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Vol. 3, 124–132, 2017
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Synergistic Neuroprotective Potential of Combined
Treatment with Vinpocetine and Minocycline Against
Streptozotocin and Lipopolysaccharide Induced
Memory Impaired Mice
Pooja Rai, Brajnandan Kishor, Rakesh Bharatia, Sanjay Kumar,
Sujeet Kumar Gupta, and Anshuman Sinha∗
Department of Pharmacology, Hygia Institute of Pharmaceutical Education and Research, Lucknow, 226020, Uttar Pradesh, India
Alzheimer’s disease, a synonym for severe dementias is characterized by oxidative stress and neuroinflammation induced
neuronal loss, impaired energy metabolism, and cholinergic deficit leading to severe cognitive impairments and other atypical neuropsychiatric changes. Tau hypothesis, cholinergic hypothesis and Amyloid hypothesis are the most established
theories explaining the pathology of AD. Enhancing cyclic nucleotide signaling by phosphodiesterase inhibition (PDEs);
inflammation and apoptosis inhibition have been reported to be advantageous in neurodegenerative disorders. Minocycline and Vinpocetine have shown protective effects in memory impairment. In the present study, memory deficit mice
model was produced by intracerebral administration of streptozotocin (0.5 mg/kg) and intraperitoneal administration of
lipopolysaccharide (250 g/kg) which was evidenced by significant increase in escape latency time, acetylcholinesterase
level and significantly altered oxidative stress parameters such as increased malondialdehyde and decreased glutathione
levels. Furthermore, combined treatment with Minocycline + Vinpocetine (25 + 5 mg/kg, p.o.) significantly attenuated streptozotocin and lipopolysaccharide-induced memory impairment as demonstrated by reduced escape latency time and
acetylcholinesterase level and significantly improvised streptozotocin and lipopolysaccharide-induced altered oxidative
stress parameters as evidenced by decreased malondialdehyde and increased glutathione levels. The results of the
present study validated the combined neuroprotective and antioxidant properties of Minocycline and Vinpocetine. Conclusively, the study supports the candidature of combined therapy of Minocycline and Vinpocetine in learning and memory
disorders like dementia of Alzheimer’s type.
KEYWORDS: Alzheimer’s Disease, Memory, Lipopolysaccharide, Streptozotocin, Minocycline, Vinpocetine.
INTRODUCTION
According to United Nations predictions approximate
370 million people will be older than 80 years by 2051
and the associated growth in patients with Alzheimer’s
disease (AD) will cause a significant socioeconomic load
(Campbell and Gowran, 2007). AD, a deliberate progressive neurodegenerative disorder, clinically categorized by
a noticeable cognitive degeneration defined by a loss of
memory and learning ability together with a reduced skill
to perform simple activities of daily living and a several
∗
Author to whom correspondence should be addressed.
Email: [email protected]
Received: 12 January 2017
Accepted: 28 February 2017
124
J. Pharm. Sci. Pharmacol. 2017, Vol. 3, No. 2
variety of neuropsychiatric symptoms such as verbal and
physical agitation, laziness, anxiety, irritability, depression,
delusions and hallucinations. A standout amongst most
vital and solid divisions of AD is the severe degeneration of cholinergic neurons from basal forebrain to cortical and hippocampal areas (Whitehouse et al., 1982). The
90% loss of cholinergic neurons has been found in AD
patients (Whitehouse et al., 1981). The neuroinflammation
is a common reason of several neurodegenerative diseases
including AD (Unger 1998) Parkinson’s disease (Nagatsu
and Sawada, 2005) and neuropsychiatric disease such as
depression (Najjar et al., 2013).
Minocycline belongs to the group of semi synthetic second generation tetracycline. Compared to the
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doi:10.1166/jpsp.2017.1084
Rai et al.
Synergistic Neuroprotective Potential of Combined Treatment with Vinpocetine and Minocycline
first-generation invention, it has a definite molecular
modification which results in a significantly improved
ability to penetrate the BBB and a longer half-time
(Stirling et al., 2005). Minocycline gives antimicrobial
actions and also possesses beneficial anti-inflammatory
and anti-apoptotic properties. Clinical studies have shown
Minocycline and related tetracycline to be useful in treating mutually rheumatoid arthritis and osteoarthritis. The
neuroprotective and anti-neuroinflammatory activities of
Minocycline were first reported in 1998 in a gerbil
model of global brain ischemia (Yrjanheikki et al., 1998).
Minocycline has been shown to have antioxidant properties that may contribute as a protection in many psychiatric diseases (Ng et al., 2008). Previously, minocycline
along with NMDA, a glutamate receptor agonist reduced
the increase in the rate of superoxide production in rat
granular neurons (Garcia-Martinez et al., 2010).
Vinpocetine (a phosphodiesterase-1 (PDE-1) inhibitor)
is a semi-synthetic drug derived from vincamine (Bonoczk
et al., 2002). Vincamine is an alkaloid extracted from the
Periwinkle plant (Medina, 2011). Vinpocetine is an herbal
supplement used in the treatment of various neurodegenerative disorders (Deshmukh et al., 2009). Vinpocetine
has also been demonstrated to inhibit Veratridine induced
opening Na+ channel activity and glutamate release
(Tretter and Adam-Vizi, 1998).
Previous reports showed that lipopolysaccharide (LPS)
excites the inflammatory stimulus in both murine BV-2 cell
line and rat primary cultured microglia through the inhibition of p38 mitogen-activated protein kinase (MAPK)
(Hu et al., 2007). It was also reported that intraperitoneal
injection of LPS induces cognitive impairment in mice.
However, underlying mechanisms involved in LPS induced
cognitive impairment are still unclear.
Streptozotocin (STZ) (2-deoxy-2({[methyl(nitroso)amino]
carbonyl}amino)--D-glucopyranose) is a naturally
occurring compound, formed by the bacterium Streptomyces achromogenes, that shows broad spectrum
antibacterial properties. Previously, intracerebroventricular
(ICV)/intracerebral (IC) injection of STZ in rats/mice
induced various pathological features of AD like impaired
brain glucose and energy metabolism (Nitsch and Hoyer,
1991), which further leads to progressive impairment
in learning and memory (Awasthi et al., 2010). Previous reports have shown the beneficial neuroprotective
role of drug combination therapy (Omote et al., 2014;
Kwon et al., 2013). Although there are reports about the
neuroprotective role of Vinpocetine and Minocycline in
memory impaired rodent models (Deshmukh and Sharma,
2013; Shang et al., 2016; Hou et al., 2016). But there are
no reports found about the neuroprotective potential of
Vinpocetine and Minocycline in combination. The present
study was therefore designed to evaluate the combined
neuroprotective effect of Minocycline and Vinpocetine in
STZ and LPS-induced memory impaired mice.
J. Pharm. Sci. Pharmacol. 3, 124–132, 2017
MATERIALS AND METHODS
Reagents
Minocycline and Donepezil were purchased from Tokyo
chemical industry, Japan. Vinpocetine was purchased from
Micro labs limited, Hosur, India. Lipolysacchride was purchased from Sigma-Aldrich, USA. The chemicals, i.e.,
Streptozotocin (STZ), Chloral hydrate, Sodium chloride
(NaCl), Sodium nitrite (NaNO2 ), Bovine serum albumin
(BSA), Acetylthiocholine iodide (ATCl), 5,5 -Dithiobis(2nitrobenzoic acid) (DTNB), 1, 1, 3, 3-Tetraethoxypropane
(TEP), 2-Thiobabituric acid (TBA) were purchased from
Sigma-Aldrich, USA. All other chemicals used in the
study were of exploratory grade. Solution of the drug and
chemicals were freshly prepared before use.
Animals
Male Swiss albino mice (20–25 g) were used in this study.
Food and water were given ad libitum and the animals
were maintained under standard conditions of temperature (23 ± 2 C) and humidity (40%) with light and dark
cycles of 12 h each. All procedures were performed in
accordance with the CPCSEA guidelines, Department of
Animal Welfare, Government of India. All experiments
depicted were checked on and affirmed by the Institutional
Animal Ethics Committee (IAEC), Hygia Institute of Pharmaceutical Education and Research, Lucknow-India (Reg.
No. 1088/PO/Re/S/2007/CPCSEA).
STZ-Induced Memory Impairment in Mice
Mice were randomly assigned into two groups for surgery.
The sham operated control group (n = 6) were injected
with isotonic saline solution. STZ dissolved in 0.9% normal saline solution was injected into the second group
of animals. Anaesthesia was induced in animals with
ketamine (100 mg/kg body weight; i.p.) and xylazine
(30 mg/kg body weight; i.p.) and STZ (0.5 mg/kg) (dissolved in normal saline) was administered in a volume
of 10 l intracerebrally (i.c.) according to the method of
Haley and McCormick (Haley and McCormick, 1957) with
a replicate dose 48 h later. The site of STZ injection in the
animal was 2 mm from each side of the centerline on a
line drawn through the anterior base of the ears. Before the
main experiment, the target region in the brain ventricular
system was identified and studied histologically by injecting 0.05 ml of a 1:10 dilution of methylene blue and further sectioning the brain slices. Animals injected with STZ
were randomly assigned to 5 groups [vehicle, Donepezil
(5 mg/kg), Minocycline (50 mg/kg each), Vinpocetine
(10 mg/kg) and Minocycline + Vinpocetine (25 mg/kg +
5 mg/kg); n = 6 for each group]. Mice in each group
were administered perorally with normal saline, Donepezil
(5 mg/kg), Minocycline (50 mg/kg each), Vinpocetine
(10 mg/kg) and Minocycline + Vinpocetine (25 mg/kg +
5 mg/kg) respectively, after one week of surgical recovery
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Synergistic Neuroprotective Potential of Combined Treatment with Vinpocetine and Minocycline
Figure 1. Overview of experimental design of streptozotocin
(STZ)-induced memory impaired rodent model.
once every day for 14 successive days, respectively
(Fig. 1).
LPS-Induced Memory Deficit Model
The animals (n = 36) were divided into six groups.
LPS (250 g/kg) was given intraperitoneally (i.p.) to
all of the groups except for the control group [normal saline] 4 hr after treatment with the standard drug
or the test compound. Donepezil (5 mg/kg), Minocycline (50 mg/kg each), Vinpocetine (10 mg/kg) and
Minocycline + Vinpocetine (25 mg/kg + 5 mg/kg) were
administered orally for 14 successive days respectively.
All the drug solutions were prepared in normal saline. For
all experiments, LPS (Escherichia coli, serotype 055:B5,
Sigma, St. Louis, MO, USA) was used to induce an inflammatory response and was injected once on day 1 of behavioral testing (Fig. 2).
Morris Water Maze (MWM) Test
The task was used to evaluate the drug [Donepezil
(5
mg/kg),
Minocycline
(50
mg/kg
each),
Vinpocetine (10 mg/kg) and Minocycline + Vinpocetine
(25 mg/kg + 5 mg/kg), p.o)] effect on learning and
Figure 2. Overview of experimental design of lipopolysaccharide (LPS)-induced memory impaired rodent model.
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Rai et al.
memory according to the method described by Morris
(Morris, 1984).
The water maze comprises of a circular pool (65 cm in
diameter, 25 cm high), filled with water (26 ± 1 C) to the
depth of 20 cm. The pool was made opaque by painting
the inside walls black. A division of the pool was made by
four quadrants. An escape platform (equidistant from the
middle of the pool and the side wall), was located in the
middle of one quadrant. The platform was placed 1.0 cm
below the water surface. During each trial same quadrant
was utilized placing the platform. Reaching on the platform is the only escape condition for the animals. In the
area of the perimeter of the pool three different starting
points were placed for animal. Animals were placed in all
the three starting points in a pseudorandom manner during
the four day training period. The animal was placed in the
water facing the wall of the pool at one of the starting
points to begin the trial. The animal was softly placed on
the platform and allowed to stay for 15 s when it failed to
reach on the platform within 120 s. A time of 5–10 min
was given to inner trial interval. The escape latency time
(ELT) was observed in all the groups. Three escape trials per day for four successive days were given to all the
animals. After removing the platform, the number of crossings over the region of the platform was also observed
for 120 s.
Neurochemical Assays
Once the Morris water maze test was done, the animals
were sacrificed. Whole brain was dissected out from the
skull, and stored at −80 C. Homogenate of the stored
brains were prepared in glass Teflon homogenizer using
homogenization buffer (400 mM NaCl and 12.5 mM
sodium phosphate buffer pH 7.0). The homogenate was
centrifuged at 1000× g for 10 min at 4 C and the enzyme
assays were performed in the supernatant. The acetylcholinesterase (AChE) activity was assessed according to
the method described by Ellman et al. (Ellman et al.,
1961). Firstly, 50 L of brain homogenate were incubated
in 96-well plates at room temperature for 30 min. Further
30 L of the substrates viz. ATCl (15 mM) was added and
incubated for 30 min. Lastly 160 L DTNB (1.5 mM) was
added and absorbance was measured at a wavelength of
415 nm using Biorad microplate reader 680XR. The AChE
level was expressed as U/ml for brain homogenates.
Malondialdehyde (MDA) and reduced glutathione
(GSH) levels were measured by the methods described
by Mihara and Uchiyama (1978) and Moron et al. (1979)
respectively.
The level of total protein in brain sample was measured
according to the method described by lowery et al. (1951).
Statistical Analysis
Graph Pad InStat software (version 5.00, San Diego, CA)
was utilized for data analysis. Every data are expressed as
J. Pharm. Sci. Pharmacol. 3, 124–132, 2017
Rai et al.
Synergistic Neuroprotective Potential of Combined Treatment with Vinpocetine and Minocycline
mean ±SEM. The mean significant difference in the experimental groups was analyzed using two way ANOVA (ELT
for Morris water maze test) and one way ANOVA followed by Bonferroni test. Values of p < 005 were judged
statistically significant.
RESULTS
Combined Effect of Minocycline and
Vinpocetine on Hippocampus-Dependent
Learning and Memory
The Morris water maze test was utilized to assess the combined effect of Vinpocetine and Minocycline using STZ
and LPS-induced neuroinflammation in mice. As demonstrated in Figures 3 and 4, the escape latency was significantly increased in STZ and LPS treated groups as
compared to the control groups amid all trial sessions.
This confirmed that STZ and LPS treatment impaired the
memory in mice. The escape latency was significantly
decreased in Donepezil (5 mg/kg) and Minocycline +
Vinpocetine (25 mg/kg + 5 mg/kg) treated groups as compared to the STZ and LPS treated groups in the probe trial
and final sessions (Figs. 3(a), 4(a)). Assessment of spatial
Figure 4. Effect of Minocycline and Vinpocetine on escape
latency in training trial and probe trial sessions (a) and the
number of crossing platform area (b) for LPS-induced memory
deficit mice. Values are expressed as mean ± SEM. (n = 6) Significant values were compared with ∗∗∗ p < 0001 versus LPS
treated group, ### p < 0001 versus control group.
working memory was made by confirming the number
of crossings over the platform. The number of crossings was significantly decreased in STZ and LPS treated
groups while Vinpocetine + Minocycline (25 mg/kg +
5 mg/kg) significantly increased the number of crossings over the platform (Figs. 3(b), 4(b)). Donepezil significantly increased the number of crossings over the
platform. Minocycline (50 mg/kg, p.o.) and Vinpocetine (10 mg/kg, p.o.) alone also showed the significant
neuroprotection but were found less effective as compared
to Minocycline + Vinpocetine treated group. Animals were
further evaluated for neurochemical analysis.
Figure 3. Effect of Minocycline and Vinpocetine on escape
latency in training trial and probe trial sessions (a) and the
number of crossing platform area (b) for STZ-induced memory
deficit mice. Values are expressed as mean ± SEM. (n = 6) Significant values were compared with ∗∗∗ p < 0001 versus STZ
treated group, ### p < 0001 versus control group.
J. Pharm. Sci. Pharmacol. 3, 124–132, 2017
Combined Anticholinesterase and Antioxidative
Effects of Minocycline and Vinpocetine in STZ
and LPS Induced Memory Impaired Mice
AChE level was significantly reduced as well as oxidative parameters were significantly modified (decreased
MDA level and increased GSH level) by Minocycline +
Vinpocetine (25 + 5 mg/kg, p.o.). To evaluate whether
Minocycline + Vinpocetine exerts better inhibitory effects
on AChE activity than alone, brain tissues of mice
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Synergistic Neuroprotective Potential of Combined Treatment with Vinpocetine and Minocycline
Rai et al.
Figure 5. Ex vivo anticholinesterase and antioxidant properties of Minocycline and Vinpocetine. AChE and MDA levels were
significantly increased and GSH level was significantly reduced in STZ treated group (a, b, c). Combined treatment with Minocycline and Vinpocetine significantly decreased acetylcholinesterase and MDA and increased GSH levels (a, b, c) as compared to
STZ treated group. Values are expressed as mean ± SEM. (n = 6) Significant values were compared with ∗∗∗ p < 0001 versus STZ
treated group, ∗∗ p < 001 versus STZ treated group, ∗ p < 005 versus STZ treated group, ### p < 0001 versus control group.
treated with Minocycline + Vinpocetine (25 + 5 mg/kg,
p.o.) were subjected to colorimetric estimation to establish
the AChE activity in the brain. Minocycline + Vinpocetine
(25 + 5 mg/kg, p.o.) reduced AChE level to 820 ±
025 U/ml (p < 001) as compared to the STZ and
LPS treated groups (1541 ± 048 and 2748 ± 093 U/ml)
(Figs. 5(a), 6(a)). This data authenticated the fact that oral
doses of Minocycline + Vinpocetine (25 + 5 mg/kg) significantly reverted AChE level, which was increased by STZ
and LPS respectively, confirming its anticholinesterase-like
activity. We then proceeded to demonstrate the combined
effects of Minocycline + Vinpocetine (25 + 5 mg/kg, p.o.)
128
on oxidative parameters. The results showed that STZ and
LPS treatment significantly (p < 0001) increased the brain
MDA levels (859 ± 013 and 865 ± 010 nM/mg protein)
as compared to the control groups (137 ± 032 and 140 ±
031 nM/mg proteins). Treatment with the standard drug
(Donepezil) and Minocycline+Vinpocetine (25 +5 mg/kg,
p.o.) significantly (p < 001) reduced brain MDA levels
[Donepezil (345 ± 038 and 370 ± 013); Minocycline +
Vinpocetine (402 ± 038 and 459 ± 038) nM/mg proteins] as compared to the corresponding STZ and LPS
treated groups (Figs. 5(b), 6(b)). Further, STZ and LPS
treatment significantly (p < 0001) decreased the brain
J. Pharm. Sci. Pharmacol. 3, 124–132, 2017
Rai et al.
Synergistic Neuroprotective Potential of Combined Treatment with Vinpocetine and Minocycline
Figure 6. Ex vivo anticholinesterase and antioxidant properties of Minocycline and Vinpocetine. AChE and MDA levels were
significantly increased and GSH level was significantly reduced in LPS treated group (a, b, c). Combined treatment with Minocycline and Vinpocetine significantly decreased acetylcholinesterase and MDA and increased GSH levels (a, b, c) as compared to
LPS treated group. Values are expressed as mean ± SEM. (n = 6) Significant values were compared with ∗∗∗ p < 0001 versus LPS
treated group, ∗∗ p < 001 versus LPS treated group, ∗ p < 005 versus LPS treated group, ### p < 0001 versus control group.
GSH levels (154 ± 028 and 149 ± 026 g/mg protein)
as compared to the respective control groups (574 ± 031
and 535 ± 004 g/mg proteins). Treatment with the standard drug (Donepezil) and the Minocycline + Vinpocetine
(25 + 5 mg/kg, p.o.) significantly (p < 001) elevated brain
GSH levels [Donepezil (399 ± 006 and 397 ± 012) and
Minocycline + Vinpocetine (331 ± 012 and 333 ± 013)
nM/mg proteins] as compared to the corresponding STZ
and LPS treated groups respectively (Figs. 5(c), 6(c)).
Minocycline (50 mg/kg, p.o.) and Vinpocetine (10 mg/kg,
p.o.) alone also showed the significant anticholinesterase
and antioxidative mechanism but were found less effective
J. Pharm. Sci. Pharmacol. 3, 124–132, 2017
as compared to Minocycline + Vinpocetine treated group.
These results revealed the combined anticholinesterase and
antioxidant properties of Minocycline + Vinpocetine.
Combined Effect of Minocycline and Vinpocetine
on Total Protein Level in STZ and LPS Induced
Memory Impaired Mice
The results showed that STZ and LPS treatment significantly (p < 0001) increased the brain total protein levels
(1839 ± 279 and 1903 ± 257 mg/g protein) as compared
to the control groups (3117 ± 271 and 3193 ± 216 mg/g
proteins). Treatment with the standard drug (Donepezil)
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Synergistic Neuroprotective Potential of Combined Treatment with Vinpocetine and Minocycline
Rai et al.
Figure 7. Combined effect of Minocycline and Vinpocetine on total protein level in STZ and LPS-induced memory deficit mice.
As compared to control group, total protein levels were significantly increased in STZ and LPS treated groups (a, b). As compared
to STZ and LPS treated groups, combined treatment with Minocycline and Vinpocetine significantly decreased total protein levels
(a, b). Values are expressed as mean ± SEM. (n = 6) Significant values were compared with ∗∗∗ p < 0001 versus STZ/LPS treated
group, ∗∗ p < 001 versus STZ/LPS treated group, ∗ p < 005 versus STZ/LPS treated group, ### p < 0001 versus control group.
and Minocycline + Vinpocetine (25 + 5 mg/kg, p.o.) significantly (p < 001) reduced brain total protein levels
(Figs. 7(a and b)) [Donepezil (8857 ± 218 and 9167 ±
227) and Minocycline + Vinpocetine (9830 ± 174 and
1013 ± 203) mg/g proteins] as compared to the corresponding STZ and LPS treated groups (Figs. 7(a and b)).
DISCUSSION
In AD several components like disease alteration, safety
and efficacy have given limitations to its monotherapy.
It appears to be far-fetched that any agent may induce
alterations in the AD pathophysiology by acting on a single molecular mechanism adequate to vary disease progression. Additionally, monotherapy requires high doses
for its efficacy, which produces more prominent or more
severe side effects. This is the situation with cholinesterase
inhibitors (ChEIs); their efficacy increases while tolerance declines in a dose-dependent manner. However, treatment with a combination of drugs having diverse modes
of action may give points of interest over monotherapy for the effective pharmacological management of
AD (Cummings, 2007; Farlow et al., 2008; Chow et al.,
2010; Ihl et al., 2011; Patel and Grossberg, 2011; Schmitt
et al., 2004). Combination therapy may improve efficacy by persuading synergistic or additive effects; improving tolerability and safety potentially permitting lower
doses to be used (Deiana et al., 2009); and interpreting
supplementary neuroprotective effects by extending the
symptomatic aids and eventually postponing the disease
progression.
130
The present study observed the combined neuroprotective effect of Minocycline + Vinpocetine on memory
impairment, oxidative stress, cholinergic dysfunction and
protein levels in intracerebral (i.c.) STZ and intraperitoneal
LPS injected mice. Pre-treatment with Minocycline +
Vinpocetine, in STZ and LPS administered mice, improved
spatial memory. Furthermore, Minocycline + Vinpocetine
also improvised the decrease of antioxidant (GSH),
increase of lipid peroxide (MDA) and the cholinergic
dysfunction in the mouse brain following STZ and LPS
injection.
Sporadic Alzheimer type dementia can be examined
by using an appropriate animal model viz. the intracerebroventricular (ICV) STZ rodent model (Nitsch and Hoyer,
1991). The present study showed that administration of
STZ in mice caused a persistent memory deficit as evidenced by significant increase in escape latency time in
the Morris water maze test. This finding authenticated
the previous experiments reporting memory impairment
by STZ injection in rodents (Tota et al., 2010). Administration of Minocycline + Vinpocetine (25 + 5 mg/kg, p.o.)
significantly attenuated STZ-induced memory impairment
as demonstrated by reduced latency to reach the platform
and increased number of crossings across the platform.
There was significant correlation in between latency time
and number of crossing of all the groups in all the sessions
demonstrating direct alteration in both the parameters.
It is familiar that mitochondria prompt oxidative stress
generates free radicals, which have been associated in neuronal damage during AD. Free radical facilitated injury
J. Pharm. Sci. Pharmacol. 3, 124–132, 2017
Rai et al.
Synergistic Neuroprotective Potential of Combined Treatment with Vinpocetine and Minocycline
leads to lipid peroxidation that produces a number of secondary products like MDA by damaging the membranes.
This elevated free radical production additionally prompts
diminished antioxidant enzymes such as GSH (Olanow,
1993). In the current study, STZ produced oxidative stress
as demonstrated by a significant rise in MDA level and a
reduction in GSH level signifying that STZ-induced learning and memory impairment is linked with oxidative stress
in rodents.
An essential part has been played by cholinergic neuronal framework in the cognitive deficits connected with
aging and neurodegenerative disorders. Previous reports
showed that there was a significant decline in the quantity
of muscarinic binding sites and acetylcholine (ACh) level
in the brains of AD patients (Quirion et al., 1986). Acetylcholinesterase (AChE) degrades ACh which is essential for
appropriate working of cholinergic transmission to control learning and memory processes. Symptomatic treatment of AD has been achieved by inhibiting AChE through
their inhibitors. In the present study, we established that
STZ treatment significantly increased AChE level in mice.
The finding is similar to the previous reports demonstrating a significant increase in AChE activity (Tota et al.,
2010) and expression (Lester-Coll et al., 2006) following treatment with STZ. Furthermore, administration of
Minocycline + Vinpocetine (25 + 5 mg/kg, p.o.) significantly attenuated STZ-induced memory impairment as
demonstrated by the reduced AChE level similar to the
standard drug Donepezil.
Besides, it was additionally stated that LPS induces
memory impairment when administered intraperitoneally
in mice (Sparkman et al., 2005). However, the underlying mechanism behind LPS-induced memory impairment
is still unclear. In our findings LPS significantly induces
memory impairment as evidenced by increased latency
time and decreased number of crossings across the platform. Furthermore, pre-treatment with Minocycline + Vinpocetine (25 + 5 mg/kg, p.o.) significantly attenuated LPS
induced memory impairment as demonstrated by significantly decreased latency time and increased number of
crossing across the platform. This suggested the combined
neuroprotective role of Minocycline and Vinpocetine in
LPS induced memory impairment.
Previous reports also showed that LPS treatment significantly increased MDA and decreased GSH in rodents
(Abdel-Salam et al., 2012). Similarly, in the present study
LPS significantly increased brain MDA and decreased
GSH in mice. However, pre-treatment with Minocycline +
Vinpocetine (25 + 5 mg/kg, p.o.) significantly attenuated
LPS-induced oxidative stress parameters as demonstrated
by a significant decrease in MDA and the increase in
GSH levels in LPS treated mice. This validated the neuroprotective and antioxidant properties of Minocycline and
Vinpocetine.
J. Pharm. Sci. Pharmacol. 3, 124–132, 2017
CONCLUSION
It can be concluded from the data that combination therapy of Minocycline and Vinpocetine might be an important
therapeutics for the treatment of Alzheimer’s type dementia. However, further studies to elaborate the mechanism
are needed.
Conflict of Interest Statement
The authors declare that there is no conflict of interest.
Acknowledgment: The authors acknowledge the management of the Hygia Institute of Pharmaceutical
Education and Research, Lucknow for providing the
research facility and financial support to perform this
study.
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