Pharmacognosy Magazine

ORIGINAL ARTICLE
Year
: 2016  |  Volume : 12  |  Issue : 45  |  Page : 63--70

Naringin and rutin alleviates episodic memory deficits in two differentially challenged object recognition tasks


Grandhi Venkata Ramalingayya, Madhavan Nampoothiri, Pawan G Nayak, Anoop Kishore, Rekha R Shenoy, Chamallamudi Mallikarjuna Rao, Krishnadas Nandakumar 
 Department of Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal University, Manipal, Karnataka, India

Correspondence Address:
Krishnadas Nandakumar
Department of Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal University, Manipal - 576 104, Karnataka
India

Abstract

Background: Cognitive decline or dementia is a debilitating problem of neurological disorders such as Alzheimer's and Parkinson's disease, including special conditions like chemobrain. Dietary flavonoids proved to be efficacious in delaying the incidence of neurodegenerative diseases. Two such flavonoids, naringin (NAR) and rutin (RUT) were reported to have neuroprotective potential with beneficial effects on spatial and emotional memories in particular. However, the efficacy of these flavonoids is poorly understood on episodic memory, which comprises an important form of autobiographical memory.Objective: This study objective is to evaluate NAR and RUT to reverse time-delay-induced long-term and scopolamine-induced short-term episodic memory deficits in Wistar rats. Materials and Methods: We have evaluated both short-term and long-term episodic memory forms using novel object recognition task. Open field paradigm was used to assess locomotor activity for any confounding influence on memory assessment. Donepezil was used as positive control and was effective in both models at 1 mg/kg, i.p. Results: Animals treated with NAR and RUT at 50 and 100 mg/kg, p.o. spent significantly more time exploring novel object compared to familiar one, whereas control animals spent almost equal time with both objects in choice trial. NAR and RUT dose-dependently increased recognition and discriminative indices in time-induced long-term as well as scopolamine-induced short-term episodic memory deficit models without interfering with the locomotor activity. Conclusion:We conclude that, NAR and RUT averted both short- and long-term episodic memory deficits in Wistar rats, which may be potential interventions for neurodegenerative diseases as well as chemobrain condition.



How to cite this article:
Ramalingayya GV, Nampoothiri M, Nayak PG, Kishore A, Shenoy RR, Rao CM, Nandakumar K. Naringin and rutin alleviates episodic memory deficits in two differentially challenged object recognition tasks.Phcog Mag 2016;12:63-70


How to cite this URL:
Ramalingayya GV, Nampoothiri M, Nayak PG, Kishore A, Shenoy RR, Rao CM, Nandakumar K. Naringin and rutin alleviates episodic memory deficits in two differentially challenged object recognition tasks. Phcog Mag [serial online] 2016 [cited 2019 Oct 13 ];12:63-70
Available from: http://www.phcog.com/text.asp?2016/12/45/63/176104


Full Text

Summary

Incidence of Alzheimer's disease is increasing globally and the current therapy is only symptomatic. Curative treatment is a major lacuna. NAR and RUT are natural flavonoids proven for their pleiotropic pharmacological effects with potential neuroprotective benefits. The study evaluated these flavonoids for their potential to improve the most common form of episodic memory (memory of autobiographical events in relation to time, places etc.) in two differential animal models assessing short-term and long-term memory, respectively. We also found that NAR and RUT were able to reverse both short-term and long-term memory deficits dose dependently in female Wistar rats.

[INLINE:1]

 Introduction



Neurodegenerative diseases are often associated with cognitive, motor and behavioral dysfunction.[1] According to WHO, by 2040, neurodegenerative disorders could be the leading cause of mortality, surpassing cancer.[2] The most common neurodegenerative disorder is the Alzheimer's disease (AD) characterized by progressive cognitive deterioration and personality changes leading to complete need for care and is found to be the most common cause of dementia in geriatric population.[3] About 5.2 million people of all age groups in the US have AD in 2013, and it is projected to nearly triple, that is, 13.8 million by 2050.[4]

Although the etiology of AD remains unclear, restoration of cholinergic transmission in the brain using cholinesterase inhibitors remains the mainstay of treatment, which is only supportive or symptomatic rather than curative or disease-modifying.[5] Satisfactory outcomes were not accomplished through pharmacological treatment by conventional drugs despite advances in understanding the pathology of the disease.[6] Hence, complementary and alternative medicines become a potential source of new drugs for the prevention of neurodegenerative diseases.[2] Earlier reports revealed that dietary intake of rich polyphenolic vegetables and fruits proved to delay the onset of dementia associated with AD.[7],[8] Animal behavioral data support the fact that, flavonoids can improve various components of cognitive processes through neuronal differentiation, long-term potentiation and also by enhancing the synaptic plasticity.[9],[10],[11],[12]

Naringin (NAR), a citrus flavanone glycoside found in grape seeds and constitutes naringenin its aglycone part. NAR possess anti-oxidant, anti-inflammatory, anti-apoptotic and cardio-protective effects in vivo.[13],[14] It was found that inflammatory mediators play a vital role in the pathogenesis of neurodegenerative anomalies.[15] Earlier works confirmed that NAR prevented the lipopolysaccharide-induced pro-inflammatory cytokines and gene expression of cyclooxygenase and nitric oxide synthase (NOS) in vitro[13],[16] as well as in vivo.[17] Moreover, NAR was reported to prevent the behavioral changes and cognitive deficits in kainic acid-induced epilepsy [18] and 3-nitropropionic acid-induced Huntington's disease animal models.[19] NAR also attenuated colchicine [20] and D-galactose [21] induced learning and memory deficits in rats.

Another flavonoid used in the present work, that is, rutin (RUT) is a flavonol glycoside (with aglycone flavanol, Quercetin) has diverse beneficial effects viz., antioxidant, anticancer, antiplatelet, antithrombotic, vasoprotective, cardioprotective and hepatoprotective effects with potential neuroprotective activities.[22],[23],[24],[25],[26] Researchers reported that RUT inhibited microglial activation and pro-inflammatory cytokines [27] and is effective against trimethyltin induced spatial memory deficits through amelioration of neuronal damage in hippocampal CA3 subregion, crucial for acquisition learning in rodents.[28] RUT also prevented scopolamine-induced cognitive deficits in inhibitory avoidance test in zebrafish.[29]

NAR and RUT have been reportedly known for their anti-inflammatory effects in many experimental studies through inhibition of NO production.[30],[31],[32] They showed potential neuroprotective effects against ischemic reperfusion induced cerebral injury by ameliorating oxidative damage, mitochondrial dysfunction, and neurological impairments.[33],[34] Moreover, researchers found that NAR and RUT alleviated AD-type neurodegeneration and the associated cognitive impairment of spatial and working memories induced by intracerebroventricularly injected streptozotocin in rats.[35],[36]

Increasing evidence shows that cholinergic neurotransmission is an essential prerequisite for the formation of recognition memories.[37] Scopolamine-induced cognitive impairment is considered as an important research tool to explore the mechanisms underlying cognitive changes observed in AD patients.[38] Due to an increased enzymatic activity of acetylcholinesterase, there will be an increased breakdown of acetylcholine in AD patients, which lead to hindered cholinergic transmission and thereby the affected memory function.[6],[39]

NAR and RUT were found to be effective in enhancing the cognitive function in behavioral tasks assessing spatial, working and emotional memories in animal models, however episodic memory deficits are often observed clinically in neurodegenerative diseases, especially in AD associated dementia [40],[41] and also in chemotherapy-induced cognitive dysfunction (chemobrain) in cancer survivors, especially in breast cancer survivors.[42] Hence, it is important to evaluate the efficacy of NAR and RUT for their potential to prevent short- and long-term episodic memory deficits, so as to contribute to the increasing knowledge about these plant-derived products and their use as medicines in treating neurodegenerative diseases.

We observed paucity for the studies assessing nootropic potential of NAR and RUT on episodic memory, hence in the current study, we focused on the efficacy of these flavonoids for episodic memory retrieval using novel object recognition task (NORT). Initially, we have evaluated both the flavonoids to reverse the time delay induced natural deficits of long-term episodic memory. This was followed by assessment in a drug-induced, that is, scopolamine-induced short-term episodic memory deficits.

 Materials and Methods



Animals

A 12-week-old, healthy (n = 72) female Wistar rats weighing 150–200 g were used in the present study. The protocol was approved by the Institutional Animal Ethics Committee (IAEC) (Approval No. IAEC/KMC/17/2013) and the experiments were performed in compliance with the Committee for the Purpose of Control and Supervision on Experimental Animals (CPCSEA) guidelines, India. Animals were maintained (with temperature, 25°C ± 2°C and relative humidity, 55% ± 5%) at Central Animal Research Facility of Manipal University, Manipal as per the directions specified by the CPCSEA guidelines. A 12/12 h of light and dark cycle was maintained, and potable water was given ad libitum throughout the study period.

Chemicals and apparatus

Pharmacological agents, donepezil (Matrix Laboratories Ltd., Secunderabad, TS, India), scopolamine hydrobromide (Sigma-Aldrich Co. LLC, St. Louis, MO, USA), NAR (Tokyo Chemical Industry Co. Ltd., Tokyo, Japan), RUT (HiMedia Laboratories, Mumbai, MH, India) and water for injection (Aculife Healthcare Pvt. Ltd., Ahmedabad, GJ, India) were used in this study.

Apparatus consisted of six square boxes (arenas) of 48 cm (length × breadth × height) which are made with plywood. Inner portions of arenas were covered with black laminate. All the six boxes were assigned for six individual animals in each group through number allocation. These boxes were also used as open field boxes for assessing the locomotor activity along with the use of ANY-maze video tracking system (Stoelting Co., Wood Dale, IL, USA). ANY-maze can be used to track the animal behavior in an open field to assess the locomotor activity by video tracking system. The system can generate the open field test parameters once we program for it. Behavioral observation of rats in arenas was monitored and recorded using a camera (model: Quickcam Pro9000, Logitech International SA, Lausanne, Switzerland) mounted above the behavioral observation arenas. Other essential accessories such as timer alarms and handheld stopwatches were procured for accurate scoring and behavioral observation. This equipment was used to assess time induced as well as scopolamine-induced episodic memory deficits in NORT. A proficient observer who was blind to the treatment groups manually scored the behavioral parameter, that is, exploration/investigation time.

Formulations and treatments

Animals were treated with either NAR and RUT at two dose levels i.e. 50 and 100 mg/kg, p.o., thatwere formulated as suspensions using 0.25% w/v sodium carboxy methyl cellulose (CMC) for 15 days in both the animal models prior to experimentation and also during the trials, 1 h prior to familiarization and/or choice trial. All the rats received a constant volume (2 ml/kg) of either vehicle or test compound/drug (donepezil/scopolamine) depending on the type of model being used. Rats were injected intraperitoneally with donepezil (1 mg/kg, i.p.), 30 min prior to familiarization trial and choice trials in time induced memory deficits model. In case of scopolamine-induced short-term memory deficits model, donepezil was given at 1 mg/kg, i.p., 30 min prior to familiarization trial and scopolamine was given at a dose of 0.5 mg/kg, i.p., 20 min prior to the familiarization trial. All the doses and the predose interval periods were selected based on the past laboratory experience and from the previous studies.[33],[43],[44]

Experimental groups

Two separate sets of animals were used for assessing short-term and long-term episodic memories. A total of six treatment groups were used in each model for assessing either time delay induced or scopolamine-induced episodic memory deficits in NORT. Vehicle group (Group 1) was treated orally with 0.25%w/v sodium carboxymethyl cellulose (CMC). Groups 2 and 3 animals were treated with NAR (50 and 100 mg/kg, p.o.), respectively whereas in Groups 4 and 5, rats were treated with RUT at (50 and 100 mg/kg, p.o.), respectively. Group 6 was positive control and received intraperitoneal donepezil at a dose of 1 mg/kg. In case of scopolamine-induced short-term episodic memory deficits model, all the six groups of animals were treated with scopolamine at a dose of 0.5 mg/kg, i.p. along with their respective treatments as mentioned above.

Change in body weight

To know the effect of test flavonoids, NAR and RUT, body weight recordings were made on alternate days throughout the study period in time induced episodic memory deficits study. The percentage increase in body weight (%IBW) was noted at the end of the study with respect to day 1 and compared among the treatment groups. This %IBW was noted for one set of animals (time induced deficits model) as the treatment for both sets of animal models was similar.

Open field test

Activation or suppression of locomotor activity will have confounding influences on any neuro-behavioral paradigm assessing cognitive functions. In the present study, to know whether NAR and RUT have influencing effects on locomotion, we have tested locomotion following the 15 days flavonoid treatment (prior to object recognition task). All the rats from diverse treatment groups in time-induced episodic memory deficits study were assessed for locomotion using an open field paradigm. Open field test is done using same square arenas (48 cm × 48 cm × 48 cm) that are used for object recognition tasks. The procedures followed were according to the earlier reports with slight modifications.[45],[46] Briefly, rats from different treatment groups were individually placed in open field boxes (following the acclimatization period). Using the system software, an imaginary 15 cm circular zone was created in the center of the arena. The locomotor activity, that is, distance (cm) travelled, mean velocity (cm/s) and time spent in the center zone during a 15 min test duration were assessed using the ANY-maze software tracking system (Version 4.82 m, Stoelting Co., Wood Dale, IL, USA). The comparison was made for the parameters monitored among different treatment groups.

Novel object recognition task

Episodic memory can be assessed typically by using NORT in rodents. Procedures were conducted as per the previous reports with some modifications.[47],[48] Briefly, the experiment was conducted in a soundproof isolated room having a light intensity of 30–40 lux over a period of 2–3 days. The experiment was carried out between 09:00 am to 04:00 pm and basically consisted of three phases, habituation, familiarization, and choice trial phases as illustrated in [Figure 1]. On day 1, animals were habituated to the behavioral observation arenas for a period of 20 min. On day 2, after treating the animals with their respective treatments and following the postdose intervals, they were subjected to familiarization trial. In scopolamine-induced short-term memory deficit model, choice trial was conducted following an inter-trial interval (ITI) of 3 min (as scopolamine produces short-term memory deficits), whereas in case of time-induced long-term memory deficits, the choice trial was conducted after an ITI of 24 h, that is, on day 3, as this time delay produces long-term episodic memory deficits. In familiarization trial, animals were allowed to explore a pair of similar objects in experimental arenas. The objects used were made of glass or plastic material, which were cylindrical in shape with dimensions of 18 cm × 5 cm. Exploration of animals was defined as sniffing, touching the objects with nose and/or any other investigative behavior directed toward object within 1 cm distance. The choice trial was performed by replacing one of the familiar objects with a novel object and cumulative exploration time of each animal toward familiar or novel object was noted for 3 min trial duration.{Figure 1}

The investigatory behavior that is, exploration of rats directed towards objects was recorded during trials using a camera mounted above the behavioral observation arenas connected to a computer system. Recognition index (RI) and discriminative index (DI), which reflects the animal's memory for the objects were determined from the exploration time in choice trial according to the earlier reports.[49] RI reflects the animal's memory for the familiar object relative to that of the novel object, whereas DI indicates the degree of discrimination showed by animals among the objects.

Hematological profiling

Following the behavioral assessment in time induced episodic memory deficits study, blood sampling was carried out using dipotassium ethylenediaminetetracetic acid (10% w/v) as an anticoagulant. Complete blood profile was performed using automated veterinary blood cell counter (model: PCE-210 VET, ERMA Inc., Tokyo, Japan) and the mean values of major parameters like, red blood cell (RBC), white blood cell (WBC), hemoglobin, platelet counts of different treatment groups were compared and analyzed.

Statistical analysis

Data are represented as mean ± standard error of the mean of time in sec spent by the animals exploring either familiar or novel objects and also as recognition and discriminative indices. Statistical analysis was carried out using Prism 6.03 trial version (GraphPad Software Inc., La Jolla, CA, USA). Exploration time was compared by paired t-test within the group and between the objects. Comparison of means for RI and DI were analyzed by using Kruskal–Wallis test followed by Dunn's post-hoc test. A value of P < 0.05 was considered as statistically significant. Locomotion was expressed as distance traveled in cm, time spent in center zone and mean velocity. %IBW, hematological parameters as well as locomotion were analyzed by using one-way ANOVA followed by Dunnett's post-hoc test.

 Results



Change in body weight

Throughout the experimental groups, we observed a gradual increase in body weight over 15 days study duration. The average %IBW for any of the treatment group was approximately 12–15%. With respect to the normal control animals, treatment with NAR and RUT at both tested doses, that is, 50 and 100 mg/kg, p.o. for 15 days did not result in any significant change in %IBW. The %IBW among different treatment groups is shown in [Figure 2].{Figure 2}

Open field test (locomotion assessment)

Like %IBW measure, locomotion assessment was carried out in one set of animals (time induced deficits model) as the treatment was same. It was found that no significant difference in the locomotor activity measures, that is, distance traveled, time spent in the center zone and mean velocity was observed among the treatment groups. Hence, it was confirmed that NAR and RUT have no confounding influence on locomotion on the improving potential for episodic memory. This is indicated by almost equal amount of distance traveled, time in the center zone and the mean velocity among the treatment groups [Figure 3].{Figure 3}

Time-induced long-term episodic memory deficits in novel object recognition task

In familiarization trial, all the animals spent nearly equal time exploring two similar objects, which indicate that there was no preference or discrimination toward any of the objects used. Vehicle treated animals spent almost equal time with both familiar and novel objects in the choice trial, indicating their inability to remember familiar object after an ITI of 24 h. Both the flavonoids, NAR and RUT dose-dependently reversed the time induced episodic memory deficits at 50 and 100 mg/kg, p.o. which is evident from the significant increase in novel object exploration time compared with familiar object. Also, we found that there was a significant improvement in recognition and discriminative indices for the groups treated with NAR and RUT as compared to vehicle control.

It was found that low dose NAR (50 mg/kg, p.o.) treated animals spent significantly more time with novel object compared to familiar one in choice trial with a comparative improvement of recognition and discriminative indices over vehicle control; however this improvement was not statistically different. Positive control, donepezil produced significant improvement of time induced memory deficits at a tested dose of 1 mg/kg, i.p. which was indicated by significant increase in time spent exploring novel object compared to familiar object in choice trial and also by significant increase in DI and RI compared to vehicle group [Figure 4]a and [Figure 4]b.{Figure 4}

Scopolamine-induced short-term episodic memory deficits in novel object recognition task

In scopolamine-induced short-term impairment of episodic memory, locomotion of the animals treated with scopolamine was comparatively increased due to the central cholinergic (M1 receptor) blockade and the resulting short-term memory deficits. Also, it was observed that the duration of the exploration time was comparatively more when compared to that of time induced deficits model. All the rats explored a pair of similar objects for the almost equal amount of time in familiarization trial. Animals of vehicle group treated with scopolamine could not able to discriminate between novel and familiar objects as they spent almost equal time with both the objects in the choice trial. However, pretreatment with NAR and RUT at 50 and 100 mg/kg for about 15 days significantly reversed the scopolamine-induced short-term episodic memory deficits in a dose-dependent manner, which was evident from significant difference in duration of exploration time directed toward either familiar or novel object.

It was also observed that discriminative and recognition indices were significantly improved for the groups treated with NAR and RUT when compared to the vehicle group. Although no significant increase in RI and DI were observed for NAR and RUT low dose (50 mg/kg) groups, rats of these groups spent significantly more time exploring the novel object compared to that of familiar one in choice trial. Animals treated with donepezil at a dose of 1 mg/kg, i.p., spent significantly more time with novel object compared to familiar object, hence prevented the scopolamine-induced short-term episodic memory deficits [Figure 5]a and [Figure 5]b.{Figure 5}

Hematological profiling

We found that NAR and RUT at tested doses of 50 and 100 mg/kg, p.o. for 15 days have improved RBC and hemoglobin levels as compared to vehicle control. However, the improvements observed were not statistically significant. Hence, these improving effects of these flavonoids on blood profile, particularly on RBC and hemoglobin may underlie as one of the reason for the procognitive like effects on episodic memory in object recognition task. Other parameters such as WBC, total granulocyte, lymphocyte, monocyte, and platelet counts were unaffected. The hematological data for one set of animals (time-induced deficits model) is represented in [Table 1].{Table 1}

 Discussion



Prefrontal cortex plays a pivotal role in encoding and retrieval of episodic memory.[50],[51] Moreover, the anatomical structures of the medial temporal lobe that are crucial for the formation of episodic memory comprise amygdala, brainstem, and hippocampus. Declarative or explicit memory is one of the two major types of memory, consisting of episodic, and semantic memories.[52] The memory of autobiographical events in relation to times, places is considered as episodic memory. Deficits of episodic memory are one of the primary cognitive complications that are observed in presenile or senile dementia associated with AD that can lead to impairment of QOL.

Finding the effective treatments requires better understanding of the physiological mechanisms underlying the disease pathogenesis.[53] Earlier studies confirmed the potential efficacy of flavonoids in formation of memory through activation of gene expression and neuronal signaling in the brain that result in changes of synaptic plasticity and neurogenesis, which can eventually influence learning and memory processes.[12] Flavonoids found to improve blood flow and possess vascular protective effects in humans.[54] These vascular effects are vital as the improved cerebrovascular function is known to enhance neurogenesis in the hippocampus based on an in vitro study.[55] New hippocampal cells near to blood vessels can multiply rapidly in the presence of vascular growth factors and can influence cognitive function.[56]

The pharmacologically active aglycone moieties of NAR and RUT, that is, naringenin and quercetin respectively showed potential anticholinesterase activity in various cognitive animal models for assessing memory patterns in rodents.[57],[58] Several studies at preclinical stage have focused mainly on the effect of NAR and RUT on spatial acquisition learning in water maze task, emotional memory in inhibitory avoidance task. However, it is essentially important to assess episodic memory and the influence of NAR and RUT on episodic memory as these deficits were known to occur frequently in AD patients and were proved clinically. Down regulation or reduction in cholinergic transmission is linked to the most common form of dementia associated with AD. Scopolamine is a muscarinic receptor blocker, which blocks central M1 receptors that are crucial for memory formation through cholinergic neurotransmission and can lead to memory deficits in humans following its administration.

Scopolamine-induced cognitive deficits through the impairment of the cholinergic neurotransmission is one of the widely used methods to investigate the underlying mechanisms for cognitive changes of different forms of memory viz., episodic, working, and spatial memory noticed in AD patients. Memory deficits can also be produced naturally by an increased time delay, that is, by increasing the time between the sample and test phases, which has a natural origin.

We have used NORT paradigm to assess the short-term as well as long-term episodic memory deficits. In time delay model, vehicle treated animals forgot the familiar object after a trial delay of 24 h, as they showed no discrimination between the objects in the choice trial. We found that NAR and RUT were effective dose-dependently, against this time induced long-term episodic memory impairment in Wistar rats at both the tested doses, i.e. 50 and 100 mg/kg, p.o., as there was a significant improvement in the recognition and discriminative indices compared to vehicle control. Donepezil at a dose of 1 mg/kg, i.p. prevented the episodic recall deficits, which showed the validity of this model for assessing episodic memory.

In scopolamine-induced cognitive deficits model, vehicle group spent almost equal time investigating both novel and familiar objects in choice trial, which indicates that, scopolamine at a dose of 0.5 mg/kg, i.p. produced short-term episodic memory deficits at an ITI of 3 min. These deficits were significantly reversed by prior administration of either donepezil at 1 mg/kg, i.p. or flavonoids, that is, NAR and RUT at a dose of 50 and 100 mg/kg, p.o. Hence, these flavonoids were able to reverse the episodic memory deficits, which were induced either by increased time delay or by scopolamine administration in a dose-dependent passion. Among the flavonoids tested, RUT showed superior activity in both short-term and long-term episodic memory models.

Further, we assessed the effect of the tested flavonoids on locomotion using open field test. The locomotion of the animals was not influenced by the treatment with NAR and RUT at any of the tested doses, which would otherwise influence the cognitive functional measurements. No significant changes were observed in view of distance moved, time spent in the central zone and mean velocity. Hence it was proved that the tested flavonoids, NAR and RUT have produced procognitive like effects without influencing the locomotor activity and reversed the time delay induced long-term as well as scopolamine-induced short-term episodic memory deficits supporting their potential use in neurodegenerative diseases, especially for the treatment of AD. No significant difference in the %IBW was noted among the treatment groups, while moderate improvement in RBC and Hemoglobin was observed for the groups treated with flavonoids at the tested doses compared to vehicle control.

Numerous mechanisms and signaling pathways may underlie the enhanced cognitive function by these flavonoids. It was found that the biological activity of flavonoids within CNS is not only attributed to their antioxidant effects,[59] but also probably through their potential to enhance surviving neuronal function by protecting vulnerable neurons, stimulating neuronal regeneration and by inducing neurogenesis.[12],[59],[60] Moreover, flavonoids were found to enhance the extracellular signal-regulated protein kinase cAMP response element-binding protein and PI3 kinase-mTOR pathways that can lead to alterations in synaptic plasticity and were also able to influence neurogenesis by activation of PI3-kinase-Akt-eNOS pathway.[61] Further, flavonoids were known to inhibit oxidative stress-induced neuronal injury by preventing the caspase-3 action, which is responsible for their neuroprotective effects.[62] Collectively, multiple mechanisms of flavonoids underlie the cognitive improving effects of NAR and RUT in rodent animal models, including the episodic memory which was assessed in the present study. Based on the current findings, studies are underway to evaluate NAR and RUT for their potential to alleviate cancer-therapy related cognitive complications such as chemobrain.

 Conclusion



NAR and RUT exhibited procognitive like effects dose-dependently by reversing the time induced long-term episodic memory deficits in NORT. Further, scopolamine-induced short-term episodic memory deficits were also found to be alleviated by prior treatment with the flavonoids at both the tested doses in a dose-dependent passion. Although, the mechanisms underlying the memory enhancement is beyond the scope of the present work, we conclude that NAR and RUT improved episodic memory impairments in Wistar rats without affecting the locomotor activity. The procognitive effects observed might have involved improved cholinergic neurotransmission by inhibition of acetylcholinesterase. In addition, potential ability of the flavonoids to interact with intracellular signaling pathways to enhance vascular function, synaptic plasticity, neurogenesis and their classical antioxidant effects also contribute to their nootropic effect in reversing the short-term as well as long-term episodic memory deficits in recognizing the objects.

Financial support and sponsorship

The author, Mr. G.V. Ramalingayya, acknowledges the fellowship (Reg. No. 120600101) received from Manipal University during the course of this work.

Conflicts of interest

There are no conflicts of interest.

References

1Woulfe J. Nuclear bodies in neurodegenerative disease. Biochim Biophys Acta 2008;1783:2195-206.
2Ip PS, Tsim KW, Chan K, Bauer R. Application of complementary and alternative medicine on neurodegenerative disorders: Current status and future prospects. Evid Based Complement Alternat Med 2012;2012:930908.
3Hampel H, Prvulovic D, Teipel S, Jessen F, Luckhaus C, Frölich L, et al . The future of Alzheimer's disease: The next 10 years. Prog Neurobiol 2011;95:718-28.
4Alzheimer's Association. 2013 Alzheimer's disease facts and figures. Alzheimers Dement 2013;9:208-45.
5Casey DA, Antimisiaris D, O'Brien J. Drugs for Alzheimer's disease: Are they effective? P T 2010;35:208-11.
6Ibach B, Haen E. Acetylcholinesterase inhibition in Alzheimer's Disease. Curr Pharm Des 2004;10:231-51.
7Dai Q, Borenstein AR, Wu Y, Jackson JC, Larson EB. Fruit and vegetable juices and Alzheimer's disease: The Kame Project. Am J Med 2006;119:751-9.
8Barberger-Gateau P, Raffaitin C, Letenneur L, Berr C, Tzourio C, Dartigues JF, et al. Dietary patterns and risk of dementia: The Three-City cohort study. Neurology 2007;69:1921-30.
9Williams RJ, Spencer JP, Rice-Evans C. Flavonoids. Flavonoids: Antioxidants or signalling molecules? Free Radic Biol Med 2004;36:838-49.
10Wang Y, Wang L, Wu J, Cai J. The in vivo synaptic plasticity mechanism of EGb 761-induced enhancement of spatial learning and memory in aged rats. Br J Pharmacol 2006;148:147-53.
11Spencer JP. The interactions of flavonoids within neuronal signalling pathways. Genes Nutr 2007;2:257-73.
12Spencer JP. The impact of flavonoids on memory: Physiological and molecular considerations. Chem Soc Rev 2009;38:1152-61.
13Kim HJ, Song JY, Park HJ, Park HK, Yun DH, Chung JH. Naringin protects against rotenone-induced apoptosis in human neuroblastoma SH-SY5Y cells. Korean J Physiol Pharmacol 2009;13:281-5.
14Raza SS, Khan MM, Ahmad A, Ashafaq M, Islam F, Wagner AP, et al. Neuroprotective effect of naringenin is mediated through suppression of NF-κB signaling pathway in experimental stroke. Neuroscience 2013;230:157-71.
15McGeer PL, Rogers J, McGeer EG. Inflammation, anti-inflammatory agents and Alzheimer disease: The last 12 years. J Alzheimers Dis 2006;9 3 Suppl: 271-6.
16Kanno S, Shouji A, Tomizawa A, Hiura T, Osanai Y, Ujibe M, et al. Inhibitory effect of naringin on lipopolysaccharide (LPS)-induced endotoxin shock in mice and nitric oxide production in RAW 264.7 macrophages. Life Sci 2006;78:673-81.
17Liu Y, Wu H, Nie YC, Chen JL, Su WW, Li PB. Naringin attenuates acute lung injury in LPS-treated mice by inhibiting NF-κB pathway. Int Immunopharmacol 2011;11:1606-12.
18Golechha M, Chaudhry U, Bhatia J, Saluja D, Arya DS. Naringin protects against kainic acid-induced status epilepticus in rats: Evidence for an antioxidant, anti-inflammatory and neuroprotective intervention. Biol Pharm Bull 2011;34:360-5.
19Kumar P, Kumar A. Protective effect of hesperidin and naringin against 3-nitropropionic acid induced Huntington's like symptoms in rats: Possible role of nitric oxide. Behav Brain Res 2010;206:38-46.
20Kumar A, Dogra S, Prakash A. Protective effect of naringin, a citrus flavonoid, against colchicine-induced cognitive dysfunction and oxidative damage in rats. J Med Food 2010;13:976-84.
21Kumar A, Prakash A, Dogra S. Naringin alleviates cognitive impairment, mitochondrial dysfunction and oxidative stress induced by D-galactose in mice. Food Chem Toxicol 2010;48:626-32.
22La Casa C, Villegas I, Alarcón de la Lastra C, Motilva V, Martín Calero MJ. Evidence for protective and antioxidant properties of rutin, a natural flavone, against ethanol induced gastric lesions. J Ethnopharmacol 2000;71:45-53.
23Janbaz KH, Saeed SA, Gilani AH. Protective effect of rutin on paracetamol- and CCl4-induced hepatotoxicity in rodents. Fitoterapia 2002;73:557-63.
24Schwedhelm E, Maas R, Troost R, Böger RH. Clinical pharmacokinetics of antioxidants and their impact on systemic oxidative stress. Clin Pharmacokinet 2003;42:437-59.
25Sheu JR, Hsiao G, Chou PH, Shen MY, Chou DS. Mechanisms involved in the antiplatelet activity of rutin, a glycoside of the flavonol quercetin, in human platelets. J Agric Food Chem 2004;52:4414-8.
26Pu F, Mishima K, Irie K, Motohashi K, Tanaka Y, Orito K, et al. Neuroprotective effects of quercetin and rutin on spatial memory impairment in an 8-arm radial maze task and neuronal death induced by repeated cerebral ischemia in rats. J Pharmacol Sci 2007;104:329-34.
27Koda T, Kuroda Y, Imai H. Rutin supplementation in the diet has protective effects against toxicant-induced hippocampal injury by suppression of microglial activation and pro-inflammatory cytokines: Protective effect of rutin against toxicant-induced hippocampal injury. Cell Mol Neurobiol 2009;29:523-31.
28Koda T, Kuroda Y, Imai H. Protective effect of rutin against spatial memory impairment induced by trimethyltin in rats. Nutr Res 2008;28:629-34.
29Richetti SK, Blank M, Capiotti KM, Piato AL, Bogo MR, Vianna MR, et al. Quercetin and rutin prevent scopolamine-induced memory impairment in zebrafish. Behav Brain Res 2011;217:10-5.
30Rotelli AE, Guardia T, Juárez AO, de la Rocha NE, Pelzer LE. Comparative study of flavonoids in experimental models of inflammation. Pharmacol Res 2003;48:601-6.
31Fang SH, Rao YK, Tzeng YM. Anti-oxidant and inflammatory mediator's growth inhibitory effects of compounds isolated from Phyllanthus urinaria. J Ethnopharmacol 2008;116:333-40.
32Wang GJ, Chen YM, Wang TM, Lee CK, Chen KJ, Lee TH. Flavonoids with iNOS inhibitory activity from Pogonatherum crinitum. J Ethnopharmacol 2008;118:71-8.
33Gaur V, Aggarwal A, Kumar A. Protective effect of naringin against ischemic reperfusion cerebral injury: Possible neurobehavioral, biochemical and cellular alterations in rat brain. Eur J Pharmacol 2009;616:147-54.
34Khan MM, Ahmad A, Ishrat T, Khuwaja G, Srivastawa P, Khan MB, et al. Rutin protects the neural damage induced by transient focal ischemia in rats. Brain Res 2009;1292:123-35.
35Javed H, Khan MM, Ahmad A, Vaibhav K, Ahmad ME, Khan A, et al. Rutin prevents cognitive impairments by ameliorating oxidative stress and neuroinflammation in rat model of sporadic dementia of Alzheimer type. Neuroscience 2012;210:340-52.
36Khan MB, Khan MM, Khan A, Ahmed ME, Ishrat T, Tabassum R, et al. Naringenin ameliorates Alzheimer's disease (AD)-type neurodegeneration with cognitive impairment (AD-TNDCI) caused by the intracerebroventricular-streptozotocin in rat model. Neurochem Int 2012;61:1081-93.
37Warburton EC, Koder T, Cho K, Massey PV, Duguid G, Barker GR, et al. Cholinergic neurotransmission is essential for perirhinal cortical plasticity and recognition memory. Neuron 2003;38:987-96.
38Klinkenberg I, Blokland A. The validity of scopolamine as a pharmacological model for cognitive impairment: A review of animal behavioral studies. Neurosci Biobehav Rev 2010;34:1307-50.
39Lane RM, Kivipelto M, Greig NH. Acetylcholinesterase and its inhibition in Alzheimer disease. Clin Neuropharmacol 2004;27:141-9.
40Bäckman L, Small BJ, Fratiglioni L. Stability of the preclinical episodic memory deficit in Alzheimer's disease. Brain 2001;124(Pt 1):96-102.
41Ergis AM, Eusop-Roussel E. Early episodic memory impairments in Alzheimer's disease. Rev Neurol (Paris) 2008;164 Suppl 3:S96-101.
42Schagen SB, Hamburger HL, Muller MJ, Boogerd W, van Dam FS. Neurophysiological evaluation of late effects of adjuvant high-dose chemotherapy on cognitive function. J Neurooncol 2001;51:159-65.
43Alam MA, Kauter K, Brown L. Naringin improves diet-induced cardiovascular dysfunction and obesity in high carbohydrate, high fat diet-fed rats. Nutrients 2013;5:637-50.
44Niture NT, Ansari AA, Naik SR. Anti-hyperglycemic activity of rutin in streptozotocin-induced diabetic rats: An effect mediated through cytokines, antioxidants and lipid biomarkers. Indian J Exp Biol 2014;52:720-7.
45Abel EL. Behavioral effects of isatin on open field activity and immobility in the forced swim test in rats. Physiol Behav 1995;57:611-3.
46Prut L, Belzung C. The open field as a paradigm to measure the effects of drugs on anxiety-like behaviors: A review. Eur J Pharmacol 2003;463:3-33.
47Ennaceur A, Delacour J. A new one-trial test for neurobiological studies of memory in rats 1: Behavioral data. Behav Brain Res 1988;31:47-59.
48Ennaceur A, Meliani K. Effects of physostigmine and scopolamine on rats' performances in object-recognition and radial-maze tests. Psychopharmacology (Berl) 1992;109:321-30.
49Antunes M, Biala G. The novel object recognition memory: Neurobiology, test procedure, and its modifications. Cogn Process 2012;13:93-110.
50Fletcher PC, Shallice T, Dolan RJ. The functional roles of prefrontal cortex in episodic memory. I. Encoding. Brain 1998;121(Pt 7):1239-48.
51Fletcher PC, Shallice T, Frith CD, Frackowiak RS, Dolan RJ. The functional roles of prefrontal cortex in episodic memory. II. Retrieval. Brain 1998;121(Pt 7):1249-56.
52Schacter DL, Gilbert DT, Wegner DM. Semantic and episodic memory. Psychology. 2nd ed. New York: Worth, Incorporated; 2011. p. 240-1.
53Mucke L. Neuroscience: Alzheimer's disease. Nature 2009;461:895-7.
54Schroeter H, Heiss C, Balzer J, Kleinbongard P, Keen CL, Hollenberg NK, et al. (-)-Epicatechin mediates beneficial effects of flavanol-rich cocoa on vascular function in humans. Proc Natl Acad Sci U S A 2006;103:1024-9.
55Gage FH. Mammalian neural stem cells. Science 2000;287:1433-8.
56Palmer TD, Willhoite AR, Gage FH. Vascular niche for adult hippocampal neurogenesis. J Comp Neurol 2000;425:479-94.
57Heo HJ, Kim MJ, Lee JM, Choi SJ, Cho HY, Hong B, et al. Naringenin from Citrus junos has an inhibitory effect on acetylcholinesterase and a mitigating effect on amnesia. Dement Geriatr Cogn Disord 2004;17:151-7.
58Jung M, Park M. Acetylcholinesterase inhibition by flavonoids from Agrimonia pilosa . Molecules 2007;12:2130-9.
59Spencer JP. Beyond antioxidants: The cellular and molecular interactions of flavonoids and how these underpin their actions on the brain. Proc Nutr Soc 2010;69:244-60.
60Spencer JP. The impact of fruit flavonoids on memory and cognition. Br J Nutr 2010;104 Suppl 3:S40-7.
61Vauzour D, Vafeiadou K, Rodriguez-Mateos A, Rendeiro C, Spencer JP. The neuroprotective potential of flavonoids: A multiplicity of effects. Genes Nutr 2008;3:115-26.
62Schroeter H, Spencer JP, Rice-Evans C, Williams RJ. Flavonoids protect neurons from oxidized low-density-lipoprotein-induced apoptosis involving c-Jun N-terminal kinase (JNK), c-Jun and caspase-3. Biochem J 2001;358(Pt 3):547-57.