RESUMO
Using real-time confocal microscopy, we have demonstrated that lysophosphatidic acid (LPA), a bioactive phospholipid existing in plasma, positively regulates fluid flow-induced [Ca(2+)](i) response in fluo 4-loaded, cultured, bovine aortic endothelial cells. The initial increase in [Ca(2+)](i) was localized to a circular area with a diameter of <4 microm and spread concentrically, resulting in a mean global increase in [Ca(2+)](i). The local increase often occurred in a stepwise manner or repetitively during constant flow. The percentage of cells that responded and the averaged level of increase in [Ca(2+)](i) were dependent on both the concentration of LPA (0.1 to 10 micromol/L) and the flow rate (25 to 250 mm/s). The response was inhibited by removing extracellular Ca(2+) or by the application of Gd(3+), an inhibitor of mechanosensitive (MS) channels, but not by thapsigargin, an inhibitor of the endoplasmic reticular Ca(2+)-ATPASE: It was also inhibited by 8-bromo-cGMP, and the inhibition was completely reversed by KT5823, an inhibitor of protein kinase G (PKG). These results suggest that the [Ca(2+)](i) response arises from Ca(2+) influx through Gd(3+)-sensitive MS channels, which are negatively regulated by the activation of PKG. The spatiotemporal properties of the [Ca(2+)](i) response were completely different from those of a Ca(2+) wave induced by ATP, a Ca(2+)-mobilizing agonist. Therefore, we called the phenomenon Ca(2+) spots. We conclude that LPA positively regulates fluid flow-induced local and oscillatory [Ca(2+)](i) increase, ie, the Ca(2+) spots, in endothelial cells via the activation of elementary Ca(2+) influx through PKG-regulating MS channels. This indicates an important role for LPA as an endogenous factor in fluid flow-induced endothelial function.
Assuntos
Cálcio/metabolismo , Endotélio Vascular/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Trifosfato de Adenosina/farmacologia , Compostos de Anilina/farmacologia , Animais , Cálcio/farmacologia , Bovinos , Células Cultivadas , Relação Dose-Resposta a Droga , Endotélio Vascular/citologia , Endotélio Vascular/metabolismo , Corantes Fluorescentes/farmacologia , Gadolínio/farmacologia , Microscopia Confocal , Estresse Mecânico , Tapsigargina/farmacologia , Fatores de Tempo , Xantenos/farmacologiaRESUMO
We have examined loss of heterozygosity on the short arm of chromosome 8 in 133 colorectal carcinomas, using 20 restriction fragment length polymorphism markers. Loss of heterozygosity was observed in 58 (44%) of 131 tumors that were informative with at least one locus. Among these 58, 32 revealed a partial or interstitial deletion of chromosome 8p. Detailed deletion mapping of chromosome 8p in these tumors identified two distinct, commonly deleted regions. One was located between markers C18-266 and pSVL-LPL at 8p23.2-8p22, and the other between CI8-319 and CI8-494 at 8p21.3-8p11.22. The genetic lengths of these two intervals were estimated to be 28 and 18 cM, respectively. The results suggest that at least two tumor suppressor genes associated with colorectal carcinomas are present on chromosome 8p. Correlation of loss of heterozygosity on 8p to the clinicopathological stage was also detected, suggesting that inactivation of a tumor suppressor gene(s) on 8p plays a role in progression of colorectal carcinomas.
Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 8 , Neoplasias Colorretais/genética , Genes Supressores de Tumor , Mapeamento Cromossômico , Heterozigoto , HumanosRESUMO
We have isolated a candidate tumor suppressor gene from a 600-kb region on chromosome 8p21.3-p22 that is commonly deleted in sporadic hepatocellular carcinomas (HCC), colorectal cancers (CRC), and non-small cell lung cancers (NSCLC). As this gene encodes a protein of 375 amino acids that bears significant sequence similarity to the extracellular (ligand-binding domain of platelet-derived growth factor receptor beta, we have designated it PRLTS (PDGF-receptor beta-like tumor suppressor). Structural rearrangement involving this gene was found in a sporadic NSCLC. In addition, somatic missense and frame-shift mutations were found in two HCCs and one CRC. These findings indicate that inactivation of the PRLTS gene may play a significant role in development of some carcinomas.
Assuntos
Cromossomos Humanos Par 8 , Genes Supressores de Tumor , Receptores do Fator de Crescimento Derivado de Plaquetas/genética , Sequência de Aminoácidos , Sequência de Bases , Carcinoma Hepatocelular/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Colorretais/genética , Humanos , Neoplasias Hepáticas/genética , Neoplasias Pulmonares/genética , Dados de Sequência Molecular , MutaçãoRESUMO
We investigated the effect of lysophosphatidic acid (LPA), a bioactive phospholipid, on the response in cytosolic free Ca2+ concentration ([Ca2+]i) to mechanical stress in cultured bovine lens epithelial cells. Spritzing of bath solution onto cells as mechanical stress caused marked increase in [Ca2+]i in the presence of LPA and this increase was concentration-dependent (1-10 microM), whereas neither addition of LPA alone nor the mechanical stress in the absence of LPA affected [Ca2+]i. The mechanical stress-induced increase in [Ca2+]i in the presence of LPA was inhibited by removing extracellular Ca2+ or by addition of Gd3+, a blocker of mechanosensitive cation channels, but not by nicardipine, thapsigargin, an inhibitor of endoplasmic reticulum-ATPase pump, or U73122, a phospholipase C inhibitor. These results show that LPA sensitises Ca2+ influx through cation-selective mechanosensitive channels, but does not sensitise Ca2+ release from intracellular stores, triggered by changes in mechanical stress. On the other hand, phosphatidic acid had less of a sensitising effect than LPA, and neither lysophosphatidylcholine nor chlorpromazine had any effect. Also Ca2+ mobilising agonists, ATP, histamine and carbachol, did not sensitise Ca2+ response to the mechanical stress. These results show that LPA sensitises mechanoreceptor-linked response in lens epithelial cells, suggesting that it plays a role in the development of cataracts due to increases in [Ca2+]i induced by mechanical stress.
Assuntos
Cálcio/metabolismo , Células Epiteliais/efeitos dos fármacos , Canais Iônicos/metabolismo , Lisofosfolipídeos/farmacologia , Animais , Bovinos , Células Cultivadas , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Cristalino/citologia , Estresse FisiológicoRESUMO
Mitochondrial free Ca2+ may regulate mitochondrial ATP production during cardiac exercise. Here, using laser scanning confocal microscopy of adult rabbit cardiac myocytes co-loaded with Fluo-3 to measure free Ca2+ and tetramethylrhodamine methylester to identify mitochondria, we measured cytosolic and mitochondrial Ca2+ transients during the contractile cycle. In resting cells, cytosolic and mitochondrial Fluo-3 signals were similar. During electrical pacing, transients of Fluo-3 fluorescence occurred in both the cytosolic and mitochondrial compartments. Both the mitochondrial and the cytosolic transients were potentiated by isoproterenol. These experiments show directly that mitochondrial free Ca2+ rises and falls during excitation-contraction coupling in cardiac myocytes and that changes of mitochondrial Ca2+ are kinetically competent to regulate mitochondrial metabolism on a beat-to-beat basis.
Assuntos
Cálcio/metabolismo , Mitocôndrias Cardíacas/metabolismo , Contração Miocárdica/fisiologia , Miocárdio/metabolismo , Compostos de Anilina , Animais , Células Cultivadas , Citosol/metabolismo , Estimulação Elétrica , Fluoresceínas , Corantes Fluorescentes , Microscopia Confocal , Miocárdio/química , Coelhos , Rodaminas , XantenosRESUMO
Several kinds of stress such as psychological stress, restraint, and foot shock inhibit feeding behavior through corticotropin-releasing factor (CRF). In contrast, a mild tail pinch increases food intake in rats. Although dopamine and opioid are thought to be involved in tail-pinch-induced food intake, it is unknown whether CRF participates in this phenomenon. Therefore, we attempted to clarify this issue using rats. A 30-s tail pinch increased food intake in 30 min after the tail pinch, and this increase was blocked by intraperitoneal injection of CRF receptor type 1 selective antagonist. CRF increased food intake in 30 min after intracerebroventricular injection at a dose of 2 or 10 ng, and this increase was also blocked by CRF receptor type 1 antagonist. Tail-pinch- or CRF-induced food intake was blocked by naloxone, pimozide, and spiperone. These results suggest that CRF, through CRF receptor type 1 as well as opioid and dopaminergic systems, are involved in the mechanism of tail-pinch-induced food intake. The results also suggest that brain CRF has dual effects on food intake, hyperphagia and anorexia, in a stress-dependent manner.
Assuntos
Hormônio Liberador da Corticotropina/farmacologia , Dopamina/fisiologia , Ingestão de Alimentos/fisiologia , Receptores Opioides/fisiologia , Estresse Fisiológico/fisiopatologia , Animais , Antagonistas de Dopamina/farmacologia , Ingestão de Alimentos/efeitos dos fármacos , Injeções Intraventriculares , Masculino , Naloxona/farmacologia , Antagonistas de Entorpecentes/farmacologia , Estimulação Física , Pimozida/farmacologia , Piridinas/farmacologia , Pirimidinas/farmacologia , Ratos , Ratos Wistar , Receptores de Hormônio Liberador da Corticotropina/antagonistas & inibidores , Receptores de Hormônio Liberador da Corticotropina/fisiologia , Espiperona/farmacologia , CaudaRESUMO
Changes in diacylglycerol kinase (DG kinase) activity in carbachol (CCh)-stimulated guinea pig taenia coli were investigated. In a mixed micellar assay system, added 1,2-dioctanoyl-sn-glycerol (diC8) and endogenous DG were competitively bound to common DG kinase isozymes from guinea pig taenia coli and phosphorylated, suggesting that diC8 is useful as a probe of agonist effects on DG kinase activity. In phosphorus-32 ([32P]Pi)- and diC8-prelabeled guinea pig taenia coli, diC8 was phosphorylated by DG kinase to [32P]dioctanoyl-phosphatidic acid ([32P]diC8-PA). CCh increased the accumulation of both [32P]diC8-PA and endogenous [32P]phosphatidic acid ([32P]PA) in a time- and dose-dependent manner (0.1-100 microM CCh). CCh-induced increases in [32P]diC8-PA and [32P]PA were inhibited by 1 microM atropine and 3 microM DG kinase inhibitor (R59022). These findings indicated the activation of DG kinase by muscarinic receptor stimulation in guinea pig taenia coli. Therefore, DG kinase activation may play an important role in CCh-induced PA formation. CCh-induced [32P]diC8-PA and [32P]PA accumulation was dependent on intracellular calcium concentrations. However, a KCl-induced increase in intracellular calcium, without receptor stimulation, was ineffective. Moreover, treatment with phorbol ester also increased accumulation of both PA species in KCl-treated tissues. These findings suggest that muscarinic receptor mediated activation of DG kinase may require both an increase in intracellular calcium and PKC activation in guinea pig taenia coli.
Assuntos
Carbacol/farmacologia , Colo/enzimologia , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Animais , Cálcio/metabolismo , Diacilglicerol Quinase , Diglicerídeos/metabolismo , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Cobaias , Ácidos Fosfatídicos/metabolismo , Fosforilação , Fosfotransferases (Aceptor do Grupo Álcool)/isolamento & purificação , Proteína Quinase C/metabolismo , Pirimidinonas/farmacologia , Tiazóis/farmacologiaRESUMO
Diacylglycerol kinase (DG kinase) is activated by various stimuli in many types of cells. We reported earlier that carbachol (CCh) induced DG kinase translocation from the cytosolic fraction to the membrane fraction in guinea pig taenia coli (Biochem. Pharmacol., 50: 591-599, 1995). In this study, the regulation mechanisms of DG kinase translocation are reported, based on the following findings: 1) CCh sustained an increase in DG kinase in the membrane fraction and a decrease in the cytosolic fraction; 2) blocking calcium influx by removing extracellular calcium did not affect the CCh-induced sustained DG kinase translocation; 3) exposing purified protein kinase C (PKC) to DG kinase increased DG kinase affinity to octylglycoside micelles only with the enzyme extracted from the cytosolic fraction; and 4) CCh-induced DG kinase translocation was reversed by removing CCh, and the serine/threonine phosphatase inhibitor, okadaic acid, blocked the reversal of the translocation. These results suggest that CCh-induced DG kinase translocation is promoted by both a transient increase in intracellular calcium, which may be released from the intracellular store, and by DG kinase phosphorylation by PKC.
Assuntos
Cálcio/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Proteína Quinase C/metabolismo , Animais , Carbacol/farmacologia , Colo/efeitos dos fármacos , Colo/metabolismo , Diacilglicerol Quinase , Ativação Enzimática , Cobaias , Técnicas In Vitro , FosforilaçãoRESUMO
The regulatory mechanisms of diacylglycerol (DG) kinase activity were studied in guinea pig taenia coli. In an octylglycoside mixed micellar assay system, DG kinase activities were distributed in both membrane and cytosolic fractions. Treatment of the tissue with carbachol (CCh) increased the activity in the membrane fraction and decreased the cytosolic fraction without affecting total DG kinase activity. The Km value of DG kinase in the membrane fraction was unchanged by treatment with CCh, although Vmax was increased. These findings suggest that DG kinase may be translocated from the cytosol to the membrane by CCh-stimulation. Increase in DG content by treatment of tissue with a cell-permeable species of DG, dioctanoyl-sn-glycerol, did not induce DG kinase translocation. Each treatment with protein kinase C (PKC) inhibitor and PKC-desensitization blocked CCh-induced DG kinase translocation; and phorbol ester induced the translocation only in intracellular calcium-accumulated tissues. Considering these results, CCh-induced DG kinase activation appears to involve DG kinase translocation from the cytosol to the membrane in association with both PKC and intracellular calcium concentration rather than cellular DG content.
Assuntos
Carbacol/farmacologia , Colo/efeitos dos fármacos , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Proteína Quinase C/metabolismo , Animais , Transporte Biológico , Colo/enzimologia , Colo/metabolismo , Diacilglicerol Quinase , Indução Enzimática , Feminino , Cobaias , Técnicas In Vitro , Masculino , Fosforilação , Frações Subcelulares/enzimologiaRESUMO
The effects of tri-n-butyltin chloride (TBT), an environmental pollutant, on the release of Ca(2+) from intracellular stores were investigated in isolated rat hepatocytes. Isolated hepatocytes permeabilized with digitonin were suspended in solution, and the concentration of extracellular Ca(2+) was measured, using a fluorescent Ca(2+) dye, fura-2. In the solution containing permeabilized hepatocytes that had been preincubated with 4.0 microM TBT for 30 min, the extracellular Ca(2+) concentration was high, but the inositol 1,4,5-trisphosphate (InsP(3))-induced increase in Ca(2+) concentration was suppressed, suggesting that the extracellular release of Ca(2+) in response to TBT treatment was from intracellular stores. Images of the Ca(2+) concentration in the intracellular stores of primary cultured hepatocytes loaded with fura-2 were obtained after digitonin-permeabilization, using digitalized fluorescence microscopy. The permeabilized hepatocytes that had been preincubated with 4.0 microM TBT for 30 min had a very low fura-2 fluorescence ratio (340/380 nm), suggesting that stored Ca(2+) was released. When the hepatocytes were treated with 4.0 microM TBT after digitonin-permeabilization, the decrease in the fura-2 fluorescence ratio was very small. However, when the permeabilized hepatocytes were incubated with 4.0 microM TBT and 2.0 microM NADPH, the decrease was enhanced, raising the possibility that TBT might be metabolized to the active form(s), thus releasing Ca(2+) from intracellular stores. When the hepatocytes were preincubated with 0.1 microM TBT for 30 min and then were permeabilized, the fura-2 fluorescence ratio was almost the same as that in the control permeabilized hepatocytes. However, the InsP(3)-induced decrease in the fluorescence ratio was suppressed significantly in the permeabilized hepatocytes. These results suggest that TBT released Ca(2+) from the intracellular stores at high concentrations, and suppressed the InsP(3)-induced Ca(2+) release at non-toxic low concentrations. It is probable that the latter effect was responsible for the previously reported suppression of Ca(2+) response induced by hormonal stimulations (Kawanish et al., Toxicol Appl Pharmacol 1999;155:54-61).
Assuntos
Cálcio/metabolismo , Hepatócitos/efeitos dos fármacos , Compostos de Trialquitina/farmacologia , Animais , Compartimento Celular , Permeabilidade da Membrana Celular , Células Cultivadas , Interações Medicamentosas , Fluorescência , Corantes Fluorescentes/metabolismo , Fura-2/metabolismo , Hepatócitos/metabolismo , Masculino , Fosfotransferases (Aceptor do Grupo Álcool)/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
During myocardial ischemia, a large reduction of tissue pH develops, and tissue pH returns to normal after reperfusion. In recent studies, we evaluated the role of pH in ischemia/reperfusion injury to cultured cardiac myocytes and perfused papillary muscles. Acidosis (pH < or = 7.0) protected profoundly against cell death during ischemia. However, the return from acidotic to normal pH after reperfusion caused myocytes to lose viability. This worsening of injury is a 'pH paradox' and was mediated by changes of intracellular pH (pH(i)), since manipulations that caused pH(i), to increase more rapidly after reperfusion accelerated cell killing, whereas manipulations that delayed the increase of pH(i) prevented loss of myocyte viability. Specifically, inhibition of the Na+/H+ exchanger with dimethylamiloride or HOE694 delayed the return of physiologic pH(i) after reperfusion and prevented reperfusion-induced cell killing to both cultured myocytes and perfused papillary muscle. Dimethylamiloride and HOE694 did not reduce intracellular free Ca2+ during reperfusion. By contrast, reperfusion with dichlorobenzamil, an inhibitor of Na+/Ca2+ exchange, decreased free Ca2+ but did not reduce cell killing. Thus, the pH paradox is not Ca(2+)-dependent. Our working hypothesis is that ischemia activates hydrolytic enzymes, such as phospholipases and proteases, whose activity is inhibited at acidotic pH. Upon reperfusion, the return to normal pH releases this inhibition and hydrolytic injury ensues. Increasing pH(i) may also induce a pH-dependent mitochondrial permeability transition and activate the myofibrillar ATPase, effects that increase ATP demand and compromise ATP supply. In conclusion, acidotic pH is generally protective in ischemia, whereas a return to physiologic pH precipitates lethal reperfusion injury to myocytes.
Assuntos
Traumatismo por Reperfusão Miocárdica/metabolismo , Miocárdio/metabolismo , Animais , Cálcio/metabolismo , Morte Celular , Hipóxia Celular , Humanos , Concentração de Íons de Hidrogênio , Líquido Intracelular/metabolismo , Modelos Cardiovasculares , Isquemia Miocárdica/terapia , Traumatismo por Reperfusão Miocárdica/etiologia , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/patologia , Músculos Papilares/metabolismo , Coelhos , RatosRESUMO
Myosin B was extracted from rabbit fast skeletal muscle. Caffeine (2-70 mM), theophylline (2-30 mM), and theobromine (2 mM) were examined for their effects on the rate and extent of myosin B superprecipitation at a low ionic strength (50 mM KCl) and a low concentration of ATP (40 microM) by the turbidimetric method. The rate of the superprecipitation was significantly (p less than 0.05 and p less than 0.01) reduced by 30-70 mM caffeine and 2-30 mM theophylline, while the extent was significantly (p less than 0.01) increased by 10-30 mM theophylline. The onset of the superprecipitation was also delayed and a clearing phase was even induced by 50-70 mM caffeine. Theobromine had no significant effect on the rate or extent of the superprecipitation at the concentration used. These results indicate that caffeine and theophylline are retardants of the myosin B superprecipitation reaction in the concentration ranges investigated.
Assuntos
Proteínas Musculares/isolamento & purificação , Músculos/análise , Miosinas/isolamento & purificação , Xantinas/farmacologia , Animais , Cafeína/farmacologia , Precipitação Química , Coelhos , Solubilidade , Teobromina/farmacologia , Teofilina/farmacologiaRESUMO
To investigate whether glucagon affects the xylitol-induced increase in the production of purine bases (hypoxanthine, xanthine, and uric acid), the present study was performed with five healthy subjects. Intravenous administration of 300 mL 10% xylitol increased the plasma concentration and urinary excretion of purine bases, erythrocyte concentrations of adenosine monophosphate (AMP) and adenosine diphosphate (ADP), and blood concentrations of glyceraldehyde-3-phosphate (GA3P) + dihydroxyacetone phosphate (DHAP), fructose-1,6-bisphosphate (FBP), and lactic acid; it decreased the blood concentration of pyruvic acid and the plasma concentration and urinary excretion of inorganic phosphate. However, intravenous administration of 1 mg glucagon together with xylitol reduced the xylitol-induced changes in oxypurines, pyruvic acid, GABP + DHAP, and FBP, whereas it promoted the xylitol-induced increase in the urinary excretion of total purine bases and did not affect the xylitol-induced increase in the plasma concentration of total purine bases. In addition, in vitro study demonstrated that sodium pyruvate prevented the xylitol-induced degradation of adenine nucleotides in erythrocytes. These results suggested that gluconeogenesis due to glucagon increased the production of pyruvic acid, accelerated the conversion of NADH to NAD, and thereby prevented both the xylitol-induced degradation of adenine nucleotides in organs similar to erythrocytes and the inhibition of xanthine dehydrogenase in the liver and small intestine, resulting in decreases in the plasma concentration and urinary excretion of oxypurines. However, it was also suggested that in the liver storing glycogen, glucagon-induced glycogenolysis accumulated sugar phosphates, resulting in purine degradation, since the xylitol-induced increase in the NADH/NAD ratio partially blocked glycolysis at the level of GABP dehydrogenase. Therefore, administration of glucagon together with xylitol may synergistically increase purine degradation more than xylitol alone, despite decreases in the plasma concentration and urinary excretion of oxypurines.
Assuntos
Glucagon/farmacologia , Purinas/metabolismo , Xilitol/farmacologia , Adulto , Glicemia/análise , Eritrócitos/metabolismo , Glucagon/sangue , Glicólise , Humanos , Masculino , Pessoa de Meia-Idade , NAD/metabolismo , Fosfatos/metabolismo , Ácido Úrico/metabolismo , Xilitol/sangueRESUMO
To determine the effect of amino acids on the plasma level and urinary excretion of uric acid and uridine, 200 mL 12% amino acid solution, and 2 weeks later, 100 mL physiological saline solution containing glucagon (1.2 microg/kg weight), was infused into five healthy men. Both increased the urinary excretion of uric acid and the concentration of glucagon, insulin, and glucose in plasma and pyruvic acid in blood, whereas they decreased the concentration of uridine and inorganic phosphate in plasma. However, neither the amino acid infusion nor glucagon infusion affected the concentration of purine bases (hypoxanthine, xanthine, and uric acid), cyclic adenosine monophosphate (cAMP) in plasma, or lactic acid in blood or the urinary excretion of oxypurines (hypoxanthine and xanthine), uridine, or sodium. These results suggest that glucagon may have an important role in the amino acid-induced increase in urinary excretion of uric acid and decrease in plasma uridine.
Assuntos
Aminoácidos/administração & dosagem , Glucagon/administração & dosagem , Ácido Úrico/sangue , Ácido Úrico/urina , Uridina/sangue , Uridina/urina , Adulto , Aminoácidos/metabolismo , Interações Medicamentosas , Glucagon/metabolismo , Humanos , Infusões Intravenosas , Masculino , Pessoa de Meia-IdadeRESUMO
To determine whether both ethanol and fructose increase the plasma concentration of uridine, we administered ethanol (0.6 g/kg) or fructose (1.0 g/kg) to seven normal subjects. Both ethanol and fructose increased the plasma concentration of uridine together with an increase in the plasma concentration of oxypurines, whereas fructose also increased the plasma concentration of uric acid, but ethanol did not. In ethanol ingestion and fructose infusion, an increase in the plasma concentration of purine bases correlated with that of uridine. These results strongly suggest that an increase in the plasma concentration of uridine is ascribable to increased pyrimidine degradation following purine degradation increased by ethanol and fructose.
Assuntos
Etanol/farmacologia , Frutose/farmacologia , Purinas/sangue , Uridina/sangue , Adulto , Consumo de Bebidas Alcoólicas , Etanol/sangue , Frutose/sangue , Humanos , Hipoxantina/sangue , Infusões Intravenosas , Lactatos/sangue , Masculino , Pessoa de Meia-Idade , Concentração Osmolar , Fosfatos/sangue , Piruvatos/sangue , Ácido Úrico/sangue , Xantina , Xantinas/sangueRESUMO
To examine whether bucladesine sodium affects the plasma concentrations of purine bases (hypoxanthine, xanthine, and uric acid) and uridine, 100 mL of physiological saline containing bucladesine sodium (6 mg/kg weight) was administered intravenously to eight healthy subjects for 1 hour after overnight fast except for water. Blood was drawn 30 minutes before, and 30 minutes and 1 hour after the beginning of the infusion, and 1-hour urine was collected before and after the beginning of the infusion. Two weeks later, 100 mL of only physiological saline was administered under the same protocol. Bucladesine sodium decreased the plasma concentrations of hypoxanthine by 36% and by 37%, and of xanthine by 16% and 33%, and of uridine by 17% and 30%, 30 minutes and 1 hour after the beginning of the infusion, respectively, and increased the urinary excretion of hypoxanthine and uric acid by 140% and 30%, respectively, after the beginning of the infusion. However, it did not affect the plasma concentration of uric acid or the urinary excretion of xanthine, and the urinary excretion of uridine was less than 0.2 micromol/h before or after bucladesine sodium infusion. On the other hand, physiological saline alone did not affect any of the values described. These results suggest that bucladesine sodium acts on the secretory process of the renal transport of hypoxanthine, resulting in the increased urinary excretion of hypoxanthine, and further suggest that bucladesine sodium enhances the uptake of uridine in plasma to liver cells.
Assuntos
Bucladesina/farmacologia , Hipoxantina/metabolismo , Ácido Úrico/metabolismo , Uridina/metabolismo , Xantina/metabolismo , Adulto , Glicemia/metabolismo , Glucagon/sangue , Humanos , Hipoxantina/sangue , Hipoxantina/urina , Insulina/sangue , Masculino , Pessoa de Meia-Idade , Ácido Úrico/sangue , Ácido Úrico/urina , Uridina/sangue , Uridina/urina , Xantina/sangue , Xantina/urinaRESUMO
To determine whether glucagon affects the plasma concentration of uridine, we administered 100 mL physiological saline containing 1 mg glucagon or 100 mL physiological saline alone intravenously over 1 hour to healthy subjects. Glucagon decreased the plasma concentration of uridine from 5.72 +/- 1.05 to 4.80 +/- 0.60 micromol/L but increased the concentrations of cyclic adenosine monophosphate (cAMP) in plasma and pyruvic acid and lactic acid in blood 59-, 1.4-, and 1.3-fold, respectively. Although glucagon increased urinary excretion of uric acid, it did not affect the plasma concentration of purine bases (hypoxanthine, xanthine, and uric acid) or urinary excretion of oxypurines and uridine, indicating that glucagon does not affect purine degradation and suggesting that glucagon does not affect adenosine triphosphate (ATP) consumption-induced pyrimidine degradation. In contrast, physiological saline did not affect any of the measured variables. These results suggest that glucagon enhanced Na+-dependent uridine uptake from the blood into the cells, since glucagon stimulates Na+-dependent uridine uptake into cells in vitro.
Assuntos
Glucagon/farmacologia , Uridina/sangue , Adulto , Glicemia/análise , AMP Cíclico/sangue , Glucagon/sangue , Humanos , Ácido Láctico/sangue , Masculino , Pessoa de Meia-Idade , Concentração Osmolar , Fosfatos/sangue , Purinas/sangue , Purinas/urina , Ácido Pirúvico/sangue , Uridina/urinaRESUMO
Urocortin (UCN), a member of the corticotropin-releasing factor (CRF) family, inhibits food intake when it is injected intracerebroventricularly in rats. To explore the site of action of UCN in feeding behavior, we examined the effects of injection of UCN into various hypothalamic nuclei on food and water intake in 24-h fasted rats. Injection of UCN into the ventromedial hypothalamic nucleus (VMH) significantly inhibited food and water intake over 3 h without sedative effect, but no significant effect was observed following injection either into the lateral hypothalamic area, or the paraventricular nucleus of the hypothalamus. To further explore the physiological significance of endogenous UCN of the VMH in feeding behavior, the effect of immunoneutralization of hypothalamic UCN on food intake was examined. Injection of anti-rat UCN rabbit gamma-globulin into the bilateral VMH in freely fed rats significantly potentiated food and water intake compared with rats that received normal rabbit gamma-globulin. These results suggest that endogenous UCN in the VMH exert inhibitory control on ingestive behavior.
Assuntos
Hormônio Liberador da Corticotropina/farmacologia , Ingestão de Líquidos/efeitos dos fármacos , Ingestão de Alimentos/efeitos dos fármacos , Comportamento Alimentar/efeitos dos fármacos , Núcleo Hipotalâmico Ventromedial/efeitos dos fármacos , Animais , Hormônio Liberador da Corticotropina/fisiologia , Ingestão de Líquidos/fisiologia , Ingestão de Alimentos/fisiologia , Comportamento Alimentar/fisiologia , Masculino , Coelhos , Ratos , Ratos Wistar , Urocortinas , Núcleo Hipotalâmico Ventromedial/fisiologia , gama-Globulinas/farmacologiaRESUMO
Stress shortens sodium pentobarbital (PbNa)-induced sleeping time through corticotropin-releasing hormone (CRH) in rats. We investigated whether this effect of brain CRH is mediated by CRH receptor type 1 (CRHR1) using a non-peptidic CRHR1 antagonist in rats. A 60 min period of restraint significantly shortened PbNa-induced sleeping time. This shortening was completely reversed by peripheral administration of CRHR1 antagonist. These results suggest that the stress-induced increase in arousal is mediated by CRHR1.
Assuntos
Hipnóticos e Sedativos/farmacologia , Pentobarbital/farmacologia , Piridinas/farmacologia , Pirimidinas/farmacologia , Receptores de Hormônio Liberador da Corticotropina/antagonistas & inibidores , Sono/efeitos dos fármacos , Estresse Fisiológico/fisiopatologia , Animais , Nível de Alerta/fisiologia , Masculino , Ratos , Ratos Wistar , Receptores de Hormônio Liberador da Corticotropina/fisiologia , Restrição Física , Sono/fisiologia , Estresse Fisiológico/etiologia , Fatores de TempoRESUMO
We previously reported that lysophosphatidic acid (LPA) sensitized mechanical stress-induced intracellular free Ca2+ concentration response (Biochem. Biophys. Res. Commun. 208, 19-25, 1995). In the present study, the signal transduction pathway of the sensitizing effect of LPA was investigated in cultured longitudinal muscle cells from guinea pig ileum. Suramin, a putative LPA receptor antagonist, did not affect the response in the presence of 30 nM LPA, suggesting that the response is induced via activation of suramin-insensitive LPA receptor. Neither pertussis toxin nor wortmannin inhibited the LPA-sensitized response, indicating that G(i/o)- and phosphatidylinositol 3-kinase (PI3-kinase)-mediated pathways are not involved in the sensitizing effect. C3 ADP ribosyltransferase had no effect on the response, whereas formation of actin-stress fiber in the presence of LPA was completely inhibited, suggesting rho-related cytoskeletal change is not involved in the response. In contrast, a phospholipase C (PLC) inhibitor, U73122, completely inhibited the response, but broad spectrum kinase inhibitors, staurosporine and H7, had no effect on the response. In addition, tyrosine kinase inhibitor, genistein, but not tyrphostin partially inhibited the response. These results suggest that LPA sensitizes the mechanical stress-induced response via activation of PLC, but not protein kinase C. Additionally, tyrphostin-insensitive tyrosine kinase, which is related to other pathway than G(i/o)- and rho-mediated pathways, may be involved in the response.