Original Article | J Adv Med Biomed Res. 2022; 30(143): 561-565
Journal of Advances in Medical and Biomedical Research | ISSN:2676-6264
Suppressive Effects of the Aerial Parts of Datura Stramonium L. Extract on
Naloxone-Precipitated Morphine Withdrawal Signs in Mice
Sobhan Kasraeifar1
, Amin Mokhtari-Zaer2
, Narges Marefati3
, Hassan Rakhshandeh4
,
1,5
Mahmoud Hosseini
1.
2.
3.
4.
5.
Psychiatry and Behavioral Sciences Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
Torbat Jam Faculty of Medical Sciences, Torbat Jam, Iran
Dept. of Physiology and Medical Physics, School of Medicine, Baqiyatallah University of Medical Sciences, Tehran,
Iran
Pharmacological Research Center of Medicinal Plants, Mashhad University of Medical Sciences, Mashhad, Iran
Applied Biomedical Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
Article Info
ABSTRACT
10.30699/jambs.30.143.561
Received: 2021/07/17;
Accepted: 2022/06/22;
Published Online: 10 Oct 2022;
Use your device to scan and read the
article online
Corresponding Information:
Narges Marefati,
Dept. of Physiology and Medical
Physics,
School
of
Medicine,
Baqiyatallah University of Medical
Sciences, Tehran, Iran
E-Mail: marefatin941@mums.ac.ir
Background & Objective: Datura stramonium L. is a medicinal herb from the
family of Solanaceae. It has been used in herbal remedies for promoting health and
treating several diseases. The current study was set up to compare the effects of
Datura stramonium L. extract on the naloxone-precipitated opiate-withdrawal in
mice.
Materials & Methods: Male BALB/c mice (30–35 g, n = 40) were arbitrarily
separated into 4 groups. The control group received morphine and normal saline and
other groups received three doses of D. stramonium extract (10, 20, or 30 mg/kg,
intraperitoneally, i.p.). Physically dependent was made by the administration of
morphine in increasing doses (50-75 mg/kg, i.p.). The withdrawal signs were elicited
by intraperitoneal injections of naloxone (5 mg/kg) 2 h after the last injection of
morphine.
Results: Administration of D. stramonium extract in doses of 20 and 30 mg/kg
markedly diminished the jumping numbers compared to the control group (P<0.05).
All three doses of D. stramonium extract could significantly suppress the increase in
climbing (P<0.05, P<0.001, and P<0.001, respectively) and diarrhea (P<0.001). D.
stramonium in higher doses (20 or 30 mg/kg) significantly decreased rearing and
itching (P<0.001).
Conclusion: The study findings suggest that D. stramonium extract is effective in
alleviating the signs of morphine withdrawal. Additional research is needed to
determine the exact mechanisms underlying D. stramonium for inhibiting morphine
withdrawal syndrome.
Keywords: Morphine Withdrawal Signs, Naloxone, Datura stramonium extract,
Opioid Addiction
Copyright © 2022, This is an original open-access article distributed under the terms of the Creative Commons Attribution-noncommercial 4.0 International License which permits
copy and redistribution of the material just in noncommercial usages with proper citation.
Introduction
Opioids, including morphine, are the most powerful
medicines now available for pain management; however,
morphine abuse often results in various behavioral and
physiological effects. The phenomenon of withdrawal is
one common feature of many abused drugs caused by the
abrupt cessation of drug administration (1). Thus, the
abstinence syndrome that results from finishing of the
drug is called withdrawal abstinence (2).
Sudden cessation of morphine usage or precipitation via
the systemic injection of an opioid receptor antagonist will
lead to the appearance of signs and symptoms of
withdrawal, such as diarrhea, rhinorrhea, abdominal
cramping, increased blood pressure, sweating, insomnia,
elevated heart rate, dysphoria, irritability, and
Volume 30, November-December 2022
hyperalgesia. The initiation and longevity of the
withdrawal signs differ and they are believed to be
associated with the pharmacokinetics of the opioid.
Indeed, it has been demonstrated that withdrawal
syndrome appeared in 24 h and lasted for 7–10 days (3).
Despite considerable research to date, the complexity of
opioid addiction and withdrawal is unclear and the
prospects for easy solutions are lacking. Currently,
buprenorphine, methadone, α 2-adrenoceptor agonists
(e.g. clonidine) are appropriate for opioid detoxification.
In addition, naltrexone (an opioid antagonist) is also
consumed (4). However, different types of medications
are used in the treatment of opioid dependence have main
limitations in safety and efficacy. Therefore, the search for
novel agents that are efficacious against opioid
Journal of Advances in Medical and Biomedical Research
562 Aerial Parts of Datura Stramonium L. Alleviate Morphine Withdrawal
dependence and withdrawal syndrome has become
increasingly important. From this point of view, the
natural extracts, which are optioned by medicinal plants,
are already being used for the treatment of drug addictions
and withdrawal management (5, 6). It has been previously
reported that medicinal plants such as Hypericum
perforatum, Avena sativa, Passiflora incarnata, and
Valeriana officinalis have analgesic, antispasmodic,
antianxiety, and hypnotic effects and can improve the
symptoms of morphine withdrawal (7). Datura
stramonium L. is an annual herb from a family of
Solanaceae that grows wild in various regions, especially
in temperate areas of the world. In traditional medicine,
the flowers and leaves of the plant are applied in the
treatment of Asthma. Interestingly, D. stramonium is used
as a sedative and hallucinogenic agent (8). The analysis of
the phytochemicals of the plant exhibited that D.
stramonium comprised of atropine, hyoscyamine, and
scopolamine. Previous studies reported that D.
stramonium has diverse biological features including antiinflammatory, antioxidant, and anticancer activities (9).
As suggested by these properties, growing evidence
indicates neuroprotective actions of Datura against a
number of insults (10, 11). However, the behavioral
effects of plant extracts of D. stramonium on the
withdrawal syndrome are not clearly understood.
Therefore, D. stramonium was hypothesized to reduce the
intensity of morphine withdrawal syndrome in rats.
Materials and Methods
Materials
Morphine sulfate was provided by Temad Company,
Tehran, Iran. Naloxone hydrochloride was kindly
provided by Tolid Daru Company, Tehran, Iran.
Preparation of extract
Datura stramonium L. was purchased from a local
herbal shop at Mashhad, Khorasan Razavi Province, Iran.
Then was identified by the Herbarium of School of
Pharmacy at Mashhad University of Medical Sciences,
Mashhad (Voucher specimen: 13261). To prepare a
hydro-alcoholic extract, 100g of the dried aerial parts of
the plant including seed, calyx, and petal was powdered
and mixed with 70% ethanol in a Soxhlet apparatus for
48h. The process is as follows: first, the dried and ground
aerial parts of the plant were packed in a thimble whit
filter paper. Solvent or 70% ethanol was added to balloon
volume which was gradually heated through the heater.
Finally, the solvent began to evaporate. The resulting
vapors flow to the condenser and the distillation operation
began in this part. Drops of solvent condensate from the
condenser were poured on the plant sample. Under these
conditions, the solvent was in direct contact with the plant
and the extraction operation began. This cycle continued
until the solvent color was clear inside the extraction
chamber, or in other words, the condition for stopping the
extraction process was that the solvent was clear in the
extraction chamber. The obtained extract was then filtered
and dehydrated in a water bath and kept at -20°C.
Volume 30, November-December 2022
Animals
Male BALB/c mice (6–8 weeks of age, 30–35 g, from
the animal house of Mashhad University of Medical
Sciences, Mashhad, Iran) were kept in pathogen-free
conditions, with a 12/12-h light/dark cycle with food and
water available ad libitum. All experiments were
approved by the Animal Research Group of Mashhad
University
of
Medical
Sciences
(NO:
IR.MUMS.MEDICAL.REC.1398.402.). All efforts were
made to minimize both the number of animals used and
the suffering caused.
Induction of morphine dependence
Continual subcutaneous injections of morphine in doses
of 50, 50, and 75 mg/kg three times daily at 9 a.m. (50
mg/kg), 1 p.m. (50 mg/kg), and 5 p.m. (75 mg/kg) for
three days were done for the generation of morphine
dependence in animals. On the fourth day, to prevent
overnight withdrawal syndrome in mice only a single
dose (50 mg/kg) of morphine was used 2 hours before
injection of naloxone (12).
Naloxone-precipitated withdrawal scoring
Two hours after the injection of the last dose of
morphine on the fourth day, a single intraperitoneal dose
of naloxone (5mg/kg, i.p.) was injected into all mice.
Following naloxone administration, animals were located
individually in a plexiglass cylinder (30 cm diameter,
30 cm high). Then withdrawal signs in animals including
jumping, climbing, itching, rearing, and the number of
times of diarrhea were recorded with a camera for 30
minutes and analyzed by the researcher. The number of
symptoms of opioid withdrawal for each mouse was
summed over 30 min (12).
Experimental design and animal grouping
40 adult male mice were randomly assigned to the four
groups (10 per group): the control group, Datura
stramonium L. (10 mg/kg), Datura stramonium L. (20
mg/kg), and Datura stramonium L. (30 mg/kg) (13). In the
Datura treatment groups, Datura stramonium L. was
administered 30 min before the injection of the last dose
of morphine (50mg/kg, i.p.) on the fourth day, whereas
the control group was treated with an equal among of
normal saline.
Statistical analysis
Data were expressed as the mean ± SEM. All analysis
was carried out using SPSS software version 19 for
Windows. Mean values were appropriately calculated and
compared using a one-way analysis of variance
(ANOVA) followed by the Bonferroni test. P<0.05 was
considered statistically significant.
Ethical Considerations
Ethical approval of our study was gotten by the ethical
committee of Mashhad University of Medical Sciences
(MUMS)
with
ethic
code
number:
IR.MUMS.MEDICAL.REC.1398.402.
Journal of Advances in Medical and Biomedical Research
Sobhan Kasraeifar et al. 563
Results
In morphine-dependent mice, i.p. injection of
naloxone elicited withdrawal signs. As shown in Figure
1, the jumping sign was induced when naloxone was
applied, however, treatment with different doses of D.
stramonium extracts showed a significant decrease in
naloxone-induced jumping. In doses of 20 and 30 mg/kg
of the extract, the occurrence of jumping significantly
diminished compared to the control group (P<0.05). In
addition, Datura extract in doses of 20 and 30 mg/kg
significantly reduced the jumping more compared to 10
mg/kg of extract (P<0.05), (Figure 1). All doses of the
extract significantly reduced climbing in comparison
with control (P<0.05, P<0.001, and P<0.001,
respectively). Just 20 and 30 mg/kg of the extract
significantly reduced climbing compared with 10 mg/kg
of Datura extract (P<0.05 and P<0.001, respectively),
(Figure 2). Only 20 and 30 mg/kg of the extract
significantly decreased rearing versus Control
(P<0.001). In addition, both 20 and 30 mg/kg showed a
significant difference revealing more decrease
(P<0.001), (Figure 3). Itching was also similar to rearing,
and 20 and 30 mg/kg of Datura were lesser than control
(P<0.001) and 10 mg/kg of Datura extract (P<0.001),
(Figure 4). The rate of diarrhea was lower in three doses
of Datura extract versus control (P<0.001). There are no
significant differences between doses of 20 and 30
mg/kg in comparison to 10 mg/kg of extract, (Figure 5).
Figure 1. Effect of the various doses of D. stramonium
on naloxone-precipitated jumping in morphine-dependent
mice. The results were expressed as mean ± SEM (N = 10),
*P<0.05 versus the control group and + P<0.05 versus D.
stramonium 10 mg/kg group.
Figure 2. Effect of the various doses of D. stramonium
on naloxone-precipitated climbing in morphine-dependent
mice. The results were expressed as mean ± SEM (N = 10),
**P<0.01, ***P<0.001 versus control group and +P<0.05,
+++P<0.001 versus D. stramonium 10 mg/kg group.
Figure 3. Effect of the various doses of D. stramonium
on naloxone-precipitated rearing in morphine-dependent
mice. The results were expressed as mean ± SEM (N = 10),
***P<0.001 versus control group and +++P<0.001 versus
D. stramonium 10 mg/kg group.
Figure 4. Effect of the various doses of D. stramonium
on naloxone-precipitated itching in morphine-dependent
mice. The results were expressed as mean ± SEM (N = 10),
***P<0.001 versus control group and +++P<0.001 versus
D. stramonium 10 mg/kg group.
Figure 5. Effect of the various doses of D. stramonium
on naloxone-precipitated diarrhea in morphine-dependent
mice. The results were expressed as mean ± SEM (N = 10),
***P<0.001 versus the control group.
Discussion
The current study is the first report describing the
effect of D. stramonium extract on the withdrawal signs
in morphine-dependent mice. This study indicates
escalating doses of morphine over a four-day period
followed by injection of naloxone induces some
morphine abstinence behavior in mice. This animal
Volume 30 November-December 2022
Journal of Advances in Medical and Biomedical Research
564 Aerial Parts of Datura Stramonium L. Alleviate Morphine Withdrawal
model was conducted to investigate the potency of the
extract to diminish morphine withdrawal behaviors
such as jumping, rearing, climbing, itching, and
diarrhea. In the present investigation, all doses of D.
stramonium extract 1 hour after the last dose of
morphine exhibited an inhibitory effect against
withdrawal symptoms of morphine in mice. The cause
and the mechanisms underlying the beneficial effects
of the D. stramonium extract on the withdrawal signs
in morphine-dependent mice were not examined in the
current study and need to be explored in details in
future studies. Nevertheless, several possible
mechanisms could be postulated to explain these
findings. The phytochemical studies showed that D.
stramonium had atropine, tropane alkaloids, and
scopolamine (14). It has been revealed that the
alcoholic extract of D. stramonium extract has
analgesic properties, which was probably mediated by
reinforcing the opioid system (15). This suggests that
the extracts of D. stramonium may cooperate with the
opioid system, a theory that is confirmed by the
evidence that the antinociceptive effects of the extracts
were suppressed by the opioid blocker naloxone.
Some studies revealed that alkaloids like tannins,
saponins, and glycosides, which are principal bioactive
constituents of most plants, could be responsible for
several plant biological activities. Phytochemical
investigations revealed that the extract of D.
stramonium included saponins, alkaloids, steroids,
tannins, phenols, flavonoids, and glycosides (11). The
findings of our study are in line with the results of prior
research that has shown that polyphenolic compounds
exert a suppressive effect on morphine withdrawal
signs (16). Thus, it may be expected that the flavonoids,
the main component of D. stramonium, are effective in
decreasing the severity of the withdrawal signs in
morphine-dependent animals.
It is possible to speculate that the effect of D.
stramonium extracts in attenuating morphine
withdrawal behavior is relevant to its antidepressant
proprieties (17). It has been described that treatment
with extract of Datura fastuosa elicits a robust
antidepressant profile in low doses and acts as an
antidepressant agent (18). In this regard, it is believed
that the D. stramonium extracts could enhance
bioavailability and increase neurotransmission of
serotonergic, noradrenergic, and monoaminergic
systems in the brain. In line with this, activation of
these systems has been implicated in limiting the
severity of morphine withdrawal (19). Besides, other
mechanisms may also be involved in reducing
withdrawal
syndrome.
D.
stramonium
via
scopolamine, a constituent in the plant, acts as an
antagonist of muscarinic cholinergic receptors (20).
For example, Large and colleagues have mentioned
that brain levels of ACh are elevated during withdrawal
(21). Thus, the extract may modulate withdrawal
syndrome by this mechanism. Furthermore, in an
experimental study, evidence showed that injections of
morphine for several days to mice led to induction of
Volume 30, November-December 2022
oxidative stress in brain tissues (21). In addition,
administration of naloxone in morphine-dependent
animals produced a significant reduction in the activity
of brain intracellular antioxidant enzymes and notable
elevations of malondialdehyde (MDA) concentration
in the brain (21). In addition, it has been reported that
the D. stramonium leaf extracts showed an effective
antioxidant activity (22). In support of this hypothesis,
it will probably be considered that the D. stramonium
extract attenuated morphine withdrawal by an
antioxidant mechanism.
Conclusion
In summary, the present study for the first time
demonstrated that D. stramonium extract could prevent
some major signs of morphine withdrawal in animal
models of addiction in mice. However, the exact
mechanism by which D. stramonium extract promotes
its beneficial effects is not completely clear and further
clinical and biochemical studies are needed to verify its
precise mechanism of action.
Acknowledgments
We thank the Medical Sciences of Mashhad, Iran, for
their continued support in carrying out this project.
Conflict of Interest
The authors declare no potential conflicts of interest.
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How to Cite This Article:
Kasraeifar S, Mokhtari-Zaer A, Marefati N, Rakhshandeh H, Hosseini M. Suppressive Effects of the Aerial Parts
of Datura Stramonium L. Extract on Naloxone-Precipitated Morphine Withdrawal Signs in Mice. J Adv Med
Biomed Res. 2022; 30(143): 561-5.
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Journal of Advances in Medical and Biomedical Research