Pranay Srivastava

Unraveling the mechanism of neuroprotection of curcumin in arsenic induced cholinergic dysfunctions in rats

Earlier, we found that arsenic induced cholinergic deficits in rat brain could be protected by curcumin. In continuation to this, the present study is focused to unravel the molecular mechanisms associated with the protective efficacy of curcumin in arsenic induced cholinergic deficits. Exposure to arsenic (20mg/kg body weight, p.o) for 28 days in rats resulted to decrease the expression of CHRM2 receptor gene associated with mitochondrial dysfunctions as evident by decrease in the mitochondrial membrane potential, activity of mitochondrial complexes and enhanced apoptosis both in the frontal cortex and hippocampus in comparison to controls. The ultrastructural images of arsenic exposed rats, assessed by transmission electron microscope, exhibited loss of myelin sheath and distorted cristae in the mitochondria both in the frontal cortex and hippocampus as compared to controls. Simultaneous treatment with arsenic (20mg/kg body weight, p.o) and curcumin (100mg/kg body weight, p.o) for 28 days in rats was found to protect arsenic induced changes in the mitochondrial membrane potential and activity of mitochondrial complexes both in frontal cortex and hippocampus. Alterations in the expression of pro- and anti-apoptotic proteins and ultrastructural damage in the frontal cortex and hippocampus following arsenic exposure were also protected in rats simultaneously treated with arsenic and curcumin. The data of the present study reveal that curcumin could protect arsenic induced cholinergic deficits by modulating the expression of pro- and anti-apoptotic proteins in the brain. More interestingly, arsenic induced functional and ultrastructural changes in the brain mitochondria were also protected by curcumin.

Scopolamine induced behavioral and biochemical modifications and protective effect of Celastrus paniculatous and Angelica glauca in rats

Introduction: Nootropic agents, including cholinesterase inhibitors are being used to improve memory, mood and behavior, but the side-effects associated with these agents have made their use limited. The present study has therefore been undertaken to assess the synergistic effects of Celastrus paniculatous and Angelica glauca on scopolamine induced dementia in rats. Materials and Methods: Rats were treated with scopolamine (1 mg/kg body weight, i.p.) alone and with donepezil (2 mg/kg body weight p.o.), C. paniculatous (150 mg/kg body weight, p.o) and A. glauca (150 mg/kg body weight, p.o.). The changes in behavioral and biochemical parameters were assessed in rats. Results: Scopolamine treated rats showed impaired learning and memory, increased activity of acetylcholinesterase (AChE) (18%), lipid peroxidation (60%), protein carbonyls (47%) and decreased levels of reduced glutathione (GSH) (35%), activity of superoxide dismutase (34%) and catalase (42%) in hippocampus as compared with control. Simultaneous treatment of C. paniculatous and A. glauca with scopolamine also caused an improvement in the learning and memory activity associated with AChE activity in hippocampus of rats as compared to those treated with scopolamine alone. Combined treatment of C. paniculatous, A. glauca and scopolamine significantly improved the learning and memory function and AChE activity (30%) associated with decreased lipid peroxidation (33%), protein carbonyls (27%) and increased levels of antioxidant enzymes like reduced GSH (46%), activity of superoxide dismutase (50%) and catalase (62%) in hippocampus of rats as compared with those treated with scopolamine alone. Conclusion: The results of the present study exhibit protective efficacy of combined treatment of C. paniculatous and A. glauca in scopolamine induced dementiaand promising as a memory enhancing agents that is associated with its strong antioxidant potential.

Nose to brain delivery of astaxanthin-loaded solid lipid nanoparticles: fabrication, radio labeling, optimization and biological studies

The present study was carried out to investigate the intranasal delivery of astaxanthin as solid lipid nanoparticles with the intention of improving brain targeting of astaxanthin for neurological disorders. The astaxanthin solid lipid nanoparticles were prepared by a double emulsion solvent displacement method. Statistical analysis using response surface methodology showed that the optimum values of stearic acid (50 mg), drug weight percentage (6.11%) and ratio of surfactant to co-surfactant (poloxamer 188 : lecithin-1 : 6) resulted in a 213.23 nm particle size and 0.367 polydispersity index for the astaxanthin solid lipid nanoparticles. Radio labeling studies were performed using technetium-99m to evaluate the biodistribution pattern after administration through different routes in experimental subjects. Radiolabeled nanoparticles were found to be 96–98% stable even after 48 h of labeling in phosphate-buffered saline (pH 7.4). Comparative biodistribution data indicated that the higher drug concentration in the brain was achieved by intranasal administration of 99mTc labeled astaxanthin solid lipid nanoparticles as compared to the intravenous route, which was also confirmed by gamma scintigraphy analysis. Furthermore, studies on the pheochromocytoma-12 cell line demonstrated the antioxidant potential of astaxanthin solid lipid nanoparticles against H2O2 induced toxicity. Our findings strongly emphasize that nanoparticle-based nasal drug delivery of astaxanthin could impart the utmost neuroprotection from oxidative stress in neurological disorders under in vitro conditions.

Brain cholinergic alterations in rats subjected to repeated immobilization or forced swim stress on lambda-cyhalothrin exposure

Role of immobilization stress (IMS), a psychological stressor and forced swim stress (FSS), a physical stressor was investigated on the neurobehavioral toxicity of lambda-cyhalothrin (LCT), a new generation type-II synthetic pyrethroid. Pre-exposure of rats to IMS (15 min/day) or FSS (3 min/day) for 28 days on LCT (3.0 mg/kg body weight, p.o.) treatment for 3 days resulted to decrease spatial learning and memory and muscle strength associated with cholinergic-muscarinic receptors in frontal cortex and hippocampus as compared to those exposed to IMS or FSS or LCT alone. Decrease in acetylcholinesterase activity, protein expression of ChAT and PKC-β1 associated with decreased mRNA expression of CHRM2, AChE and ChAT in frontal cortex and hippocampus was also evident in rats pre-exposed to IMS or FSS on LCT treatment, compared to rats exposed to IMS or FSS or LCT alone. Interestingly, changes both in behavioral and neurochemical endpoints were marginal in rats subjected to IMS or FSS for 28 days or those exposed to LCT for 3 days alone, compared to controls. The results suggest that stress is an important contributor in LCT induced cholinergic deficits.

PI3K/Akt/GSK3β induced CREB activation ameliorates arsenic mediated alterations in NMDA receptors and associated signaling in rat hippocampus: Neuroprotective role of curcumin

HIGHLIGHTSProtective role of curcumin in arsenic induced NMDA receptor alterations investigated.Sodium arsenite decreased expression of BDNF, pAkt, pErk ½, pGSK3&bgr; and pCREB.Curcumin protected sodium arsenite induced changes via PI3/Akt/MAPK pathway in hippocampus.Impaired cognitive deficits by sodium arsenite in rats also recovered by curcumin. ABSTRACT Protective efficacy of curcumin in arsenic induced NMDA receptor dysfunctions and PI3K/Akt/ GSK3&bgr; signalling in hippocampus has been investigated in vivo and in vitro. Exposure to sodium arsenite (in vivo – 20mg/kg, body weight p.o. for 28 days; in vitro – 10&mgr;M for 24h) and curcumin (in vivo – 100mg/kg body weight p.o. for 28 days; in vitro – 20&mgr;M for 24h) was carried out alone or simultaneously. Treatment with curcumin ameliorated sodium arsenite induced alterations in the levels of NMDA receptors, its receptor subunits and synaptic proteins ‐ pCaMKII&agr;, PSD‐95 and SynGAP both in vivo and in vitro. Decreased levels of BDNF, pAkt, pERK1/2, pGSK3&bgr; and pCREB on sodium arsenite exposure were also protected by curcumin. Curcumin was found to decrease sodium arsenite induced changes in hippocampus by modulating PI3K/Akt/GSK3&bgr; neuronal survival pathway, known to regulate various cellular events. Treatment of hippocampal cultures with pharmacological inhibitors for ERK1/2, GSK3&bgr; and Akt individually inhibited levels of CREB and proteins associated with PI3K/Akt/GSK3&bgr; pathway. Simultaneous treatment with curcumin was found to improve sodium arsenite induced learning and memory deficits in rats assessed by water maze and Y‐maze. The results provide evidence that curcumin exercises its neuroprotective effect involving PI3K/Akt pathway which may affect NMDA receptors and downstream signalling through TrK&bgr; and BDNF in arsenic induced cognitive deficits in hippocampus.

Prenatal exposure to lambda-cyhalothrin impairs memory in developing rats: Role of NMDA receptor induced post-synaptic signalling in hippocampus

HIGHLIGHTSPrenatal exposure to LCT altered expression of NMDA receptors in developing rats.Alteration in postsynaptic signaling in hippocampus of developing rats also evident.Changes associated with impairment in memory of rats on PD22 and PD45.Changes found to persist in developing rats exposed at high dose (3 mg/kg bw). ABSTRACT Effect of prenatal exposure to lambda‐cyhalothrin (LCT) has been assessed on the integrity of NMDA receptors and associated post‐synaptic signalling in hippocampus of developing rats. Decrease in the binding of [3H]‐MK 801, known to label NMDA receptors was observed in hippocampus of rats prenatally exposed to LCT (1 and 3 mg/kg body weight) on PD22, compared to controls. Consistent with this, decrease in the mRNA and protein expression of NR1 and NR2B subunits of NMDA receptors was evident in rats prenatally exposed to LCT (1 and 3 mg/kg body weight) on PD22. There was no change in mRNA and protein expression of NR2A subunit of NMDA receptors. Prenatal exposure to LCT (1 and 3 mg/kg body weight) decreased the expression of positive regulators (PSD95, pERK1/2, CaMKII&agr; & pCREB) and increased the expression of negative regulators (Cdk5 & SynGAP) associated with NMDA receptor dependent synaptic plasticity in hippocampus and impaired learning and memory of rats on PD22. The neurobehavioral changes continued to persist in rats exposed to LCT at high dose (3 mg/kg body weight) while exhibited trend of recovery in those exposed at moderate dose (1 mg/kg body weight) on PD45, compared to controls. No change in any of the neurobehavioral endpoint was observed in developing rats prenatally exposed to LCT at low dose (0.5 mg/kg body weight) on PD22 and PD45. The results exhibit that alterations in NMDA receptors on prenatal exposure to LCT may affect postsynaptic signalling associated with impaired learning and memory in developing rats.

Influence of immobilization and forced swim stress on the neurotoxicity of lambda-cyhalothrin in rats: Effect on brain biogenic amines and BBB permeability.

HIGHLIGHTSInfluence of stress in the neurotoxicity of LCT investigated in rats.Marginal change in plasma corticosterone & BBB on pre‐exposure to stress or LCT alone.No significant change in brain neurotransmitters evident on exposure to stress or LCT.Intense neurochemical changes observed in rats co‐exposed to stress and LCT.Pre‐exposure to stress enhanced vulnerability of rats to neurotoxic response of LCT. ABSTRACT Experimental studies have been carried out on rats to understand the influence of immobilization stress (IMS), a psychological stressor and forced swim stress (FSS), a physical stressor in the neurotoxicity of lambda‐cyhalothrin (LCT), a new generation type II synthetic pyrethroid with extensive applications. No significant change in plasma corticosterone levels and blood brain barrier (BBB) permeability was observed in rats subjected to IMS (one session of 15 min/day), FSS (one session of 3 min/day) for 28 days or LCT treatment (3.0 mg/kg body weight, p.o. suspended in groundnut oil) for 3 days (26th, 27th and 28th day) as compared to controls. Marginal changes in the levels of biogenic amines and their metabolites (NE, EPN, DA, HVA, DOPAC, 5‐HT) in hypothalamus, frontal cortex, hippocampus, and corpus striatum were observed in rats subjected to IMS or FSS or LCT alone as compared to controls. It was interesting to note that pre‐exposure to IMS or FSS followed by LCT treatment for 3 days caused a marked increase in plasma corticosterone levels associated with disruption in the BBB permeability as compared to rats exposed to IMS or FSS or LCT alone. Pre‐exposure to IMS or FSS followed by LCT treatment for 3 days resulted to alter the levels of biogenic amines and their metabolites in hypothalamus, frontal cortex, hippocampus, and corpus striatum as compared to rats exposed to IMS or FSS or LCT alone. Although neurochemical changes were more intense in rats pre‐exposed to IMS as compared to those subjected to FSS on LCT treatment, the results indicate that both psychological and physical stress could be important influencing factors in the neurotoxicity of LCT.

Efficacy of Natural Compounds in Neurodegenerative Disorders

Neurodegenerative disorders represent clusters of serious diseases that results in progressive deterioration of normal structure and physiology of central nervous system. Pathophysiology of Alzheimers, Parkinsons or other neurodegenerative disorders involves multifaceted permutation of genetic and environmental factors. Combinations of lifestyle modification linked with environmental factor jointly or alone represent the largest share of cases of these disorders. Etiology of such neuronal degeneration involves manifestation of toxic reaction in the form of functional anomalies leading to dysfunction of the ubiquitin-proteasome system, activated inflammatory cascade, compromised neuronal survival pathway, mitochondrial dysfunction and finally neuronal apoptosis/necrosis and cell death. Furthermore, evidences from various studies exhibited role of oxidative stress and compromised anti-oxidant defense system as one of the prime factors associated with activation of various signal transduction pathways that would ultimately lead to the formation of amyloid beta or alpha synuclein in the brain. Keeping in view of complex etiology and pathophysiology along with a miniscule of available treatment options associated with these neurodegenerative disorders, the role of natural agents and herbal extracts as therapeutic alternatives alone or in combination with synthetic drugs could not be ruled out. In the same context the present chapter has been aimed to investigate the role of selected natural plants like Withania somnifera, Bacopa monnieri, Curcuma longa, Centella asiatica, Ocimum sanctum, Nardostachys jatamansi and Emblica officinalis in various neurodegenerative disorders and explore their targets to ameliorate neurotoxicity in various experimental models. The rationale for selection of these plants was based on their strong anti-inflammatory and anti-oxidant potential and large body of evidence that suggest their efficacy in preclinical as well as in clinical studies. Active constituents if these herbals might play an important role in preserving the integrity of various neurotransmitters and their receptor in the brain influencing its functions at the molecular level.

Involvement of PKA/DARPP-32/PP1α and β- arrestin/Akt/GSK-3β Signaling in Cadmium-Induced DA-D2 Receptor-Mediated Motor Dysfunctions: Protective Role of Quercetin

Given increasing risk of cadmium-induced neurotoxicity, the study was conducted to delineate the molecular mechanisms associated with cadmium-induced motor dysfunctions and identify targets that govern dopaminergic signaling in the brain involving in vivo, in vitro, and in silico approaches. Selective decrease in dopamine (DA)-D2 receptors on cadmium exposure was evident which affected the post-synaptic PKA/DARPP-32/PP1α and β-arrestin/Akt/GSK-3β signaling concurrently in rat corpus striatum and PC12 cells. Pharmacological inhibition of PKA and Akt in vitro demonstrates that both pathways are independently modulated by DA-D2 receptors and associated with cadmium-induced motor deficits. Ultrastructural changes in the corpus striatum demonstrated neuronal degeneration and loss of synapse on cadmium exposure. Further, molecular docking provided interesting evidence that decrease in DA-D2 receptors may be due to direct binding of cadmium at the competitive site of dopamine on DA-D2 receptors. Treatment with quercetin resulted in the alleviation of cadmium-induced behavioral and neurochemical alterations. This is the first report demonstrating that cadmium-induced motor deficits are associated with alteration in postsynaptic dopaminergic signaling due to a decrease in DA-D2 receptors in the corpus striatum. The results further demonstrate that quercetin has the potential to alleviate cadmium-induced dopaminergic dysfunctions.

Secretome of Differentiated PC12 Cells Restores the Monocrotophos-Induced Damages in Human Mesenchymal Stem Cells and SHSY-5Y Cells: Role of Autophagy and Mitochondrial Dynamics

A perturbed cellular homeostasis is a key factor associated with xenobiotic exposure resulting in various ailments. The local cellular microenvironment enriched with secretory components aids in cell–cell communication that restores this homeostasis. Deciphering the underlying mechanism behind this restorative potential of secretome could serve as a possible solution to many health hazards. We, therefore, explored the protective efficacy of the secretome of differentiated PC12 cells with emphasis on induction of autophagy and mitochondrial biogenesis. Monocrotophos (MCP), a widely used neurotoxic organophosphate, was used as the test compound at sublethal concentration. The conditioned medium (CM) of differentiated PC12 cells comprising of their secretome restored the cell viability, oxidative stress and apoptotic cell death in MCP-challenged human mesenchymal stem cells and SHSY-5Y, a human neuroblastoma cell line. Delving further to identify the underlying mechanism of this restorative effect we observed a marked increase in the expression of autophagy markers LC3, Beclin-1, Atg5 and Atg7. Exposure to autophagy inhibitor, 3-methyladenine, led to a reduced expression of these markers with a concomitant increase in the expression of pro-apoptotic caspase-3. Besides that, the increased mitochondrial fission in MCP-exposed cells was balanced with increased fusion in the presence of CM facilitated by AMPK/SIRT1/PGC-1α signaling cascade. Mitochondrial dysfunctions are strongly associated with autophagy activation and as per our findings, cellular secretome too induces autophagy. Therefore, connecting these three potential apices can be a major breakthrough in repair and rescue of xenobiotic-damaged tissues and cells.