RESUMO
This study examines the ability of the grass species Andropogon virginicus to alter the subsurface transport and redistribution of a suite of radionuclides (99Tc, 133Cs (stable analog for 135Cs and 137Cs), 237Np, 238U) with varying chemical behaviors in a Savannah River Site soil via the use of vegetated and unvegetated soil columns. After an acclimation period, a small volume of solution containing all radionuclides was introduced into the columns via Rhizon© pore water sampling tubes. Plants were grown for an additional 4 weeks before shoots were harvested, and columns were prepared for sampling. Plant presence led to decreased radionuclide release from the columns, mainly due to radionuclide specific combinations of system hydrology differences resulting from plant transpiration as well as plant uptake. For the most mobile radionuclides, 99Tc followed by 237Np, plant presence resulted in significantly different soil concentration profiles between vegetated and unvegetated columns, including notable upward migration for 237Np in columns with plants. Additionally, plant uptake of 99Tc was the greatest of all the radionuclides, with plant tissues containing an average of 44 % of the 99Tc, while plant uptake only accounted for <2 % of 237Np and <0.5 % of 133Cs and 238U in the system. Although overall plant uptake of 133Cs and 238U were similar, the majority of 133Cs taken up by plants was associated with 133Cs already available in the aqueous phase while 238U uptake was mainly associated with the solid phase, meaning that plant activity resulted in a fraction of the native 238U being mobilized and thus, made available for plant uptake. Overall, this study quantified the influence of several plant-mediated physical and biogeochemical factors that have significant influence on radionuclide mobility and transport in this complex system which can be further utilized in future system or site-specific environmental transport and risk assessment models.
Assuntos
Andropogon , Netúnio , Poluentes Radioativos do Solo , Urânio , Poluentes Radioativos do Solo/análise , Urânio/análise , Poaceae , Solo , Radioisótopos de Césio/análise , PlantasRESUMO
OBJECTIVE: To investigate antidiabetic efficacy of the extract of field grown and in vitro raised leaves of Solanum xanthocarpum (S. xanthocarpum) against alloxan induced diabetic rats. METHODS: The antidiabetic activity of the crude methanol extracts of the field grown and in vitro raised leaves of S. xanthocarpum at different concentrations (100-200 mg/kg bw) was tested against alloxan induced diabetic rats. The antidiabetic efficacy was validated through various biochemical parameters and the antioxidant effect was also determined. The phytochemical analyses of field grown S. xanthocarpum and in vitro rasied S. xanthocarpum leaves were done by estimating their chlorophyll, carotenoids, total sugar, protein, amino acid and minerals contents. RESULTS: The results revealed that the methanol extracts of both the leaves (field grown and in vitro raised) of S. xanthocarpum was efficient anti hyperglycemic agents at a concentration of 200 mg/kg bw and posses potent antioxidant activity. However, the extracts of in vitro rasied S. xanthocarpum raised leaves exhibit higher efficacy than the field grown leaves in all tested concentrations. Proximal composition and mineral analysis of S. xanthocarpum revealed higher concentration of contents in in vitro rasied S. xanthocarpum than field grown S. xanthocarpum. CONCLUSIONS: From the results it can be concluded that the leaves extracts of S. xanthocarpum can be a potential candidate in treating the hyperglycemic conditions and suits to be an agent to reduce oxidative stress.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Hiperglicemia/tratamento farmacológico , Fitoterapia , Extratos Vegetais/uso terapêutico , Folhas de Planta/química , Solanum/química , Animais , Biomarcadores/sangue , Diabetes Mellitus Experimental/complicações , Hiperglicemia/etiologia , Masculino , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Ratos , Ratos Wistar , Resultado do TratamentoRESUMO
Studies involving regulation of corticotropin-releasing hormone (CRH) in vitro have been used to validate findings obtained in vivo and more importantly have been used as model systems to better understand signalling mechanisms responsible for the expression of the CRH gene and peptide. Many in vitro studies examining CRH have utilized hypothalamic tissue while a few have focused on the amygdala. Clonal cell lines have also been utilized as models of central nervous system CRH neurons. Stimuli that have been implicated in regulating hypothalamic CRH regulation in vitro include protein kinase A (PKA) and protein kinase C (PKC) activators, glucocorticoids, biogenic amines, cytokines and the gaseous neurotransmitters. Amygdalar CRH levels in vitro are affected by some of the same stimuli that regulate hypothalamic CRH; however there is evidence supporting differential regulation of CRH in these two brain regions by some of the same stimuli. Only a few studies in aggregate have investigated signal transduction mechanisms and these studies have focused on PKA- and glucocorticoid-mediated changes in CRH expression. Thus, much more investigative work in better understanding CRH regulation in vitro is needed.
Assuntos
Tonsila do Cerebelo , Hormônio Liberador da Corticotropina/genética , Regulação da Expressão Gênica , Hipotálamo , Transdução de Sinais/fisiologia , Tonsila do Cerebelo/citologia , Tonsila do Cerebelo/enzimologia , Tonsila do Cerebelo/metabolismo , Células Cultivadas , Hormônio Liberador da Corticotropina/biossíntese , Hipotálamo/citologia , Hipotálamo/enzimologia , Hipotálamo/metabolismo , Transdução de Sinais/genéticaRESUMO
Corticotropin-releasing hormone (CRH) is a 41 amino acid neuropeptide which plays an important role in the stress response in the hypothalamus. We describe the development of an immortalized hypothalamic cell line which expresses CRH. We hypothesized that this cell line would possess the relevant characteristics of parvocellular CRH-expressing neurones such as glucocorticoid receptor (GR) expression and vasopressin (VP) coexpression. For production of hypothalamic cells, embryonic day 19 rat pup hypothalami were dissected and dissociated into tissue culture dishes. They were immortalized by retrovirus-mediated transfer of the SV40 large T antigen gene at 3 days of culture and then screened for expression of CRH following dilution cloning. One cell line was chosen (IVB) which exhibited CRH-like immunoreactivity (CRH-LI) and expressed CRH, VP and CRH1 receptor RNA via the reverse transcriptase-polymerase chain reaction. In addition, the cell line expressed the neuronal marker, microtubule-associated protein-2. We verified that the CRH-LI from IVB cell lysates coeluted with CRH standard via reversed-phase high-performance liquid chromatography (HPLC). Furthermore, oxidation of the lysate converted its HPLC profile to that identical with oxidized CRH standard. In addition, IVB cells exhibited high affinity binding to CRH. Incubation of IVB cells with CRH lead to increases in cAMP levels and protein kinase A activity in a concentration-dependent manner. Incubation of IVB cells with CRH also resulted in increases in phospho-cyclic-AMP response element binding protein (CREB) immunostaining as detected by immunocytochemical analysis. Finally, CRH treatment of IVB cell lines has been linked to CREB-mediated gene expression as determined via the PathDetect CREB trans-reporting system. The characteristics of IVB cells, such as CRH and VP coexpression, GR expression and a biologically active CRH-R1-mediated signalling pathway, suggest that this neuronal cell line may serve as model of parvocellular CRH neurones.
Assuntos
Hormônio Liberador da Corticotropina/genética , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Expressão Gênica , Hipotálamo/metabolismo , Receptores de Hormônio Liberador da Corticotropina/metabolismo , Transdução de Sinais , Animais , Antígenos Transformantes de Poliomavirus/genética , Western Blotting , Linhagem Celular Transformada , Cromatografia Líquida de Alta Pressão , Hormônio Liberador da Corticotropina/metabolismo , Hormônio Liberador da Corticotropina/farmacologia , AMP Cíclico/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Dexametasona/farmacologia , Expressão Gênica/efeitos dos fármacos , Hipotálamo/química , Fosforilação , Pró-Opiomelanocortina/genética , RNA Mensageiro/análise , Ratos , Ratos Sprague-Dawley , Receptores de Glucocorticoides/análise , Receptores de Glucocorticoides/genética , Transfecção , Vasopressinas/genéticaRESUMO
Although second messenger systems subserving neuropeptide Y (NPY)-mediated behaviors have been identified for a variety of receptors in several tissues, downstream signaling events are not well known. The nuclear binding protein, cyclic AMP response element binding protein (CREB) appears to be a transcription factor that is activated following injection of NPY into rat hypothalamus. To allow determination of the functional nature of CREB mediation of NPY-induced feeding, injection cannulae were implanted into the perifornical hypothalamus of 18 rats. Treatment of seven rats with CREB antisense oligonucleotide (15 ug) significantly antagonized NPY feeding for up to one week after treatment, while similar injections of CREB sense oligonucleotide (15 ug) had no significant effect on NPY-induced feeding. Two weeks after the antisense oligonucleotide treatment, feeding was once again elicited by the injection of NPY. Hypothalamic CREB protein was also reduced significantly two days after the CREB antisense oligonucleotide treatment. These results suggest that activation of CREB, probably through phosphorylation, may be a necessary event for the signal transduction of NPY stimulation into feeding behavior.
Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Comportamento Alimentar/fisiologia , Hipotálamo/fisiologia , Neuropeptídeo Y/farmacologia , Oligodesoxirribonucleotídeos Antissenso/farmacologia , Animais , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Comportamento Alimentar/efeitos dos fármacos , Hipotálamo/efeitos dos fármacos , Masculino , Microinjeções , Neuropeptídeo Y/administração & dosagem , Neuropeptídeo Y/antagonistas & inibidores , Oligodesoxirribonucleotídeos Antissenso/administração & dosagem , Ratos , Ratos Sprague-Dawley , Tionucleotídeos , Fatores de TempoAssuntos
Ingestão de Alimentos/fisiologia , Hipotálamo/efeitos dos fármacos , Neuropeptídeo Y/fisiologia , Oligonucleotídeos Antissenso/farmacologia , Animais , Proteína de Ligação a CREB , AMP Cíclico/metabolismo , Masculino , Proteínas Nucleares/metabolismo , Ligação Proteica , Ratos , Ratos Sprague-Dawley , Elementos de Resposta , Transdução de Sinais , Tionucleotídeos , Transativadores/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Maintaining tumor-bearing rats on total parenteral nutrition (TPN) for eight days significantly reduced mass, protein, and DNA in small intestine and colon. Coinfusion of glucagon-like peptide 2 (GLP-2) significantly increased each of these variables in the duodenum, jejunum, and ileum, but not in the colon. Histological analysis of tissue revealed normal mucosa thickness and villus height in the small intestine of GLP-2-treated rats, whereas non-treated rats maintained on TPN exhibited villus shortening and thinning of the mucosa. Compared with TPN alone, no significant effects of GLP-2 were noted on tumor growth, liver weight, or heart weight. Coinfusion of GLP-2 with TPN had no significant effect on TPN-associated immunosuppression, as measured by mitogen-induced proliferation of cultured splenocytes. Although translocation of bacteria to the mesenteric lymph nodes appeared to be reduced in GLP-2-treated rats, the difference between groups was not statistically significant. These results suggest that hormonal alterations may be more important than an absence of luminal nutrition in TPN-associated mucosa changes in tumor-bearing rats. Additionally, maintenance of gut integrity during TPN does not appear to be a sufficient condition for the avoidance of the negative sequelae associated with this route of supplemental nutrition.
Assuntos
Enteropatias/prevenção & controle , Intestino Delgado/patologia , Nutrição Parenteral/efeitos adversos , Peptídeos/uso terapêutico , Animais , Translocação Bacteriana/efeitos dos fármacos , DNA , Peptídeo 2 Semelhante ao Glucagon , Peptídeos Semelhantes ao Glucagon , Enteropatias/etiologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Masculino , Microvilosidades , Neoplasias , Ratos , Ratos Endogâmicos F344RESUMO
Although reduced biological activity of the obese gene product, leptin, has been associated with obesity, little information is available concerning leptin alterations during anorexia. Therefore, we measured circulating leptin concentrations and hypothalamic leptin binding in anorectic tumor-bearing and pair-fed control rats. Plasma concentrations of leptin decreased in tumor-bearing rats early in the course of tumor growth, and fell to nearly non-detectable levels during severe anorexia. The pair-fed control rats that ate the same amount of food as did the anorectic tumor-bearing rats exhibited a 50% decrease in plasma leptin concentration. Concentrations of free fatty acids were elevated in both tumor-bearing and pair-fed groups, while circulating levels of triglycerides were increased only in anorectic tumor-bearing rats. Leptin receptor density was doubled in the hypothalamus of tumor bearing rats, while binding affinity was decreased by 50%. These results suggest that peripheral leptin production is down-regulated, perhaps due to increased lipolysis in tumor-bearing rats. It appears that hypothalamic leptin systems up-regulate receptor numbers in response to decreased blood leptin level, however, the decrease in binding affinity may compensate for these alterations. Therefore, the influence of leptin on hypothalamic neuropeptide Y feeding systems may be minimal in anorectic tumor-bearing rats.
Assuntos
Anorexia/metabolismo , Proteínas de Transporte/metabolismo , Hipotálamo/metabolismo , Proteínas/metabolismo , Receptores de Superfície Celular , Sarcoma Experimental/metabolismo , Animais , Anorexia/sangue , Peso Corporal , Ingestão de Alimentos/fisiologia , Jejum , Ácidos Graxos não Esterificados/sangue , Hipotálamo/fisiologia , Leptina , Masculino , Obesidade/metabolismo , Ligação Proteica , Ratos , Ratos Endogâmicos F344 , Receptores para Leptina , Sarcoma Experimental/sangueRESUMO
Modifications of (D-Trp32) neuropeptide Y (NPY) led to the development of potential peptide-based lower molecular weight (500-800 Da) NPY feeding antagonists. One compound, WRYamide (N-Ac-Trp-Arg-Tyr-NH2), blocked NPY-induced feeding for 1 to 4 h when injected intrahypothalamically (i.h.t.) at 1 to 40 microgram. Schedule-induced feeding was also antagonized for up to 24 h by 20 microgram of WRYamide, i.h.t. Injection of 2.5 mg/kg (1 mg/rat) of WRYamide, i.v., also reduced significantly schedule-induced feeding for 4 h. A conditioned taste aversion could not be classically conditioned to saccharin using WRYamide as the unconditioned stimulus. These results may lead to the development of systemically active anti-obesity drugs.
Assuntos
Depressores do Apetite/farmacologia , Ingestão de Alimentos/efeitos dos fármacos , Neuropeptídeo Y/análogos & derivados , Neuropeptídeo Y/química , Oligopeptídeos/farmacologia , Animais , Cateterismo , Relação Dose-Resposta a Droga , Hipotálamo/efeitos dos fármacos , Injeções Intravenosas , Masculino , Neuropeptídeo Y/administração & dosagem , Neuropeptídeo Y/farmacologia , Oligopeptídeos/administração & dosagem , Ratos , Ratos Sprague-Dawley , Sacarina/farmacologia , Fatores de TempoRESUMO
Intrahypothalamic (IHT) administration of neuropeptide Y (NPY) induces a robust feeding response in rats. We have shown previously that NPY-induced feeding is mediated by a pertussis-toxin-sensitive G protein in rats. NPY receptors are coupled to cAMP and Ca2+. Because these second messengers are known to activate cAMP response element binding proteins, (CREB), cAMP response element modulators, or activating transcription factor 1, we investigated the involvement of these transcription factors in NPY-induced feeding in rats. Compared with control injections of cerebrospinal fluid (1 microl), IHT administration of NPY increased cAMP response element (CRE) binding to rat hypothalamic nuclear extracts in a time-dependent manner, as detected by an electrophoretic mobility shift assay. In contrast, IHT administration of the anorectic neuropeptide, pituitary adenylate cyclase activating polypeptide, strongly inhibited the CRE binding. Food deprivation for 48 hr also increased CRE binding, whereas 8 hr of refeeding normalized CRE activity. Preincubation of the hypothalamic nuclear extracts of NPY-treated and unfed rats with antibody specific to CREB blocked CRE binding, whereas preincubation with phosphoCREB antibody retarded the migration of CRE-protein complex, indicating that phosphoCREB is involved in this process. Consistently, immunohistochemical studies with food-deprived rats showed an intense phosphoCREB signal in the paraventricular nuclei and ventromedial hypothalamus in comparison to rats fed ad libitum. Hypothalamic calcium/calmodulin-dependent protein kinase II activity was also increased by IHT-NPY. These results suggest that calcium/calmodulin-dependent protein kinase II induced phosphorylation of CREB may be involved in regulating feeding behavior induced by NPY.
Assuntos
Estimulantes do Apetite/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Privação de Alimentos/fisiologia , Hipotálamo/efeitos dos fármacos , Hipotálamo/metabolismo , Neuropeptídeo Y/farmacologia , Animais , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Proteínas Quinases Dependentes de Cálcio-Calmodulina/efeitos dos fármacos , Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Núcleo Celular/metabolismo , Vias de Administração de Medicamentos , Masculino , Neuropeptídeos/farmacologia , Neurotransmissores/farmacologia , Fosforilação , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase , Ligação Proteica , Ratos , Ratos Sprague-DawleyAssuntos
Neuropeptídeo Y/análogos & derivados , Neuropeptídeo Y/farmacologia , Receptores de Neuropeptídeo Y/antagonistas & inibidores , Animais , Comportamento Alimentar/efeitos dos fármacos , Hipotálamo/efeitos dos fármacos , Hipotálamo/fisiologia , Cinética , Neuropeptídeo Y/química , Neuropeptídeo Y/metabolismo , Ratos , Relação Estrutura-AtividadeRESUMO
Neuropeptide Y (NPY) is a potent orexigenic peptide. Structure-activity studies have revealed that nearly the entire sequence of NPY is required to elicit feeding responses. Therefore, in order to develop antagonistic peptides for NPY-induced feeding, we synthesized full-length analogs of NPY, substituting D-Trp in the C-terminal receptor binding region, and screened their activity in rat hypothalamus. Although [D-Trp36]NPY and [D-Trp34]NPY inhibited isoproterenol-stimulated hypothalamic membrane adenylate cyclase activity, [D-Trp32]NPY exhibited no intrinsic activity. Furthermore, [D-Trp32]NPY inhibited [125I]NPY binding to rat hypothalamic membranes with a potency comparable to that of NPY. The presence of 30 and 300 nM concentrations of [D-Trp32]NPY shifted the inhibitory dose-response curve of NPY on isoproterenol-stimulated hypothalamic membrane adenylate cyclase activity parallel to the right with comparable KB values. Moreover, in vivo experiments in rats revealed that [D-Trp32]NPY (10 micrograms) significantly attenuated the 1-h feeding response induced by NPY (1 microgram). Several other substitutions at position 32 including 2-D-Nal resulted in agonist activity, suggesting that there are strict structural requirements to induce the antagonistic property in NPY. These findings confirm that [D-Trp32]NPY is a competitive antagonist of NPY in both in vitro and in vivo systems. Analogs based on [D-Trp32]NPY may have potential clinical application, since NPY has been implicated in the pathophysiology of a number of feeding disorders including obesity, anorexia, and bulimia.
Assuntos
Hipotálamo/efeitos dos fármacos , Neuropeptídeo Y/análogos & derivados , Neuropeptídeo Y/antagonistas & inibidores , Inibidores de Adenilil Ciclases , Adenilil Ciclases/metabolismo , Sequência de Aminoácidos , Animais , Relação Dose-Resposta a Droga , Ingestão de Alimentos/efeitos dos fármacos , Hipotálamo/enzimologia , Conformação Molecular , Dados de Sequência Molecular , Neuropeptídeo Y/síntese química , Neuropeptídeo Y/farmacologia , Ratos , Ratos Sprague-Dawley , EstereoisomerismoRESUMO
The efficacy of NPY to elicit feeding in TB rats was reduced prior to the onset of overt anorexia, with the feeding response decreasing further as anorexia developed. Hypothalamic concentration of NPY was reduced in TB rats, with the magnitude of the decrease paralleling the degree of anorexia. Binding affinity of NPY to hypothalamic membranes taken from TB rats suggested decreased binding affinity with no change in receptor number. Infusing ammonium salts at a concentration and rate necessary to increase blood ammonia levels to the degree observed in TB rats, produced anorexia and decreased NPY feeding. These results suggest that NPY feeding systems are abnormal in TB rats and that hyperammonemia may be of primary importance in this dysfunction.
Assuntos
Anorexia/etiologia , Encéfalo/metabolismo , Neuropeptídeo Y/fisiologia , Sarcoma Experimental/complicações , Amônia/farmacologia , Animais , Ingestão de Alimentos/efeitos dos fármacos , Hipotálamo/metabolismo , Masculino , Metilcolantreno , Transplante de Neoplasias , Neuropeptídeo Y/farmacologia , Ratos , Ratos Endogâmicos F344 , Receptores de Neuropeptídeo Y/metabolismo , Sarcoma Experimental/induzido quimicamente , Sarcoma Experimental/patologia , Fatores de TempoRESUMO
Neuropeptide Y (NPY) is the most powerful peptide drug stimulating feeding in rats. Rats with paraventricular hypothalamic (PVH) cannulae were used to investigate the mechanisms involved in NPY-induced feeding. Consistent with previous reports, injection of 2 micrograms of NPY into the PVH significantly increased the cumulative food intake over 1-, 2- and 4-hr periods. Ad lib feeding decreased significantly two days after pertussis toxin (PT) administration, but recovered to nearly normal levels on the fourth day. PT had no immediate effect on NPY-induced feeding; however, four days after PT was injected NPY (2 micrograms) did not increase the food intake compared to control. In vitro investigations showed that isoproterenol-stimulated adenylate cyclase activity in the hypothalamus of control rats was inhibited by NPY. In PT-treated rats, however, no inhibition of cAMP production was observed. These results suggest that cAMP may mediate NPY-induced feeding and that a PT-sensitive G protein may be involved in this signal transduction.