High Fructose Negatively Impacts Proliferation of NSC-34 Motor Neuron Cell Line.

Objectives The main aim of this study is to identify the deleterious effects of indiscriminately consumed high fructose on motor neurons that are critically affected in many neurological conditions causing movement disorders including paralysis. Materials and Methods Neuroblastoma x mouse spinal cord motor neuron cell line (NSC-34) motor neuron cell lines were treated with high fructose and oxygen supplementation (18.8%) and assayed for cell proliferation/death, reactive oxygen species (ROS) generation, and oxidative stress response induction Statistical Analysis Mean and standard deviation, significance with and without high fructose (F)-5%, were estimated by t -tests using GraphPad Prism ver. 8.2.1 Results F-5% along with O 2 (18.8%) annihilates the cells (∼85%) by day10 and inhibits cell division as observed by the presence of multinucleated cells. Unexpectedly, 1 to 2% of cells that survived, differentiated and displayed progressive neurite extension. Though not healthy, they were viable up to 80 days. F-5% increased ROS levels (∼34%) not accompanied by concomitant enhanced expression of oxidative stress response regulator, the transcription factor, nrf-2 , or downstream effector, sod-1. Conclusion High fructose is extremely harmful to NSC-34 motor neuron cell line.

Detrimental effects of fructose on mitochondria in mouse motor neurons and on C. elegans healthspan.

BACKGROUND: Fructose-common sweetener, consumed in large quantities, is now known to be associated with various metabolic diseases. Recent reports suggest fructose's involvement in neurodegeneration, neurotoxicity, and neuroinflammation. But, its impact at cellular and subcellular level and on energy metabolism, especially, mitochondrial bioenergetics, in neurons is not known. OBJECTIVES: To study the adverse effects of high fructose in general, and on the mitochondria in a spinal cord motor neuron cell line, NSC-34, in vitro, and Caenorhabditis elegans in vivo. METHODS: NSC-34 was treated with 0.5%-5% of fructose for different time periods. Fructose's effect on cell viability (MTT assay), metabolic activity (XF24 Seahorse assays) and C. elegans, chronically fed with 5% fructose and alteration in healthspan/mitochondria was monitored. RESULTS: In NSC-34: Fructose at 4-5% elicits 60% cell death. Unlike 1%, 5% fructose (F5%) decreased mitochondrial membrane potential by 29%. Shockingly, 6hours F5% treatment almost abolished mitochondrial respiration - basal-respiration (∨123%), maximal-respiration (∨ 95%) and spare-respiratory-capacity (∨ 83%) and ATP production (∨98%) as revealed by XF 24- Seahorse assays. But non - mitochondrial respiration was spared. F5% treatment for 48hrs resulted in the total shutdown of respiratory machinery including glycolysis. Chronic feeding of wildtype C.elegans to F5% throughout, shortened lifespan by ~3 days (∨ 17%), progressively reduced movement (day-2 -∨10.25%, day-5 -∨25% and day-10 -∨56%) and food intake with age (day-5-∨9% and day-10 -∨48%) and instigated mitochondrial swelling and disarray in their arrangement in adult worms body-wall muscle cells. CONCLUSION: Chronic exposure to high fructose negatively impacts cell viability, mitochondrial function, basal glycolysis, and healthspan.

DMSO Delays Alzheimer Disease Causing Aß-induced Paralysis in C. elegans Through Modulation of Glutamate/Acetylcholine Neurotransmission.

BACKGROUND: Alzheimer's disease (AD), a prevalent neurodegenerative disease with progressive dementia and neurotransmission (NT)-dysfunction-related complications in older adults, is known to be caused by abnormal Amyloid-ß (Aß) peptide and associated amyloid plaques in the brain. Drugs to cure AD are not in sight. Two major excitatory neurotransmitters, glutamate (Glu) and acetylcholine (ACh), and their signaling systems are implicated in AD. OBJECTIVE: To determine the effect of various NT-altering compounds including fenobam, quisqualic acid, and dimethyl sulfoxide (DMSO) in the protection against Aß toxicity. Further, to identify the potential mechanism through which the protection happens. METHODS: The well-known C. elegans AD model, CL4176, in which human Aß expression is turned on upon a temperature shift to 25 °C that leads to paralysis, was screened for protection/delay in paralysis because of Αß toxicity. While screening the compounds, dimethyl sulfoxide (DMSO), a universal solvent used to solubilize compounds, was identified to provide protection. Aldicarb and levamisole assays were performed to identify the contribution of ACh neurotransmission in Αß toxicity protection by DMSO. RESULTS: One percent and two percent DMSO delayed paralysis by 48% and 90%, respectively. DMSO was dominant over one of the Glu-NT pathway-related compounds, Fenobam-Group I mGluR antagonist. But DMSO provided only 30% to 50% protection against Quisqualic acid, the Glu-agonist. DMSO (2%) delayed ACh-NT, both presynaptic acetylcholine esterase inhibitor (AchEi)-aldicarb and postsynaptic-iAChR-agonst-levamisole induced paralysis, by ∼70% in CL4176. DMSO seems to be altering Ca2+ ion permeability essential for NT as EthyleneDiamine Tetra-Acetic acid (EDTA) and DMSO provided similar aldicarb resistance either combined or alone in wildtype worms. But postsynaptic Ca2+ depletion by EDTA could reverse DMSO-induced levamisole hypersensitivity. Surprisingly, the absence of FOrkhead boXO (FOXO) transcription factor homolog, daf-16 (loss-of-function mutant), a critical transcription factor in the reduced IIS-mediated longevity in C. elegans, abolished DMSO-mediated AldR. CONCLUSION: DMSO and Fenobam protect against Aß toxicity through modulation of NT.

Absence of metabotropic glutamate receptor homolog(s) accelerates acetylcholine neurotransmission in Caenorhabditis elegans.

Glutamate (Glu) and Acetylcholine (ACh), are excitatory neurotransmitters, acting through ionotropic (iR) and metabotropic receptors (mR). Importantly, both neurotransmitters and their signalling are impaired in the prevalent neurodegenerative disease-Alzheimer disease (AD). Glu and its signalling cascade's influence on ACh-neurotransmission (NT) are sparsely understood. The mGluRs coupled to G-protein signalling acting through PI3K cascade (GrpI) or inhibition of adenylate cyclase-cAMP cascade (GrpII and GrpIII) brings about long-lasting structural/functional changes. These complexities are challenging to decipher. Here, we report that human/mouse mGluRs when compared with their Caenorhabditis elegans homologs, MGL-1-3 showed overall of homology of ∼31-39 %. Phylogeneitc analysis revealed homology of MGL-2 to GrpI, MGL-3 with Grp1 &II and GRM6 of GrpIII and MGL-1, a low homology that falls between GrpI & GrpII. Then, alteration of ACh-NT in C. elegans loss-of-function mutants of mgl-1, mgl-2, mgl-3, PI3K (age-1) and iGluR (NMDA)(nmr-1) was estimated by well-established acute aldicarb (Ald), that increases ACh at synapse, and levamisole (Lev) (postsynaptic activation of levamisole sensitive iAChR) induced time-dependent paralysis assays. Surprisingly, all of them were hypersensitive to Ald and Lev compared to wildtype (in percentage), namely, mgl-1 -17, 54; mgl-2 - 7.2, 24; mgl-3 -52, 64; age-1 - 27, 32; nmr-1- 24, 48; respectively. Of the three, mgl-3 contributes to maximal overall acceleration of ACh-NT. Adenylate cyclase, acy-1 gain-of-function mutant showed less hypersensitivity, Ald - 7% and Lev- 25 %. Together, Glu receptors and signalling cascades are altering ACh-NT permanently, thus establishing the interplay between them thereby provide potential drug targets to be considered for AD.

Organic solvents can influence acetylcholine neurotransmission in Caenorhabditis elegans.

BACKGROUND: Identification of novel drugs by bio-prospecting natural products like various parts of the plants, or other extracts and drug discovery requires differential fractionation with various organic solvents followed by their concentration through evaporation under nitrogen gas, which is a standard practice. PURPOSE: Determination of contribution of vehicle control of organic solvents (chloroform, ethanol, ethyl acetate and n-hexane) processed in the similar manner in the modulation of acetylcholine(ACh) neurotransmission in Caenorhabditis elegans, Aldicarb induced paralysis assay. METHODS: The organic solvents concentrated as described in background was used to identify their contribution in ACh modulation through ACh esterase inhibitor, Aldicarb, treatment of C. elegans, which leads to time dependent paralysis of the worms. RESULTS: The vehicle, organic solvents, control itself bestows modulation of acetylcholine release as Aldicarb resistance in C. elegans. CONCLUSION: Given the exorbitant cost and time taken for drug discovery, identification of efficacy of bioactive molecules fractionated through organic solvents and concentrated under nitrogen gas should have appropriate vehicle control as described above to avoid the rate of false positives. This is universally applicable whether the drug is chemically synthesized or purified from natural products.

Reserpine requires the D2-type receptor, dop-3, and the exoribonuclease, eri-1, to extend the lifespan in C. elegans.

Lifespan extension is an all systems encompassing event. Involvement of reduced insulin/IGF1 signalling is well worked out, first in the model organism Caenorhbaditis elegans followed by other systems including humans. But the role of neuronal component in lifespan extension is not well understood due to the refractory nature of neurons to small RNA interference (sRNAi) in C. elegans. Earlier, we have demonstrated that an antihypertensive drug, reserpine, extends lifespan through modulation of neurotransmitter release, especially, acetylcholine, in C. elegans. Intriguingly, the reserpine mediated lifespan extension (RMLE) does not happen through the known longevity pathways. Here, we report that the D2-type dopamine receptor (DOP-3), which acts through the inhibitory Gprotein coupled (G alpha i) pathway mediated signalling is partly required for RMLE. In the dop-3 loss of function mutant RMLE is shortened. DOP-3 acts through Gαo (goa-1). One of the downstream targets of G protein signalling is the transcription factor, jun-1. MRP-1, an ATP binding cassette transporter, belonging to the multidrug resistance protein family is one of the genes turned on by JUN-1. RMLE is shortened in dop-3-->goa-1-->jun1-->mrp-1 loss of function mutants, elucidating the contribution of dop-3 signalling. The dop-3 receptor system is known to inhibit acetylcholine release. This suggests dopamine receptor, dop-3 could be contributing to the modulation of acetylcholine release by reserpine. ERI-1 is a 3'-5' exoribonuclease, one of the negative regulators of sRNAi, whose loss of function makes neurons amenable to siRNA. In the absence of eri-1, RMLE is shortened. In the dop-3 loss-of-function background, lack of eri-1 completely abolishes RMLE. This suggests that dop-3 and eri-1 act in independent parallel pathways for RMLE and these two pathways are essential and sufficient for the longevity enhancement by reserpine in C. elegans.

A Novel Way of Amelioration of Amyloid Beta Induced Toxicity in Caenorhabditis elegans.

BACKGROUND: With an incidence of 1 in 85 persons above the age of 60 years succumbing to the disease, Alzheimer's disease (AD), has been predicted to create havoc globally. In spite of enormous efforts and exhaustive research, no cure is in sight. Hence, it is critical to unravel the mechanism of AD development/protection and identification of a cure soon. PURPOSE: This study is aimed at investigating the mechanism of reserpine action, which alleviates the toxicity of amyloid beta (Aß) (AD-causing peptide) in Caenorhabditis elegans [1, 2]. METHODS: Determination of alleviation of Aß toxicity with reserpine manifested as reduction in progressive paralysis, in the background of GFP reporter driven by the promoter of the FMRFamide neuropeptide, FLP-11 (AD; Pflp-11::GFP) and acetylcholine contribution through aldicarb (which inhibits acetylcholine esterase) treatment. RESULTS: The most significant protection against Aß toxicity was obtained in the background of Pflp-11::GFP. This protection had 2 components. The promoter of FLP-11 with the reporter GFP, Pflp-11::GFP, per se gave significant protection. Further reserpine treatment provided additional alleviation. Together they could almost eliminate Aß toxicity. These 2 components of Aß toxicity alleviation are dependent on acetylcholine levels, as an increase in acetylcholine by aldicarb treatment reduces the protective effect. CONCLUSION: A unique way to alleviate Aß toxicity is reserpine treatment in combination with Pflp-11::GFP. Reserpine should be evaluated as a potential drug in a pilot study in AD patients. Furthermore, identification of the mechanism of Pflp-11::GFP-mediated reduction in Aß toxicity is a potential pathway to develop therapeutics for AD.

Impact on cognitive functions following gamma knife radiosurgery for cerebral arteriovenous malformations.

BACKGROUND: Radiosurgery is an alternative to surgical resection of arteriovenous malformation (AVM). Very few studies have addressed the concern of radiation injury to the brain and its attendant adverse effects on cognitive function. MATERIALS AND METHODS: This prospective study included all patients who underwent gamma knife radiosurgery (GKRS) at our institute for cerebral AVM between 2006 and December 2008 (n = 34). All patients underwent neuropsychological evaluation before the procedure. Neuropsychological evaluation was repeated in eighteen patients 2 years following GKRS. Clinical outcome, AVM obliteration, and factors influencing outcome were analyzed in these eighteen patients. RESULTS: Before GKRS, more than 50% had significant impairment of neuropsychological functions compared to normal population norms. 66.6% achieved the excellent radiosurgical outcome. At 2 years follow-up, patients showed varied improvement in neuropsychological function in various categories. Pretherapeutic median value for percentage perseverative responses was 26.5 and at follow-up, it reduced to 18.2 (P = 0.039). Set shifting improved in 11 patients (61.1%), remained same in 5 patients (27.7%), and deteriorated in two patients (11.1%). Patients with a higher Spetzler-Martin grade AVM demonstrated a significantly more favorable shift in follow-up test values for set shifting function (P = 0.021). Patients with postradiation imaging changes had lesser tendency to improve in neuropsychological performance at follow-up. CONCLUSIONS: GKRS has no clinically harmful effect on cognitive and neuropsychological functioning in patients with brain AVM. On the contrary, there is an improvement in majority of patients at 2 years following radiosurgery when nidus is obliterated.

Clinical, electrophysiological, imaging, pathological and therapeutic observations among 18 patients with Rasmussen's encephalitis.

We studied the clinical, electrophysiological, imaging and pathological features of 18 patients with Rasmussen's encephalitis (RE). This descriptive study included 18 patients (six males, 12 females) with RE who were evaluated for demographic and phenotypic details, electroencephalogram (EEG) results, MRI results, pathological features, virological markers and outcome. Radiological staging as per Bien et al. and pathological staging in accordance with Robitaille et al. were performed. Simple partial seizures were the most common initial manifestation. During the disease course, epilepsia partialis continua (EPC) developed in 15/18 (83.3%) and hemiparesis in 17/18 (94.4%) patients. EEG revealed hemispheric slowing (100%), interictal epileptiform discharges (100%) and ictal pattern (44.4%). Brain MRI revealed unihemispheric focal cortical atrophy (100%), white matter changes (88.2%), basal ganglia-ipsilateral caudate and putamen involvement (50.0%) and progression of atrophy on serial MRI (100%). Unusual presentations in this series included late onset (n=1), and isolated lingual EPC (n=1). Diagnostic biopsies in two patients revealed Robitaille stage 3 disease. The six hemispherotomy specimens showed stage 2 disease in one, stage 3 in three and stage 4 in two cases. Heterogeneity in disease stage in the different neuroanatomical regions and within the same cortical segment reflected progression of immune-mediated damage. Immunomodulation provided only temporary benefit. Patients who underwent functional hemispherotomy had reduction in seizure frequency and improved quality of life. The clinical, EEG and MRI findings are in accordance with the established literature. MRI staging was concordant with Robitaille pathological staging. Immunomodulation did result in transient reduction in seizure frequency while surgery in six produced reasonable benefit.

Secreted trophic factors of Human umbilical cord stromal cells induce differentiation and neurite extension through PI3K and independent of cAMP pathway.

BACKGROUND: Trophic factors (TFs) play important role during development and adult tissue maintenance. In neurodegenerative diseases (ND) TF supplementation provides protection. Stromal cells (HUMS) derived from the human umbilical cord matrix provide neuroprotection in the ND models of mice. PURPOSE: Though TF mediated protection is known, the exact mechanism of protection is not clear. So, here the essential TFs (secreted by HUMS cells) and the pathway of induction of neurite extension, differentiation and networking is addressed. METHODS: The HUMS cells from the human umbilical cord matrix were derived and the mouse spinal cord motor neuron cell line, NSC-34 was extensively used. Flow cytometry, immunohistochemistry, RT- PCR, western blot, ELISA and antibody/inhibitor treatment were carried out to figure out the TF pathway. RESULTS: The HUMS cells secrete six neurotrophic factors (sTFs), namely, NT-3, NGF, BDNF, VEGF, IGF-1 and GDNF (TFs). These TFs are sufficient to induce differentiation, neurite extension and neural networking in a motor neuron cell line, NSC34. All the 5 TFs need to be neutralized simultaneously with their antibodies to abrogate neurite extension. These motor neurons express the concomitant receptors, which are either receptor tyrosine kinase (TrK) coupled or to the receptor followed by the TrKs, for the above trophic factors (except for BDNF). The tyrosine kinase inhibitor, K252a, drastically reduces neurite extension. In NSC34, the TFs are coupled to the PI3K-Akt-pathway and the RAS-MAP kinase signaling through phosphorylation of ERK1 and ERK2. PI3K inhibitor, Ly 294002, abolishes neural differentiation and neurite extension. Thus, differentiation, neurite extension and networking could be achieved through the PI3K pathway. Intriguingly, the cAMP second messenger system coupling was not required. H89, PKA-inhibitor caused extensive cell death. But, had no effect in the presence of HUMS-secreted-TFs(HSTFs) suggesting a pathway switch for cell survival itself. CONCLUSION: HUMS cells and their secreted factors could be of great use in regenerative medicine (RM). The activators of PI3K pathway, the major route of these HUMS-TFs action could be explored in RM and in the neurobiology of neural differentiation and extension.

Cognitive and sexual functions in patients with traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) has an immense psychosocial impact on an individual as well as on the close relatives. Sexuality is one among the functions which are usually found compromised post injury. The aim of present study was to examine cognitive and sexual functions post TBI. The objective of the study was to explore these domains and their relationship with each other. TOOLS: The tools used were sociodemographics record sheet, Edinburg handedness inventory, brief sexual function inventory, depression anxiety stress scales-21 and NIMHANS head injury battery. The sample consisted of 30 patients with mild-to-moderate TBI. All the subjects were tested individually in their regional language. RESULTS: On cognitive domain, patients performed inadequately on all the tests; however, the percentage was higher in mental speed (43.3%), sustained attention (26.7%), verbal working memory (30%), response inhibition (36.7%), verbal memory (immediate and delayed) (43%) and visual (immediate, 23.3% and delayed, 26.7%). On the domain of sexual functions, all the four domains (sexual drive, erection, ejaculation and problem assessment) were affected however overall satisfaction (93.3%) was adequate. Among the four domains higher percentage of involvement was noted on problem assessment (70%), ejaculation (56.7%), and erection (46.7%). Significant correlation was found between mental speed, verbal working memory, planning, and visual memory with sexual drive, erection, ejaculation and overall satisfaction domains of sexual functioning. Negative correlation was found between motor speed and sustained attention with sexual drive, erection and ejaculation. CONCLUSION: Both cognitive and sexual functioning were found effected post TBI. However less emphasis is given to sexual functioning by the professionals. Educational intervention is needed to sensitize professional about this area and to include this area for better management.

Clinico-pathological factors influencing surgical outcome in drug resistant epilepsy secondary to mesial temporal sclerosis.

BACKGROUND: Mesial temporal sclerosis (MTS) is the most common cause of drug resistant epilepsy amenable for surgical treatment and seizure control. METHODS: This study analyzed the outcome of patients with MTS following anterior temporal lobectomy and amygdalohippocampectomy (ATL-AH) over 10 years and correlated the electrophysiological and radiological factors with the post operative seizure outcome. RESULTS: Eighty seven patients were included in the study. Sixty seven (77.2%) patients had an Engel Class 1 outcome, 9 (11.4%) had Class 2 outcome. Engel's class 1 outcome was achieved in 89.9% at 1 year, while it reduced slightly to 81.9% at 2 years and 76.2% at 5 year follow up. Seventy seven (88.5%) patients had evidence of hippocampal sclerosis on histopathology. Dual pathology was observed in 19 of 77 specimens with hippocampal sclerosis, but did not influence the outcome. Factors associated with an unfavorable outcome included male gender (p=0.04), and a higher frequency of pre-operative seizures (p=0.005), whereas the presence of febrile seizures (p=0.048) and loss of hippocampal neurons in CA4 region on histopathology (p=0.040) were associated with favorable outcome. The effect of CA4 loss on outcome is probably influenced by neuronal loss in other subfields as well since isolated CA4 loss was rare. Abnormal post operative EEG at the end of 1 week was found to be a significant factor predicting unfavorable outcome (p=0.005). On multivariate analysis, the pre-operative seizure frequency was the only significant factor affecting outcome. CONCLUSIONS: The present study observed excellent seizure free outcome in a carefully selected cohort of patients with MTS with refractory epilepsy. The presence of dual pathology did not influence the outcome.

Evolution of MRI changes in Rasmussen's encephalitis.

PURPOSE: We studied the MRI findings in 16 patients with Rasmussen's encephalitis (RE), further analysed serial MRI changes in 11 of them and correlated it with clinical features. METHODOLOGY: The diagnosis of RE was based on the European consensus statement (Brain, 128, 2005, 454). Details related to demographical, clinical, MRI observations were analysed. RESULTS: Forty MRIs of brain of 16 patients were reviewed. Eleven patients had undergone serial brain MRIs ranging from two to five occasions. All the patients had unihemispheric focal cortical atrophy, predominantly in the perisylvian region (n = 13). Other features were white matter signal changes (n = 14), and ipsilateral caudate (n = 6) and putamen (n = 4) atrophy. Signal alterations in putamen and caudate were noted in four each. In all the 11 patients with serial MRI, there was progression of cerebral atrophy and a trend towards increase in MRI staging. The MRI signal changes remained same in five patients, resolved in three patients, differential change in two patients and increased in one patient. Diffusion-weighted imaging showed facilitated diffusion (n = 5), and MR spectroscopy showed reduced N-acetyl-aspartate and elevated lactate (n = 2). CONCLUSIONS: Pattern recognition of MRI findings and the changes in serial MRI might serve as a surrogate marker of disease viz. unihemispheric progressive focal cortical atrophy and signal changes predominantly in the perisylvian distribution and caudate followed by putamen involvement. This might assist in understanding and monitoring of the disease progression.

Outcome of lesionectomy in medically refractory epilepsy due to non-mesial temporal sclerosis (non-MTS) lesions.

OBJECTIVES: To analyze the seizure outcome of lesionectomy for refractory epilepsy secondary to non-mesial temporal sclerosis (non-MTS) lesions. METHODS: Sixty-eight patients with non-MTS lesions (M:F=42:26; age at onset: 11.7±9.6 years; age at surgery: 21.1±9.4 years), who underwent lesionectomy for refractory epilepsy were analyzed. The age at onset, frequency/type of seizure, MRI findings, video-EEG, histopathology and Engel's grading at 1 year/last follow up were recorded. RESULTS: The duration of epilepsy at surgery was 9.9±6.9 years. The location of lesions were: temporal: 41 (60.3%); frontal: 21 (30.9%); parietal: 6 (8.8%). The type of lesionectomies performed were temporal 41 (60.3%), extra-temporal: 25 (36.8%), temporo-frontal and temporo-parietal: 1 (1.5%) patient each. The histopathological diagnosis were neoplastic: 32 (47.1%), cortical dysplasia: 19 (27.9%), other focal lesions: 17 (25%). At mean follow up of 2.9±2.1 years (median: 2.6 years), outcome was - Engel's class I: 43 (63.2%), IIa: 14 (20.6%), III: 7 (10.3%), IV: 4 (5.9%). Good seizure control (Engel's class I/IIa) was achieved in 57 (83.8%) patients. The good prognostic markers included temporal seizures, extended lesionectomy and AEDs after surgery while poor prognostic marker was gliotic lesion on histopathology. CONCLUSION: Following lesionectomy due to non-MTS lesions, seizure freedom (Engel I) was noted in about 63.2% of patients, which is comparable to other series and reiterates the effectiveness of lesionectomy for seizure control.

Variation at diabetes- and obesity-associated Loci may mirror neutral patterns of human population diversity and diabetes prevalence in India.

South Asian populations harbor a high degree of genetic diversity, due in part to demographic history. Two studies on genome-wide variation in Indian populations have shown that most Indian populations show varying degrees of admixture between ancestral north Indian and ancestral south Indian components. As a result of this structure, genetic variation in India appears to follow a geographic cline. Similarly, Indian populations seem to show detectable differences in diabetes and obesity prevalence between different geographic regions of the country. We tested the hypothesis that genetic variation at diabetes- and obesity-associated loci may be potentially related to different genetic ancestries. We genotyped 2977 individuals from 61 populations across India for 18 SNPs in genes implicated in T2D and obesity. We examined patterns of variation in allele frequency across different geographical gradients and considered state of origin and language affiliation. Our results show that most of the 18 SNPs show no significant correlation with latitude, the geographic cline reported in previous studies, or by language family. Exceptions include KCNQ1 with latitude and THADA and JAK1 with language, which suggests that genetic variation at previously ascertained diabetes-associated loci may only partly mirror geographic patterns of genome-wide diversity in Indian populations.

Withania somnifera root extract extends lifespan of Caenorhabditis elegans.

BACKGROUND: In the ancient Indian herbal medicine system several ayurvedic preparations are claimed to have longevity enhancing effects. But, so far, no clear scientific evidence has been provided. One among them, is the roots of the plant, commonly known as Ashwagandha (Withania somnifera Dunal- WSD), which is supposed to have myriad of beneficial effects including long life. PURPOSE: Here, we evaluated both the root extract (RE) and its purified ingredients (PI-RE) with a similar composition as in RE obtained from the roots of WSD for lifespan extension in the well established model system, C. elegans. PI-RE could extend the lifespan of C. elegans. METHODS: We used wild type C. elegans (N2) or RB918: acr-16 (ok789); andNL2099: rrf-3 (pk1426) mutant worms and analysed their lifespan assay in Ashwagandha extract spreaded on plates containing Bacterial Lawns. RESULTS: Strangely, while there was no effect on the wild type worms, the mutant for the human nicotinic acetylcholine receptor, nAchR, α7 equivalent, acr-16, showed around ~20% lifespan extension when treated with PI-RE. CONCLUSION: Thus, we are able to show that one of the age old healthy longlife supplements, Ashwagandha does extend lifespan of C. elegans.

Reserpine modulates neurotransmitter release to extend lifespan and alleviate age-dependent Aß proteotoxicity in Caenorhabditis elegans.

Aging is a debilitating process often associated with chronic diseases such as diabetes, cardiovascular and neurodegenerative diseases like Alzheimer's disease (AD). AD occurs at a very high incidence posing a huge burden to the society. Model organisms such as C. elegans become essential to understand aging or lifespan extension - the etiology, molecular mechanism and identification of new drugs against age associated diseases. The AD model, manifesting Aß proteotoxicity, in C. elegans is well established and has provided valuable insights. Earlier, we have reported that Reserpine, an FDA-approved antihypertensive drug, increases C. elegans lifespan with a high quality of life and ameliorates Aß toxicity in C. elegans. But reserpine does not seem to act through the known lifespan extension pathways or inhibition of its known target, vesicular monoamine transporter, VMAT. Reserpine's mode of action and the pathways it activates are not known. Here, we have evaluated the presynaptic neurotransmitter(s) release pathway and identified acetylcholine (ACh) as the crucial player for reserpine's action. The corroborating evidences are: i) lack of lifespan extension in the ACh loss of function (hypomorphic) - synthesis (cha-1) and transport (unc-17) mutants; ii) mitigation of chronic aldicarb effect; iii) lifespan extension in dopamine (cat-2) and dopamine and serotonin (bas-1) biosynthetic mutants; iv) no rescue from exogenous serotonin induced paralysis in the AD model worms; upon reserpine treatment. Thus, modulation of acetylcholine is essential for reserpine's action.

ALS associated mutant SOD1 impairs the motor neurons and astrocytes and wild type astrocyte secreted-factors reverse the impaired motor neurons.

BACKGROUND: Amyotrophic Lateral Sclerosis, in which motor neurons degenerate, leading to paralysis, not only the affected motor neurons, but the surrounding non-neuronal cells also contribute significantly to the disease. However, the disease mechanism is not known. PURPOSE: In this study we have addressed the disease mechanism by expressing the ALS associated mutant SOD1(G37R) in the motor neurons (mMN) and astrocytes (mA) cell lines. METHODS: A series of cell culture assays, immunostaining, RT-PCR and Western blot analysis were performed. RESULTS: We noticed impairments in both these cell types. The mMN motor neurons were insensitive to forskolin, a known activator of adenylate cyclase, which leads to motor neuron death. In addition, less number of mMN were positive for phosphorylated neurofilament-H (pNFH) unlike the normal motor neurons. Similarly, the mutant SOD1 expressing astrocytes (mA) had two impairments: The inability to activate the oxidative stress protection and the absence of secretory factor(s). Normal astrocytes and their secreted factors could restore the pNFH in the mMN but not the mA. In addition, we show that pNFH restoration is a specific function since the insensitivity of mMN to forskolin could be rescued by neither normal astrocytes nor their secreted factors. CONCLUSION: Thus we demonstrate some of the abnormalities caused by the ALS associated mutant SOD1(G37R) and a potential way, to reverse an abnormality through cell replacement.

Cerebrospinal fluid from sporadic amyotrophic lateral sclerosis patients induces degeneration of a cultured motor neuron cell line.

We investigated the effect of Cerebrospinal Fluid (CSF) from sporadic Amyotrophic Lateral Sclerosis patients (SALS-CSF) on motor neuron-like cells to delineate the pathomechanism of SALS. Exposure of NSC-34 cells to SALS-CSF caused lower viability, reduction in differentiation and enhanced lactate dehydrogenase activity. Additionally, reduced choline acetyl transferase expression alongside intracellular aggregation of phosphorylated neurofilaments was prominently seen. The aggregates were immunopositive for ubiquitin. These findings are comparable to the pathological changes seen in the postmortem tissue of ALS patients. Unlimited supply of NSC-34 cells and their vulnerability to SALS-CSF render them to be a good bioassay system to identify new therapeutic agents conferring protection to motor neurons.

Reserpine ameliorates Abeta toxicity in the Alzheimer's disease model in Caenorhabditis elegans.

Earlier we have reported that reserpine, an antihypertensive drug, known to downregulate biogenic amines through inhibition of the vesicular monoamine transporter (VMAT), increases longevity of Caenorhabditis elegans with a high quality of life, namely, enhanced and prolonged mobility (Srivastava et al., 2008). As neurodegenerative diseases are of adult onset, we addressed the protective ability of reserpine against neurodegenerative diseases, especially Alzheimer's disease (AD). In the well established AD model in C. elegans, Amyloid beta (Abeta) is expressed in the muscles and Abeta toxicity is manifested as paralysis (Link, 1995). In this model, reserpine significantly delayed paralysis and increased the longevity. In addition, reserpine provided thermotolerance, but interestingly the Abeta transcript and expression levels remains grossly unchanged.