RESUMO
We studied the effects of vitamin A deficiency and repletion on rat insulin release and islet cellular retinol binding protein (CRBP) and cellular retinoic acid binding protein (CRABP). Biphasic insulin release from vitamin A-deficient perifused islets was markedly impaired. Release remained impaired with retinoic acid (RA) repletion, 2 micrograms/g diet compared to release from islets of rats repleted with retinol in the form of retinyl palmitate, 4 micrograms/g diet. Release normalized with RA, 8 micrograms/g diet. Vitamin A deficiency did not affect islet insulin content, cell size, number or structure. In vivo, vitamin A-deficient rats had impaired glucose-induced acute insulin release and glucose intolerance, which improved with repletion. Normal islets had greater concentrations of CRBP than CRABP; vitamin A deficiency reduced CRBP but not CRABP levels. We conclude retinol is required for normal insulin secretion. Retinoic acid may substitute for retinol in this function.
Assuntos
Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Deficiência de Vitamina A/metabolismo , Animais , Glicemia/metabolismo , Peso Corporal , Proteínas de Transporte/metabolismo , Ingestão de Energia , Técnicas In Vitro , Secreção de Insulina , Ilhotas Pancreáticas/patologia , Proteínas/metabolismo , Ratos , Receptores do Ácido Retinoico , Proteínas de Ligação ao Retinol/metabolismo , Proteínas Celulares de Ligação ao Retinol , Tretinoína/deficiência , Tretinoína/farmacologia , Vitamina A/farmacologiaRESUMO
Leptin is a peptide hormone produced by adipose tissue which acts centrally to decrease appetite and increase energy expenditure. Although leptin increases norepinephrine turnover in thermogenic tissues, the effects of leptin on directly measured sympathetic nerve activity to thermogenic and other tissues are not known. We examined the effects of intravenous leptin and vehicle on sympathetic nerve activity to brown adipose tissue, kidney, hindlimb, and adrenal gland in anesthetized Sprague-Dawley rats. Intravenous infusion of mouse leptin over 3 h (total dose 10-1,000 microg/kg) increased plasma concentrations of immunoreactive murine leptin up to 50-fold. Leptin slowly increased sympathetic nerve activity to brown adipose tissue (+286+/-64% at 1,000 microg/kg; P = 0.002). Surprisingly, leptin infusion also produced gradual increases in renal sympathetic nerve activity (+228+/-63% at 1,000 microg/kg; P = 0.0008). The effect of leptin on sympathetic nerve activity was dose dependent, with a threshold dose of 100 microg/kg. Leptin also increased sympathetic nerve activity to the hindlimb (+287+/-60%) and adrenal gland (388+/-171%). Despite the increase in overall sympathetic nerve activity, leptin did not increase arterial pressure or heart rate. Leptin did not change plasma glucose and insulin concentrations. Infusion of vehicle did not alter sympathetic nerve activity. Obese Zucker rats, known to possess a mutation in the gene for the leptin receptor, were resistant to the sympathoexcitatory effects of leptin, despite higher achieved plasma leptin concentrations. These data demonstrate that leptin increases thermogenic sympathetic nerve activity and reveal an unexpected stimulatory effect of leptin on overall sympathetic nerve traffic.
Assuntos
Proteínas/farmacologia , Sistema Nervoso Simpático/fisiologia , Potenciais de Ação/efeitos dos fármacos , Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Marrom/inervação , Glândulas Suprarrenais/efeitos dos fármacos , Glândulas Suprarrenais/inervação , Animais , Glicemia/metabolismo , Modelos Animais de Doenças , Hemodinâmica/efeitos dos fármacos , Membro Posterior/efeitos dos fármacos , Membro Posterior/inervação , Insulina/sangue , Rim/efeitos dos fármacos , Rim/inervação , Leptina , Masculino , Obesidade/genética , Proteínas/metabolismo , Ratos , Ratos Sprague-Dawley , Ratos Zucker , Sistema Nervoso Simpático/efeitos dos fármacosRESUMO
In our previous studies, perifused islets from vitamin D-deficient (D-def) rats showed marked impairment of glucose-induced biphasic release, accounted for at least in part by a decrease in food intake. In studies reported here, we test whether D-def rat islets have an impaired response to 5.6 mM glucose or tolbutamide, (T), and if so, whether this impairment is related to a decrease in food intake or a defect in islet calcium metabolism. We isolated islets of normal rats, D-def rats, and rats pair fed (PF) to D-def rats. Biphasic insulin release from perifused islets and net 45Ca retention in lot-incubated islets were measured in response to 5.6 mM glucose, 0.37 mM T, or both. Compared with secretion from normal islets, biphasic insulin release from islets of both D-def rats and PF rats was diminished by greater than 50% in response to 5.6 mM glucose alone or 5.6 mM glucose plus T. Insulin secretion was not significantly different between islets of D-def rats and islets of PF rats. In contrast, net calcium retention in islets of D-def rats was decreased to 68% of retention in islets of PF rats. However, net calcium retention in islets of both PF and D-def rats increased in response to T. The pair-feeding experiments suggest that the decrease in insulin release from islets of D-def rats is due to the decrease in food intake associated with the D-def state.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Cálcio/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Deficiência de Vitamina D/metabolismo , Animais , Glucose/farmacologia , Secreção de Insulina , Ilhotas Pancreáticas/efeitos dos fármacos , Ratos , Tolbutamida/farmacologiaRESUMO
The central role of mitochondria in most pathways leading to programmed cell death (PCD) has focused our investigations into the mechanisms of glucose-induced neuronal degeneration. It has been postulated that hyperglycemic neuronal injury results from mitochondria membrane hyperpolarization and reactive oxygen species formation. The present study not only provides further evidence to support our model of glucose-induced PCD but also demonstrates a potent ability for uncoupling proteins (UCPs) to prevent this process. Dorsal root ganglion (DRG) neurons were screened for UCP expression by Western blotting and immunocytochemistry. The abilities of individual UCPs to prevent hyperglycemic PCD were assessed by adenovirus-mediated overexpression of UCP1 and UCP3. Interestingly, UCP3 is expressed not only in muscle, but also in DRG neurons under control conditions. UCP3 expression is rapidly downregulated by hyperglycemia in diabetic rats and by high glucose in cultured neurons. Overexpression of UCPs prevents glucose-induced transient mitochondrial membrane hyperpolarization, reactive oxygen species formation, and induction of PCD. The loss of UCP3 in DRG neurons may represent a significant contributing factor in glucose-induced injury. Furthermore, the ability to prevent UCP3 downregulation or to reproduce the uncoupling response in DRG neurons constitutes promising novel approaches to avert diabetic complications such as neuropathy.
Assuntos
Apoptose/fisiologia , Gânglios Espinais/fisiologia , Glucose/farmacologia , Neurônios/fisiologia , Estresse Oxidativo/fisiologia , Animais , Apoptose/efeitos dos fármacos , Proteínas de Transporte/genética , Modelos Animais de Doenças , Gânglios Espinais/citologia , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/embriologia , Hiperglicemia/fisiopatologia , Canais Iônicos , Potenciais da Membrana/fisiologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Proteínas Mitocondriais , Neurônios/citologia , Neurônios/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Desacopladores , Proteína Desacopladora 3RESUMO
We studied the effects of retinoids on islet cell-to-cell adhesiveness and glucose-induced insulin release from rat islets. For adhesion studies, islets were dispersed using low concentrations of trypsin. Thirteen cis-retinoic acid (13 cis-RA) was added to a suspension of 15 X 10(5) islet cells and adhesion of cells was quantitated using a hemocytometer. For functional studies, we measured biphasic insulin release from collagenase-isolated perifused islets, dispersed cells, and single large aggregates (clumps) of islet cells. Thirteen cis-RA (10(-4) M) stimulated insulin secretion at 9.7, 12.5, 16.7, and 27.7 mM glucose. Maximal effects of 13 cis-RA (174% of control) were evident during second phase release at 9.7 mM glucose. Thirteen cis-RA (10(-7) and 10(-6) M) caused cells to adhere to each other, and at higher concentrations, 13 cis-RA caused dispersed cells to reaggregate into a single clump. These retinoid-induced clumps were perifused in a Bio-Gel P-2 gel column. Secretion from the clump was twofold greater than from an equal number of perifused dispersed cells. Electron microscopic and freeze-fracture examination of the clump showed reaggregated cells to be intact and the presence of gap junctions between cells. In conclusion, 13 cis-RA has marked effects on islet cell-to-cell adhesiveness. Trypsin-dispersed cells reaggregated by 13 cis-RA have anatomical contacts and secrete more insulin as an aggregate than as dispersed cells. Thirteen cis-RA increases insulin release possibly by increasing adhesion or interactions between beta-cells.
Assuntos
Adesão Celular/efeitos dos fármacos , Insulina/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Retinoides/farmacologia , Animais , Comunicação Celular/efeitos dos fármacos , Glucose/farmacologia , Técnicas In Vitro , Secreção de Insulina , Ilhotas Pancreáticas/ultraestrutura , Isotretinoína , Masculino , Ratos , Tretinoína/farmacologiaRESUMO
Adrenergic responsiveness to acute hypoglycemia is impaired after prior episodes of hypoglycemia. Although circulating epinephrine responses are blunted, associated alterations in adrenal sympathetic nerve activity (SNA) have not been reported. We examined adrenal nerve traffic in normal conscious rats exposed to acute insulin-induced hypoglycemia compared with insulin with (clamped) euglycemia. We also examined adrenal SNA and catecholamine responses to insulin-induced hypoglycemia in normal conscious rats after two antecedent episodes of hypoglycemia (days -2 and -1) compared with prior episodes of sham treatment. Acute insulin-induced hypoglycemia increased adrenal sympathetic nerve traffic compared with insulin administration with clamped euglycemia (165 +/- 12 vs. 118 +/- 21 spikes/s [P < 0.05]; or to 138 +/- 8 vs. 114 +/- 10% of baseline [P < 0.05]). In additional experiments, 2 days of antecedent hypoglycemia (days -2 and -1) compared with sham treatment significantly enhanced baseline adrenal SNA measured immediately before subsequent acute hypoglycemia on day 0 (180 +/- 11 vs. 130 +/- 12 spikes/s, respectively; P < 0.005) and during subsequent acute hypoglycemia (229 +/- 17 vs. 171 +/- 16 spikes/s; P < 0.05). However, antecedent hypoglycemia resulted in a nonsignificant reduction in hypoglycemic responsiveness of adrenal SNA when expressed as percent increase over baseline (127 +/- 5% vs. 140 +/- 14% of baseline). Antecedent hypoglycemia, compared with sham treatment, resulted in diminished epinephrine responsiveness to subsequent hypoglycemia. Norepinephrine responses to hypoglycemia were not significantly altered by antecedent hypoglycemia. In summary, prior hypoglycemia in normal rats increased adrenal sympathetic tone, but impaired epinephrine responsiveness to acute hypoglycemia. Hence, these data raise the intriguing possibility that increased sympathetic tone resulting from antecedent hypoglycemia downregulates subsequent epinephrine responsiveness to hypoglycemia. Alternatively, it is possible that the decrease in epinephrine responsiveness after antecedent hypoglycemia could be the result of reduced adrenal sympathetic nerve responsiveness.
Assuntos
Glândulas Suprarrenais/inervação , Epinefrina/sangue , Hipoglicemia/fisiopatologia , Insulina/farmacologia , Norepinefrina/sangue , Sistema Nervoso Simpático/fisiologia , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Pressão Sanguínea/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , Hipoglicemia/induzido quimicamente , Ratos , Ratos Sprague-Dawley , Sistema Nervoso Simpático/fisiopatologia , Fatores de TempoRESUMO
Analysis of glucose transporter mRNA levels in adipose tissue from streptozotocin (STZ)-induced diabetic rats demonstrated a specific decrease (10-fold) in adipose tissue GLUT-4 mRNA with no significant effect on GLUT-1 mRNA levels. Treatment of STZ-diabetic rats with twice daily injections of insulin for 1-3 days resulted in a 16-fold increase in the relative amount of GLUT-4 mRNA to levels approximately 2-fold greater than those in control animals. However, after 7 days of insulin therapy the amount of GLUT-4 mRNA decreased approximately 2-fold back to the levels in the control animals. Normalization of the STZ-induced serum hyperglycemia by phlorizin treatment, which inhibits renal tubular reabsorption of glucose, had no effect on GLUT-4 mRNA in the absence of insulin. Similar to STZ-diabetes, fasting for 48 h also reduced adipose GLUT-4 mRNA levels. Parenteral administration of insulin with glucose over 7.5 h, but not glucose alone, increased the levels of the GLUT-4 mRNA 3- to 4-fold. These studies demonstrate that the relative glycemic state does not influence GLUT-4 glucose transporter mRNA expression in vivo and strongly suggests that insulin is a major factor regulating the levels of GLUT-4 mRNA in adipose tissue.
Assuntos
Tecido Adiposo/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Insulina/farmacologia , Proteínas de Transporte de Monossacarídeos/biossíntese , Tecido Adiposo/metabolismo , Animais , Glicemia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Epididimo , Jejum , Hiperglicemia/metabolismo , Masculino , Proteínas de Transporte de Monossacarídeos/genética , Florizina/farmacologia , RNA Mensageiro/biossíntese , Ratos , Ratos Endogâmicos , Estimulação Química , EstreptozocinaRESUMO
We used a novel adaptation of the polymerase chain reaction to examine relative levels of mRNA encoding two members of the facilitative glucose transporter gene family, the GLUT1 or erythrocyte/HepG2/brain isoform and the GLUT4 or insulin-regulatable isoform. The method was fast (vs. hybridization methods), required no specific probe, and used total RNA samples of less than 1 microgram. Taking advantage of regions of structural similarity and differences between the two isoforms, we designed a single set of oligonucleotide primers capable of amplifying both GLUT1 and GLUT4 cDNAs such that their respective products could be resolved on the basis of a 12 base pair size differential. Hence, reverse transcription and complementary DNA amplification could be carried out for both transcripts using identical primers in the same reaction tube. Using this methodology, we examined the relative amounts of GLUT4 and GLUT1 mRNAs in several rat tissues. As expected based on prior reports using Northern analysis, rat brain contained only GLUT1 mRNA and skeletal muscle contained a large predominance of GLUT4 mRNA. Both isoform mRNAs were found in adipose tissue whereas adipose cells, heart and diaphragm contained predominantly GLUT4 mRNA. Induction of diabetes with streptozocin decreased the GLUT4 to GLUT1 ratio in adipose tissue 4-fold and 24 h of insulin treatment of the diabetic rats increased this ratio 9- to 10-fold. Insulin treatment of normal rats increased this ratio by 70%. Hindlimb skeletal muscle GLUT4 mRNA was quantified in diabetic and insulin-treated diabetic rats as a function of brain GLUT1 mRNA added as an internal standard. Using this methodology, no significant difference in muscle GLUT4 mRNA was noted as a result of 24 h of insulin therapy. In summary, quantitative PCR may be used to compare mRNA levels encoding specific members of a gene family either within given cells or tissues or as affected by physiological perturbations. Subject to certain limitations discussed within, this methodology may be useful in future measurements of glucose transporter mRNA, especially when only small tissue or cell samples are available.
Assuntos
Regulação da Expressão Gênica , Proteínas de Transporte de Monossacarídeos/genética , Reação em Cadeia da Polimerase , Sequência de Aminoácidos , Animais , DNA/genética , Diabetes Mellitus Experimental/metabolismo , Insulina/farmacologia , Masculino , Dados de Sequência Molecular , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos , Transcrição GênicaRESUMO
The brown adipose tissue uncoupling protein 1 (UCP1) catalyzes proton reentry without ATP synthesis, thereby dissipating energy as heat. In contrast, the function(s) of the recently described homologs, UCP2 and UCP3, are less clear. The aim of the present study was to determine whether overexpressed UCP subtypes affect mitochondrial respiration and substrate oxidation in cultured insulin-secreting INS-1 insulinoma cells. Adenoviral overexpression of UCP2 significantly decreased the ADP/O ratio by 31% and 39% in comparison to beta-galactosidase (beta-gal) or the mitochondrial protein manganese superoxide dismutase (MnSOD), respectively, and increased state 4 respiration in the presence of succinate and oligomycin by 52% and 59% in comparison to beta-gal or MnSOD, respectively. Adenoviral overexpression of UCP3 also decreased the ADP/O ratio by 18% (nonsignificant) and increased state 4 respiration by 24% (nonsignificant) in comparison to ss-gal and significantly decreased the ADP/O ratio by 32% and increased state 4 respiration by 35% in comparison to MnSOD. Both UCP2 and UCP3 expression significantly increased whole cell lipid oxidation by 34% (P < 0.01) and 30% (P < 0.05), respectively, compared with cells expressing Ad5CMVlacZ. However, glucose oxidation was not significantly altered by UCP2 or UCP3 expression. Adenoviral UCP2 expression, but not UCP3 (compared with beta-gal), significantly inhibited insulin secretion in the presence of 15 mM glucose [6.17 +/- 0.42 ng/mg cell protein for beta-gal compared with 4.69 +/- 0.39 for UCP2 (P < 0.05) and 5.51 +/- 0.50 for UCP3]. Both overexpressed UCPs significantly reduced INS-1 cell ATP content. Within certain limitations, which are discussed, these data are the first to demonstrate increased respiration and impaired coupling of oxidative phosphorylation as a result of UCP homolog expression in isolated mammalian mitochondria. Our results also suggest an important role for UCP in lipid metabolism and, possibly, insulin secretion.
Assuntos
Proteínas de Transporte/metabolismo , Proteínas de Membrana Transportadoras , Mitocôndrias/metabolismo , Proteínas Mitocondriais , Fosforilação Oxidativa , Consumo de Oxigênio , Proteínas/metabolismo , Desacopladores/metabolismo , Adenoviridae , Animais , Proteínas de Transporte/genética , Vetores Genéticos , Insulinoma , Canais Iônicos , Peroxidação de Lipídeos , Ácido Oleico/metabolismo , Proteínas/genética , Proteínas Recombinantes/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Transfecção , Células Tumorais Cultivadas , Proteína Desacopladora 2 , Proteína Desacopladora 3 , beta-Galactosidase/genética , beta-Galactosidase/metabolismoRESUMO
Leptin is believed to act through hypothalamic centers to decrease appetite and increase energy utilization, in part through enhanced thermogenesis. In this study, we examined the effects of fasting for 2 days and exogenous s.c. leptin, 200 microg every 8 h for 2 days, on the regulation of uncoupling protein (UCP) subtypes in brown adipose tissue (BAT) and gastrocnemius muscle. Northern blot analysis (UCP-1) and ribonuclease protection (UCP-2 and 3) were used for quantitative messenger RNA (mRNA) analysis, and specific antibodies were used to measure UCP-1 and UCP-3 total protein expression. Leptin, compared with vehicle, did not alter BAT UCP-1 or UCP-3 mRNA or protein expression when administered to normal ad libitum fed rats. Fasting significantly decreased BAT UCP-1 and UCP-3 mRNA expression, to 31% and 30% of ad libitum fed controls, respectively, effects which were prevented by administration of leptin to fasted rats. Fasting also significantly decreased BAT UCP-1 protein expression, to 67% of control; however, that effect was not prevented by leptin treatment. Fasting also decreased BAT UCP-3 protein, to 85% of control, an effect that was not statistically significant. Fasting, with or without leptin administration, did not affect BAT UCP-2 mRNA; however, leptin administration to ad libitum fed rats significantly increased BAT UCP-2 mRNA, to 138% of control. Fasting significantly enhanced gastrocnemius muscle UCP-3 mRNA (411% of control) and protein expression (168% of control), whereas leptin administration to fasted rats did not alter either of these effects. In summary, UCP subtype mRNA and protein are regulated in tissue- and subtype-specific fashion by leptin and food restriction. Under certain conditions, the effects of these perturbations on UCP mRNA and protein are discordant.
Assuntos
Tecido Adiposo/química , Jejum/fisiologia , Expressão Gênica , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Músculo Esquelético/química , Proteínas/farmacologia , Desacopladores/análise , Animais , Northern Blotting , Proteínas de Transporte/análise , Proteínas de Transporte/genética , Canais Iônicos , Leptina , Masculino , Proteínas de Membrana/análise , Proteínas de Membrana/genética , Proteínas/administração & dosagem , Proteínas/análise , Proteínas/genética , RNA Mensageiro/análise , Ratos , Ratos Sprague-Dawley , Proteína Desacopladora 1 , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
To determine whether leptin has insulin sensitizing effects in normal rodents, we measured plasma glucose and insulin concentrations in male Sprague-Dawley rats treated with leptin or vehicle by continuous s.c. infusion for 48 h. In additional experiments, we examined the acute effect of i.v. leptin upon insulin sensitivity under conditions of clamped glycemia. Subcutaneous leptin was administered at 10.0 and 1.0 microg/h. To avoid confounding effects of differences in food intake, both leptin- and vehicle-treated rats were fasted during the 48-h period of infusion. Infusion of leptin, 10 microg/h, significantly reduced both plasma glucose and insulin. Leptin, 1.0 microg/h, also decreased plasma glucose and insulin, although the effects on insulin did not achieve statistical significance. Leptin at either dose did not alter body weight or epididymal fat mass compared with vehicle treated controls. Leptin, 10 microg/h, decreased circulating insulin-like growth factor-1 levels. No differences in GLUT-4 content in either in brown or epididymal fat were observed as a result of leptin-treatment. Leptin, 10 microg/h, significantly decreased urine osmolality, increased water intake, and reduced renal potassium excretion compared with vehicle-infused rats. In additional rats, we measured the acute effect of i.v. leptin on insulin sensitivity determined as whole body glucose utilization during hyperinsulinemic glucose clamps performed at glucose targets of 60 and 90 mg/100 ml. Glucose utilization was increased by 29% during the last 135 min of glycemia clamped at 60 mg/100 ml (P < 0.05) and by 30% during the last 135 min of glycemia clamped at 90 mg/dl (P < 0.01) in rats infused with leptin compared with vehicle. In summary, leptin increased insulin sensitivity in normal rats both under fasting conditions and in the presence of hyperinsulinemia at clamped glucose. These effects did not appear dependent on altered body weight. Leptin also altered salt and water metabolism under fasting conditions resulting in increased water intake and more dilute urine.
Assuntos
Resistência à Insulina/fisiologia , Proteínas Musculares , Proteínas/farmacologia , Tecido Adiposo Marrom/química , Animais , Glicemia/análise , Glicemia/metabolismo , Peso Corporal/fisiologia , Epididimo/química , Glucose/metabolismo , Transportador de Glucose Tipo 4 , Hiperinsulinismo/sangue , Hiperinsulinismo/fisiopatologia , Bombas de Infusão Implantáveis , Insulina/sangue , Fator de Crescimento Insulin-Like I/análise , Fator de Crescimento Insulin-Like I/metabolismo , Leptina , Masculino , Proteínas de Transporte de Monossacarídeos/análise , Proteínas de Transporte de Monossacarídeos/metabolismo , Proteínas/administração & dosagem , Proteínas/metabolismo , Ratos , Ratos Sprague-Dawley , Fatores de Tempo , Equilíbrio HidroeletrolíticoRESUMO
To determine whether impaired insulin release from perifused rat islets of vitamin D-deficient (D-def) rats is a result of vitamin D-deficiency specifically or an associated decrease in food intake, we: 1) compared insulin release from islets of vitamin D-def rats with insulin release from islets of pair fed (pf) normal rats, and 2) measured the effects of 1,25(OH)2D3 treatment on food intake and insulin secretion from islets of D-def rats. Both vitamin D-def and pf normal rat islets showed significantly diminished insulin release in comparison with normal controls but were not different from each other. When D-def rats were repleted with 1,25(OH)2D3, food intake increased and insulin secretion improved during perifusion of rat islets. When D-def rats treated with 1,25(OH)2D3 were prevented from increasing their food intake in response to 1,25(OH)2D3 by pair feeding to a group of untreated D-def rats, insulin release from islets of treated rats was not significantly different from untreated D-def rats. To separate the effects of vitamin D deficiency from hypocalcemia, a group of vitamin D-def hypocalcemic rats was compared with a group of D-def normocalcemic rats. Normocalcemia did not reverse the defect in insulin release. In studies of cellular calcium uptake, both pf and D-def rat islets took up less calcium than normal islets but calcium uptake was not different between pf and D-def rat islets. Our studies suggest that vitamin D deficiency is associated with marked impairment of biphasic insulin release and that the decrease in food intake may account for this impairment at least in part.
Assuntos
Calcitriol/farmacologia , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Deficiência de Vitamina D/fisiopatologia , Animais , Cálcio/sangue , Cálcio/metabolismo , Ingestão de Alimentos/efeitos dos fármacos , Privação de Alimentos/fisiologia , Glucose/farmacologia , Técnicas In Vitro , Secreção de Insulina , Masculino , RatosRESUMO
Body weight is tightly regulated physiologically. The recent discovery of the peptide hormone leptin has permitted more detailed evaluation of the mechanisms responsible for control of body fat. Leptin is almost exclusively produced by adipose tissue and acts in the CNS through a specific receptor and multiple neuropeptide pathways to decrease appetite and increase energy expenditure. Leptin thus functions as the afferent component of a negative feedback mechanism to control adipose tissue mass. Increasing evidence suggests that leptin may have wider actions influencing autonomic, cardiovascular, and endocrine function. Intravenous leptin increases norepinephrine turnover and sympathetic nerve activity to thermogenic brown adipose tissue. Studies from our laboratory suggest that leptin also increases sympathetic nerve activity to kidney, hindlimb, and adrenal gland. However, systemic administration of leptin does not acutely increase arterial pressure or heart rate in anesthetized animals. Thus, longer-term exposure to hyperleptinemia may be necessary for full expression of the expected pressor effect of renal sympathoexcitation. Alternatively, leptin may have additional cardiovascular actions to oppose sympathetically mediated vasoconstriction. Leptin in high doses increases renal sodium and water excretion, apparently through a direct tubular action. In addition, leptin appears to increase systemic insulin sensitivity, even in the absence of weight loss. Although we are at an early stage of understanding, we speculate that abnormalities in the actions of leptin may have implications for the sympathetic, cardiovascular, and renal changes associated with obesity.
Assuntos
Pressão Sanguínea/efeitos dos fármacos , Rim/efeitos dos fármacos , Proteínas/farmacologia , Sistema Nervoso Simpático/efeitos dos fármacos , Tecido Adiposo/efeitos dos fármacos , Animais , Humanos , Insulina/metabolismo , Secreção de Insulina , Leptina , Obesidade/fisiopatologiaRESUMO
Leptin plays an important role in regulation of body weight through regulation of food intake and sympathetically mediated thermogenesis. The hypothalamic melanocortin system, via activation of the melanocortin-4 receptor (MC4-R), decreases appetite and weight, but its effects on sympathetic nerve activity (SNA) are unknown. In addition, it is not known whether sympathoactivation to leptin is mediated by the melanocortin system. We tested the interactions between these systems in regulation of brown adipose tissue (BAT) and renal and lumbar SNA in anesthetized Sprague-Dawley rats. Intracerebroventricular administration of the MC4-R agonist MT-II (200 to 600 pmol) produced a dose-dependent sympathoexcitation affecting BAT and renal and lumbar beds. This response was completely blocked by the MC4-R antagonist SHU9119 (30 pmol ICV). Administration of leptin (1000 microg/kg IV) slowly increased BAT SNA (baseline, 41+/-6 spikes/s; 6 hours, 196+/-28 spikes/s; P=0.001) and renal SNA (baseline, 116+/-16 spikes/s; 6 hours, 169+/-26 spikes/s; P=0.014). Intracerebroventricular administration of SHU9119 did not inhibit leptin-induced BAT sympathoexcitation (baseline, 35+/-7 spikes/s; 6 hours, 158+/-34 spikes/s; P=0.71 versus leptin alone). However, renal sympathoexcitation to leptin was completely blocked by SHU9119 (baseline, 142+/-17 spikes/s; 6 hours, 146+/-25 spikes/s; P=0.007 versus leptin alone). This study demonstrates that the hypothalamic melanocortin system can act to increase sympathetic nerve traffic to thermogenic BAT and other tissues. Our data also suggest that leptin increases renal SNA through activation of hypothalamic melanocortin receptors. In contrast, sympathoactivation to thermogenic BAT by leptin appears to be independent of the melanocortin system.
Assuntos
Tecido Adiposo Marrom/inervação , Ventrículos Cerebrais/fisiologia , Rim/inervação , Proteínas/farmacologia , Receptores da Corticotropina/fisiologia , Sistema Nervoso Simpático/fisiologia , Animais , Ventrículos Cerebrais/efeitos dos fármacos , Infusões Intravenosas , Injeções Intraventriculares , Leptina , Masculino , Hormônios Estimuladores de Melanócitos/farmacologia , Proteínas/administração & dosagem , Ratos , Ratos Sprague-Dawley , Receptor Tipo 4 de Melanocortina , Receptores da Corticotropina/agonistas , Receptores da Corticotropina/antagonistas & inibidores , Proteínas Recombinantes/farmacologia , Sistema Nervoso Simpático/efeitos dos fármacosRESUMO
OBJECTIVE: Administration of leptin to animals increases sympathetic nerve activity and heart rate. We therefore tested the hypothesis that plasma leptin is linked independently to muscle sympathetic nerve activity (MSNA) and heart rate in healthy humans. METHODS: We measured plasma leptin, plasma insulin, body mass index (BMI), percent body fat, waist: hip ratio, MSNA, heart rate and blood pressure in 88 healthy individuals (50 men and 38 women). RESULTS: In men, plasma leptin concentration correlated significantly with BMI (r = 0.75, P < 0.001), percent body fat (r = 0.70, P< 0.001), waist: hip ratio (r = 0.69, P < 0.001), insulin (r = 0.37, P = 0.009), and age (r = 0.38, P = 0.006). Only BMI and waist: hip ratio were linked independently to plasma leptin concentration (r = 0.78, P < 0.001). Plasma leptin concentrations also correlated with heart rate (r = 0.39, P = 0.006) and mean arterial pressure (MAP; r = 0.38, P = 0.007), but not with MSNA (r = 0.17, P = 0.24). After adjustment for BMI and waist: hip ratio, plasma leptin concentration correlated significantly only with heart rate (r = 0.29, P = 0.04), and not with MAP (r = 0.21, P = 0.14). Individuals were divided into high-leptin and low-leptin subgroups on the basis of plasma leptin concentrations adjusted for BMI and waist: hip ratio. Those with high leptin concentrations had significantly faster heart rates than those with low leptin. MAP and MSNA were similar in both subgroups. No relationship between leptin and either heart rate or MSNA was evident in women. CONCLUSIONS: In normal men, heart rate, but not MSNA, is linked to plasma leptin concentration. This sex-specific relationship between heart rate and plasma leptin is independent of plasma insulin, BMI, waist:hip ratio and percentage body fat.
Assuntos
Frequência Cardíaca/fisiologia , Leptina/fisiologia , Sistema Nervoso Simpático/fisiologia , Adulto , Animais , Pressão Sanguínea/fisiologia , Constituição Corporal , Índice de Massa Corporal , Feminino , Humanos , Insulina/sangue , Leptina/sangue , Masculino , Pessoa de Meia-Idade , Músculos/inervação , Caracteres SexuaisRESUMO
OBJECTIVE: To examine the influence of genetic factors on plasma leptin levels. SUBJECTS AND METHODS: We measured plasma leptin levels, body mass index and body fat distribution in healthy young female monozygotic (n = 19) and dizygotic (n = 14) twins. The twin zygosity was verified by determination of short tandem repeat and amplified fragment length polymorphism systems. The genetic analysis included analysis of variance-based and maximum likelihood-based methods. RESULTS: Plasma leptin levels were correlated significantly with body mass index (r = 0.59, P < 0.001), waist circumference (r = 0.54, P < 0.001) and hip circumference (r = 0.63, P < 0.001), but not with age (r = -0.17) or the waist:hip ratio (r = 0.02). The heritability estimates derived from intraclass correlations were significant for body mass index (P = 0.001), waist circumference (P = 0.004), hip circumference (P = 0.01) and plasma leptin levels (P = 0.005), but not for the waist:hip ratio (P = 0.22). In the maximum likelihood-based path analysis, heritability was estimated at 79% for body mass index and at 73% for plasma leptin levels. After adjustment for body mass index, the heritability estimate for leptin levels from the model-fitting approach was 55%. CONCLUSIONS: Genetic factors are major determinants of plasma leptin levels in humans and may account for as much as half of the variance in leptin levels.
Assuntos
Ligação Genética , Proteínas/metabolismo , Gêmeos Dizigóticos/genética , Gêmeos Monozigóticos/genética , Tecido Adiposo/fisiologia , Adolescente , Adulto , Constituição Corporal , Índice de Massa Corporal , Feminino , Humanos , Leptina , Obesidade/sangue , Obesidade/genética , Polimorfismo de Fragmento de Restrição , Valores de ReferênciaRESUMO
Leptin is a protein produced by adipocytes. Leptin is known to markedly and rapidly increase sympathetic nerve activity to the kidney and hindlimb of experimental animals. Recent studies suggest that leptin may stimulate endothelial production of nitric oxide, which could oppose sympathetically induced vasoconstriction. We tested the hypothesis that such actions of leptin may produce peripheral functional sympatholysis. In Sprague-Dawley rats, we intermittently stimulated the abdominal sympathetic trunk and measured renal and hindlimb blood flows before and after 3 h of infusion of leptin (1000 microg/kg, n = 7) or vehicle (n = 7). Leptin did not change arterial pressure, heart rate, or renal or hindlimb conductance over the course of 3 h. In addition, leptin did not significantly alter sympathetically mediated vasomotor responses to electrical stimulation, as compared with vehicle. Thus, we conclude that leptin does not change regional blood flows, and that leptin also does not appear to have vascular or neural actions to cause peripheral functional sympatholysis.
Assuntos
Leptina/farmacologia , Sistema Nervoso Simpático/efeitos dos fármacos , Animais , Pressão Sanguínea/efeitos dos fármacos , Estimulação Elétrica , Frequência Cardíaca/efeitos dos fármacos , Membro Posterior/irrigação sanguínea , Masculino , Ratos , Ratos Sprague-Dawley , Fluxo Sanguíneo Regional/efeitos dos fármacos , Nervos Esplâncnicos/efeitos dos fármacos , Nervos Esplâncnicos/fisiologia , Sistema Nervoso Simpático/fisiologia , Vasodilatação/efeitos dos fármacosRESUMO
Streptozocin (STZ) administration (125 mg/kg) to normal rats resulted in a rapid (24-hour) decrease in circulating insulin levels, marked hyperglycemia, and weight loss. Adipose tissue glucose transporter 4 (GLUT4) mRNA levels decreased approximately eightfold, whereas GLUT4 protein levels were unchanged. However, GLUT4 protein levels decreased approximately 30% by 48 hours and fivefold by 72 hours of insulin deficiency. Although GLUT4 mRNA levels were rapidly restored by insulin therapy (twofold above control levels within 12 hours), GLUT4 protein levels increased only gradually, reaching peak values of 1.5-fold control levels following 7 to 10 days of insulin treatment. Insulin treatment in normal rats increased adipose GLUT4 mRNA levels nearly 100% by 24 hours, while GLUT4 protein levels increased in a more gradual fashion. The delay in GLUT4 protein induction relative to its mRNA was shorter in normal rats treated with insulin than in insulin-treated diabetic rats. These data demonstrate that insulin-induced changes in adipose GLUT4 protein are considerably delayed relative to its mRNA, and that the diabetic state enhances this difference. The known in vivo time-dependent effects of insulin treatment on adipocyte glucose transport activity can be at least partly explained by altered specific expression of GLUT4 protein.
Assuntos
Insulina/farmacologia , Proteínas de Transporte de Monossacarídeos/genética , Animais , Diabetes Mellitus Experimental , Masculino , Proteínas de Transporte de Monossacarídeos/efeitos dos fármacos , RNA Mensageiro/análise , Ratos , Ratos Sprague-Dawley , Estreptozocina , Fatores de TempoRESUMO
Adipose tissue leptin mRNA levels are decreased by food deprivation or induction of insulin-deficient diabetes. To determine whether plasma leptin concentrations are similarly affected, whether treatment of diabetes with insulin restores plasma leptin, and whether this requires restoration of body weight (lost as a result of diabetes) and/or normalization of glycemia, we measured plasma leptin concentrations in control, untreated streptozotocin (STZ)-diabetic, and insulin-treated STZ-diabetic rats. Plasma leptin was markedly reduced in untreated STZ-diabetic rats. Insulin treatment for 4 to 17 days increased plasma leptin approximately twofold above control levels. However, despite the hyperleptinemia, insulin-treated diabetic rats gained weight at a rate equal to that of sham-treated controls. Epididymal adipose tissue leptin mRNA levels in 17-day insulin-treated diabetic rats were equal to but did not exceed sham-control levels, unlike plasma leptin. Plasma glucose concentrations in insulin-treated STZ-diabetic rats were lower than in sham controls. Therefore, to determine whether hypoglycemia may be important in increasing plasma leptin, we measured plasma leptin levels in diabetic rats infused with insulin for 3 hours along with a variable-rate glucose infusion targeting glycemia to 200 or 40 mg/100 mL. Plasma leptin rapidly increased in these rats irrespective of target glycemia. Plasma leptin also increased rapidly in normal rats infused with insulin and glucose (target glycemia, 200 mg/100 mL). We conclude that plasma leptin concentrations are markedly reduced under conditions of insulin deficiency and rapidly increased by insulin treatment. The increase in plasma leptin does not require restoration of body weight and, under glucose clamp conditions, does not depend on target glycemia. Hyperleptinemia in insulin-treated diabetic rats is not explained on the basis of steady-state leptin mRNA levels, at least as reflected in epididymal fat.
Assuntos
Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus/sangue , Hipoglicemiantes/uso terapêutico , Insulina/uso terapêutico , Obesidade , Proteínas/metabolismo , Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/metabolismo , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Peso Corporal/efeitos dos fármacos , Peso Corporal/fisiologia , Epididimo/química , Epididimo/efeitos dos fármacos , Epididimo/metabolismo , Glucose/administração & dosagem , Hipoglicemiantes/administração & dosagem , Insulina/administração & dosagem , Leptina , Masculino , Proteínas/efeitos dos fármacos , Proteínas/genética , RNA Mensageiro/análise , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Estreptozocina/administração & dosagemRESUMO
We studied the immunogenicity of human insulin in 11 diabetic mothers and their newborns. Serum antibody formation was assayed by two different methods. Upon switching four patients from beef/pork insulin to human insulin, we found that elevated baseline antibody levels in three women decreased, in two to undetectable levels at term. The fourth patient had undetectable antibody levels at baseline and borderline levels at term. Only one of their four newborns had antibodies. Upon initiation of insulin treatment in another five diabetics without detectable antibodies at baseline, only two developed antibodies, and only one of their newborns developed antibodies. Two other patients, initially not on insulin, had baseline elevations of antibody that decreased with administration of human insulin; both of their newborns had antibodies. Overt diabetes evolved subsequently in both mothers after pregnancy. We conclude the following: 1) Upon transfer from beef/pork insulin to human insulin, mothers and their newborns show a decrease in insulin antibodies; 2) new patients initiated on insulin develop low levels of antibodies, if any, and their newborns also have low levels of antibodies if any; and 3) the decreased or absent immunogenicity of human insulin supports its use in pregnant diabetics.