LL-00066471, a novel positive allosteric modulator of α7 nicotinic acetylcholine receptor ameliorates cognitive and sensorimotor gating deficits in animal models: Discovery and preclinical characterization.

α7 nicotinic acetylcholine receptor (α7 nAChR) is an extensively validated target for several neurological and psychiatric conditions namely, dementia and schizophrenia, owing to its vital roles in cognition and sensorimotor gating. Positive allosteric modulation (PAM) of α7 nAChR represents an innovative approach to amplify endogenous cholinergic signaling in a temporally restricted manner in learning and memory centers of brain. α7 nAChR PAMs are anticipated to side-step burgeoning issues observed with several clinical-stage orthosteric α7 nAChR agonists, related to selectivity, tolerance/tachyphylaxis, thus providing a novel dimension in therapeutic strategy and pharmacology of α7 nAChR ion-channel. Here we describe a novel α7 nAChR PAM, LL-00066471, which potently amplified agonist-induced Ca2+ fluxes in neuronal IMR-32 neuroblastoma cells in a α-bungarotoxin (α-BTX) sensitive manner. LL-00066471 showed excellent oral bioavailability across species (mouse, rat and dog), low clearance and good brain penetration (B/P ratio > 1). In vivo, LL-00066471 robustly attenuated cognitive deficits in both procognitive and antiamnesic paradigms of short-term episodic and recognition memory in novel object recognition task (NORT) and social recognition task (SRT), respectively. Additionally, LL-00066471 mitigated apomorphine-induced sensorimotor gating deficits in acoustic startle reflex (ASR) and enhanced antipsychotic efficacy of olanzapine in conditioned avoidance response (CAR) task. Further, LL-00066471 corrected redox-imbalances and reduced cortico-striatal infarcts in stroke model. These finding together suggest that LL-00066471 has potential to symptomatically alleviate cognitive deficits associated with dementias, attenuate sensorimotor gating deficits in schizophrenia and correct redox-imbalances in cerebrovascular disorders. Therefore, LL-00066471 presents potential for management of cognitive impairments associated with neurological and psychiatric conditions.

Mechanism of Oxidative Stress and Synapse Dysfunction in the Pathogenesis of Alzheimer's Disease: Understanding the Therapeutics Strategies.

Synapses are formed by interneuronal connections that permit a neuronal cell to pass an electrical or chemical signal to another cell. This passage usually gets damaged or lost in most of the neurodegenerative diseases. It is widely believed that the synaptic dysfunction and synapse loss contribute to the cognitive deficits in patients with Alzheimer's disease (AD). Although pathological hallmarks of AD are senile plaques, neurofibrillary tangles, and neuronal degeneration which are associated with increased oxidative stress, synaptic loss is an early event in the pathogenesis of AD. The involvement of major kinases such as mitogen-activated protein kinase (MAPK), extracellular receptor kinase (ERK), calmodulin-dependent protein kinase (CaMKII), glycogen synthase-3ß (GSK-3ß), cAMP response element-binding protein (CREB), and calcineurin is dynamically associated with oxidative stress-mediated abnormal hyperphosphorylation of tau and suggests that alteration of these kinases could exclusively be involved in the pathogenesis of AD. N-methyl-D-aspartate (NMDA) receptor (NMDAR) activation and beta amyloid (Aß) toxicity alter the synapse function, which is also associated with protein phosphatase (PP) inhibition and tau hyperphosphorylation (two main events of AD). However, the involvement of oxidative stress in synapse dysfunction is poorly understood. Oxidative stress and free radical generation in the brain along with excitotoxicity leads to neuronal cell death. It is inferred from several studies that excitotoxicity, free radical generation, and altered synaptic function encouraged by oxidative stress are associated with AD pathology. NMDARs maintain neuronal excitability, Ca(2+) influx, and memory formation through mechanisms of synaptic plasticity. Recently, we have reported the mechanism of the synapse redox stress associated with NMDARs altered expression. We suggest that oxidative stress mediated through NMDAR and their interaction with other molecules might be a driving force for tau hyperphosphorylation and synapse dysfunction. Thus, understanding the oxidative stress mechanism and degenerating synapses is crucial for the development of therapeutic strategies designed to prevent AD pathogenesis.

Ameliorative effect of Noni fruit extract on streptozotocin-induced memory impairment in mice.

This study evaluated the effects of a standardized ethyl acetate extract of Morinda citrifolia L. (Noni) fruit on impairment of memory, brain energy metabolism, and cholinergic function in intracerebral streptozotocin (STZ)-treated mice. STZ (0.5 mg/kg) was administered twice at an interval of 48 h. Noni (50 and 100 mg/kg, postoperatively) was administered for 21 days following STZ administration. Memory function was evaluated using Morris Water Maze and passive avoidance tests, and brain levels of cholinergic function, oxidative stress, energy metabolism, and brain-derived neurotrophic factor (BDNF) were estimated. STZ caused memory impairment in Morris Water Maze and passive avoidance tests along with reduced brain levels of ATP, BDNF, and acetylcholine and increased acetylcholinesterase activity and oxidative stress. Treatment with Noni extract (100 mg/kg) prevented the STZ-induced memory impairment in both behavioral tests along with reduced oxidative stress and acetylcholinesterase activity, and increased brain levels of BDNF, acetylcholine, and ATP level. The study shows the beneficial effects of Noni fruit against STZ-induced memory impairment, which may be attributed to improved brain energy metabolism, cholinergic neurotransmission, BDNF, and antioxidative action.

Protective effect of Cnestis ferruginea and its active constituent on scopolamine-induced memory impairment in mice: a behavioral and biochemical study.

CONTEXT: Cnestis ferruginea Vahl ex DC (Connaraceae) (CF) is used in traditional African medicine in the management of CNS disorders. The degeneration and dysfunction of cholinergic neurons is closely associated with the cognitive deficits of Alzheimer's disease (AD) and oxidative stress has been implicated in its pathogenesis. However, the influence of C. ferruginea on the cholinergic system and oxidative stress parameters has not been explored. OBJECTIVE: The present study investigates the effect of methanol root extract of C. ferruginea and its active constituent amentoflavone (CF-2) on memory, oxidative stress and acetylcholinesterase (AChE) activity in scopolamine-induced amnesia. MATERIALS AND METHODS: Mice were orally treated with CF (25-200 mg/kg), CF-2 (6.25-25 mg/kg) for three days and memory impairment was induced by intraperitoneal injection of scopolamine (3 mg/kg). Memory function was evaluated by passive avoidance and Morris water maze tests. Biochemical parameters of oxidative stress and cholinergic function were estimated in brain after the completion of behavioral studies. RESULTS: Scopolamine caused memory impairment along with increased AChE activity and oxidative stress in mice brain. Oral administration of CF and CF-2 significantly prevented scopolamine-induced memory impairment, inhibited AChE and enhanced antioxidant enzyme activity in the brain following scopolamine injection as compared to vehicle administration in scopolamine (i.p.)-treated mice that were comparable to the effect of tacrine. DISCUSSION AND CONCLUSION: The study demonstrated that C. ferruginea and its constituent have significant protective effect against scopolamine-induced memory deficits in mice that can be attributed to their antioxidant and antiAChE activity.

Effect of angiotensin II on spatial memory, cerebral blood flow, cholinergic neurotransmission, and brain derived neurotrophic factor in rats.

RATIONAL: Studies have shown the involvement of angiotensin II (Ang II) in neurobehavioral aspects, but the exact role of Ang II in memory is still ambiguous. OBJECTIVE: This study explored the effect of central Ang II on spatial memory along with cholinergic neurotransmission, brain energy metabolism, cerebral blood flow (CBF), and brain-derived neurotrophic factor (BDNF) in rats. METHODS: Spatial memory was evaluated by Morris water maze (MWM) after Ang II (ICV) administration in male Sprague-Dawley rats. CBF was measured by laser Doppler flowmetry. Oxidative stress adenosine triphosphate (ATP), BDNF, acetylcholinesterase (AChE), and acetylcholine (ACh) were estimated in the cortex and hippocampus at 1, 24, and 48 h after Ang II administration. The effect of AT1 and AT2 receptor blocker (candesartan and PD123,319, respectively), AChE inhibitor (donepezil), and antioxidant melatonin was studied on memory, CBF, and biochemical parameters. RESULTS: Ang II caused spatial memory impairment by affecting acquisition, consolidation, and recall in the MWM test along with a significant reduction in CBF. Ang II significantly reduced ACh level and caused oxidative stress in the rat brain 1 h post-injection. No significant change was observed in BDNF, AChE, and ATP level. Candesartan and donepezil prevented Ang II-induced memory impairment, reduction in CBF and ACh level. However, PD123,319 and melatonin failed to prevent Ang II-induced memory impairment but improved CBF partially. CONCLUSION: This study suggests that Ang II, via the AT1 receptor, affects spatial memory formation, CBF, and ACh level while AT2 receptor has no significant role.

Combretum mucronatum and Capparis thonningii prevent scopolamine-induced memory deficit in mice.

CONTEXT: Roots of Combretum mucronatum Schumach. & Thonn. (Combretaceae) and Capparis thonningii Schum. (Capparaceae) are used in southwest Nigeria in the treatment of inflammatory disorders and mental illness. OBJECTIVE: This study evaluated the antidementic effect of the methanol root extracts of C. mucronatum and C. thonningii on scopolamine (3 mg/kg, i.p.) induced memory impairment in mice. MATERIALS AND METHODS: The effect of C. mucronatum and C. thonningii (50-200 mg/kg) administered orally for 3 days on memory impairments induced in mice by scopolamine was assessed in the passive avoidance and Morris water maze test and compared with that of tacrine (5 mg/kg, i.p.). The activities of acetylcholinesterase (AchE) and antioxidant enzymes were estimated in the brain after the completion of behavioral studies. RESULTS: C. mucronatum and C. thonningii root extracts (50-200 mg/kg) reversed scopolamine-induced memory deficit with significant (p < 0.05) increase in transfer latency in passive avoidance test. Similarly, the extracts (200 mg/kg) ameliorated memory deficit as a result of significant (p < 0.001) decrease in escape latency and path length in Morris water maze test. The increased AChE activity induced by scopolamine was significantly (p < 0.05) inhibited by C. mucronatum and C. thonningii (100 and 200 mg/kg) treatment which was similar to the effect of tacrine. Both extracts significantly (p < 0.05) attenuated scopolamine-induced increase in oxidative stress parameters as well as restoration of glutathione activity. DISCUSSION AND CONCLUSION: C. mucronatum and C. thonningii extracts possess significant anticholinesterase, antioxidant and antidementic properties, which may be useful in the management of Alzheimer's disease.

Lead optimization studies towards the discovery of novel carbamates as potent AChE inhibitors for the potential treatment of Alzheimer's disease.

The optimization of our previous lead compound 1 (AChE IC(50)=3.31 µM) through synthesis and pharmacology of a series of novel carbamates is reported. The synthesized compounds were evaluated against mouse brain AChE enzyme using the colorimetric method described by Ellman et al. The three compounds 6a (IC(50)=2.57µM), 6b (IC(50)=0.70 µM) and 6i (IC(50)=2.56 µM) exhibited potent in vitro AChE inhibitory activities comparable to the drug rivastigmine (IC(50)=1.11 µM). Among them, the compound 6b has been selected as possible optimized lead for further neuropharmacological studies. In addition, the AChE-carbamate Michaelis complexes of these potent compounds including rivastigmine and ganstigmine have been modeled using covalent docking protocol of GOLD and important direct/indirect interactions contributing to stabilization of the AChE-carbamate Michaelis complexes have been investigated.

Antidepressant and anxiolytic effects of amentoflavone isolated from Cnestis ferruginea in mice.

The root decoction of Cnestis ferruginea (CF) Vahl DC (Connaraceae) is used in traditional African medicine in the management of psychiatric disorders. This study presents the antidepressant and anxiolytic effects of amentoflavone (CF-2) isolated from the root extract of C. ferruginea. The antidepressant effect was studied using the forced swimming (FST) and tail suspension tests (TST) while the hole-board, elevated plus maze (EPM) and light/dark tests were used to evaluate the anxiolytic effect. Acute treatment with CF extract and amentoflavone significantly (p<0.001) reduced the duration of immobility in FST and TST with peak effects observed at 100 and 50mg/kg respectively in comparison to control treated. Antidepressant effects of CF and amentoflavone were significantly higher (p<0.05) when compared to imipramine in FST but comparable to the fluoxetine treated group in TST. The pretreatment of mice with metergoline (4mg/kg, i.p., a 5-HT2 receptor antagonist), prazosin (62.5µg/kg, i.p., an α1-adrenoceptor antagonist), and yohimbine (1mg/kg, i.p., an α2-adrenoceptor antagonist), but not sulpiride (50mg/kg, i.p., a dopamine D2 receptor antagonist), cyproheptadine (3mg/kg, i.p., a 5-HT2 receptor antagonist), atropine (1mg/kg, i.p., a muscarinic receptor antagonist) 15mins before the administration of amentoflavone (50mg/kg; p.o.) significantly prevented its antiimmobility effect in the FST. CF extract and CF-2 significantly (p<0.05) attenuated anxiety by increasing the number of head-dips in the hole-board test, the time spent on the open arms in the EPM, and the exploration of the light chamber in the light/dark test. Pretreatment with flumazenil (3mg/kg, i.p., ionotropic GABA receptor antagonist) 15min before oral administration of amentoflavone (25mg/kg) significantly reduced the time spent in the open arms in EPM. It is concluded from the results obtained that amentoflavone produces its antidepressant effect through interaction with 5-HT2 receptor and α1-, and α2-adrenoceptors while the anxiolytic effect involved the ionotropic GABA receptor.

Okadaic acid induced neurotoxicity leads to central cholinergic dysfunction in rats.

Central cholinergic system is involved in regulation of memory and disturbances in these results in memory loss. Previously, we examined the effect of okadaic acid, OKA (200ng, i.c.v.) on memory impairment and mitochondrial dysfunction in rats. In the present study, we investigated effect of OKA (i.c.v) on cholinergic function by observing acetylcholine level (ACh), acetylcholinestrase (AChE) activity, and mRNA expression of acetylcholinestrase and α7nicotinic receptor (α7-nAChR) as a cholinergic markers in brain areas (cerebellum, striatum cortex and hippocampus). In present work OKA, caused a significant decrease in acetylcholine level, acetylcholinestrase activity and mRNA expression of acetylcholinestrase and α7-nicotinic receptor in rat but these changes were mainly observed in cortex and hippocampus. Further, histopathological study by cresyl violet staining showed neuronal loss in cortex and hippocampus after OKA administration indicating neurotoxicity. Pretreatment with anti-dementic drugs donepezil (AChE inhibitor; 5mg/kg, p.o) and memantine (NMDA receptor antagonist; 10mg/kg, p.o) daily for 13 day prevented cholinergic dysfunction and neuronal loss in cortex and hippocampus of OKA treated rat. Daily per se treatment for 13 day with donepezil decreased acetylcholinestrase activity and increased mRNA expression of acetylcholinestrase and α7-nicotinic receptor. Whereas, per se treatment with memantine daily for 13 day did not affect acetylcholinestrase activity, mRNA expression of acetylcholinestrase and α7-nicotinic receptor. Findings of this work shows that OKA (i.c.v.), apart from memory impairment and mitochondrial dysfunction, as our previous study showed, also induced cholinergic dysfunction and neuronal loss, which can be addressed by antidementic drugs like donepezil and memantine.

A study on neuroinflammatory marker in brain areas of okadaic acid (ICV) induced memory impaired rats.

AIMS: The aim of the present study is to investigate the status of proinflammatory cytokine in the brain of intracerebroventricular (i.c.v.) okadaic acid (OKA) induced memory impaired rat. MAIN METHODS: OKA (200 ng) intracerebroventricular (i.c.v.) was administered in rats. Memory was assessed by Morris water maze test. Biochemical marker of neuroinflammation (TNF-α, IL-ß), total nitrite, mRNA (RT PCR) and protein expression (WB) of iNOS and nNOS were estimated in rat brain areas. KEY FINDINGS: OKA caused memory-impairment in rats with increased expression of proinflammatory cytokine TNF-α and IL-1ß and total nitrite in brain regions hippocampus and cortex. The expression of mRNA and protein of iNOS was increased while; the expressions were decreased in case of nNOS. Pretreatment with antidementic drugs donepezil (5 mg/kg, p.o.) and memantine (10 mg/kg, p.o) for 13 days protected i.c.v. OKA induced memory impairment and changes in level of TNF-α, IL-ß, total nitrite and expressions of iNOS and nNOS in OKA treated rat. SIGNIFICANCE: This study suggests that neuroinflammation may play a vital role in OKA induced memory impairment.

Inhibition of central angiotensin converting enzyme ameliorates scopolamine induced memory impairment in mice: role of cholinergic neurotransmission, cerebral blood flow and brain energy metabolism.

Evidences indicate that inhibition of central Renin angiotensin system (RAS) ameliorates memory impairment in animals and humans. Earlier we have reported involvement of central angiotensin converting enzyme (ACE) in streptozotocin induced neurodegeneration and memory impairment. The present study investigated the role of central ACE in cholinergic neurotransmission, brain energy metabolism and cerebral blood flow (CBF) in model of memory impairment induced by injection of scopolamine in mice. Perindopril (0.05 and 0.1 mg/kg, PO) was given orally for one week before administration of scopolamine (3mg/kg, IP). Then, memory function was evaluated by Morris water maze and passive avoidance tests. CBF was measured by laser Doppler flowmetry. Biochemical and molecular parameters were estimated after the completion of behavioral studies. Scopolamine caused impairment in memory which was associated with reduced CBF, acetylcholine (ACh) level and elevated acetylcholinesterase (AChE) activity and malondialdehyde (MDA) level. Perindopril ameliorated scopolamine induced amnesia in both the behavioral paradigms. Further, perindopril prevented elevation of AChE and MDA level in mice brain. There was a significant increase in CBF and ACh level in perindopril treated mice. However, scopolamine had no significant effect on ATP level and mRNA expression of angiotensin receptors and ACE in cortex and hippocampus. But, perindopril significantly decreased ACE activity in brain without affecting its mRNA expression. The study clearly showed the interaction between ACE and cholinergic neurotransmission and beneficial effect of perindopril can be attributed to improvement in central cholinergic neurotransmission and CBF.

Role of central angiotensin receptors in scopolamine-induced impairment in memory, cerebral blood flow, and cholinergic function.

RATIONAL: Inhibition of renin-angiotensin system (RAS) improves cognitive functions in hypertensive patients. However, role of AT1 and AT2 receptors in memory impairment due to cholinergic hypofunction is unexplored. OBJECTIVE: This study investigated the role of AT1 and AT2 receptors in cerebral blood flow (CBF), cholinergic neurotransmission, and cerebral energy metabolism in scopolamine-induced amnesic mice. METHODS: Scopolamine was given to male Swiss albino mice to induce memory impairment tested in passive avoidance and Morris water maze tests after a week long administration of blocker of AT1 receptor, candesartan, and AT2 receptor, PD123, 319. CBF was measured by laser Doppler flowmetry. Biochemical and molecular studies were done in cortex and hippocampus of mice brain. RESULTS: Scopolamine caused memory impairment, reduced CBF, acetylcholine (ACh) level, elevated acetylcholinesterase (AChE) activity, and malondialdehyde (MDA). Administration of vehicle had no significant effect on any parameter in comparison to control. Candesartan prevented scopolamine-induced amnesia, restored CBF and ACh level, and decreased AChE activity and MDA level. In contrast, PD123, 319 was not effective. However, the effect of AT1 receptor blocker on memory, CBF, ACh level, and oxidative stress was blunted by concomitant blockade of AT2 receptor. Angiotensin-converting enzyme (ACE) activity, ATP level, and mRNA expression of AT1, AT2, and ACE remained unaltered. CONCLUSION: The study suggests that activation of AT1 receptors appears to be involved in the scopolamine-induced amnesia and that AT2 receptors contribute to the beneficial effects of candesartan. Theses finding corroborated the number of clinical studies that RAS inhibition in hypertensive patients could be neuroprotective.

Central angiotensin converting enzyme facilitates memory impairment in intracerebroventricular streptozotocin treated rats.

Preclinical and clinical studies indicated involvement of renin angiotensin system (RAS) in memory functions. However, exact role of RAS in cognition is still ambiguous. Our aim was to explore how angiotensin converting enzyme (ACE) modulates memory in experimental model of memory impairment. Memory deficit was induced by intracerebroventricular administration of streptozotocin (STZ, 3mg/kg) in rats. Perindopril, an ACE inhibitor, was given for 21 days and memory function was evaluated by Morris water maze test. Cerebral blood flow (CBF) was measured by laser doppler flowmetry. The biochemical and expression studies were done in cortex and hippocampus of rat brain after the completion of behavioral studies. STZ caused impairment in memory along with significant reduction in CBF, ATP level and elevated oxidative and nitrosative stress. The activity and mRNA expression of acetylcholinesterase (AChE) and ACE were also increased in rat brain regions following STZ administration. However, serum ACE activity remained unaffected. Treatment with perindopril dose dependently improved memory by increasing energy metabolism and CBF. Perindopril also decreased oxidative and nitrosative stress, activity and mRNA expression of AChE and ACE in STZ treated rat. Further, ACE inhibition mitigated STZ induced neurodegeneration as observed in histopathological studies. Moreover, perindopril per se improved memory and CBF, decreased oxidative stress with no effect on AChE activity and expression. However, perindopril per se significantly reduced ACE activity but increased mRNA expression of ACE in rat brain. These results suggest that ACE occupies a pivotal role in STZ induced memory deficit thus implicating central RAS in cognition.

Protective effect of fruits of Morinda citrifolia L. on scopolamine induced memory impairment in mice: a behavioral, biochemical and cerebral blood flow study.

ETHNOPHARMACOLOGICAL RELEVANCE: Noni (Morinda citrifolia L.) is widely used for different illnesses including CNS disorders. Recently Noni has been reported to prevent amyloid beta induced memory impairment in mice. However, the influence of Noni on cholinergic system has not been explored so far. Therefore, present study was designed to investigate effect of Noni fruit on memory, cerebral blood flow (CBF), oxidative stress and acetylcholinesterase (AChE) activity in scopolamine induced amnesia model. MATERIALS AND METHODS: Mice were orally treated with ethanolic extract of Noni fruit and chloroform, ethyl acetate and butanol fractions of ethanolic extract for three days. Scopolamine was administered 5 min prior to acquisition trial and memory function was evaluated by passive avoidance test. CBF was measured by laser doppler flowmetry. AChE activity and oxidative stress parameters were estimated in mice brain at the end of behavioral studies. Further, effect of ethanolic extract and its fractions (5-400 µg/ml) on AChE activity was measured in vitro. RESULTS: Scopolamine caused memory impairment along with reduced CBF, increased AChE activity and oxidative stress in mice brain. Ethanolic extract of Noni fruits and its chloroform and ethyl acetate fractions significantly improved memory and CBF. However, butanol fraction had no effect. Further, increased oxidative stress and AChE activity following scopolamine was significantly attenuated by ethanolic extract of Noni and its fractions. Moreover ethanolic extract and its fractions showed dose dependent inhibition of AChE activity in vitro. CONCLUSION: These observations suggest that Noni may be useful in memory impairment due to its effect on CBF, AChE and oxidative stress.

Effect of pomegranate juice on the pharmacokinetics of nitrendipine in rabbits.

Pomegranate juice (PJ) is known to be a potent inhibitor of human cytochrome enzymes. The purpose of this study was to investigate the effect of acute and chronic PJ on the pharmacokinetics of oral nitrendipine (10 mg/kg) in rabbits. Male New Zealand rabbits were pretreated with PJ for 1 week and on the last day, a single dose of nitrendipine was given orally. In another group, both PJ and nitrendipine were co-administered to evaluate the acute effect of PJ on nitrendipine pharmacokinetics. The control group received oral distilled water for 1 week and administered with nitrendipine on the last day. Blood samples were collected at different time points and nitrendipine concentration was estimated by high-performance liquid chromatography. Relative to control, the area under the concentration-time curve and peak plasma concentration of nitrendipine were 2.03- and 2-fold, respectively, greater in the PJ-pretreated group. However, co-administration of PJ had no significant effect on these parameters. Further, there was no significant change in the elimination rate constant and elimination half-life of nitrendipine in both PJ co-administered and pretreated groups in comparison with control. These results suggest that PJ inhibits the intestinal metabolism of nitrendipine without affecting hepatic metabolism in rabbits. Although this potential interaction needs to be explored further, the concomitant use of PJ and nitrendipine should be avoided.

Mitochondrial dysfunction: a crucial event in okadaic acid (ICV) induced memory impairment and apoptotic cell death in rat brain.

Mitochondrial abnormalities have been identified in a large proportion of neurodegenerative diseases. Recently we have reported that intracerebroventricular (ICV) administration of okadaic acid (OKA) causes memory impairment in rat. However involvement of mitochondrial function in OKA induced memory impairment and neuronal damage has not been determined. OKA (200 ng) was administered by ICV route. After 13th day of OKA administration memory function was evaluated by Morris Water Maze test. Following completion of behavioral studies on 16th day, mitochondrial membrane potential, Ca(2+) and reactive oxygen species were evaluated in mitochondrial preparation of cortex, hippocampus, striatum and cerebellum of rat brain. While ATP, mitochondrial activity, lipid peroxidation and nitrite were investigated in synaptosomal preparation of rat brain areas. The activities and mRNA expression of apoptotic factors, caspase-3 and caspase-9, were studied in rat brain regions. The neuronal damage was also confirmed by histopathological study. OKA treated rats showed memory impairment including increased Ca(2+) and reactive oxygen species and decreased mitochondrial membrane potential, ATP and mitochondrial activity in mitochondrial preparation. There was a significant increase in lipid peroxidation and nitrite in synaptosomal preparations. Preventive treatment daily for 13 days with antidementic drugs, donepezil (5 mg/kg, p.o) and memantine (10 mg/kg, p.o), significantly attenuated OKA induced mitochondrial dysfunction, apoptotic cell death, memory impairment and histological changes. Mitochondrial dysfunction appeared as a key factor in OKA induced memory impairment and apoptotic cell death. This study indicates that clinically used antidementic drugs are effective against OKA induced adverse changes at behavioral, cellular, and histological levels and mitochondrial dysfunction.

Improvement of brain energy metabolism and cholinergic functions contributes to the beneficial effects of silibinin against streptozotocin induced memory impairment.

Recently, silibinin, a clinically used hepatoprotectant, has been reported to prevent amyloid beta induced memory impairment by reducing oxidative stress and inflammation in mice brain. However, the exact mechanism of neuroprotective effect of silibinin has not been properly studied especially in context of brain energy metabolism and cholinergic functions, the essential factors that undergo impairment in Alzheimer's disease. Therefore, the present study investigated the effect of silibinin on impairment in memory, brain energy metabolism and cholinergic function following intracerebral (IC) streptozotocin (STZ) administration in mice. STZ (0.5mg/kg), administered twice at an interval of 48h, caused significant memory impairment tested by Morris water maze. Further, STZ significantly decreased ATP and increased synaptosomal calcium level in mice brain. Increased oxidative and nitrosative stress was also observed in IC STZ injected mice brain. STZ IC induced memory impairment is associated with increased activity and mRNA expression of acetylcholinesterase (AChE) and decreased α-7 nicotinic acetylcholine receptor (α-7-nAChR) mRNA expression in mice brain. Pretreatment with silibinin (100 and 200mg/kg, po) attenuated STZ induced memory impairment by reducing oxidative and nitrosative stress and synaptosomal calcium ion level. Further, silibinin dose dependently restored ATP level indicating improvement in brain energy metabolism. The activity and mRNA expression of AChE was restored by silibinin. Moreover, α-7-nAChR mRNA expression was significantly increased by silibinin in STZ treated mice brain. The present study clearly demonstrates that beneficial effects of silibinin in STZ induced memory impairment in mice is due to improvement in brain energy metabolism and cholinergic function.

Protective effect of quercetin against intracerebral streptozotocin induced reduction in cerebral blood flow and impairment of memory in mice.

The aim of the present study is to investigate the effect of quercetin, a naturally occurring flavonoid, on cerebral blood flow (CBF), brain energy metabolism, memory impairment, oxidative stress and cholinergic dysfunction in brain following intracerebral (i.c.) streptozotocin (STZ) administration in mice. STZ (0.5mg/kg, i.c.) was administered twice at an interval of 48h. We found a significant reduction in CBF as measured by Laser Doppler Flowmetry (LDF). The brain energy metabolism was also altered as evidenced by significant reduction in brain ATP content. Daily treatment with quercetin (2.5, 5 and 10mg/kg, p.o.) starting from the first dose of STZ showed a dose-dependent restoration of CBF and ATP content. Further, quercetin prevented STZ induced memory impairment as assessed by Morris water maze and passive avoidance tests. Biochemical analysis revealed that STZ significantly increased malondialdehyde (MDA), nitrite and depleted glutathione (GSH) levels in the mice brain. Quercetin decreased oxidative and nitrosative stress as evidenced by a significant decrease in MDA, nitrite and increase in GSH levels. Quercetin also attenuated elevated acetylcholinesterase activity in the STZ-treated mice. Neither STZ (i.c.) nor quercetin showed any change in locomotor activity and blood glucose level. The present study demonstrates the beneficial effects of quercetin in improving CBF along with preventing memory impairment, oxidative stress, altered brain energy metabolism and cholinergic dysfunction caused by STZ in mice. Therefore, consumption of dietary stuff rich in quercetin should be encouraged to ward off dementia associated with vascular and neurodegenerative disorders.

Okadaic acid (ICV) induced memory impairment in rats: a suitable experimental model to test anti-dementia activity.

Okadaic acid (OKA) is a potent and selective inhibitor of protein phosphatases, PP2A and PP1. In the present study, we evaluated effect of intracerebroventricular (ICV) bilateral injection of OKA (100 and 200 ng) on memory function and oxidative stress in rats. ICV injection of OKA (200 ng) produced memory impairment as evidenced by no significant decrease in latency time to reach the hidden platform in water maze test. It produced increase in malondialdehyde (MDA), nitrite level, reactive oxygen species (ROS) generation, mitochondrial calcium ion [Ca(2)](i) level and decreased glutathione (GSH) level in rat brain areas, indicating oxidative stress. Furthermore, we evaluated the effect of anti-dementia drugs memantine, a NMDA antagonist, and donepezil, a cholinesterase inhibitor, on OKA ICV induced memory impairment. Administration of memantine (10 mg/kg, p.o.) and donepezil (5 mg/kg, p.o.) for 13 days starting from the OKA injection improved performance in memory tests and also significantly restored GSH, MDA, nitrite levels, ROS generation and [Ca(2+)](i) level. This study demonstrates that the clinically used anti-dementic drugs are effective in OKA induced free radical generation and memory impairment in rats. Thus, OKA ICV induced memory impairment in rat appeared as a useful test model to screen anti-dementia drugs.

Protective effect of curcumin against intracerebral streptozotocin induced impairment in memory and cerebral blood flow.

AIMS: The aim of the present study is to investigate the effect of curcumin on cerebral blood flow (CBF), memory impairment, oxidative stress and cholinergic dysfunction in intracerebral (IC) streptozotocin (STZ) induced memory impairment in mice. MAIN METHODS: Memory impairment was induced by STZ (0.5mg/kg, IC) administered twice with an interval of 48h in mice. Memory function was assessed by Morris water maze and passive avoidance test. CBF was measured by Laser Doppler Flowmetry (LDF). To study the preventive effect, curcumin (10, 20 and 50mg/kg, PO) was administered for 21days starting from the first dose of STZ. In another set of experiment, curcumin was administered for 7days from 19th day after confirming STZ induced dementia to observe its therapeutic effect. Biochemical parameters of oxidative stress and cholinergic function were estimated in brain on day 21. KEY FINDINGS: The major finding of this study is that STZ (IC) caused a significant reduction in CBF along with memory impairment, cholinergic dysfunction and enhanced oxidative stress. Curcumin dose dependently improved CBF in STZ treated mice together with amelioration of memory impairment both in preventive and therapeutic manner. SIGNIFICANCE: The present study clearly demonstrates the beneficial effects of curcumin, the dietary staple of India, on CBF, memory and oxidative stress which can be exploited for dementia associated with age related vascular and neurodegenerative disorders.