Brain Glutamate Dehydrogenase Changes In Streptozotocin Diabetic Rats As A Function Of Age

Kinetic parameters of brain glutamate dehydrogenase (GDH) were compared in the brain stem, cerebellum and cerebral cortex of three weeks and one year old streptozotocin (STZ) induced four day diabetic rats with respective controls. A single intrafemoral dose of STZ (60mg/Kg body weight) was administered to induce diabetes in both age groups. After four days the blood glucose levels showed a significant increase in the diabetic animals of both age groups compared with the respective controls. The increase in blood glucose was significant in one year old compared to the three weeks old diabetic rats. The Vmm of the enzyme was decreased in all the brain regions studied, of the three weeks old diabetic rats without any significant change in the Km. In the adult the Vmax of GDH was increased in cerebellum and brain stem but was unchanged in the cerebral cortex. The K. was unchanged in cerebellum and cerebral cortex but was increased in the brain stem. These results suggest there may be an important regulatory role of the glutamate pathway in brain neural network disturbances and neuronal degeneration in diabetes as a function of age.

Glutamate Receptor Gene Expression: IP3, cAMP, cGMP Functional Regulation in Hypoglycaemic and Diabetic Rats

Department of Biotechnology, Cochin University of Science and Technology

Serotonin- 5-HT2A and Glutamate Receptors Gene Expression in Streptozotocin induced diabetic rats:Effects of pyridoxine and Aegle marmelose leaf extract

Department of Biotechnology, Cochin University of Science and Technology

Bone marrow cells differentiation to neurons in the rat brain using Serotonin and GABA:Their role on glutamate receptors, IP3, cAMP and cGMP functional regulation in unilateral Parkinsonism induced by 6-hydroxydopamine

Parkinson's disease is a chronic progressive neurodegenerative movement disorder characterized by a profound and selective loss of nigrostriatal dopaminergic neurons. Our findings demonstrated that glutamatergic system is impaired during PD. The evaluations of these damages have important implications in understanding the molecular mechanism underlying motor, cognitive and memory deficits in PD. Our results showed a significant increase of glutamate content in the brain regions of 6- OHDA infused rat compared to control. This increased glutamate content caused an increase in glutamatergic and NMDA receptors function. Glutamate receptor subtypes- NMDAR1, NMDA2B and mGluR5 have differential regulatory role in different brain regions during PD. The second messenger studies confirmed that the changes in the receptor levels alter the IP3, cAMP and cGMP content. The alteration in the second messengers level increased the expression of pro-apoptotic factors - Bax and TNF-α, intercellular protein - α-synuclein and reduced the expression of transcription factor - CREB. These neurofunctional variations are the key contributors to motor and cognitive abnormalities associated with PD. Nestin and GFAP expression study confirmed that 5-HT and GABA induced the differentiation and proliferation of the BMC to neurons and glial cells in the SNpc of rats. We also observed that activated astrocytes are playing a crucial role in the proliferation of transplanted BMC which makes them significant for stem cell-based therapy. Our molecular and behavioural results showed that 5-HT and GABA along with BMC potentiates a restorative effect by reversing the alterations in glutamate receptor binding, gene expression and behaviour abnormality that occur during PD. The therapeutic significance in Parkinson’s disease is of prominence.

Muscarinic MI and Glutamate Receptor Gene Expression and their Functional Role in Pilocarpine Induced Temporal Lobe Epilepsy in rats: Regulation by Bacoside A and Bacopa monnieri Extracts

The present work is to understand the alterations of total muscarinic. muscarinic MI and glutamate receptors in the brain regions of pilocarpine induced epileptic rats. The work focuses on the evaluation of the anti epileptic activity of extracts of Bacopa monnieri, Bacoside A and Carbamazepine in vivo. The molecular changes in the muscarinic M I receptors in the pre- and post-treated epileptic model with Bacopa monnieri, Bacoside A and Carbamazepine were also studied. These studies will help us to elucidate the functional role of muscarinic and glutamate receptors in epilepsy.

Dopamine D1 and D2 Receptor Functional Regulation:Glutamate Receptor Gene Expression in the Brain of Hypoglycaemic and Hyperglycaemic Rats

In the present study a detailed investigation on the alterations of dopamine and its receptors in the brain regions of streptozotocin induced diabetic and insulin induced hypoglycaemic rats were carried out. Glutamate receptor, NMDARI gene expression in the hypoglycaemic and hyperglycaemic brain was also studied. EEG recording in hypoglycaemic and hyperglycaemic will be carried out to measure brain activity. in vitro studies on glucose uptake and insulin secretion, with and without specific antagonists were carried out to confirm the specific receptor subtypes - DA D1, DA D2 and NMDA involved in the functional regulation during hyperglycaemic and hypoglycaemic brain damage. The molecular studies on the brain damage through dopaminergic and glutamergic receptors will elucidate the therapeutic role in the corrective measures of the damage to the brain during hypoglycaemia and hyperglycaemia. This has importance in the management of diabetes and antidiabetic treatment for better intellectual functioning of the individual.

Neurotransmitter Functional Role in Neurodegenerative Diseases: human Health

Neuroscience is the study of'tbe ne rvous system , including the i - ; . in, spinal cord and peripheral nerves . Neurons are the basic cells of the brain and nervous system which exerts its functional role through various neurotransmitters and receptor systems . The activity of a nen ren depends on the balance between the number of excitatory and inhibito r y processes affecting it, both processes occurring individually and sin ,tlte-' ,ieously. The functional bal,ince of different neurotransmitters such as Acct >>lcholine (Ach), Dopamine (DA), Serotonin (5-1-17), Nor epinepbri,te (N.1 j, Epinephrine (LPI), Glutamate and Gamma amino butyric acid (GA BA) regulates the growth , division and other vital functions ofa normal cell / organisin (Sudha, 1 998). The micro-environ ; nertt of the cell is controlled / the macro-environment that surrounds the individual. Any change in the cell environment causes imbalance in cell homeostasis and f,ntction. Pollution is a significant cause of imbalance caused iii the inacYcenvironment. Interaction with polluted environments can have an adverse impact on the health of humans. The alarming rise in enviromilmieil cont.iniin :rtion has been linked to rises in levels of pesticides, ndltstr al effluents, domestic Waste, car exhausts and other anthropogenic activities. Persistent exposures to contaminant cause a negative imp,-, on brain health and development . Pollution also causes a change in the neurotransmitters and their receptor function leading to earl.;' recurrence of neurodcge,terative disorders such as flypoxia , Alzbeimers's and Huntington 's disease early in life.

Neurotransmitters Functional Balance in Neurodegenerative Disease Management:recent Advances

The recent developments in neurobiology have rendered new prominence and potential to study about the structure and function of brain and related disorders. Human behaviour is the net result of neural control of the communication between brain cells. Neurotransmitters are chemicals that are used to relay, amplify and modulate electrical signals between neurons and/or another cell. It mediates rapid intercellular communication through the nervous system by interacting with cell surface receptors. These receptors often trigger second messenger signaling pathways that regulate the activity of ion channels. The functional balance of different neurotransmitters such as Acetylcholine (Ach), Dopamine (DA), Serotonin (5-HT), Norepinephrine (NE), Epinephrine (EPI), Glutamate and Gamma amino butyric acid (GABA) regulates the growth, division and other vital functions of a normal cell / organism (Sudha, 1998). Any change in neurotransmitters' functional balance will result in the failure of cell function and may lead to the occurrence of diseases. Abnormalities in the production or functioning of neurotransmitters have been implicated in a number of neurological disorders like Schizophrenia, Alzheimer's, Epilepsy, Depression and Parkinson's disease. Changes in central and peripheral neuronal signaling system is also noted in diabetes, cancer, cell proliferation, alcoholism and aging. Elucidation of neurotransmitters receptor interaction pathways and gene expression regulation by second messengers and transcriptional factors in health and disease conditions can lead to new small molecules for development of therapeutic agents to improve neurological disease conditions. Increased awareness of the global effects of neurological disorders should help health care planners and the neurological community set appropriate priorities in research, prevention, and management of these diseases.

STUDIES ON HAEMOLYMPH CONSTITUENTS OF PILA VIRENS (LAMARCK) AND THE EFFECTS OF PESTICIDES ON THE ACTIVITY PATTERN OF SELECTED ENZYMES

The present study is an attempt to understand the physiological responses of a freshwater gastropod, in terms of haematological parameters, in normal conditions as well as in various natural and man made altered conditions of the environment.Pila virens, a freshwater prosobranch,commonly found in paddy fields, ponds, and streams of Kerala is selected as the test animal for the present investigation. Various haemolymph constituents such as total carbohydrate, glycogen, total protein, total lipid, urea,ammonia,sodium,potasium, calcium, and chloride which are directly involved in the control and maintenance of different physiological systems, were analysed in the present study. Selected haematological parameters like total haemocyte number, and packed cell volume were also determined. Besides , the activity pattern of selected haemolymph enzymes such as acid phosphatase (ACP), alkaline phosphatase (ALP),Glutamate-oxaloacetate transaminase (GOT), and glutamate-pyruvate transaminase (GPT), all having diagnostic value in terms of internal defence system and metabolism of the organism, were also studied.

Adrenergic and Glutamergic Receptor Gene Expression and Their Functional regulation in the Cerebral Cortex of Hypoxia Induced Neonatal Rats:Role of Oxygen,Epinephrine and Glucose Supplementation

In the present work, the role of oxygen, epinephrine and glucose supplementation in regulating neurotransmitter contents, adrenergic and glutamate receptor binding parameters in the cerebral cortex of experimental groups of neonatal rats were investigated. The study of neurotransmitters and their receptors in the cerebral cortex and the EEG pattern in the brain regions of neonatal rats were taken as index for brain damage due to hypoxia, oxygen and epinephrine. Real-Time PCR work was done to confirm the binding parameters. Second messenger, cyclic Adenosine Monophosphate (cAMP) was assayed to find the functional correlation of the receptors. Behavioural studies were carried out to confirm the biochemical and molecular studies. The efficient and timely supplementation of glucose plays a crucial role in correcting the molecular changes due to hypoxia, oxygen and epinephrine. The addictive neuronal damage effect due to oxygen and epinephrine treatment is another important observation. The corrective measures from the molecular study brought to practice will lead to maintain healthy intellectual capacity during the later developmental stages, which has immense clinical significance in neonatal care.

Muscarinic M1 and M3 Receptors,IP3 and cGMP Functional Regulation in Streptozotocin Induced Diabetic Rats: Insulin and Somatotropin Induced Rejuvenation as a Function of Age

The present study describes that acetylcholine through muscarinic Ml and M3 receptors play an important role in the brain function during diabetes as a function of age. Cholinergic activity as indicated by acetylcholine esterase, a marker for cholinergic function, decreased in the brain regions - the cerebral cortex, brainstem and corpus striatum of old rats compared to young rats. in diabetic condition, it was increased in both young and old rats in cerebral cortex, and corpus striatum while in brainstem it was decreased. The functional changes in the muscarinic receptors were studied in the brain regions and it showed that muscarinic M I receptors of old rats were down regulated in cerebral cortex while in corpus striatum and brainstem it was up regulated. Muscarinic M3 receptors of old rats showed no significant change in cerebral cortex while in corpus striatum and brainstem muscarinic receptors were down regulated. During diabetes, muscarinic M I receptors were down regulated in cerebral cortex and brainstem of young rats while in corpus striatum they were up regulated. In old rats, M I receptors were up regulated in cerebral cortex, corpus striatum and in brainstem they were down regulated. Muscarinic M3 receptors were up regulated in cerebral cortex and brainstem of young rats while in corpus striatum they were down regulated. In old rats, muscarinic M l receptors were up regulated in cerebral cortex, corpus striatum and brainstem. In insulin treated diabetic rats the activity of the receptors were reversed to near control. Pancreatic muscarinic M3 receptor activity increased in the pancreas of both young and old rats during diabetes. In vitro studies using carbachol and antagonists for muscarinic Ml and M3 receptor subtypes confirmed the specific receptor mediated neurotransmitter changes during diabetes. Calcium imaging studies revealed muscarinic M I mediated Ca2 + release from the pancreatic islet cells of young and old rats. Electrophysiological studies using EEG recording in young and old rats showed a brain activity difference during diabetes. Long term low dose STH and INS treated rat brain tissues were used for gene expression of muscarinic Ml, M3, glutamate NMDARl, mGlu-5,alpha2A, beta2, GABAAa1 and GABAB, DAD2 and 5-HT 2C receptors to observe the neurotransmitter receptor functional interrelationship for integrating memory, cognition and rejuvenating brain functions in young and old. Studies on neurotransmitter receptor interaction pathways and gene expression regulation by second messengers like IP3 and cGMP in turn will lead to the development of therapeutic agents to manage diabetes and brain activity.From this study it is suggested that functional improvement of muscarinic Ml, M3, glutamate NMDAR1, mGlu-5, alpha2A, beta2, GABAAa1 and GABAB, DAD2 and 5-HT 2C receptors mediated through IP3 and cGMP will lead to therapeutic applications in the management of diabetes. Also, our results from long term low dose STH and INS treatment showed rejuvenation of the brain function which has clinical significance in maintaining healthy period of life as a function of age.

L-Glutaminase production by Marine Bacteria

L-glutaminases (L—glutamine amidohydrolase EC.3.5.l.2) is proposed as a prospective candidate for enzyme therapy cnf cancer and also as zui important additive during enzymatic digestion of shoyu koji since it could enhance glutamate content of soysauce. Commercial production of glutaminase could make possible its wide application in these areas, which would demand availability of potential sources and suitable fermentation techniques. The ‘present investigation highlighted marine environment as a potential source of efficient glutaminase producing bacteria mainly species of pseudomonas, aeromonas ,vibrio,alcaligenes, acinetobacter bacillus and planococci.Among them pseudomonas fluorescens ACMR 267 and v.cholerae ACMR 347 were chosen as the ideal strains for glutaminase production.Extracellular glutaminase fraction from all strains were in higher titres than intracellular enzymes during growth in mineral media, nutrient broth and nutrient broth added with glutamine.Glutaminase from all strains were purified employing (NH4)2SO4 fractionation followed tnr dialysis and ion exchange chromatography. The purified glutaminase from all strains were observed to be active and stable over a wide range of gfii and temperature.Optimization studies cflf environmental variables that normally influence time yiehi of glutaminase indicated that the optimal requirements of these bacteria for maximal glutaminase production remained stable irrespective of the medium, they are provided with for enzyme production. However, solid state fermentation technique was observed to be the most suitable process for the production of Glutaminase.

Withania somnitera and withanolide a mediated restoration of AMPA and NMDA receptor function in pilocarpine induced temporal lobe epilepsy

The onset of spontaneous seizures triggers a cascade of molecular and cellular events that eventually leads to neuronal injury and cognitive decline. The present study investigated the effect of Withania somnifera (WS) root extract and Withanolide A (WA) in restoring behavioural deficit by inhibiting oxidative stress induced alteration in glutamergic neurotransmission. The subdued performance in behavioural tests shows impaired motor coordination and memory. Histopathological investigations revealed significant neuronal loss in hippocampus of epileptic rats indicating glutamate mediated excitotoxicity. The treatment with WS and WA restored behavioural deficit and ameliorated neuronal loss. An altered redox homeostasis leading to oxidative stress is a hallmark of TLE. The antioxidant potential was afflicted in epileptic rats, evident from altered activity of SOD and CAT, down regulation of SOD and GPX expression and enhanced lipid peroxidation. The antioxidant property of WS and WA restored altered antioxidant capacity. Alteration in GDH activity and down regulation of GLAST expression resulted in enhanced glutamate content in the brain regions. The metabolism of glutamate was altered in the form of down regulated GAD expression. The alteration in synthesis, transport and metabolism resulted in further increase of the glutamate concentration at the synapse leading to glutamate mediated excitotoxicity. The decreased NMDA and AMPA receptor binding and down regulated NMDA R1, NMDA 2B and AMPA (GluR2) mRNA expression indicated altered glutamergic receptor function. The treatment with WS and WA reversed altered glutamergic receptor function, synthesis, transport and metabolism. The enhanced levels of second messenger IP3 responsible for Ca2+ mediated toxicity was reversed after treatment with WS and WA. Neurotoxics concentration of glutamate resulted in up regulation of pro apoptotic factors Bax and Caspase 8 and down regulation of anti apoptotic factor Akt resulting in neuronal death. The treatment with WS and WA resulted in activation of Akt and down regulation of Bax and caspase 8 leading to blocking of apoptotic pathway. The treatment with WS and WA resulted in reduced seizure frequency and amelioration of associated alterations suggesting the therapeutic role of Withania somnifera in temporal lobe epilepsy

Glutamatergic Nmda And Ampa Receptors Functional Regulation In Streptozotocin Induced Diabetic Rats: Effect Of Vitamin D3 And Curcumin Supplementation

The present study was designed to investigate the protective effect of curcumin and vitamin D3 in the functional regulation of glutamatergic NMDA and AMPA receptors in streptozotocin (STZ) induced diabetic rats. Alterations in glutamatergic neurotransmission in the brain were evaluated by analyzing the glutamate content, glutamate receptors - NMDA and AMPA receptors binding parameters and gene expression, GAD and GLAST gene expression. Immunohistochemistry studies using confocal microscope were carried out to confirm receptor density and gene expression results of NMDA and AMPA receptors. The role of glutamatergic receptors in pancreas was studied using the following parameters; glutamate content, GLAST expression, glutamate receptors - NMDA and AMPA receptor binding and gene expression. Increasing evidence in both experimental and clinical studies suggests that oxidative stress plays a major role in the pathogenesis of diabetes. In the present study SOD assay and GPx gene expression were done to evaluate the activity of antioxidant enzymes in the brain regions and pancreas. NeuroD1 and Pdx1 gene expression were performed in pancreas of experimental rats to evaluate pancreatic islet survival. Gene expression profiles of caspase 8, Bax, and Akt in brain regions and pancreas were studied to understand the possible mechanism behind curcumin and vitamin D3 mediated neuroprotection and islet survival. Gene expression studies of vitamin D3 receptor localisation in the pancreas was done to understand the mechanism of vitamin D3 in insulin secretion. Curcumin and vitamin D3 mediated insulin secretion via Ca2+ release were studied using confocal microscope.