parts of
the apical dendrities of pyramidal neurons in hippocampal CA1
and CA3 fields (Safiulina et al., 2005). The striking differences
in the structural changes of hippocampal CA1 and CA3 fields
suggested that the anxiety-like symptoms may be related to
morphological changes in rats, associated with morphine and
buspirone administrations. It is a good confirmation with our
results that anterograde and retrograde alteration related to
synaptic plasticity are necessary for memory formation (Morris
et al., 2003). It is also supporting our results that tianeptine
administrition for several hours is able to reverse the effects of
stress when the stress and anesthesia onset is remarkable
(Shakesby et al., 2002). In an attempt to isolate the principal
driving force(s) of structural plasticity from combined influ-
ences of age, behavior, and hormonal control, it is found that
structural changes in the antennal lobes of the bee coincide not
ory. Based on correlation between thickness of the postsynaptic
nucleus density and excitatory or inhibitory function of a syn-
apse holding in the nucleus, it argued that the changes in mor-
phology of synapses contributed some inhibitory synapses to
excitatory contacts (Guldner & Ingham, 1980). Recent evi-
dences demonstrated that synaptic plasticity in hippocampal-
nucleus accumbens (NAc) was disrupted by repeated cocaine
treatment (Goto & Grace, 2005). Taken together these data, we
suggested that, related to functional roles of hippocampal CA1
and CA3 fields, the changes of these excitatory synapses on
dopaminergic cells were a neural adaption common to all major
classes of addictive substances and therefore might play a criti-
cal role in the development of addiction.
In conclusion, we showed that anxiety-like symptoms of rats
significantly occurred to the rats after acute morphine with-
drawal, but buspirone administration could reverse these indexes.
We also proved that the appearance/disappe
lated symptoms was related to the ultrastructural changes/re-
versibility of synapses in the hippocampus with morphine and
buspirone administrations. So, these results are theoretically
related the psychological symptoms with neural circuitry regu-
lation. Future studies will focus on the hippocampal neuron
signal pathway of the buspirone administration after the mor-
phine acute withdrawal. It is helpful to pursue the effective
therapeutic methods of morphine addiction.
The authors greatly acknowledge the financial support of this
project by the National Natural Science Foundation
he Special Foundation for
College in China (No. 2005
oject of the Chinese Ministry of Education (No. 206136).
Anseloni, V. C., Coimbra, N. C., Morato, S., & Brandao, M. L. (19
mbens of rats sub
. Experimental brain re
Hirnforschung, 129, 260-268.
als-Kubik, R., Herz, A., & Shippenberg, T. S. (1989). Evidence that
the aversive effects of opioid antagonists and kappa-agonists are cen-
trally mediated. Psychopharmacology, 98, 203-206.
Copyright © 2013 SciRes. 17
J. L. GAO ET AL.
Blcantara, A. A., & Green-
United States of
lack, J. E., Isaacs, K. R., Anderson, B. J., A
ough, W. T. (1990). Learning causes synaptogenesis, whereas motor
activity causes angiogenesis, in cerebellar cortex of adult rats. Pro-
ceedings of the National Academy of Sciences of the
America, 87, 5568-5572.
rown, S. M., Napper, R. M., Thompson, C. M., & Mercer, A. R. (2002).
Stereological analysis reveals striking differences in the structural
plasticity of two readily identifiable glomeruli in the antennal lobes
of the adult worker hone
ybee. Journal of Neuroscience, 22, 8514-
Devoogd, T. J., Nixdorf, B., & Nottebohm, F. (1985). Synaptogenesis
and changes in synaptic morphology related to acquisition of a new
behavior. Brain Research, 329, 304-308.
yson, S.D E., & Jones, D. G. (1980). Quantitation of terminal parame-
ters and their inter-relationships in maturing central synapses: A per-
spective for experimental studies. Brain Research, 183, 43-59.
E. L., & Hjeds, H. bert, B., Thorkildsen, C., Andersen, S., Christrup, L
(1998). Opioid analgesics as noncompetitive N-methyl-D-aspartate
(NMDA) antagonists. Biochemical Pharmacology, 56, 553-559.
ttenberg, A., & Bernardi, R. E. (2007). Effects oEf buspirone on the
immediate positive and delayed negative properties of intravenous
cocaine as measured in the conditioned place preference test. Phar-
macology, Biochemistry, and Behavior, 87, 171-178.
Goto, Y., & Grace, A. A. (2005). Dopamine-dependent interactions
between limbic and prefrontal cortical plasticity in the nucleus ac-
cumbens: Disruption by cocaine sensitization. Neuron, 47, 255-266.
uldner, F. H., & Ingham, C. A. (1980). IncreGase in postsynaptic den-
sity material in optic target neurons of the rat suprachiasmatic nu-
cleus after bilateral enucleation. Neuroscience Letters, 17, 27-31.
arris, K. M., & Sultan, P. (1995).Variation in tHhe number, location
and size of synaptic vesicles provides an anatomical basis for the
nonuniform probability of release at hippocampal CA1 synapses.
Neuropharmacology, 34, 1387-1395.
Jensen, F. E., & Harris, K. M. (1989). Preservation of neuronal ultra-
structure in hippocampal slices using rapid microwave-enhanced
fixation. Journal o f Neuroscience Methods, 29, 217-230.
Joural study into the nes, D. G., & Devon, R. M. (1978). An ultrastruct
effects of pentobarbitone on synaptic organization. Brain Research,
147, 47-63. http://dx.doi.org/10.1016/0006-8993(78)90771-0
lintsova, A. Y., Goodlett, C. R., & Greenough, W. T. (20
peutic motor training ameliorates cerebellar effect
s of postnatal binge
alcohol. Neurotoxicology and Teratology, 22, 125-132.
oob, G. F. (1992). Drugs of abuse: anatomy, pharmacology anKd func-
tion of reward pathways. Trends in Pharmacological Sciences, 13,
auk, M. D., Peroutka, S. J., & Kocsis, J. D. (1988). Busp
ates synaptic activation of hippocampal pyramidal
cells. Journal of
M S., Rozenfeld, R., Dolios, G., Wang, R., &
Neuroscience, 8, 1-11.
orinville, A., Cahill, C. M., Esdaile, M. J., Aibak, H., Collier, B.,
Kieffer, B. L., & Beaudet, A. (2003). Regulation of δ-opioi
trafficking via µ-opioid receptor stimulation: Evidence from µ-opioid
receptor knock-out mice. Journal of Neuroscience, 2 3, 4888-4898.
oron, J. A., Abul-Husn, N.
Devi, L. A. (2007). Morphine administration alters the profile of hip-
pocampal postsynaptic density-associated proteins: A proteomics
study focusing on endocytic proteins. Molecular & Cellular Proteo-
mics, 6, 29-42. http://dx.doi.org/10.1074/mcp.M600184-MCP200
Morris, R. G., Moser, E. I., Riedel, G. Martin, S. J., Sandin, J., Day, M.,
& O'Carroll, C. (2003). Elements of a neurobiological theory of the
hippocampus: The role of activity-dependent synaptic plasticity in
memory. Philosophical Transactions of the Royal Society of London,
358, 773-786. http://dx.doi.org/10.1098/rstb.2002.1264
ucha, R. F., & Herz, A. (1985). Motivational properties of kappa and
mu opioid receptor agonists studied with place and taste preference
conditioning. Psychopharmacology, 86, 274-280.
Riblet, L. A., Eison, A. S., Eison, M. S., Taylor, D. P., Temple, D. L.,
& VanderMaelen, C. P. (1986). Neuropharmacology of buspirone.
Psychopathology, 17, 69-78. http://dx.doi.org/10.1159/000284133
Sevich, V. A., Bogdanova, O.
afiulina, V. F., Kas'yanov, A. M., Mark
G., Dvorzhak, A. Y., Zosimovskii, V. A., & Ezrokhi, V. L. (2005).
Studies of the synaptic plasticity of field CA3 of the hippocampus
during tetanization of the perforant path. Neuroscience and Beha
ioral Physiology, 35, 693-698.
almanzadeh, F., Fathollahi, Y., Semnanian, S., Shafizadeh, M., & Ka-
zemnejad, A. (2003). Dependence on morphine leads to a prominent
sharing among the different mech
the CA1 region of rat hippocampus. Brain rese
anisms of long-term potentiation in
arch, 963, 93-100.
cheuermann, D. W., Krammer, H. J., Timmermans, J. P., Stach, W.,
Adriaensen, D., & De Groodt-Lasseel, M. H. (1991). Fine structure
of morphologically well-defined type II neurons in the enteric nerv
ous system of the porcine small intestine revealed
tivity for calcitonin gene-related peptide. Acta Anatomica, 142, 236-
hakesby, A. C., Anwyl, R., & Rowan, M. J. (2002). Overcoming the
effects of stress on synaptic plasticity in the intact hippocampus:
rapid actions of serotonergic and antidepressant agents. Journal of
Neuroscience, 22, 3638-3644.
Shepherd, G. M., & Harris, K. M. (1998). Three-dimensional structure
and composition of CA3-->CA1 axons in rat hippocampal slices:
Implications for presynaptic connectivity and compartmentalization.
Journal of Neuroscience, 18, 830
Sterio, D. C. (1984). The unbiased estimation of number and sizes of
arbitrary particles using the disector. Journal of Microscopy, 134,
wift, R. M., & Stout, R. L. (1992). The relationship bSetween craving,
anxiety, and other symptoms in opioid withdrawal. Journal of Sub-
stance Abuse, 4, 19-26.
Turner, A. M., & Greenough, W. T. (1985). Differential rearing effects
on rat visual cortex synapses. I. Synaptic and neuronal density and
synapses per neuron. Brain Research, 329, 195-203.
Trulson, M. E., & Arasteh, K. (1986). Buspirone decreases the activity
of 5-hydroxytryptamine-containing dorsal raphe neurons in-vitro.
The Journal of Pharmacy and Phar ma col ogy , 38, 380-382.
Weibel, E. R., Kistler, G. S., & Scherle, W. F. (1966). Practical stereo-
logical methods for morphometric cytology. The Journal of Cell Bi-
ology, 30, 23-38. http://dx.doi.org/10.1083/jcb.30.1.23
ithers, G. S., Fahrbach, S. E., & Robinson, S. E. (19W93). Selective
neuroanatomical plasticity and division of labour in the honeybee.
Nature, 364, 238-240. http://dx.doi.org/10.1038/364238a0
occa, F. D. (1990). Neurochemistry and neurophysioloYgy of buspi-
rone and gepirone: Interactions at presynaptic and postsynaptic 5-
HT1A receptors. Journal of clinical psychopharmacology, 10, 6S-
Yuan, X. R., Madamba, S., & Siggins, G. R. (1992). Opioid peptides
reduce synaptic transmission in the nucleus accumbens. Neurosci-
ence Letters, 134, 223-228.
Zhang, Z., & Schulteis, G. (2008). Withdrawal from acute morphine
dependence is accompanied by increased anxiety-like behavior in the
elevated plus maze. Pharma
cology, Biochemistry, and Behavior, 89,
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