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Passiflora incarnata attenuation of neuropathic allodynia and vulvodynia apropos GABA-ergic and opioidergic antinociceptive and behavioural mechanisms

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  • CECOS University Peshawar

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Background Passiflora incarnata is widely used as an anxiolytic and sedative due to its putative GABAergic properties. Passiflora incarnata L. methanolic extract (PI-ME) was evaluated in an animal model of streptozotocin-induced diabetic neuropathic allodynia and vulvodynia in rats along with antinociceptive, anxiolytic and sedative activities in mice in order to examine possible underlying mechanisms. MethodsPI-ME was tested preliminary for qualitative phytochemical analysis and then quantitatively by proximate and GC-MS analysis. The antinociceptive property was evaluated using the abdominal constriction assay and hot plate test. The anxiolytic activity was performed in a stair case model and sedative activity in an open field test. The antagonistic activities were evaluated using naloxone and/or pentylenetetrazole (PTZ). PI-ME was evaluated for prospective anti-allodynic and anti-vulvodynic properties in a rat model of streptozotocin induced neuropathic pain using the static and dynamic testing paradigms of mechanical allodynia and vulvodynia. ResultsGC-MS analysis revealed that PI-ME contained predominant quantities of oleamide (9-octadecenamide), palmitic acid (hexadecanoic acid) and 3-hydroxy-dodecanoic acid, among other active constituents. In the abdominal constriction assay and hot plate test, PI-ME produced dose dependant, naloxone and pentylenetetrazole reversible antinociception suggesting an involvement of opioidergic and GABAergic mechanisms. In the stair case test, PI-ME at 200 mg/kg increased the number of steps climbed while at 600 mg/kg a significant decrease was observed. The rearing incidence was diminished by PI-ME at all tested doses and in the open field test, PI-ME decreased locomotor activity to an extent that was analagous to diazepam. The effects of PI-ME were antagonized by PTZ in both the staircase and open field tests implicating GABAergic mechanisms in its anxiolytic and sedative activities. In the streptozotocin-induced neuropathic nociceptive model, PI-ME (200 and 300 mg/kg) exhibited static and dynamic anti-allodynic effects exemplified by an increase in paw withdrawal threshold and paw withdrawal latency. PI-ME relieved only the dynamic component of vulvodynia by increasing flinching response latency. Conclusions These findings suggest that Passiflora incarnata might be useful for treating neuropathic pain. The antinociceptive and behavioural findings inferring that its activity may stem from underlying opioidergic and GABAergic mechanisms though a potential oleamide-sourced cannabimimetic involvement is also discussed.
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R E S E A R C H A R T I C L E Open Access
Passiflora incarnata attenuation of
neuropathic allodynia and vulvodynia
apropos GABA-ergic and opioidergic
antinociceptive and behavioural mechanisms
Urooj Aman
1
, Fazal Subhan
1*
, Muhammad Shahid
1
, Shehla Akbar
1
, Nisar Ahmad
1
, Gowhar Ali
1
, Khwaja Fawad
1
and Robert D. E. Sewell
2
Abstract
Background: Passiflora incarnata is widely used as an anxiolytic and sedative due to its putative GABAergic
properties. Passiflora incarnata L. methanolic extract (PI-ME) was evaluated in an animal model of streptozotocin-
induced diabetic neuropathic allodynia and vulvodynia in rats along with antinociceptive, anxiolytic and sedative
activities in mice in order to examine possible underlying mechanisms.
Methods: PI-ME was tested preliminary for qualitative phytochemical analysis and then quantitatively by proximate
and GC-MS analysis. The antinociceptive property was evaluated using the abdominal constriction assay and hot
plate test. The anxiolytic activity was performed in a stair case model and sedative activity in an open field test. The
antagonistic activities were evaluated using naloxone and/or pentylenetetrazole (PTZ). PI-ME was evaluated for
prospective anti-allodynic and anti-vulvodynic properties in a rat model of streptozotocin induced neuropathic pain
using the static and dynamic testing paradigms of mechanical allodynia and vulvodynia.
Results: GC-MS analysis revealed that PI-ME contained predominant quantities of oleamide (9-octadecenamide),
palmitic acid (hexadecanoic acid) and 3-hydroxy-dodecanoic acid, among other active constituents. In the
abdominal constriction assay and hot plate test, PI-ME produced dose dependant, naloxone and pentylenetetrazole
reversible antinociception suggesting an involvement of opioidergic and GABAergic mechanisms. In the stair case
test, PI-ME at 200 mg/kg increased the number of steps climbed while at 600 mg/kg a significant decrease was
observed. The rearing incidence was diminished by PI-ME at all tested doses and in the open field test, PI-ME
decreased locomotor activity to an extent that was analagous to diazepam. The effects of PI-ME were antagonized
by PTZ in both the staircase and open field tests implicating GABAergic mechanisms in its anxiolytic and sedative
activities. In the streptozotocin-induced neuropathic nociceptive model, PI-ME (200 and 300 mg/kg) exhibited static
and dynamic anti-allodynic effects exemplified by an increase in paw withdrawal threshold and paw withdrawal
latency. PI-ME relieved only the dynamic component of vulvodynia by increasing flinching response latency.
Conclusions: These findings suggest that Passiflora incarnata might be useful for treating neuropathic pain. The
antinociceptive and behavioural findings inferring that its activity may stem from underlying opioidergic and
GABAergic mechanisms though a potential oleamide-sourced cannabimimetic involvement is also discussed.
Keywords: Passiflora incarnata, Allodynia, Vulvodynia, GABA receptors, Opioid receptors, Neuropathic pain
* Correspondence: fazal_subhan@upesh.edu.pk
1
Department of Pharmacy, University of Peshawar, Peshawar 25120, Khyber
Pakhtunkhwa, Pakistan
Full list of author information is available at the end of the article
© 2016 Aman et al. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77
DOI 10.1186/s12906-016-1048-6
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Background
Pain is an unpleasant sensory and emotional experience
associated with actual or potential tissue damage or de-
scribed in terms of such damage [1]. The phenomenon
of pain may be nociceptive or neuropathic in nature,
and caused by damage to non-neural or neuronal tissues
respectively [2, 3]. Neuropathic pain is a major cause of
morbidity and has a profound impact on patient well-
being. It involves the sensation of allodynia; a painful
sensation to a normally non-noxious stimulus and
hyperalgesia; an exaggerated pain response to a normally
noxious stimulus [4]. Neuropathic pain results from
various causes that affect the central nervous system in-
cluding multiple sclerosis, post stroke or spinal cord
pain. Alternatively, it may be associated with damage to
the peripheral nervous system, for instance, diabetic
neuropathy and trigeminal or post-herpetic neuralgia
[5]. Management of neuropathic pain poses an enor-
mous challenge due to the restricted efficacy of assorted
pharmacotherapies including both natural treatments
[68] and synthetic medicaments [9, 10] which are lim-
ited by the occurrence of side effects and the extent of
pain inhibition [11].
Passiflora incarnata L. (Additional file 1: Figure S1)
from the genus Passiflora (family: Passifloraceae) com-
monly known as Passion flower, is a fast growing peren-
nial vine widely spread in tropical and warm temperate
regions [12]. Phytochemical analysis of P. incarnata has
demonstrated that flavonoids constitute about 2.5 % of the
total phyto-constituents [13, 14] mainly present in the
leaves, the greatest concentration of flavonoid being
vitexin compared to the other species of its genus
[12, 15]. P. incarnata has been studied for its anal-
gesic [16], anxiolytic [1720], anticonvulsant [21], an-
titussive [22], aphrodisiac [23], anti-asthmatic [24],
anti-diabetic and hypolipidemic properties [25] along
with efficacy in the treatment of cannabinoid [26],
morphine [27], nicotine [28] and alcohol dependence
[29]. Traditionally, P. incarnata has been used for
curing various ailments like anxiety, insomnia, convul-
sions, sexual dysfunction, cough and cancer [30] and
is well known in relieving neuropathic conditions
[12]. In this regard, an eye wipe test has been con-
ducted suggesting a potential application in relieving
trigeminal neuralgia [31]. Clinical investigations on P.
incarnata have indicated effectiveness in the treat-
ment of anxiety [32, 33], insomnia [34], opioid with-
drawal [35], attention deficit hyperactivity disorder
[36] and postmenopausal symptoms [37].
Neuropathic pain results from a cascade of neurobio-
logical events that induces electrical hyperexcitability in
somatosensory conduction pathways and results in
hyperesthesia, dysesthesia, hyperalgesia, paresthesia or
allodynia [38]. Currently, the most common choices of
therapy for neuropathic pain are tricyclic antidepressants
and anticonvulsants [39, 40]. However, these therapies
are only partially effective and are usually accompanied
by a variety of side effects [41]. The use of complemen-
tary and alternative medicine has been shown to pro-
duce some beneficial effects in the management of
painful neuropathy [42] and several herbal medicines ex-
hibit promise in different types of experimentally in-
duced neuropathic pain models [6, 8, 4345]. Thus there
is some scope for new herbal medicines to combat neuro-
pathic pain syndromes [46]. The present study was there-
fore designed to evaluate the ameliorative effect of P.
incarnata methanolic extract (PI-ME) in an animal model
of streptozotocin-induced diabetic neuropathic allodynia
and vulvodynia [47] in rodents. Additionally, PI-ME in-
duced antinociceptive, anxiolytic and sedative activities
were also investigated using naloxone and pentylenetetra-
zole (PTZ) to probe its possible underlying mechanisms.
Methods
Chemicals
Morphine (Punjab Drug House, Lahore, Pakistan), diclo-
fenac sodium (98 %, Continental Chemicals Company
Pvt. Ltd. Pakistan), naloxone (98 %, Hangzhou Uniwise
International Co., Ltd, China), gabapentin (99 %, MKB
Pharmaceuticals Pvt Ltd Peshawar, Pakistan), diazepam
(Valium 10 mg/ 2 ml, Roche, Pakistan), pentylenetetra-
zole (98 %, Sigma Aldrich, UK) , streptozotocin (98 %,
Sigma Aldrich, UK) and commercial grade methanol
(Haq Chemicals Ltd Peshawar, Pakistan).
Preparation of Passiflora incarnata methanolic extract
P. incarnata whole plant was collected from the botanical
garden of the Department of Pharmacy, University of
Peshawar. It was authenticated by Prof. Dr. Mohammad
Ibrar of the Department of Botany, University of Peshawar
and a specimen was deposited in the herbarium with a vou-
cher number 20062 (PUP). The aerial parts were separated,
shade dried, and coarsely powdered (1000 g). It was macer-
ated for 7 days with commercial grade methanol (5 L). The
extract was filtered and concentrated under reduced pres-
sure at 60 °C in a rotary evaporator until a semisolid extract
containing no methanol was obtained (yield: 31.20 %).
Phytochemical analysis
PI-ME was preliminary evaluated by qualitative phyto-
chemical analysis [47] and was further screened by quanti-
tative analysis of flavonoids, alkaloids, saponins and tannins
[48, 49]. It was also subjected to gas chromatography/mass
spectrometry (GC/MS) analysis which was carried out on a
6890 N Agilent gas chromatograph coupled to a JMS
600 H JEOL mass spectrometer. The compound mixture
was separated on a fused silica capillary SPBI column,
30 m × 0.32 mm, 0.25 μm film thickness, in a temperature
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 2 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
programfrom50to25Cwitharateof4°C/minwith
2 min hold. The injector was at 260 °C and the flow rate of
the carrier gas, helium was 1 mL/min. The EI mode of the
JMS 600 H JEOL mass spectrometer had an ionization volt-
age of 70 eV, electron emission of 100 μA, ion source
temperature of 250 °C and analyzer temperature of 250 °C.
Samples were injected manually in split mode and the total
elution time was 90 min. MS scanning was performed from
m/z 85 to 390. Identification of the active constituents was
based on the computer evaluation of mass spectra of the
sample through NIST-based AMDIS (automated mass
spectral deconvolution and identification software), direct
comparison of peaks and retention times with those of
standard compounds as well as by following the character-
istic fragmentation patterns of the mass spectra of particu-
lar classes of compounds.
Animals
BALB/c mice (1826 g) and female Sprague Dawley rats
(150-200 g) maintained in a 12 h light/dark cycle at 22
± 2 °C were used in the experiments. Food and water
were provided ad libitum. Experiments on animals were
performed in accordance with the UK Animals (Scientific
Procedures) Act 1986 and according to the rules and
ethics set forth by the Ethical Committee of the Department
of Pharmacy, University of Peshawar. Approval for this
study was granted with the registration number: 06/EC-14/
Pharm (dated: April 06, 2014). The animal control groups
used in experiments were given normal saline which was
also the vehicle for all the drugs administered throughout
all the experiments.
Abdominal constriction assay
BALB/c mice (1822 g, n= 8 mice per group) of either
sex were injected with 0.6 % acetic acid (10 mL/kg, i.p)
to induce an abdominal constriction response [50, 51].
In the abdominal constriction assay, the mean incidence
of constrictions expressed as % protection across all
experiments was normalized relative to untreated con-
trols. PI-ME (150, 200 and 250 mg/kg, p.o), morphine
(5 mg/kg, i.p) or diclofenac (50 mg/kg, i.p) were
administered 30 min before acetic acid injection. In
the opioid antagonism study, the animals were pre-
treated with naloxone (0.5 mg/kg, s.c), 5 min before
acetic acid administration. Percentage protection was
calculated as:
%Protection ¼1Number of abdominal constrictions after
treatment=Number of abdominal constrictions
of untreated control100
Hot plate test
BALB/c mice (1822 g, n= 8 mice per group) of either
sex were pretested for their response latencies on a hot
plate (Harvard apparatus, USA) maintained at 54.0 ±
0.1 °C. The response end-point was signified by hind
limb flick, lick or jumping at which point animals were
immediately removed from the thermal nociceptive
stimulus in order to avoid any tissue damage or possibil-
ity of subsequent hyperalgesia. A cut-off time of 30 s
was imposed such that if they did not respond within
this latency period then they were immediately removed
from the hot plate stimulus [51]. Thirty minutes after
pretesting, the animals were administered PI-ME (100,
150, 200 mg/kg; p.o), morphine (5 mg/kg; i.p) or diclofe-
nac (50 mg/kg, i.p). In the antagonism studies, naloxone
(1.0 mg/kg, s.c) or PTZ (10 mg/kg, i.p) were administered
10 or 30 min respectively before treatment and the animal
response latencies were measured at 30, 60, 90 and
120 min. The percentage antinociception was calculated as:
%Antinociception ¼Test latency control latencyðÞ=
Cutoff time control latencyðÞ100
Anxiolytic activity (Staircase test)
BALB/c mice (1824 g, n= 8 mice per group) of either
sex were administered PI-ME (200, 400 and 600 mg/kg,
p.o) or diazepam (2 mg/kg, i.p). In the drug combination
experiments, PTZ (10 mg/kg, i.p) was administered
30 min prior to drug treatment. The number of rears
and steps climbed by each animal was observed for
3 min using the staircase apparatus and the methods de-
scribed by Simiand and coworkers [52]. A step was con-
sidered to be climbed only if the criterion was met
whereby an animal placed all four paws on the step.
Locomotor activity
BALB/c mice (1826 g, n= 6 mice per group) of either
sex were administered with PI-ME (200, 400 and
600 mg/kg, p.o) or diazepam (4 mg/kg, i.p). In the drug
combination experiments, PTZ (10 mg/kg, i.p) was ad-
ministered 30 min prior to drug treatment. Thirty min
later, the animals were placed in the recording apparatus
with a floor area of 50 × 40 cm divided into four equal
quadrants by lines. The number of lines crossed by each
animal was recorded for 30 min using a digital camera
(Cats Eye IR IP Camera, Taiwan) [53].
Streptozotocin induced neuropathic pain
Induction of mechanical allodynia and vulvodynia
Female Sprague Dawley rats (150200 g, n= 6 rats
per group) food withdrawn for 16 h were administered
streptozotocin (50 mg/kg, i.p) and food was immediately
provided. On the 5
th
day, animals exhibiting random
blood glucose levels greater than 270 mg/dl were in-
cluded in the study [54]. Body weights and blood glucose
were measured at specified time periods. The bedding
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 3 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
material was frequently changed to avoid any infection
due to excessive urination. On the 29
th
day post strepto-
zotocin administration, animals were transferred to wire
mesh cages and acclimatized for 1545 min. They were
then assessed for mechanical allodynia or vulvodynia be-
fore and after PI-ME or standard gabapentin administra-
tion using the von Frey up-down method [55].
Treatment schedule
Animals were divided into five groups. Group I received
normal saline and served as control. Group II remained
as the streptozotocin positive control group. Group III
received a single intraperitoneal dose of gabapentin
(75 mg/kg) and served as the standard. Group IV and V
were treated with PI-ME at doses of 200 and 300 mg/kg
respectively. The therapeutic doses of PI-ME for evalu-
ation in neuropathic pain were selected on the basis of
its analgesic, anxiolytic, locomotor and respective antag-
onistic activities.
Assessment of static and dynamic allodynia
Static allodynia was assessed using a series of von Frey fila-
ments (0.4, 0.6, 1, 1.4, 2, 4, 6, 8, 10, 15 g), starting with a
2.0 g force applied perpendicularly to the plantar surface of
the right hind paw for 5 s or until the animal displayed a
withdrawal response (lifting of the paw). Animals respond-
ing to 3.63 g force or below were included in the study and
15 g was selected as the cut-off force [54].
Dynamic allodynia was assessed by lightly stroking the
plantar surface of the hind paw with a cotton bud. Lift-
ing or licking the paw was considered as a withdrawal
response and the time taken to show a withdrawal reac-
tion was considered as the paw withdrawal latency
(PWL). Animals responding to the cotton bud within 8 s
were included in the study and 15 s was selected as the
cut off time [54].
Assessment of static and dynamic vulvodynia
Static vulvodynia was assessed by shaving the anogenital
area including the mons pubis. A series of von Frey fila-
ments (0.008, 0.02, 0.04, 0.07, 0.16, 0.4, 0.6, 1 g), were
applied perpendicularly to the mucous membrane of the
anogenital region for 4 s starting with a 0.04 g force,
until a flinching response occurred. Animals responding
to a 0.16 g force or below were included in the study
and a 1.0 g force was selected as the cut-off force [56].
Dynamic vulvodynia was assessed by lightly brushing a
cotton bud over the mucous membrane of the anogeni-
tal region for 10 s or until a flinching response occurred.
Animals showing a flinching response within 5 s were
included in the study and 10 s was selected as the cut-
off time [56].
Statistical analysis
Data were expressed as mean ± SEM. Statistical compari-
sons were carried out by one way ANOVA followed by
Dunnetts, Bonferroni or Tukeys multiple comparison tests
where appropriate using GraphPad Prism 5 (GraphPad
Software Inc. San Diego CA, USA). Statistical significance
was deduced at P0.05.
Results
Phytochemical analysis of Passiflora incarnata
Preliminary qualitative analysis of PI-ME disclosed the
presence of flavonoids, alkaloids, carbohydrates, tannins,
glycosides, fixed oils and saponins (Table 1). Subsequent
more detailed quantitative analysis revealed the presence
of flavonoids (72 %), saponins (10 %) and alkaloids
(13.4 %) in PI-ME. The major compounds obtained from
GC-MS analysis of PI-ME included: 9-Octadecenamide
(Oleamide) (C
18
H
35
NO, MW: 281), n-Hexadecanoic acid
(Palmitic acid) (C
16
H
32
O
2
, MW: 256), dodecanoic acid,
3-hydroxy- (C
12
H
24
O
3
, MW: 216), 4H-Pyran-4-one, 2,3-
dihydro-3,5-dihydroxy-6-methyl- (C
6
H
8
O
4
, MW: 144),
vitamin-E (C
29
H
50
O
2
, MW: 430), cis,cis,cis-7,10,13-Hexa-
decatrienal (C
16
H
26
O, MW: 234), β-Sitosterol (C
29
H
50
O,
MW: 414), 9,10-Secocholesta-5,7,10(19)-triene-3,24,25-triol,
(3β,5Z,7E)- (C
27
H
44
O
3
, MW: 416), pregnane-3,11,20,21-
tetrol, cyclic 20,21-(butyl boronate), (3α,5β,11β,20R)-
(C
25
H
43
BO
4
, MW: 418), ethyl 9-hexadecenoate (C
18
H
34
O
2
,
MW: 282), stigmasterol (C
29
H
48
O, MW: 412), octade-
canoic acid (C
18
H
36
O
2
, MW: 284), 2H-1-Benzopyran-
6-ol, 3,4-dihydro-2,8-dimethyl-2- (4,8,12-trimethyltri-
decyl)-, [2R-[2R*(4R*,8R*)]]- (C
27
H
46
O
2
, MW: 402),
and phytol (C
20
H
40
O, MW: 296) among other import-
ant constituents (Table 2 and Fig. 1).
Antinociceptive activity of Passiflora incarnata
Abdominal constriction assay (tonic visceral chemically-
induced nociception)
A significant attenuation (F
(5,42)
= 91.99, P< 0.001) of
acetic acid incited abdominal constriction was produced
by PI-ME at doses of 200 mg/kg (P< 0.01) and 250 mg/
kg (P< 0.001) compared to saline control. Similarly, a
significant increase (P< 0.001) in the percentage protec-
tion against abdominal constriction was observed with
diclofenac (50 mg/kg) and morphine (5 mg/kg) (Fig. 2a).
Naloxone (0.5 mg/kg) (F
(9,70)
= 44.75, P< 0.001) signifi-
cantly reversed the antinociceptive activity of morphine
(P< 0.001) and PI-ME (200 and 250 mg/kg) (P< 0.05)
but not that of diclofenac (50 mg/kg) as shown in
Fig. 2b.
Hot plate test (acute phasic thermal nociception)
In the hot plate test, 30 min after drug administration
(F
(5,42)
=200.2,P< 0.001) a marked increase in percentage
antinociception was observed with morphine (5 mg/kg)
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 4 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
(P< 0.001), diclofenac (50 mg/kg) (P< 0.05) and PI-ME at
adoseof200mg/kg(P< 0.05). After 60 min (F
(5,42)
=
55.36, P< 0.001), the increase in percentage response was
less significant (P< 0.05) for morphine whilst it was highly
significant (P< 0.001) for PI-ME (150 and 200 mg/kg), the
activity being retained in the latter case up to 90 min
(F
(5,42)
=36.61, P< 0.001, not shown). However, after
120 min (F
(5,42)
=4.352, P< 0.01) it was only PI-ME at
doses of 150 mg/kg (P< 0.05) and 200 mg/kg (P<0.01)
that afforded protection against thermal nociception
(Fig. 3). Naloxone (1.0 mg/kg) (F
(7,56)
=46.60, P<0.001)
reduced the % antinociceptive effect of both morphine
(P< 0.001) and PI-ME (150 and 200 mg/kg) (P<0.01)
(Fig. 4a). Pentylenetetrazole (10 mg/kg) (F
(7,56)
= 35.91,
P< 0.001) by way of contrast, significantly reduced
the antinociceptive effect of PI-ME only at the
150 mg/kg dose (P< 0.05) (Fig. 4b).
Anxiolytic-like activity of Passiflora incarnata
In the staircase test, there was a substantial increase in
the number of steps climbed (F
(4,25)
= 21.04, P< 0.001) in
response to both diazepam (2 mg/kg, P< 0.001) and PI-
ME (200 mg/kg, P< 0.05) versus the animal control
group treated with saline vehicle. However, at the high-
est dose (600 mg/kg) the passiflora extract significantly
reduced (P< 0.05) the number of steps climbed in com-
parison with the controls (Fig. 5a). In contrast, the num-
ber of rears (F
(4,25)
= 5.403, P< 0.01) was inhibited not
only by treatment with diazepam (P< 0.01) but also by
all three doses of PI-ME (200 and 400 mg/kg, P< 0.05;
600 mg/kg, P< 0.01) in comparison with the saline ve-
hicle controls (Fig. 5b). The post hoc test revealed that
there was no significant effect of pentylenetetrazole
(10 mg/kg) by itself on step climbing nor was there any
modification of the stair climbing responses when it was
administered in combination with diazepam or PI-ME
(Fig. 6a). However, it did reverse the decrement in rears
initiated by PI-ME (200, 400 and 600 mg/kg) and actu-
ally augmented (P< 0.05) the overall rearing incidence
(F
(9,50)
= 6.497, P< 0.001) as shown in Fig. 6b.
Sedative activity of Passiflora incarnata
Locomotor activity
In the locomotor activity study, there was a pronounced
reduction in cage line crossing instigated by both (F
(4,25)
=
15.39, P< 0.001) diazepam (4.0 mg/kg, P< 0.001) and PI-
ME at 400 mg/kg (P< 0.01) and 600 mg/kg (P<0.001)
though there was no significant motoric decline at the
lowest PI-ME dose (200 mg/kg, P> 0.05) (Fig. 7a). Penty-
lenetetrazole (10 mg/kg) (F
(7,40)
=26.88,P<0.001)blocked
(P< 0.05) the reduced locomotor effect of PI-ME (400
and 600 mg/kg) by increasing the incidence of line
crossing but it did not modify the diazepam loco-
motor regression (Fig. 7b).
Effect of Passiflora incarnata on mechanical allodynia and
vulvodynia
Animals administered a single streptozotocin (50 mg/kg)
treatment developed both static and dynamic allodynia
in their hind paws when tested 29 days later (Fig. 8).
Hence, there was a substantial decrease (P< 0.001) in
PWT and PWL in comparison with saline treated ani-
mals. One hour after PI-ME dosing in STZ-pretreated
animals on the test day, there was an ensuing increase in
PWT (F
(4,25)
= 31.41, P< 0.001) and PWL (F
(4,25)
= 20.25,
P< 0.001) observed for PI-ME at doses of 200 mg/kg
Table 1 Preliminary qualitative phytochemical analysis of Passiflora incarnata methanolic extract (PI-ME)
Sample Test Observation Result
1. Aqueous solution of PI-ME + 10 % ammonium hydroxide solution Appearance of yellow
coloration
Flavonoids present
2. A portion of PI-ME + few drops of Wagners reagent Reddish brown
precipitate
Alkaloids present
3. A small volume of PI-ME + 12 drops of Mayers reagent Creamy or white
precipitate
Alkaloids present
4. 0.5 ml PI-ME + 0.5 ml benedicts reagent mixed and boiled for 2 min Characteristic colored
precipitate
Carbohydrates
present
5. 1 ml PI-ME + 1 ml Barfoeds reagent boiled for 2 min Red precipitate Carbohydrates
present
6. 50 mg PI-ME + 5 ml distilled water + small amount of 5 % ferric chloride solution Intense green
coloration
Tannins and phenolic
compounds present
7. 50 mg PI-ME + conc. HCL heated on water bath for 2 h resultant hydrolysate filtered
2 ml hydrolysate + 3 ml chloroform chloroform layer separated out + 10 % ammonia
solution
Pink coloration Glycosides present
8. A small amount of PI-ME compressed between two pieces of filter paper Formation of oil spot
on filter paper
Fixed oils present
9. 50 mg PI-ME + 20 ml distilled water shaken for 15 min Formation of 2 cm
thick layer of foam
Saponins present
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 5 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
(P< 0.001, P<0.01)and300mg/kg(P<0.001). Simi-
larly, 2 h following treatment with PI-ME on the test
day in the STZ-pretreated group, there was a sizeable
increase in PWT (F
(4,25)
= 17.92, P< 0.001) noted at
doses of 200 mg/kg (P< 0.01) and 300 mg/kg (P<
0.001) of PI-ME. However, at the 2 h test, PWL
(F
(4,25)
= 59.63, P< 0.001) was increased only by the
300 mg/kg (P< 0.001) rather than the 200 mg/kg PI-
ME dose. Gabapentin (75 mg/kg) administered as a
positive control, also generated an alleviation of
mechanical allodynia by elevating (P< 0.001) PWT
and PWL compared to the 29-day streptozotocin
alone pretreated animals at both the 1 and 2 h testing
times (Fig. 8).
The animal group pretreated with streptozotocin by it-
self 29 days earlier expressed mechanical static and dy-
namic vulvodynia (P< 0.001) compared to the saline
vehicle treated controls on the test day (Fig. 9). It was
notable that PI-ME (200 and 300 mg/kg) did not modify
the diminished FRT (streptozotocin induced static vul-
vodynia) at either the 1 h (F
(4,25)
= 49.85, P< 0.001) or
2h(F
(4,25)
= 17.12, P< 0.001) test day readings. However,
there was a significant increase in streptozotocin-
shortened FRL (dynamic vulvodynia) within 1 h (F
(4,25)
= 27.38, P< 0.001) and 2 h (F
(4,25)
= 10.08, P< 0.001) of
PI-ME treatment at 200 mg/kg (P< 0.001, P< 0.05) and
300 mg/kg (P< 0.001, P< 0.01). The single test day posi-
tive control dose of gabapentin (75 mg/kg) alleviated
both mechanical static and dynamic vulvodynia at the 1
and 2 h readings as evidenced by significant increases in
FRT (P< 0.001, P< 0.01) and FRL (P< 0.001) versus the
streptozotocin alone pretreated animals.
Discussion
The antinociceptive activity of P. incarnata methanolic
extract (PI-ME) was evaluated in mice using the hot
Table 2 GC/MS analysis of Passiflora incarnata methanolic extract
Chemical constituent Formula Molecular weight R.T. (min) Percent abundance
10-Undecen-1-al, 2-methyl- C
12
H
22
O 182 8.465 0.377
1,3-Pentanediamine C
5
H
14
N
2
102 8.809 0.353
4H-Pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl- C
6
H
8
O
4
144 10.27 5.477
1-Pentanol, 2-methyl-, acetate C
8
H
16
O
2
144 10.83 0.621
1,2,6-Hexanetriol C
6
H
14
O
3
134 11.10 0.623
4-Cyclopropylcarbonyloxytridecane C
17
H
32
O
2
268 11.32 0.664
5-Cyclopropylcarbonyloxypentadecane C
19
H
36
O
2
296 11.60 0.796
9-Tetradecen-1-ol, acetate, (E)- C
16
H
30
O
2
254 12.87 0.488
trans-2-undecenoic acid C
11
H
20
O
2
184 15.98 0.438
Dodecanoic acid, 3-hydroxy- C
12
H
24
O
3
216 15.99 13.64
4-((1E)-3-Hydroxy-1-propenyl)-2-methoxyphenol C
10
H
12
O
3
180 18.64 0.393
d-Mannose C
6
H
12
O
6
180 18.86 0.378
7-Methyl-Z-tetradecen-1-ol acetate C
17
H
32
O
2
268 19.34 0.395
l-Gala-l-ido-octose C
8
H
16
O
8
240 19.54 0.406
n-Hexadecanoic acid; (Palmitic acid) C
16
H
32
O
2
256 23.68 21.86
Phytol C
20
H
40
O 296 30.20 1.004
9-Hexadecyn-1-ol C
16
H
30
O 238 31.33 0.956
cis,cis,cis-7,10,13-Hexadecatrienal C
16
H
26
O 234 31.81 2.175
Octadecanoic acid C
18
H
36
O
2
284 33.09 1.209
9-Octadecenamide, (Z)-; (Oleamide) C
18
H
35
NO 281 42.11 33.52
9,10-Secocholesta-5,7,10(19)-triene-3,24,25-triol, (3β,5Z,7E)- C
27
H
44
O
3
416 42.79 1.762
Pregnane-3,11,20,21-tetrol, cyclic 20,21-(butyl boronate), (3α,5β,11β,20R)- C
25
H
43
BO
4
418 42.98 1.422
2H-1-Benzopyran-6-ol, 3,4-dihydro-2,8-dimethyl-2-(4,8,12-trimethyltridecyl)-,
[2R-[2R*(4R*,8R*)]]-
C
27
H
46
O
2
402 43.69 1.033
Ethyl 9-hexadecenoate C
18
H
34
O
2
282 45.78 1.390
Vitamin E C
29
H
50
O
2
430 45.99 2.579
Stigmasterol C
29
H
48
O 412 48.01 1.229
β-Sitosterol C
29
H
50
O 414 49.08 1.776
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Content courtesy of Springer Nature, terms of use apply. Rights reserved.
plate test which is suitable for assessing centrally medi-
ated acute phasic nociception [57] and the acetic acid in-
duced abdominal constriction assay for tonic visceral
nociception [58, 59]. Mice were selected as the species
of choice in these specific tests because they are mani-
festly sensitive not only to opioid mediated effects but
also to coexistent non-steroidal anti-inflammatory drug
(NSAID) activity [51]. What is more, accumulating evi-
dence indicates that GABAergic transmission plays a
pivotal role in the inhibitory regulation of the nocicep-
tive process, and the murine abdominal constriction
assay as well as the hot plate test both detect dose
dependent GABA agonist antinociception in this species
[60, 61]. In both tests, diclofenac as a standard anti-
inflammatory analgesic and PI-ME produced antinocicep-
tive activity consistent with previous studies [16, 31, 62]. It
was notable that the antinociceptive effect of PI-ME was
reversed by the opioid- and GABA
A
- receptor antagonists,
naloxone and pentylenetetrazole (PTZ) respectively, sug-
gesting an involvement of opioidergic and GABAergic
mechanisms in the mediation of the antinociception. Opi-
oid agonists decrease pain transmission by activating de-
scending nerve fibers from the periaqueductal gray and
raphe nuclei supraspinally and also by inhibition of affer-
ent nerve transmission by binding to pre- and postsynap-
tic opioid receptors within the spinal cord dorsal horn
[63]. Furthermore, GABAergic neurons and receptors that
are intercalated within the spinal cord and higher brain
pathways are important for the origination, transmission,
and modification of pain impulses in such a way that alter-
ation of GABA transmission yields antinociception [64]. P.
incarnata has been shown to modulate the activity of
GABAergic and opioid systems [21] to produce central
analgesic activity as evaluated by a reduced duration of
paw licking in neurogenic and inflammatory nociceptive
phases in the formalin test [31]. Due to a prevalence of
GABA as a non-α-amino acid constituent of P. incarnata
extract [65], several of its pharmacological effects have
been ascribed to mediation via the GABA system. These
include not only affinity for GABA
A
but also GABA
B
re-
ceptors in addition to modification of GABA uptake [66].
The antinociceptive effects of both GABA
A
and GABA
B
receptor agonists are known to involve activation or inhib-
ition of other neurotransmitter or neuromodulator path-
ways [64] and it is evident that central GABAergic
systems are involved in opioid-mediated analgesia [67].
Thus, it is possible that administration of GABA receptor
agonists in combination with other agents may yield
GABA receptor-related therapies for the treatment of
acute and chronic pain [64].
The anxiolytic-like activity of PI-ME was assessed by
the incidence of rears or steps climbed in the stair case
test. The extract at a dose of 200 mg/kg significantly in-
creased the number of climbed steps, although at a
Fig 1 MS chromatogram of Passiflora incarnata methanolic extract
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 7 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Fig. 2 aAntinociceptive activity of Passiflora incarnata (PI-ME) in the mouse abdominal constriction assay. ***P< 0.001, **P< 0.01 compared to
saline vehicle control (ANOVA followed by Dunnettspost hoc test), (n= 8 mice per group). bEffect of naloxone (NXL-0.5) on the antinociceptive
activity of PI-ME in the mouse abdominal constriction assay. *P< 0.05, ***P< 0.001 compared to morphine (MOR-5), diclofenac (DIC-50) or PI-ME
(ANOVA followed by Bonferronis multiple comparison post hoc test), (n= 8 mice per group)
Fig. 3 Antinociceptive activity of Passiflora incarnata (PI-ME) in the mouse hot plate test. *P< 0.05, **P< 0.01, ***P< 0.001 compared to saline
vehicle control (ANOVA followed by Dunnettspost hoc test), (n= 8 mice per group)
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 8 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
higher dose (600 mg/kg) it decreased this parameter.
Similarly, the frequency of rears was diminished by the
extract at all three doses tested and this outcome was
blocked by PTZ. Anxiolytic-like activity has been shown
to be associated with an increase in the number of steps
climbed by mice whilst sedative activity is thought to be
linked to a decrease in the frequency of rears [18] and
this is the very reason why this paradigm was chosen in
this species to evaluate P. incarnarta. Other studies have
attributed an increased rearing incidence to an anxiety-
like behavior and a decreased number of steps climbed
to a sedative effect [68]. In conjunction with this, anxio-
lytic activity has been coupled with lower doses while
sedative effects have been related to higher doses of
plant extracts or reference drugs [69]. In this context,
PI-ME displayed an anxiolytic-like effect at 200 mg/kg,
while at 600 mg/kg it exhibited sedative activity. This
was also confirmed in the open field test where it was
observed that PI-ME decreased the number of lines
crossed at doses of 400 and 600 mg/kg comparable to
that of diazepam and these findings concur with the lit-
erature [1719, 70]. Since PTZ reversed the anxiolytic-
like and sedative actions of PI-ME, underlying GABA
mediated mechanisms may well be implicated. In a se-
lection of studies, the sedative and anxiolytic properties
of P. incarnata have been attributed to benzodiazepine
and GABA receptor mediated biochemical processes in
the body [18, 19, 71, 72], binding to GABA
A/B
sites and
Fig. 4 aEffect of naloxone (NXL-1); or bpentylenetetrazole (PTZ-10) on the antinociceptive effect of Passiflora incarnata (PI-ME) in the mouse hot
plate test. *P< 0.05, **P< 0.01, ***P< 0.001 compared to morphine (MOR-5) or PI-ME (150 or 200 mg/kg) (ANOVA followed by Bonferronis mul-
tiple comparison post hoc test), (n= 8 mice per group)
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 9 of 17
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inhibition of GABA uptake being of particular conse-
quence [66].
The modulatory effect of P. incarnata on GABAergic
and opioid systems may provide some insight into its
beneficial effect in various painful neuropathic condi-
tions. Neuropathy induced hypersensitivity is known to
involve disruption of tonic GABAergic transmission [73]
and GABA agonists and metabolic inhibitors have been
shown to be effective in various neuropathic nociceptive
models [7476]. Neuropathic pain has been reported to
be refractory to opioids [77, 78]. However, several studies
have shown that neuropathic pain can be attenuated by
morphine and other μ-opioid receptor agonists [7981]
and these reports suggest that local μ-opioid receptors on
the terminals of uninjured primary afferent nociceptive
neurons are an essential target for alleviating mechanical
allodynia. In the current study we have evaluated the
methanolic extract of P. incarnata in a novel streptozoto-
cin induced diabetic animal model of neuropathic pain
established exclusively in rats [56]. The results showed
that PI-ME (200 and 300 mg/kg) induced mechanical
anti-allodynic activity exemplified by an increase in paw
withdrawal threshold (PWT) and paw withdrawal latency
(PWL) 1 and 2 h post treatment. Similarly, PI-ME also re-
lieved dynamic vulvodynia by increasing the flinching re-
sponse latency (FRL) although the extract was devoid of
activity on the static component of vulvodynia. The inten-
sity of the PI-ME dynamic anti-vulvodynia response was
comparable to that of gabapentin which was used as a ref-
erence drug due to the fact that it has proven pain reliev-
ing effects in various neuropathic pain models [54].
Gabapentin also exhibits an established propensity to alle-
viate both static and dynamic components of allodynia
and vulvodynia [56] and the current study corroborates
this assertion. The present findings also indicate that the
behavioural and antinociceptive effects of PI-ME involve
GABAergic and opioidergic mechanisms because they
were reversed by PTZ and naloxone respectively. Conse-
quently, it might be inferred that analogous processes are
implicated in PI-ME anti-allodynic/vulvodynic activity and
this requires a direct focus of further study. In relation to
this notion, Ingale and Kasture [31] suggested that opioi-
dergic as well as the nicotinic cholinergic system are in-
volved in the central analgesic activity of butanolic P.
incarnata extract in the eye wipe test. This paradigm is
used to study trigeminal pain because corneal nociceptive
receptors have a large representation in the trigeminal
ganglion through the ophthalmic branch of the tri-
geminal nerve [82]. Moreover, it has been suggested
from radioligand binding studies that it is very un-
likely that P. incarnata acts via the benzodiazepine-site of
the GABA
A
-receptor [66]. In this connection, it has been
postulated that GABA
A
α1-sparing benzodiazepine-site li-
gands might constitute a class of analgesics suitable for
the treatment of chronic pain syndromes [83]. Further-
more, there is considerable evidence implicating an im-
portant role for diminished GABAergic tone in the
development of central sensitization and hyperalgesia in
neuropathic pain models [8486].
The phytochemical screening results of our study ver-
ify the presence of a preponderance of flavonoids as well
as alkaloids in P. incarnata as described elsewhere
[25, 87, 88]. Flavonoids are reported to be the major
phytoconstituents of P. incarnata and include chrysin,
vitexin, isovitexin, orientin, isoorientin, apigenin and
kampferol [14, 30, 89]. These polyphenolic metabo-
lites may play a role in the neuropharmacological ac-
tivity of several plants [9092] including P. incarnata
[18, 93, 94]. Additionally, flavonoids have been reported
to elicit an analgesic effect through opioid [95] as well as
GABAergic systems [96] and have a beneficial role in
relieving neuropathic pain conditions [9799]. Some
flavonoids like quercetin have also been identified in
P. incarnata extract [100] and are believed to be ef-
fective in diabetes mellitus induced peripheral neur-
opathy [101, 102] the activity being mediated through
an opioidergic mechanism [103]. The GCMS analysis
Fig. 5 Effect of Passiflora incarnata (PI-ME), and diazepam (2 mg/kg)
on athe number of steps climbed in the staircase test and bthe
incidence of rears in mice. *P< 0.05, **P< 0.01, ***P< 0.001 compared
to saline vehicle control (ANOVA followed by Dunnettspost hoc test),
(n= 8 mice per group)
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 10 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
in this study revealed that P. incarnata contains a pre-
dominance of the fatty acid amide 9-octadecenamide (also
known as oleamide), which has hypnotic, analgesic, and
anxiolytic actions [104]. Many of oleamides behavioural
effects stem from its activity on various receptor systems
including GABA
A
[105107], 5HT
1A
,5HT
2A
,5HT
2C
,
5HT
7
[108110], G-proteins [111], voltage gated sodium
channels [107, 112] and CB
1
receptors [113]. In this re-
spect, oleamide enhances GABA receptor activity and spe-
cifically potentiates the peak chloride current when applied
with GABA to benzodiazepine-sensitive GABA
A
receptors
[106]. The cannabimimetic action of oleamide resulting
from its agonist action at CB
1
receptors [110, 113] gives
rise to cannabinoid antagonist reversible antinociception
which is also sensitive to blockade by the GABA
A
antagonist bicuculline [104]. It has been posited that en-
dogenous fatty acid derivatives such as oleamide interact
with endocannabinoids like anandamide in the modulation
of pain sensitivity [114] which may well contribute to the
inhibitory effect of P. incarnata on allodynia and vulvody-
nia observed in this study.
Other important constituents present in P. incarnata
include hexadecanoic acid (palmitic acid), 3-hydroxy-
dodecanoic acid, 2,3-dihydro-3,5-dihydroxy-6-methyl-
4H-Pyran-4-one, and vitamin-E, that have strong anti-
oxidant and neuroprotective activities and/or modulate
the GABAergic system [115119].
The modulation of GABAergic and/or opioidergic sys-
tems by P. incarnata reported in this study may consti-
tute a proportion of the mechanisms implicated in the
Fig. 6 Effect of pentylenetetrazole (PTZ-10) on diazepam (DIZ-2) or Passiflora incarnata (PI-ME, 200 and 600 mg/kg) with respect to athe number
of steps climbed in the staircase test and bthe incidence of rears in mice. *P< 0.05 compared to PI-ME alone (200, 400 or 200 mg/kg) (ANOVA
followed by Dunnettspost hoc test), (n= 8 mice per group)
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 11 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
Fig. 7 aEffect of diazepam and Passiflora incarnata (PI-ME) on mouse locomotor activity. **P< 0.01, ***P< 0.001 compared to saline vehicle
control (ANOVA followed by Dunnettspost hoc test), (n= 6 mice per group). bEffect of pentylenetetrazole (PTZ-10) on the mouse locomotor
activity induced by Passiflora incarnata (PI-ME 400 and 600 mg/kg) or diazepam (DIZ-4). *P< 0.05 compared to PI-ME (400 or 600 mg/kg) alone
(ANOVA followed by Bonferronis multiple comparison post hoc test), (n= 6 mice per group)
Fig. 8 Effect of Passiflora incarnata (PI-ME) and gabapentin on astatic or bdynamic allodynia, at 1 or 2 h post-treatment times in female rats on
day 29 in a streptozotocin (STZ) induced neuropathic pain model.
###
P< 0.001 compared to saline control, *P< 0.01, **P< 0.05, ***P< 0.001
compared to streptozotocin alone treated animals, (n= 6 rats per group)
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 12 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
amelioration of diabetic neuropathy. Additional pro-
cesses however, like a cannabimimetic action [110, 113,
114] cannot be ignored inasmuch as P. incarnata ex-
hibits antihyperglycemic and hypolipidemic activities in
streptozotocin induced diabetes mellitus [25] which
would otherwise lead to neuropathic allodynia and
vulvodynia [56]. Hyperglycemia and dyslipidaemia driven
oxidative stress is a major contributor to reduced nerve
function [120, 121] and diabetes mellitus is a major
cause of peripheral neuropathy, commonly manifested
as distal symmetrical polyneuropathy [122]. Further-
more, diabetes mellitus has been reported to be linked
Fig. 9 Effect of Passiflora incarnata (PI-ME) and gabapentin on astatic or bdynamic vulvodynia at 1 or 2 h post-treatment times in female
rats on day 29 in the streptozotocin (STZ) induced neuropathic pain model.
###
P< 0.001 compared to saline, *P< 0.01, **P< 0.05, ***P< 0.001
compared to streptozotocin alone treated animals, (n= 6 rats per group)
Fig. 10 Scheme summarizing the anti-allodynic/anti-vulvodynic properties of Passiflora incarnarta plus its antinociceptive, anxiolytic-like and
higher dose sedative activities
Aman et al. BMC Complementary and Alternative Medicine (2016) 16:77 Page 13 of 17
Content courtesy of Springer Nature, terms of use apply. Rights reserved.
with vulvodynia either as an isolated symptom or as part
of a constellation of other neuropathic abnormalities.
Such neuropathic morbidity has been termed diabetic
vulvopathyand it profoundly affects patients quality of
life and management needs in order to address the phys-
ical, psychological and relationship problems caused by
the pain [123]. Our study showed that the methanolic ex-
tract of P. incarnata significantly alleviated only the dy-
namic component of vulvodynia which has been reported
more likely to be provoked by contact with clothing
among other triggers [124] and the cotton swab test is
usually used to localize painful areas in vulvodynia [125].
Conclusion
In conclusion, the methanolic extract of P. incarnata
possesses peripheral and central phasic as well as tonic
antinociceptive activity mediated through modulation of
GABA
A
and opioid receptors (GABAergic and opioider-
gic mechanisms shown in Fig. 10) which are disclosed
by their naloxone and PTZ reversibility. The findings
also manifest anxiolytic-like and higher dose sedative ac-
tivity of the extract, resulting from GABAergic stimula-
tion as indicated by their sensitivity to PTZ inhibition.
The extract also exhibited significant mechanical anti-
allodynic and dynamic anti-vulvodynic effects (Fig. 10)
that may be attributable at least in part to the oleamide
content and a cannabinoid-like action [110, 113, 114].
The outcomes from our study advocate an effectiveness
of P. incarnata in the treatment of various neuropathic
pain conditions. However, further studies are warranted
in order to determine a more precise association be-
tween the active constituents responsible for the anal-
gesic, anxiolytic and sedative effects of P. incarnata as
well as the specific molecular mechanisms underlying its
actions on allodynia and vulvodynia.
Additional file
Additional file 1: Passiflora incarnata plant, grown in the botanical
garden of the Department of Pharmacy, University of Peshawar,
Pakistan. (DOCX 637 kb)
Competing interests
The authors declare that they have no competing interests.
Authorscontributions
FS initiated the idea and guided the research group as supervisor in
planning and conducting experiments throughout the research project. UA
conducted the experiments and carried out calculations and statistical
analysis. She also prepared the initial draft of the manuscript. MS helped in
the analysis and interpretation of data as well as in preparing the final
version of the manuscript. SA provided her help during pharmacological
experiments throughout the study. NA assisted in the extraction of plant
material and other pharmacological activities. GA helped in the neuropathic
pain related experiments. KF attributed materials during pharmacological
screening of the plant extract. RDES guided the research group and
interpreted the results as well as critically revised the manuscript for
important intellectual content. All authors read and approved the final
manuscript.
Author details
1
Department of Pharmacy, University of Peshawar, Peshawar 25120, Khyber
Pakhtunkhwa, Pakistan.
2
Cardiff School of Pharmacy and Pharmaceutical
Sciences, Cardiff University, Cardiff CF10 3NU, UK.
Received: 15 August 2015 Accepted: 11 February 2016
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... Thus, the search for new therapeutic alternatives is very relevant, and, in this context, therapy research based on the use of plants has increased worldwide (Hou et al., 2020;Kim and Song, 2012;Kim et al., 2017;Rahbardar and Hosseinzadeh, 2020). In this sense, there is consistent evidence of the effectiveness of several plants in the treatment of psychiatric disorders (Sarris, 2018), highlighting the genus Passiflora (Aman et al., 2016;da Cunha et al., 2021;Dantas et al., 2017;Giovannini and Howes, 2017;Janda et al., 2020;Kim et al., 2019), especially Passiflora incarnata, used to treat insomnia and anxiety (Kim et al., 2019;Miroddi et al., 2013;Patel et al., 2009;Schäfer et al., 2021). The anxiolytic activity seems to be due to flavonoids, which would act on the GABAergic system (Fonseca et al., 2020). ...
... The anxiolytic effect appears to be due to an action on the gabaergic system (Fonseca et al., 2020;Jawna-Zboinska et al., 2016;Kim et al., 2019b), showing that flavonoids play a similar role to GABA (gamma-Aminobutyric acid) (Otify et al., 2015). Thus, the genus Passiflora has been considered to treat anxiety, a component present in depression and anxiety and post-traumatic stress disorders (Aman et al., 2016;Kim et al., 2019a). ...
... Male BALB/c albino mice 18-24 g were administered vehicle (VEH), morphine 5 mg/kg (MOR-5), diclofenac sodium (50 mg/kg; i.p) (DIC-50), GBP (100 mg/kg; i.p), and GPS (25, 50, 75, and 100 mg/kg; i.p). Abdominal constriction behavior was induced by injection of low-dose (1%) acetic acid (i.p) solution 30 min after drug treatment (Aman et al. 2016). Disregarding the initial 5 min after acetic acid injection, the incidence of abdominal constrictions was then counted over a period of 20 min. ...
... To substantiate the potential of GPS in other pain and inflammatory models, it was first evaluated in the acetic acid-induced abdominal constriction assay which is considered to signify tonic visceral nociception (Ness 1999). Next, it was assessed in a thermal phasic pain model (Aman et al. 2016) and finally studied against inflammatory models Fig. 8 Anti-inflammatory response of GPS in the carrageenan (A), histamine (B), and serotonin (C) induced paw edema assays in mice. Each bar represents the mean paw volume in mL ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, versus VEH control (two-way RM ANOVA followed by Bonferroni's post hoc test) (n = 8) Fig. 9 Antipyretic response of GPS in the brewer's yeastinduced pyrexia model in mice. ...
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Gabapentinoids are effective drugs in most animal models of pain and inflammation with variable effects in humans. The current study evaluated the pharmacological activity of gabapentin (GBP) and its salicylaldehyde derivative (gabapentsal; [2-(1-(((2-hydroxybenzylidene) amino) methyl) cyclohexyl) acetic acid]; GPS) in well-established mouse models of nociceptive pain, inflammatory edema, and pyrexia at doses of 25–100 mg/kg. GPS allayed tonic visceral pain as reflected by acetic acid-induced nociception and it also diminished thermally induced nociception as a mimic of phasic thermal pain. Antagonism of GPS-induced antinociceptive activities by naloxone (NLX, 1.0 mg/kg, subcutaneously, s.c), beta-funaltrexamine (β-FNT, 5.0 mg/kg, s.c), naltrindole (NT, 1.0 mg/kg, s.c), and nor-binaltorphimine (NOR-BNI, 5.0 mg/kg, s.c), and pentylenetetrazole (PTZ-15 mg/kg, intraperitoneally, i.p) implicated an involvement of both opioidergic and GABAergic mechanisms. Tail immersion test was conducted in order to delineate the mechanistic insights of antinociceptive response. Inflammatory edema induced by carrageenan, histamine, or serotonin was also effectively reversed by GPS in a fashion analogous to aspirin (150 mg/kg, i.p), chlorpheniramine (1.0 mg/kg, i.p), and mianserin (1.0 mg/kg, i.p), respectively. Additionally, yeast-induced pyrexia was decreased by GPS in a comparable manner to acetaminophen (50 mg/kg, i.p). These observations suggest that GPS possesses ameliorative properties in tonic, phasic, and tail immersion tests of nociception via opioidergic and GABAergic mechanisms, curbs inflammatory edema, and is antipyretic in nature.
... To study antagonism, naloxone (1 mg kg −1 ) was administered via subcutaneous route 10 min before administration of test compound and tramadol. 22 Finally, anti-nociceptive activity (%) was determined via formula; % anti À nociceptive activity ¼ latency of test À latency of control cut À off time À latency of control ...
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Purpose: Gastric ulcer induced by NSAIDs is the major medical concern and researchers are utilizing several approaches to combat this medical issue. In the current study, we investigated the efficacy of thiadiazinethione derivative (2,2'(2-thioxo-1,3,5-thiadiazinane-3,5-diyl) diacetic acid, as new less ulcerogenic compound. Methods: 2,2'(2-thioxo-1,3,5-thiadiazinane-3,5-diyl) diacetic acid was evaluated using standard animal models including hot plate, writhing test and formalin induced nociceptive models. Anti-inflammatory activity was assessed via carrageenan-induced paw oedema model. Involvement of opioidergic nociceptive mechanism was confirmed via naloxone administration in hot plat assay. The gastro-ulcerogenic potential of test and standard compounds were evaluated via NSAID-induced pyloric ligation model followed by standard histopathological and biochemical analysis. Results: In acetic acid-induced writhing test, our compound significantly reduced abdominal constrictions at the tested doses of 15 (p < 0.05), 30 (p < 0.01) and 45 mg kg-1 (p < 0.001) as compared to control (p < 0.001). In hot plate test, after 30 min of administration, our test compound showed significant anti-nociceptive potential (p < 0.05 at 15 and 30 mg kg-1 and p < 0.01 at 45 mg kg-1) and tramadol (p ˂ 0.001) at 30 mg kg-1 dose. After 60 min tramadol (30 kg-1) and test sample (30, 45 mg kg-1) exhibited significant anti-nociceptive activity p < 0.001. In Formalin-induced nociceptive response, a significant decline (p ˂ 0.001) was observed for aspirin and test compound during acute and chronic phases. Decline in the anti-nociceptive potential of tramadol and test sample via administration of naloxone indicate the involvement of opioidergic mechanism. Our compound exhibited significant anti-inflammatory activity in second phase of carrageenan induced paw oedema model. Histological and biochemical parameters exhibited less ulcerogenic potential as compared to aspirin. Conclusion: Our findings suggests that our test compound has desirable anti-nociceptive and anti-inflammatory potentials with less propensity to cause gastric ulcer.
... Passiflora incarnata L., referred to as purple passionflower, belongs to the Passifloraceae family, widely spread in tropical and warm temperate regions. Traditionally, the plant has been used for the treatment of insomnia, anxiety, cough, sexual dysfunction, convulsion, and cancer [64]. Phytochemical analysis has demonstrated the presence of phenolic compounds, alkaloids, and cyanogenic compounds, with flavonoids being the most common phytoconstituents [65,66]. ...
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The pivotal role of childhood nutrition has always roused a growing interest from the scientific community. Plant extracts and bioactive dietary components play a significant role in the maintenance of human health and wellness, with the potential to modulate risk factors and manage symptoms for a large number of common childhood disorders such as memory impairment, respiratory illnesses, gastrointestinal disorders, metabolic derangements, and pathologies related to the oral cavity. This review is designed to highlight the health benefits of botanical extracts and bioactive dietary components in children as evidenced by clinical trials, considering their safety with regards to childhood sensibilities. The supplementation of children with the herbal extracts or bioactive components mentioned in this review leads to the conclusion that they are useful for treating various ailments, with no serious adverse events being reported. However, for the limited number of investigations specifically focused on the safety of such products in children, time is needed to expand the literature data covering the safety of childhood supplementation with botanical extract and bioactive food components.
... In the literature, flavonoids and phenolic acids from Passiflora are reported to mediate their neuropharmacological action through the GABA-ergic pathway [69,71,72], promoting neuroinhibitory and tranquil state indole group in indole alkaloids such as harmalol, haramaline and their derivative glycosides (e.g., vitexin) has been correlated the endogenous neurotransmitter serotonin, suggesting their effectiveness as neuromodulators with antidepressant effects [73,74]. Other studies have shown that the extracts and metabolites involve the cholinergic system, inhibiting AChE as well as decreasing neuroinflammatory process [75,76]. ...
Chapter
Insomnia and sleep apnea represent the most prevalent sleep disorders worldwide. Incidences are discouraging, with perhaps less attention in some parts, and the complexity in the pathophysiologies of the two diseases pose a major challenge to clinicians and researchers. Mainstream therapeutic regimens are afflicted by adverse side effects, laborious, demanding enormous amounts of money, and are in particular, less accessible to the vast majority. Traditional medicine remains critically the “Hobsons’s choice” and stands out as a scaffold for the development of novel drugs with more desirable pharmacological attributes. This chapter covers the pathophysiological attributes of insomnia and sleep apnea, pointing out their interplay, possible biomarkers, and targets for both conventional and alternative therapeutic approaches. The chapter further delves more into the role of selected most important plant species in the management of insomnia and sleep apnea. Realizing the importance of phytochemistry in the drug discovery endeavor, the chapter, as well, underscores the phytochemical profile underlying the neuropharmacological relevance of each plant to its possible effectiveness in improving sleep quality.
... Although most of Passiflora pharmacological effects have been shown to be GABA mediated [42], the participation of other neurotransmitter systems in these effects should not be excluded. In fact, opioidergic and GABAergic mechanisms were reported to be associated with antinociceptive, anxiolytic and sedative effects of P. incarnata [60]; moreover, the antidepressive-like effects of P. edulis seems to recruit monoaminergic pathways [61]. S100B, a neurotrophic factor essential for neuronal survival, is a marker of glial activation [62]. ...
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Chronic opioid use changes brain chemistry in areas related to reward processes, memory, decision-making, and addiction. Both neurons and astrocytes are affected, ultimately leading to dependence. Passiflora incarnata L. (Passifloraceae) is the basis of frequently used herbals to manage anxiety and insomnia, with proven central nervous system depressant effects. Anti-addiction properties of P. incarnata have been reported. The aim of this study was to investigate the effect of a commercial extract of Passiflora incarnata (Sintocalmy®, Aché Laboratory) in the naloxone-induced jumping mice model of morphine withdrawal. In addition, glial fibrillary acidic protein (GFAP) and S100 calcium-binding protein B (S100B) levels were assessed in the frontal cortex and hippocampus, and DNA damage was verified on blood cells. In order to improve solubilization a Sintocalmy methanol extract (SME) was used. SME is mainly composed by flavonoids isovitexin and vitexin. The effects of SME 50, 100 and 200 mg/kg (i.p.) were evaluated in the naloxone-induced withdrawal syndrome in mice. SME 50 and SME 100 mg/kg decreased naloxone-induced jumping in morphine-dependent mice without reducing locomotor activity. No alterations were found in GFAP levels, however SME 50 mg/kg prevented the S100B increase in the frontal cortex and DNA damage. This study shows anti-addiction effects for a commercial standardized extract of P. incarnata and suggests the relevance of proper clinical assessment. Graphic Abstract
... Aman et al. carried out research on mice which indicated that P. incarnata may be useful in treating neuropathic pain. The authors suggested that these properties may result from underlying opioid and GABA-ergic mechanisms, but also pointed to the potential involvement of oleamid-based cannabimimetics [41]. The mechanism of action cannot, at present, be regarded as clarified; however, more recent studies imply that the anxiolytic effects may be mediated via modulation of the GABA system [42][43][44] This systematic review has some limitations. ...
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Background: Stress is a natural response of the body, induced by factors of a physical (hunger, thirst, and infection) and/or psychological (perceived threat, anxiety, or concern) nature. Chronic, long-term stress may cause problems with sleep, concentration, and memory, as well as affective disorders. The passionflower (Passiflora incarnata) is a perennial plant with documented therapeutic properties. The literature data suggest that the passionflower itself, as well as its preparations, helps reduce stress and can therefore be helpful in the treatment of insomnia, anxiety, and depression. The objective of this systematic review was to evaluate Passiflora incarnata in terms of its neuropsychiatric effects. Methods: The scientific databases PubMed, ClinTrials.gov, and Embase were searched up to 22 October 2019. The search identified randomized clinical trials describing the effects of Passiflora incarnata in neuropsychiatric disorders. Results: The systematic review included nine clinical trials. The duration of the studies included in the analysis varied widely, from one day up to 30 days. Study participants were no less than 18 years old. In each of the papers, the effects of passionflower were measured by using a number of different tests and scales. The majority of studies reported reduced anxiety levels following the administration of Passiflora incarnata preparations, with the effect less evident in people with mild anxiety symptoms. No adverse effects, including memory loss or collapse of psychometric functions, were observed. Conclusion: Passiflora incarnata may be helpful in treating some symptoms in neuropsychiatric patients.
... These neuropathic pain animals models are extensively utilized in the behavioral pharmacological research (Kumar et al. 2018). The animal models of peripheral nerve injury combined with testing of pain hypersensitivity remain the gold standard for the identification of novel therapeutic agents in order to treat chronic neuropathic pain conditions (Ali et al. 2015;Aman et al. 2016;Shahid et al. 2017b;Nawaz et al. 2018;Akbar et al. 2020;Rukh et al. 2020). Using the CCI experimental model of neuropathic pain, we have measured the paradigms of allodynia and hyperalgesia as readouts for the neuropathic pain. ...
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The current pharmacotherapy of neuropathic pain is inadequate as neuropathic pain involves varied clinical manifestations with multifactorial etiology, modulated by a cascade of physical and molecular events leading to different clinical presentations of pain. There is an accumulating evidence of the involvement of oxidative stress in neuropathy, and antioxidants have shown promise in mitigating neuropathic pain syndromes. To explore the evidence supporting this beneficial proclivity of antioxidants, this study investigated the antinociceptive effectiveness of N-(2-mercaptopropionyl)glycine or tiopronin, a well-recognized aminothiol antioxidant, in a refined chronic constriction injury (CCI) rat model of neuropathic pain. Tiopronin (10, 30, and 90 mg/kg, i.p.) and pregabalin (30 mg/kg, i.p.) were administered daily after CCI surgery. The neuropathic paradigms of mechanical/cold allodynia and mechanical/heat hyperalgesia were assessed on days 3, 7, 14, and 21 post-nerve ligation. At the end of study, malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) levels were estimated in the sciatic nerve, dorsal root ganglion, and spinal cord for assessing the extent of oxidative stress. The expression of neuropathic nociception was attenuated by tiopronin which was observed as a significant attenuation of CCI-induced allodynia and hyperalgesia. Tiopronin reversed the neuronal oxidative stress by significantly reducing MDA, and increasing SOD, CAT, and GSH levels. Pregabalin also showed similar beneficial propensity on CCI-induced neuropathic aberrations. These findings suggest prospective neuropathic pain attenuating efficacy of tiopronin and further corroborated the notion that antioxidants are effective in mitigating the development and expression of neuropathic pain and underlying neuronal oxidative stress.
Chapter
Conventional treatment of complex regional pain syndrome (CRPS) involves a pharmacologic regimen that supplements intensive physical and cognitive-behavioral therapy. Pharmacotherapies available for CRPS are at times insufficient for pain control or carry with them intolerable side effects. Frequently, patients and practitioners then turn to adjuvant therapies to facilitate treatment and symptom relief and reduce prescription pain medication use. This chapter offers an overview of adjuvant treatments, their proposed mechanisms of action, and a review of existing literature. Overall, there was a paucity of literature investigating the use of nonconventional therapies for CRPS, suggesting the need for further evidence-based research in this area.
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With a large and increasing elderly population, neurodegenerative diseases such as Parkinson’s disease (PD), Huntington disease (HD), Alzheimer’s disease (AD), Amyotrophic lateral sclerosis (ALS) and Multiple sclerosis (MS) have become a major and growing health problem. During the past few decades, the elderly population has grown 2.5 % every year. Unfortunately, there are no specific therapeutic remedies available to slow the onset or development of these diseases. An aging brain causes many pathophysiological changes and is the major risk factor for most of the neurodegenerative disorders. Polyphenolic compounds such as flavonols have shown therapeutic potential and can contribute to the treatment of these diseases. In this review, evidence for the beneficial neuroprotective effect of multiple flavonols is discussed and their multifactorial cellular pathways for the progressions of age-associated brain changes are identified. Moreover, the animal models of these diseases support the neuroprotective effect and target the potential of flavonols in the treatment of neurodegenerative diseases.
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Opioids constitute the first-line treatment for pain and they provide a potent analgesic effect, but they are also responsible for various adverse effects such as nausea, vomiting, sedation, constipation and respiratory depression, which seriously limit their use. The purpose of this study was to evaluate whether a GABA agonist which was given along with opioids at a minimal dose could elicit an anti-nociceptive activity in albino mice or not, as compared to morphine. Analgesia evaluation by using the acute pain model hotplate method was employed. The GABA agonists Gabapentin, Baclofen, Tiagabine and Vigabatrine with opioids Morphine and Tramadol, separately, at minimal doses, were given to mice and they were compared with a morphine analgesic dose. Morphine 3mg/kg showed a significant analgesic effect. This dose hot plate latency was taken as a standard and this was compared with all the test drugs which were used in this study. Morphine 1mg/kg (low dose) alone showed minimal antinociception, whereas in combination with a low dose GABA agonist, it showed significant antinociception. Tramadol 20mg/kg showed a significant analgesic effect and Tramadol 10mg/kg (low dose) showed a minimal analgesic effect, whereas the low dose Tramadol with the low dose GABA agonists in a combination showed a significant analgesic effect as that of Morphine 3mg/kg. The combination of a minimal dose of opioid and a GABA agonist has a significant anti nociceptive activity.
Article
The anxiolytic effects of passion flower (Passiflora incarnata L.) have been confirmed in several pharmacological studies [1–3], however, the compounds responsible for this effect are still a matter of debate. The purpose of this study was to characterize the putative anxiolytic-like activity of fractions prepared from a hydroethanolic extract using the elevated plus-maze (EPM) in mice. The fractions were prepared as published recently [4], yielding into a butanol, petroleum ether and chloroform fraction. Male BL6/C57J mice were either treated orally with each fraction in three different concentrations according to their percent amount in the extract or the positive control diazepam (1.5mg/kg). From the tested fractions, the butanol fraction showed significant increases in the number of open arm entries in the EPM in concentrations of 2.1mg/kg and 4.2mg/kg corresponding to 150 and 300mg/kg of the original extract. The highest activity was found for the chloroform fraction in doses of 0.17mg/kg (10.0±1.9, p<0.001) and 0.34mg/kg (6.6±0.86; p<0.05) which corresponds to a total extract dose of 150 and 300mg/kg respectively. Interestingly, the petroleum ether fraction did not show any effects in the elevated plus maze. A sedative effect of each of the fractions could be excluded, since none of the compounds had an influence on the total distance that the animals covered during the observation period. Our results suggest that the active principle of passion flower seems to be in the chloroform fraction and to a lower extend in the butanol fraction. References: 1. Soulimani, R. et al. (1997)J Ethnopharmacol 57(1): 11–20. 2. Grundmann, O. et al. (2009) Pharmazie 64: 63–64. 3. Grundmann et al. (2008) Planta Medica 74(15): 1769–1773. 4. Holbik, M. (2010) Diploma Thesis, University of Vienna.
Article
A naturally occurring brain lipid, cis-9,10-octadeceamideoleamide (OA), is found in increased concentrations in the cerebrospinal fluid of sleep-deprived cats, which suggests that it may be an endogenous sleep-inducing substance. We studied the effects of this fatty-acid derivative on the function of cloned gamma-aminobutyric acid (GABAA) receptors expressed in Xenopus oocytes. Oocytes were injected with cRNA synthesized in vitro to express simple GABAA receptors (alpha 1 beta 1, alpha 3 beta 1, alpha 5 beta 1, and alpha 1 beta 2 subunit combinations) and receptors in which the GABA-induced chloride currents were potentiated in the presence of benzodiazepines (alpha 1 beta 1 gamma 2s and alpha 1 beta 2 gamma 2s subunit combinations). OA only produced significant potentiation of the peak Cl⁻ current when applied with GABA to benzodiazepine-sensitive GABAA receptors. The peak currents of the simple GABAA receptors in the presence of OA were either unaffected or slightly inhibited by OA, but the overall mean currents were not significantly altered. Oleic acid was also capable of potentiating benzodiazepine-sensitive GABAA receptor function. The function of other ligand-gated ion channels, such as the N-methyl-D-aspartate receptor (NR1 + NR2A or 2C) and the 5-HT3 receptor expressed in Xenopus oocytes, were unaffected by OA. Sprague-Dawley rats receiving intraperitoneal injections of oleamide (10, 20, or 100 mg/kg) showed no change in the minimum alveolar anesthetic concentration (MAC) of desflurane required to abolish movement in response to noxious (tail clamp) stimulation (control MAC 6.48% +/- 1.28% atm; 100 mg/kg OA MAC 7.05% +/- 0.42% atm). These results reinforce the view that oleyl compounds may be natural modulators of inhibitory ion channel function, but that these effects contribute little to the central nervous system depression produced by volatile anesthetics as measured by MAC. Implications: The putative sleep-inducing substance, oleamide, potentiates benzodiazepinesensitive gamma-aminobutyric acid receptor function but does not alter desflurane minimum alveolar anesthetic concentration in rats. (Anesth Analg 1998;86:1294-300)
Article
Tiagabine HCl [(R]-N-[4,4-bis(3-methyl-2-thienyl]-3-butenyl] nipecotic acid hydrochloride], a potent and selective GABA uptake inhibitor, was evaluated for potential anti-allodynic effects in a rodent model of neuropathic pain and for antinociceptive activity in rodent models of acute and persistent pain. The effect of tiagabine on neuropathic pain was evaluated in rats that developed allodynia after tight ligation of L5 and L6 spinal nerves. The anti-allodynic effects of tiagabine were dose-dependent, with significant increases in response threshold to tactile stimulation occurring at 72.8 mu moles/kg, ip, but not at lower doses of 7.2 and 24.3 mu moles/kg, ip. In the hot-plate test in mice, tiagabine significantly increased foot-licking latency at 7.2 and 24.3 mu moles/kg, ip, and jump latency at 24.3 mu moles/kg, ip. After twice daily dosing with 72.8 mu moles/kg, ip, of tiagabine for 4 days, mice showed a tolerance to the antinociceptive effect of the 7.2 mu moles/kg, ip, dose of tiagabine in the hot-plate test. Tolerance did not occur after twice daily dosing of the 7.2 mu moles/kg, ip, dose of tiagabine. Tiagabine did not have significant effects on the tail-flick latency of rats at doses of 0.7 to 72.8 mu moles/kg, ip. Doses of 7.2 and 24.3 mu moles/kg, ip, of tiagabine significantly reduced the number of acetic acid-induced stretches in mice. In rats, tiagabine significantly decreased the number of paw flinches in the early phase of the formalin test at 24.3 and 72.8 mu moles/kg, ip, and in the late phase of the test at 72.8 mu moles/kg, ip. Tiagabine had no significant effects on carrageenan-induced paw edema at doses of 0.7 to 72.8 mu moles/kg, ip. Taken together, the results of these experiments revealed the potential anti-allodynic and antinociceptive pharmacology of tiagabine. Drug Dev. Res. 44:106-113, 1998. (C) 1998 Wiley-Liss, Inc.
Article
Background: Crocus sativus L. (saffron) is used in folk medicine, for example as an antiedematogenic agent. We aimed to evaluate the antinociceptive and anti-inflammatory activity of saffron extracts in mice.
Article
We applied and validated a quantitative allodynia assessment technique, using a recently developed rat surgical neuropathy model wherein nocifensive behaviors are evoked by light touch to the paw. Employing von Frey hairs from 0.41 to 15.1 g, we first characterized the percent response at each stimulus intensity. A smooth log-linear relationship was observed, with a median 50% threshold at 1.97 g (95% confidence limits, 1.12-3.57 g). Subsequently, we applied a paradigm using stimulus oscillation around the response threshold, which allowed more rapid, efficient measurements. Median 50% threshold by this up-down method was 2.4 g (1.81-2.76). Correlation coefficient between the two methods was 0.91. In neuropathic rats, good intra- and inter-observer reproducibility was found for the up-down paradigm; some variability was seen in normal rats, attributable to extensive testing. Thresholds in a sizable group of neuropathic rats showed insignificant variability over 20 days. After 50 days, 61% still met strict neuropathy criteria, using survival analysis. Threshold measurement using the up-down paradigm, in combination with the neuropathic pain model, represents a powerful tool for analyzing the effects of manipulations of the neuropathic pain state.