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.

Adrenergic and muscarinic receptor subtypes functional regulation during diabetogenesis: Effect of curcumin and vitamin D3 pre-treatment

In the present study, the initial phase was directed to confirm the effects of curcumin and vitamin D3 in preventing or delaying diabetes onset by studying the blood glucose and insulin levels in the pre-treated and diabetic groups. Behavioural studies were conducted to evaluate the cognitive and motor function in experimental rats. The major focus of the study was to understand the cellular and neuronal mechanisms that ensure the prophylactic capability of curcumin and vitamin D3. To elucidate the mechanisms involved in conferring the antidiabetogenesis effect, we examined the DNA and protein profiles using radioactive incorporation studies for DNA synthesis, DNA methylation and protein synthesis. Furthermore the gene expression studies of Akt-1, Pax, Pdx-1, Neuro D1, insulin like growth factor-1 and NF-κB were done to monitor pancreatic beta cell proliferation and differentiation. The antioxidant and antiapoptotic actions of curcumin and vitamin D3 were examined by studying the expression of antioxidant enzymes - SOD and GPx, and apoptotic mediators like Bax, caspase 3, caspase 8 and TNF-α. In order to understand the signalling pathways involved in curcumin and vitamin D3 action, the second messengers, cAMP, cGMP and IP3 were studied along with the expression of vitamin D receptor in the pancreas. The neuronal regulation of pancreatic beta cell maintenance, proliferation and insulin release was studied by assessing the adrenergic and muscarinic receptor functional regulation in the pancreas, brain stem, hippocampus and hypothalamus. The receptor number and binding affinity of total muscarinic, muscarinic M1, muscarinic M3, total adrenergic, α adrenergic and β adrenergic receptor subtypes were studied in pancreas, brain stem and hippocampus of experimental rats. The mRNA expression of muscarinic and adrenergic receptor subtypes were determined using Real Time PCR. Immunohistochemistry studies using confocal microscope were carried out to confirm receptor density and gene expression results. Cell signalling alterations in the pancreas and brain regions associated with diabetogenesis and antidiabetogenesis were assessed by examining the gene expression profiles of vitamin D receptor, CREB, phospholipase C, insulin receptor and GLUT. This study will establish the anti-diabetogenesis activity of curcumin and vitamin D3 pre-treatment and will attempt to understand the cellular, molecular and neuronal control mechanism in the onset of diabetes.Administration of MLD-STZ to curcumin and vitamin D3 pre-treated rats induced only an incidental prediabetic condition. Curcumin and vitamin D3 pretreated groups injected with MLD-STZ exhibited improved circulating insulin levels and behavioural responses when compared to MLD-STZ induced diabetic group. Activation of beta cell compensatory response induces an increase in pancreatic insulin output and beta cell mass expansion in the pre-treated group. Cell signalling proteins that regulate pancreatic beta cell survival, insulin release, proliferation and differentiation showed a significant increase in curcumin and vitamin D3 pre-treated rats. Marked decline in α2 adrenergic receptor function in pancreas helps to relent sympathetic inhibition of insulin release. Neuronal stimulation of hyperglycemia induced beta cell compensatory response is mediated by escalated signalling through β adrenergic, muscarinic M1 and M3 receptors. Pre-treatment mediated functional regulation of adrenergic and cholinergic receptors, key cell signalling proteins and second messengers improves pancreatic glucose sensing, insulin gene expression, insulin secretion, cell survival and beta cell mass expansion in pancreas. Curcumin and vitamin D3 pre-treatment induced modulation of adrenergic and cholinergic signalling in brain stem, hippocampus and hypothalamus promotes insulin secretion, beta cell compensatory response, insulin sensitivity and energy balance to resist diabetogenesis. Pre-treatment improved second messenger levels and the gene expression of intracellular signalling molecules in brain stem, hippocampus and hypothalamus, to retain a functional neuronal response to hyperglycemia. Curcumin and vitamin D3 protect pancreas and brain regions from oxidative stress by their indigenous antioxidant properties and by their ability to stimulate cellular free radical defence system. The present study demonstrates the role of adrenergic and muscarinic receptor subtypes functional regulation in curcumin and vitamin D3 mediated anti-diabetogenesis. This will have immense clinical significance in developing effective strategies to delay or prevent the onset of diabetes.

Hypothalamic 5-HT functional regulation through 5-HTIA and 5-HT2c receptors during pancreatic regeneration

nerv5o-HusT s ryescteempto. rTshaer oeb pjercetdiovme oinfa tnhtilsy s ltoucdaytewd ains ttoh ein bvreasitnig aanted tahree rionlveo olvf ehdy ipno tphaanlacrmeiact i5c- fHuTn,c t5i-oHn TaInAd acneldl p5r-HolTif2ercatrieocnepttohrr obuingdh isnygm apnadt hgeetniec pphtrqsHehepayaxuTevnepnpa cecIocnrhAirarettfyehilies pfceaasaai tdiolnetaoiae tcddnndmhc tr etab5aiueoncly-ggsamHr oermeHnndiaTasPeuituse2s rsLremdtca id oC tn[orri 3fegoa.5d H c n7t5-epseH.]2l- a mpHro nThtefeoTcv IsprrApeIueaAralga nesnaeterccninrdgrcrdeei e erntc aae5oeeettxg -pie.npHc ectTe rnotrTahoereersme2 rgas acseeiisthosnsxienaaoeprdmynrer a eicr wniestani pstalot iestrhsonov.aen r ted5u shloo-sm..yHifn nT pOe5RTgoh -u bINtH6iAhrys0AT a r%saluIe ta neussA mdxupidn plauya5tgnrnss - ei csHdssospr u sfT5teeg hia-s2cogHehticneef aT fisc.rmc it2teTr oacsc htmot gehr eppoteey oentc 5oh. rei -iarysTpdHsttpthee oTwonde[rt3I ,t ehp AgiH7rfaaeey2 ]lnnaa8 ce5nhmd-r O- doweaiHw caHn5atTnds-i sDc H I-ea7rAPrnT reodaA eg2atalguoyTncyelnz dan.sr eete5 ee5drrp-cdg a-HaebH itincpyTino Tc tr2nRore2cterThccaswee-r trpPe eahecctgCscyoet eRoeperpnmv tpo.feo autt5i rohlsen-ueraxHacdalpstaTtigsremor aeedcanynsuot asbs esnwli.y. o t er5e Ran5ex-nsgTH-pt Hudi-rTnPlwoeTa Csncatt sciohesRioo n oehnb ntna i ey7tgdn ne i huaundntel rs tywartii,nshn y ai5igngesss-

Enhanced /3-adrenergic receptors in the brain and pancreas during pancreatic regeneration in weanling rats

Adrenergic stimulation has an inyortant role in the pancreatic It-cell proliferation and insulin secretion. In the present study. we have investigaled how sympathetic system mgulales the panrrealic n I rnerui nr ht an:ilyiing I'pinephi inn 1111 ), Norepinephrinc (NE) and /1-adrenergic receptor changes in the brain as (%eli is in the I swirls. Fill and NII showed a significant decrease in the brain regions, pancreas and plasma :rt 72Ius iller partial prurcrealectonty. We observed an increase in the circulating insulin levels at 72 hrs. Scatchard analysis using I CHI propranolol showed a significant increase in the number of loth the low affinity and high affinity t-adrenergic receplors in cerebral cortex and hypothalamus of partially pancreatectornised rats during peak DNA synthesis. The affinity of the receptors decrea,ed significantly in the low and high affinity receptors of cerebral cortex and the high affinity hypothalamic receptors. In file brain stein, low affinity receptors were increased significantly during regeneration whereas there was no change in the high affinity receptors. The pancreatic ff-adrenergic receptors were also up regulated at 72 firs after partial panerealectony. In vitro studies showed that /i-adrenergic receptors are positive regulators of islet cell proliferation and insulin secretion. Thus our results suggest that the t-adrenergic receptors are functionally enhanced during pancreatic regeneration, which in turn increases pancreatic ft-cell proliferation an(hilisulin secretion in wean hug rats.

Decreased [ 3 H] YM-09151-2 binding to dopamine D2 receptors in the hypothalamus, brainstem and pancreatic islets of streptozotocin -induced diabetic rats

In the present study dopamine was measured in the hypothalamus, brainstem, pancreatic islets and plasma, using HPLC. Dopamine D2 receptor changes in the hypothalamus, brainstem and pancreatic islets were studied using [3H] YM-09151-2 in streptozotocin-induced diabetic and insulintreated diabetic rats. There was a significant decrease in dopatnine content in the hypothalamus (P<0.001), brainstem (P<0.001), pancreatic islets (P<0.001) and plasma (P<0.00I) in diabetic rats when compared to control. Scatchard analysis of [3H] YM-09151-2 in the hypothalamus of diabetic rats showed a significant decrease in Bax (P<0.001) and Kd, showing an increased affinity of D2 receptors when compared to control. Insulin treatment did not completely reverse the changes that occurred during diabetes. There was a significant decrease in B,nax (P<0.01) with decreased affinity in the brainstem of diabetic rats. The islet membrane preparation of diabetic rats showed a significant decrease (P<0.001) in the binding of [3H] YM-09 151-2 with decreased Kd (P<0.001) compared to control. The increase in affinity of D2 receptors in hypothalamus and pancreatic islets and the decreased affinity in brainstem were confirmed by competition analysis. Thus our results suggest that the decreased dopamine D, receptor function in the hypothalamus, brainstem and pancreas affects insulin secretion in diabetic rats, which has immense clinical relevance to the management of diabetes.

Enhancement of [m-methoxy 3H]MDL100907 binding to 5HT 2A receptors in cerebral cortex and brain stem of streptozotocin induced diabetic rats

5-Hydroxytryptamine2A (5-HT2A) receptor kinetics was studied in cerebral cortex and brain stem of streptozotocin (STZ) induced diabetic rats. Scatchard analysis with [3H] (±) 2,3dimethoxyphenyl-l-[2-(4-piperidine)-methanol] ([3H]MDL100907) in cerebral cortex showed no significant change in maximal binding (Bmax) in diabetic rats compared to controls. Dissociation constant (K) of diabetic rats showed a significant decrease (p < 0.05) in cerebral cortex, which was reversed to normal by insulin treatment. Competition studies of [3H]MDL100907 binding in cerebral cortex with ketanserin showed the appearance of an additional low affinity site for 5-HT2A receptors in diabetic state, which was reversed to control pattern by insulin treatment. In brain stem, scatchard analysis showed a significant increase (p < 0.05) in Bmax accompanied by a significant increase (p < 0.05) in Kd. Competition analysis in brain stem also showed a shift in affinity towards a low affinity State for 5-HT2A receptors. All these parameters were reversed to control level by insulin treatment. These results show that in cerebral cortex there is an increase in affinity of 5-HT2A receptors without any change in its number and in the case of brain stem there is an increase in number of 5HT2A receptors accompanied by a decrease in its affinity during diabetes. Thus, from the results we suggest that the increase in affinity of 5-HT2A receptors in cerebral cortex and upregulation of 5-HT2A receptors in brain stem may lead to altered neuronal function in diabetes.

Dopaminergic Regulation of Glucose-induced Insulin Secretion through Dopamine D2 Receptors in the Pancreatic Islets In Vitro

The stimulatory effect of dopamine through dopamine 1)2 receptor on glucose - induced insulin secretion was studied in the pancreatic islets in nitro. I)oparnilie signifieanlly stimula(ed insulin secretion at a concentration of 10 a N1 in the presence of high,glucose ( 20 nii1 ). ' fhe higher concentrations of dopamine (111 -1() 4) inhibited glucose- induced insulin secretion in the presence of both 4 mM1 and 20 m M glucose. Stimulatory and inhibitory effect of dopamine on glucose - induced insulin secretion was reverted by the addition of dopamine 1)2 receptor antagonists such as butaclamol and sulpiride . Norepinephrine (NE) at 111 4 11 concentration inhibited the dopamine uptake as well as its stimulatory effect at 11) - 8 IN1 concentration on glucose induced insulin secretion. Our results suggest that dopamine exerts a differential effect on glucose -induced insulin secretion through dopamine D2 receptor and it is essential for the regulation of glucose-induced insulin secretion by pancreatic islets.

Effect of Pyridoxine Deficiency in Young-Rats on High-Affinity Serotonin and Dopamine Receptors

The high-affinity bindings of [3H]-5-hydroxytryptamine to serotonin S-1 receptors, [3H]-ketanserin to serotonin S-2 receptors in the cerebral cortex, [3H]- fluphenazine to dopamine D-1 receptors, and [3H]-spiroperidol to dopamine D-2 receptors in the corpus striatum were studied in pyridoxine-deficient rats and compared to pyridoxine-supplemented controls. There was a significant increase in the maximal binding (Bmax) of serotonin S-1 and S-2 receptors with a significant decrease in their binding affinities (Kd). However, there were no significant changes either in the maximal binding or binding affinity of striatal dopamine D- 1 and D-2 receptors. Receptor sensitivity seems to correlate negatively with the corresponding neurotransmitter concentrations in the pyridoxine-deficient rats.

Age-related and sex-related alterations in f3-adrenergic receptors in different regions of rat brain

The binding of (-)[ 3H ]dihydroalprenolol , an antagonist of norepinephrine , to $-adrenergic receptors in different regions of the brain of male and female rats of various ages was measured . The binding to the synaptosomal fraction of corpus striatum , hypothalamus, cerebral cortex, cerebellum and the brainstems shows a significant decrease in the binding in old rats of both sexes . Only in the female corpus striatal region, the binding in the adult and the old is the same . In the case of females, the highest binding is seen in the young. In the male, an increase in binding occurs up to adulthood , after which it declines, suggesting a definite sex-related difference in the Q-adrenergic receptor.

Dopamine Receptor Gene Expression In Streptozotocin Induced Diabetic Rats And Its Role In Insulin Secretion

Diabetes Mellitus is a metabolic disorder associated with insulin deficiency, which not.only affects the carbohydrate metabolism but also is associated with various central and peripheral complications. Chronic hyperglycemia during diabetes mellitus is a major initiator of diabetic microvascular complications like retinopathy, neuropathy, The central nervous system (CNS) neurotransmitters play an important role in the regulation of glucose homeostasis. These neurotransmitters mediate rapid intracellular communications not only within the central nervous system but also in the peripheral tissues. They exert their function through receptors present in both neuronal and non neuronal cell surface that trigger second messenger signaling pathways. Dopamine is a neurotransmitter that has been implicated in various central neuronal degenerative disorders like Parkinson's disease and behavioral diseases like Schizophrenia. Dopamine is synthesised from tyrosine, stored in vesicles in axon terminals and released when the neuron is depolarised. Dopamine interacts with specific membrane receptors to produce its effect. Dopamine plays an important role both centrally and peripherally. The recent identification of five dopamine receptor subtypes provides a basis for understanding dopamine's central and peripheral actions . Dopamine receptors are classified into two major groups : DA D1 like and DA D2 like. Dopamine D1 like receptors consists of DA D1 and DA D5 receptors . Dopamine D2 like receptors consists of DA D2, DA D3 and DA D4 receptors. Stimulation of the DA D1 receptor gives rise to increased production of cAMP. Dopamine D2 receptors inhibit cAMP production, but activate the inositol phosphate second messenger system . Impairment of central dopamine neurotransmission causes muscle rigidity, hormonal regulation , thought disorder and cocaine addiction. Peripheral dopamine receptors mediate changes in blood flow, glomerular filtration rate, sodium excretion and catecholamine release. The dopamine D2 receptors increased in the corpus striatum and cerebral cortex but decreased in the hypothalamus and brain stem indicating their involvement in regulating insulin secretion. Dopamine D2 receptor which has a stimulatory effecton insulin secretion decreased in the pancreatic islets during diabetes. Our in vitro studies confirmed the stimulatory role of dopamine D2 receptors in stimulation of glucose induced insulin secretion. A detailed study at the molecular level on the mechanisms involved in the role of dopamine in insulin secretion, its functional modification could lead to therapeutic interventions that will have immense clinical importance.

Microwave Dielectric Resonator Based on Ba[(Bi0.2 D3+0.3)Nb0.5]O3(D3+=Y,Pr,Sm,Gd,Dy,Er)

Dielectric resonator ceramics with composition formula Ba[(D3+0.3 Bi0.2)Nb0.5]O3,where D3+=Y,Pr,Sm,Gd,Dy and Er,were prepared by the conventional ceramic preparation route

Down-regulation of cerebellar 5-HT2C receptors in pilocarpine-induced epilepsy in rats: Therapeutic role of Bacopa monnieri extract

Epilepsy is a syndrome of episodic brain dysfunction characterized by recurrent unpredictable, spontaneous seizures. Cerebellar dysfunction is a recognized complication of temporal lobe epilepsy and it is associated with seizure generation, motor deficits and memory impairment. Serotonin is known to exert a modulatory action on cerebellar function through 5HT2C receptors. 5-HT2C receptors are novel targets for developing anticonvulsant drugs. In the present study, we investigated the changes in the 5-HT2C receptors binding and gene expression in the cerebellum of control, epileptic and Bacopa monnieri treated epileptic rats. There was a significant down regulation of the 5-HT content (pb0.001), 5-HT2C gene expression (pb0.001) and 5-HT2C receptor binding (pb0.001) with an increased affinity (pb0.001). Carbamazepine and B. monnieri treatments to epileptic rats reversed the down regulated 5-HT content (pb0.01), 5-HT2C receptor binding (pb0.001) and gene expression (pb0.01) to near control level. Also, the Rotarod test confirms the motor dysfunction and recovery by B. monnieri treatment. These data suggest the neuroprotective role of B. monnieri through the upregulation of 5-HT2C receptor in epileptic rats. This has clinical significance in the management of epilepsy

Decreased GABAA Receptors Functional Regulation 3 in the Cerebral Cortex and Brainstem of Hypoxic Neonatal Rats: 4 Effect of Glucose and Oxygen Supplementation

Hypoxia in neonates can lead to biochemical and molecular alterations mediated through changes in neurotransmitters resulting in permanent damage to brain. In this study, we evaluated the changes in the receptor status of GABAA in the cerebral cortex and brainstem of hypoxic neonatal rats and hypoxic rats supplemented with glucose and oxygen using binding assays and gene expression of GABAAa1 and GABAAc5. In the cerebral cortex and brainstem of hypoxic neonatal rats, a significant decrease in GABAA receptors was observed, which accounts for the respiratory inhibition. Hypoxic rats sup- plemented with glucose alone and with glucose and oxygen showed, respectively, a reversal of the GABAA receptors, andGABAAa1 and GABAAc5 gene expression to control. Glucose acts as an immediate energy source thereby reducing the ATP-depletion-induced increase in GABA and oxygenation, which helps in encountering anoxia. Resuscitation with oxygen alone was less effective in reversing the receptor alterations. Thus, the results of this study suggest that reduction in the GABAA receptors functional regulation during hypoxia plays an important role in mediating the brain damage. Glucose alone and glucose and oxygen supplementation to hypoxic neonatal rats helps in protecting the brain from severe hypoxic damage.

5-HT2C and NMDA Receptors, IP3, cGMP and cAMP Functional Regulation in Pilocarpine Induced Temporal Lobe Epilepsy in Rats: Neuroprotective Role of Bacopa monnieri

Department of Biotechnology, Cochin University of Science and Technology