RESUMO
Among the strategies for bioavailability improvement of poorly soluble drugs, co-amorphous systems have revealed to have a significant impact in the increase of the aqueous solubility of the drug, and at the same time increasing the amorphous state stability and dissolution rate when compared with the neat drug. Tolbutamide (TBM) is an oral hypoglycemic drug largely used in the treatment of type II Mellitus diabetes. TBM is a class II drug according to the Biopharmaceutical Classification System, meaning that it has low solubility and higher permeability. The aim of this study was to synthesize a co-amorphous material of tolbutamide (TBM) with tromethamine (TRIS). Density functional theory (DFT), allowed to study the structural, electronic, and thermodynamic properties, as well as solvation effects. In same theory level, several interactions tests were performed to obtain the most thermodynamically favorable drug-coformer intermolecular interactions. The vibrational spectra (mid infrared and Raman spectroscopy) are in accordance with the theoretical studies, showing that the main molecular interactions are due to the carbonyl, sulfonyl, and amide groups of TMB and the alcohol and amine groups of TRIS. X-ray powder diffraction was used to study the physical stability in dry condition at 25 °C of the co-amorphous system, indicating that the material remained in an amorphous state up to 90 days. Differential scanning calorimetry and thermogravimetric results showed a high increase of the Tg when compared with the amorphous neat drug, from 4.3 °C to 83.7 °C, which generally translated into good physical stability. Solubility studies demonstrated an increase in the solubility of TBM by 2.5 fold when compared with its crystalline counterpart.
Assuntos
Diabetes Mellitus , Tolbutamida , Varredura Diferencial de Calorimetria , Sistemas de Liberação de Medicamentos , Estabilidade de Medicamentos , Humanos , Solubilidade , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios XRESUMO
Insulin resistance during pregnancy is counteracted by enhanced insulin secretion. This condition is aggravated by obesity, which increases the risk of gestational diabetes. Therefore, pancreatic islet functionality was investigated in control nonpregnant (C) and pregnant (CP), and cafeteria diet-fed nonpregnant (Caf), and pregnant (CafP) obese rats. Isolated islets were used for measurements of insulin secretion (RIA), NAD(P)H production (MTS), glucose oxidation ((14)CO(2) production), intracellular Ca(2+) levels (fura-2 AM), and gene expression (real-time PCR). Impaired glucose tolerance was clearly established in Caf and CafP rats at the 14th wk on a diet. Insulin secretion induced by direct depolarizing agents such as KCl and tolbutamide and increasing concentrations of glucose was significantly reduced in Caf, compared with C islets. This reduction was not observed in islets from CP and CafP rats. Accordingly, the glucose oxidation and production of reduced equivalents were increased in CafP islets. The glucose-induced Ca(2+) increase was significantly lower in Caf and higher in CafP, compared with all other groups. CP and CafP islets demonstrated an increased Ca(2+) oscillation frequency, compared with both C and Caf islets, and the amplitude of oscillations was augmented in CafP, compared with Caf islets. In addition, Ca(v)alpha1.2 and SERCA2a mRNA levels were reduced in Caf islets. Ca(v)alpha1.2, but not SERCA2a, mRNA was normalized in CafP islets. In conclusion, cafeteria diet-induced obesity impairs insulin secretion. This alteration is related to the impairment of Ca(2+) handling in pancreatic islets, in especial Ca(2+) influx, a defect that is reversed during pregnancy allowing normalization of insulin secretion.
Assuntos
Dieta , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Obesidade/metabolismo , Prenhez/fisiologia , Animais , Glicemia/metabolismo , Peso Corporal/fisiologia , Cálcio/metabolismo , Canais de Cálcio/biossíntese , Canais de Cálcio/genética , DNA/biossíntese , DNA/genética , Feminino , Expressão Gênica/fisiologia , Teste de Tolerância a Glucose , Homeostase/fisiologia , Hipoglicemiantes/farmacologia , Lipídeos/sangue , Obesidade/etiologia , Tamanho do Órgão/fisiologia , Oxirredução , Cloreto de Potássio/farmacologia , Gravidez , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/biossíntese , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/genética , Tolbutamida/farmacologiaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Tecoma stans aqueous extract (TAE) is widely used as a traditional antidiabetic remedy in Mexico; its rational use is controversial. We provide evidence of its main antidiabetic activities. AIM OF THE STUDY: To evaluate in vivo and in vitro intestinal alpha-glucosidases inhibition as the possible mode of action of TAE on type 2 diabetes mellitus (DM2) animal models, and to test the effects of its sub-chronic administration on lipids and glucose blood levels. MATERIALS AND METHODS: In healthy and streptozotocin (STZ)-induced diabetic male Sprague-Dawley rats, glucose or cornstarch was administered after an oral dose of TAE, acarbose, tolbutamide or vehicle, in order to build starch and glucose tolerance curves (STC and GTC). An intestinal brush border preparation was used to evaluate the TAE alpha-glucosidases inhibitory activity. Moreover, in STZ-induced diabetic rats TAE, tolbutamide or vehicle was administered for 21 days for evaluate their effects on fasting glucose cholesterol and triglycerides. Also, TAE total phenolic compounds were quantified. RESULTS: In STC, TAE decreased hyperglycemic peak values in both healthy and STZ-treated rats, in a magnitude similar to that of acarbose. The in vitro preparation showed a dose-dependent inhibition of glucose release from starch. Sub-chronic administration of TAE significantly reduced cholesterol and triglycerides levels. Moreover, we confirmed that acute and sub-chronic administration of TAE (500mg/kg) in both rat models did not diminish fasting glucose and did not modify the GTC. CONCLUSIONS: The study present evidence that the main antidiabetic effect of TAE is due to intestinal alpha-glucosidase inhibition by decreasing the postprandial hyper-glycaemia peak; in addition, TAE sub-chronic administration reduces triglycerides and cholesterol, without modifying fasting glucose.
Assuntos
Bignoniaceae , Colesterol/sangue , Diabetes Mellitus Experimental/tratamento farmacológico , Hipoglicemiantes/uso terapêutico , Fitoterapia , Extratos Vegetais/uso terapêutico , Triglicerídeos/sangue , Acarbose/farmacologia , Animais , Bignoniaceae/química , Glicemia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Feminino , Teste de Tolerância a Glucose , Inibidores de Glicosídeo Hidrolases , Hipoglicemiantes/farmacologia , Mucosa Intestinal/metabolismo , Intestinos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fenóis/análise , Extratos Vegetais/farmacologia , Ratos , Ratos Sprague-Dawley , Amido , Tolbutamida/farmacologiaRESUMO
Islet Neogenesis Associated Protein (INGAP) increases pancreatic beta-cell mass and potentiates glucose-induced insulin secretion. Here, we investigated the effects of the pentadecapeptide INGAP-PP in adult cultured rat islets upon the expression of proteins constitutive of the K(+)(ATP) channel, Ca(2+) handling, and insulin secretion. The islets were cultured in RPMI medium with or without INGAP-PP for four days. Thereafter, gene (RT-PCR) and protein expression (Western blotting) of Foxa2, SUR1 and Kir6.2, cytoplasmic Ca(2+) ([Ca(2+)](i)), static and dynamic insulin secretion, and (86)Rb efflux were measured. INGAP-PP increased the expression levels of Kir6.2, SUR1 and Foxa2 genes, and SUR1 and Foxa2 proteins. INGAP-PP cultured islets released significantly more insulin in response to 40 mM KCl and 100 muM tolbutamide. INGAP-PP shifted to the left the dose-response curve of insulin secretion to increasing concentrations of glucose (EC(50) of 10.0+/-0.4 vs. 13.7+/-1.5 mM glucose of the controls). It also increased the first phase of insulin secretion elicited by either 22.2 mM glucose or 100 microM tolbutamide and accelerated the velocity of glucose-induced reduction of (86)Rb efflux in perifused islets. These effects were accompanied by a significant increase in [Ca(2+)](i) and the maintenance of a considerable degree of [Ca(2+)](i) oscillations. These results confirm that the enhancing effect of INGAP-PP upon insulin release, elicited by different secretagogues, is due to an improvement of the secretory function in cultured islets. Such improvement is due, at least partly, to an increased K(+)(ATP) channel protein expression and/or changing in the kinetic properties of these channels and augmented [Ca(2+)](i) response. Accordingly, INGAP-PP could potentially be used to maintain the functional integrity of cultured islets and eventually, for the prevention and treatment of diabetes.
Assuntos
Cálcio/metabolismo , Citocinas/farmacologia , Ilhotas Pancreáticas/efeitos dos fármacos , Canais KATP/metabolismo , Fragmentos de Peptídeos/farmacologia , Animais , Western Blotting , Expressão Gênica/efeitos dos fármacos , Glucose/farmacologia , Fator 3-beta Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Canais KATP/genética , Proteínas Associadas a Pancreatite , Canais de Potássio Corretores do Fluxo de Internalização/genética , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Cloreto de Potássio/farmacologia , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Técnicas de Cultura de Tecidos , Tolbutamida/farmacologiaRESUMO
Neonatal diabetes is a rare condition characterized by hyperglycemia, requiring insulin treatment, diagnosed within the first months of life. The disorder may be either transient, resolving in infancy or early childhood with possible relapse later, or permanent in which case lifelong treatment is necessary. Both conditions are genetically heterogeneous; however, the majority of the cases of transient neonatal diabetes are due to abnormalities of an imprinted region of chromosome 6q24. For permanent neonatal diabetes, the most common causes are heterozygous activating mutations of KCNJ11, the gene encoding the Kir6.2 sub-unit of the ATP-sensitive potassium channel. In this article we discuss the clinical features of neonatal diabetes, the underlying genetic defects and the therapeutic implications.
Assuntos
Diabetes Mellitus/genética , Mutação , Diabetes Mellitus/tratamento farmacológico , Proteínas de Homeodomínio/genética , Humanos , Hipoglicemiantes/uso terapêutico , Recém-Nascido , Insulina/genética , Insulina/uso terapêutico , Canais KATP/genética , Canais de Potássio Corretores do Fluxo de Internalização/genética , Canais de Potássio Corretores do Fluxo de Internalização/uso terapêutico , Tolbutamida/uso terapêutico , Transativadores/genéticaRESUMO
O diabetes neonatal (DN) é uma condição rara caracterizada por hiperglicemia, que necessita de tratamento com insulina, diagnosticado nos primeiros meses de vida. Clinicamente pode ser classificado em DN transitório quando ocorre remissão da doença em poucos meses, podendo haver recorrência posterior; ou permanente quando, como o nome indica, não ocorre remissão. Ambas as condições são geneticamente heterogêneas; entretanto a maioria dos casos de DN transitório é decorrente de anormalidades da região de imprinted no cromossomo 6q24. Mutações ativadoras em heterozigose no gene KCNJ11, que codifica a subunidade Kir6.2 do canal de potássio ATP-sensível, são a causa mais comum de DN permanente. No presente artigo, discutimos as características clínicas do DN, os mecanismos moleculares envolvidos e suas implicações terapêuticas.
Neonatal diabetes is a rare condition characterized by hyperglycemia, requiring insulin treatment, diagnosed within the first months of life. The disorder may be either transient, resolving in infancy or early childhood with possible relapse later, or permanent in which case lifelong treatment is necessary. Both conditions are genetically heterogeneous; however, the majority of the cases of transient neonatal diabetes are due to abnormalities of an imprinted region of chromosome 6q24. For permanent neonatal diabetes, the most common causes are heterozygous activating mutations of KCNJ11, the gene encoding the Kir6.2 sub-unit of the ATP-sensitive potassium channel. In this article we discuss the clinical features of neonatal diabetes, the underlying genetic defects and the therapeutic implications.
Assuntos
Humanos , Recém-Nascido , Diabetes Mellitus/genética , Mutação , Diabetes Mellitus/tratamento farmacológico , Proteínas de Homeodomínio/genética , Hipoglicemiantes/uso terapêutico , Insulina/genética , Insulina/uso terapêutico , Canais KATP/genética , Canais de Potássio Corretores do Fluxo de Internalização/genética , Canais de Potássio Corretores do Fluxo de Internalização/uso terapêutico , Tolbutamida/uso terapêutico , Transativadores/genéticaRESUMO
A series of chalcone derivatives from 3,4-methylenedioxybenzaldehyde and substituted acetophenones have been synthesized and investigated as antihyperglycemic agents in a glucose loaded animal model. Chalcones with biological activity were compared with lispro, regular insulin and tolbutamide effects on serum glucose levels. Compound 01, without substituent in the A-ring was not able to change glycemic levels. On the other hand, compounds 03, 04, 05, 09 and 10 with substitutions at position 3' and/or 4' in the A-ring caused significant reduction in serum glucose levels. Concerning the antihyperglycemic effect, compounds 03 and 05 (methoxy substituent) inhibited the hyperglycemia induced by glucose around 96% similar to that demonstrated for lispro insulin and tolbutamide at 60 min. A rapid and lasting antihyperglycemic effect was found with compound 09 and 10 (nitro substituent). In conclusion, besides the nature of the functional groups electron-donor substituent, as methoxy and hydroxyl or electron-acceptor, as nitro groups, the position of the group may be mandatory for biological activity.
Assuntos
Glicemia/efeitos dos fármacos , Chalcona/análogos & derivados , Chalcona/administração & dosagem , Hiperglicemia/tratamento farmacológico , Hipoglicemiantes/administração & dosagem , Administração Oral , Animais , Glicemia/análise , Chalcona/síntese química , Chalcona/química , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Glucose/administração & dosagem , Hiperglicemia/sangue , Hipoglicemiantes/síntese química , Hipoglicemiantes/química , Insulina/administração & dosagem , Masculino , Estrutura Molecular , Ratos , Ratos Wistar , Estereoisomerismo , Fatores de Tempo , Tolbutamida/administração & dosagemRESUMO
The effect of the crude extract, ethyl acetate and n-butanol fractions from Vitex megapotamica (Spreng) Moldenke on glycemia was investigated in diabetic rats. Oral administration of crude extract significantly reduced serum glucose levels in both normal and diabetic animals. In normal rats, serum glucose lowering was observed with 400 and 800 mg/kg at 2 and 2-3h, respectively after oral crude extract treatment. Nevertheless, the hypoglycemic effect of Vitex megapotamica in diabetic rats was evident at 1 and 2h and from 1 to 3h after treatment with 400 and 800 mg/kg, respectively. The ethyl acetate as well as n-butanol fractions were able to diminish glycemia in diabetic animals. The ethyl acetate fraction (400 and 800 mg/kg) produced the maximum hypoglycemic effect (28 and 20%, respectively) in diabetic rats and the same dose of the n-butanol fraction reduced the hyperglycemia only by 11% at 1h after treatment. Additionally, in hyperglycemic normal rats neither crude extract nor ethyl acetate fraction modified the glucose tolerance and the known tolbutamide effect on insulin release was clearly observed in this group. Thus, this study shows that Vitex megapotamica has an anti-hyperglycemic action, is able to ameliorate the diabetic state and, probably, is a source of hypoglycemic compounds.
Assuntos
Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/tratamento farmacológico , Hiperglicemia/sangue , Hiperglicemia/tratamento farmacológico , Hipoglicemiantes/farmacologia , Fitoterapia , Vitex/química , Acetatos , Animais , Glicemia/metabolismo , Teste de Tolerância a Glucose , Hipoglicemiantes/química , Insulina/farmacologia , Masculino , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Folhas de Planta/química , Ratos , Ratos Wistar , Solventes , Espectrofotometria Infravermelho , Espectrofotometria Ultravioleta , Espectroscopia de Infravermelho com Transformada de Fourier , Tolbutamida/farmacologiaRESUMO
BACKGROUND AND PURPOSE: Central anti-nociceptive actions of baclofen involve activation of K+ channels. Here we assessed what types of K+ channel might participate in the peripheral anti-nociception induced by baclofen. EXPERIMENTAL APPROACH: Nociceptive thresholds to mechanical stimulation in rat paws treated with intraplantar prostaglandin E2.(PGE2) to induce hyperalgesia were measured 3 h after PGE2 injection. Other agents were also given by intraplantar injection. KEY RESULTS: Baclofen elicited a dose-dependent (15 - 240 microg per paw) anti-nociceptive effect. An intermediate dose of baclofen (60 microg) did not produce antinociception in the contralateral paw, showing its peripheral site of action. The GABAB receptor antagonist saclofen (12.5 - 100 microg per paw) antagonized, in a dose-dependent manner, peripheral antinociception induced by baclofen (60 microg), suggesting a specific effect. This antinociceptive action of baclofen was unaffected by bicuculline, GABAA receptor antagonist (80 microg per paw), or by (1,2,5,6 tetrahydropyridin-4-yl) methylphosphinic acid, GABAC receptor antagonist (20 microg per paw). The peripheral antinociception induced by baclofen (60 microg) was reversed, in a dose-dependent manner, by the voltage-dependent K+ channel blockers tetraethylammonium (7.5 - 30 microg per paw) and 4-aminopyridine (2.5 - 10 microg per paw). The blockers of other K+ channels, glibenclamide (160 microg), tolbutamide (320 microg), charybdotoxin (2 microg), dequalinium (50 microg) and caesium (500 microg) had no effect. CONCLUSIONS AND IMPLICATIONS: This study provides evidence that the peripheral antinociceptive effect of the GABAB receptor agonist baclofen results from the activation of tetraethylammonium-sensitive K+ channels. Other K+ channels appear not to be involved.
Assuntos
Analgésicos/farmacologia , Baclofeno/farmacologia , Agonistas GABAérgicos/farmacologia , Agonistas dos Receptores de GABA-B , Canais de Potássio/efeitos dos fármacos , Tetraetilamônio/farmacologia , Animais , Baclofeno/análogos & derivados , Bicuculina/farmacologia , Césio/farmacologia , Charibdotoxina/farmacologia , Dequalínio/farmacologia , Glibureto/farmacologia , Masculino , Ratos , Ratos Wistar , Tolbutamida/farmacologiaRESUMO
The aim of the present study was to determine the effect of pertussis toxin (PTX) on inflammatory hypernociception measured by the rat paw pressure test and to elucidate the mechanism involved in this effect. In this test, prostaglandin E(2) (PGE(2)) administered subcutaneously induces hypernociception via a mechanism associated with neuronal cAMP increase. Local intraplantar pre-treatment (30 min before), and post-treatment (5 min after) with PTX (600 ng/paw1, in 100 microL) reduced hypernociception induced by prostaglandin E(2) (100 ng/paw, in 100 microL, intraplantar). Furthermore, local intraplantar pre-treatment (30 min before) with PTX (600 ng/paw, in 100 microL) reduced hypernociception induced by DbcAMP, a stable analogue of cAMP (100 microg/paw, in 100 microL, intraplantar), which indicates that PTX may have an effect other than just G(i)/G(0) inhibition. PTX-induced analgesia was blocked by selective inhibitors of nitric oxide synthase (L-NMMA), guanylyl cyclase (ODQ), protein kinase G (KT5823) and ATP-sensitive K(+) channel (Kir6) blockers (glybenclamide and tolbutamide). In addition, PTX was shown to induce nitric oxide (NO) production in cultured neurons of the dorsal root ganglia. In conclusion, this study shows a peripheral antinociceptive effect of pertussis toxin, resulting from the activation of the arginine/NO/cGMP/PKG/ATP-sensitive K(+) channel pathway.
Assuntos
Analgésicos/metabolismo , Arginina/metabolismo , Proteínas Quinases Dependentes de GMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Óxido Nítrico/metabolismo , Toxina Pertussis/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Trifosfato de Adenosina/metabolismo , Analgesia , Animais , Bucladesina/metabolismo , Carbazóis/metabolismo , Células Cultivadas , Dinoprostona/administração & dosagem , Dinoprostona/imunologia , Inibidores Enzimáticos/metabolismo , Gânglios Espinais/citologia , Glibureto/metabolismo , Indóis/metabolismo , Canais KATP , Masculino , Neurônios/citologia , Neurônios/metabolismo , Oxidiazóis/metabolismo , Dor/metabolismo , Medição da Dor , Quinoxalinas/metabolismo , Ratos , Ratos Wistar , Transdução de Sinais/fisiologia , Tolbutamida/metabolismo , ômega-N-Metilarginina/metabolismoRESUMO
The effect of modulators of the nitric oxide-cyclic GMP-protein kinase G-K+ channels pathway on the local peripheral antinociceptive action induced by gabapentin was assessed in the rat 1% formalin test. Local peripheral administration of gabapentin produced a dose-dependent antinociception in the second phase of the test. Gabapentin-induced antinociception was due to a local action as its administration in the contralateral paw was ineffective. Local peripheral pretreatment of the paws with NG-L-nitro-arginine methyl ester (L-NAME, a nitric oxide synthesis inhibitor), 1H-(1,2,4)-oxadiazolo(4,2-a)quinoxalin-1-one (ODQ, a soluble guanylyl cyclase inhibitor) and KT-5823 (a protein kinase G inhibitor) dose-dependently reduced gabapentin-induced antinociception. Likewise, glibenclamide or tolbutamide (ATP-sensitive K+ channel inhibitors), 4-aminopyridine or tetraethylammonium (non-selective inward rectifier K+ channel inhibitors) or charybdotoxin (large-conductance Ca2+-activated-K+ channel blocker), but not apamin (small-conductance Ca2+-activated-K+ channel blocker) or naloxone (opioid receptor antagonist), reduced the antinociception induced by gabapentin. Our data suggest that gabapentin could activate the nitric oxide-cyclic GMP-protein kinase G-K+ channels pathway in order to produce its peripheral antinociceptive effect in the rat 1% formalin test.
Assuntos
Aminas/farmacologia , Analgésicos/farmacologia , Proteínas Quinases Dependentes de GMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Ácidos Cicloexanocarboxílicos/farmacologia , Óxido Nítrico/metabolismo , Canais de Potássio/efeitos dos fármacos , Ácido gama-Aminobutírico/farmacologia , 4-Aminopiridina/farmacologia , Animais , Apamina/farmacologia , Carbazóis/farmacologia , Charibdotoxina/farmacologia , Proteínas Quinases Dependentes de GMP Cíclico/antagonistas & inibidores , Feminino , Gabapentina , Glibureto/farmacologia , Indóis/farmacologia , NG-Nitroarginina Metil Éster/farmacologia , Naloxona/farmacologia , Medição da Dor , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio/metabolismo , Ratos , Ratos Wistar , Tolbutamida/farmacologiaRESUMO
OBJECTIVES: To characterize the clinical features and insulin regulation in infants with hypoglycemia due to prolonged neonatal hyperinsulinism. STUDY DESIGN: Data were collected on 26 infants with hypoglycemia due to neonatal hyperinsulinism that later resolved. Acute insulin response (AIR) tests to calcium, leucine, glucose, and tolbutamide were performed in 11 neonates. Results were compared to children with genetic hyperinsulinism due to mutations of the adenosine triphosphate-dependent potassium (K(ATP)) channel and glutamate dehydrogenase (GDH). RESULTS: Among the 26 neonates, there were significantly more males, small-for-gestational-age infants, and cesarean deliveries. Only 5 of the 26 had no identifiable risk factor. Hyperinsulinism was diagnosed at a median age of 13 days (range, 2 to 180 days) and resolved by a median age of 181 days (range, 18 to 403 days). Diazoxide was effective in 19 of the 21 neonates treated. In the 11 neonates tested, the AIRs to calcium, leucine, glucose, and tolbutamide resembled those in normal controls and differed from genetic hyperinsulinism due to K(ATP) channel and GDH mutations. CONCLUSIONS: We define a syndrome of prolonged neonatal hyperinsulinism that is responsive to diazoxide, persists for several months, and resolves spontaneously. AIR tests suggest that both the K(ATP) channel and GDH have normal function.
Assuntos
Anti-Hipertensivos/uso terapêutico , Diazóxido/uso terapêutico , Hiperinsulinismo/terapia , Hipoglicemia/terapia , Cálcio/farmacologia , Estudos de Casos e Controles , Nutrição Enteral , Jejum , Feminino , Glucose/farmacologia , Humanos , Hiperinsulinismo/complicações , Hiperinsulinismo/diagnóstico , Hipoglicemia/etiologia , Hipoglicemiantes/farmacologia , Lactente , Recém-Nascido , Infusões Intravenosas , Insulina/sangue , Insulina/metabolismo , Secreção de Insulina , Leucina/farmacologia , Masculino , Estudos Prospectivos , Fatores de Risco , Edulcorantes/farmacologia , Tolbutamida/farmacologiaRESUMO
Mammalian sperm must undergo a series of physiological changes after leaving the testis to become competent for fertilization. These changes, collectively known as capacitation, occur in the female reproductive tract where the sperm plasma membrane is modified in terms of its components and ionic permeability. Among other events, mouse sperm capacitation leads to an increase in the intracellular Ca(2+) and pH as well as to a hyperpolarization of the membrane potential. It is well known that ion channels play a crucial role in these events, though the molecular identity of the particular channels involved in capacitation is poorly defined. In the present work, we report the identification and potential functional role of K(ATP) channels in mouse spermatogenic cells and sperm. By using whole-cell patch clamp recordings in mouse spermatogenic cells, we found K(+) inwardly rectifying (K(ir)) currents that are sensitive to Ba(2+), glucose and the sulfonylureas (tolbutamide and glibenclamide) that block K(ATP) channels. The presence of these channels was confirmed using inhibitors of the ATP synthesis and K(ATP) channel activators. Furthermore, RT-PCR assays allowed us to detect transcripts for the K(ATP) subunits SUR1, SUR2, K(ir)6.1 and K(ir)6.2 in total RNA from elongated spermatids. In addition, immunoconfocal microscopy revealed the presence of these K(ATP) subunits in mouse spermatogenic cells and sperm. Notably, incubation of sperm with tolbutamide during capacitation abolished hyperpolarization and significantly decreased the percentage of AR in a dose-dependent fashion. Together, our results provide evidence for the presence of K(ATP) channels in mouse spermatogenic cells and sperm and disclose the contribution of these channels to the capacitation-associated hyperpolarization.
Assuntos
Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Capacitação Espermática/fisiologia , Espermatogênese/fisiologia , Espermatozoides/metabolismo , Transportadores de Cassetes de Ligação de ATP/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Bário/metabolismo , Bário/farmacologia , Diazóxido/metabolismo , Diazóxido/farmacologia , Relação Dose-Resposta a Droga , Glibureto/metabolismo , Glibureto/farmacologia , Canais KATP , Masculino , Potenciais da Membrana , Camundongos , Microscopia Confocal , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Pinacidil/metabolismo , Pinacidil/farmacologia , RNA Mensageiro/metabolismo , Receptores de Droga , Espermatozoides/citologia , Receptores de Sulfonilureias , Fatores de Tempo , Tolbutamida/metabolismo , Tolbutamida/farmacologiaRESUMO
The possible participation of K+ channels in the antinociceptive action of meloxicam was assessed in the 1% formalin test. Local peripheral administration of meloxicam produced a dose-dependent antinociception only during the second phase of the formalin test. K+ channel blockers alone did not modify formalin-induced nociceptive behavior. However, local peripheral pretreatment of the paw with charybdotoxin and apamin (large- and small-conductance Ca2+-activated K+ channel inhibitors, respectively), 4-aminopyridine and tetraethylammonium (non-selective voltage-dependent K+ channel inhibitors), but not glibenclamide or tolbutamide (ATP-sensitive K+ channel inhibitors), dose-dependently prevented meloxicam-induced antinociception. It is concluded that meloxicam could open large- and small-conductance Ca2+-activated K+ channels, but not ATP-sensitive K+ channels, in order to produce its peripheral antinociceptive effect in the formalin test. The participation of voltage-dependent K+ channels was also suggested, but since non-selective inhibitors were used the data await further confirmation.
Assuntos
Analgésicos não Narcóticos/farmacologia , Dor/prevenção & controle , Canais de Potássio Cálcio-Ativados/fisiologia , Tiazinas/farmacologia , Tiazóis/farmacologia , 4-Aminopiridina/farmacologia , Análise de Variância , Animais , Apamina/farmacologia , Comportamento Animal/efeitos dos fármacos , Charibdotoxina/farmacologia , Relação Dose-Resposta a Droga , Feminino , Formaldeído , Glibureto/farmacologia , Meloxicam , Dor/induzido quimicamente , Dor/fisiopatologia , Medição da Dor/métodos , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio Cálcio-Ativados/antagonistas & inibidores , Canais de Potássio Corretores do Fluxo de Internalização/antagonistas & inibidores , Canais de Potássio Corretores do Fluxo de Internalização/fisiologia , Ratos , Ratos Wistar , Tetraetilamônio/farmacologia , Tolbutamida/farmacologiaRESUMO
The mechanism of the antinociceptive action of the phosphodiesterase 5 inhibitor, sildenafil, was assessed in the formalin test. Local peripheral ipsilateral, but not contralateral, administration of sildenafil (50-200 microg/paw) produced a dose-related antinociception during both phases of the formalin test. The local peripheral pretreatment with protein kinase G inhibitor peptide (PKG inhibitor, 0.01-1 microg/paw), charybdotoxin (large- and intermediate-conductance Ca2+-activated K+ channel blocker, 0.01-1 microg/paw), apamin (small-conductance Ca2+-activated K+ channel blocker, 0.1-2 microg/paw), tolbutamide (ATP-sensitive K+ channel blocker, 12.5-50 microg/paw), and tetraethylammonium (non-selective voltage-dependent K+ channel blocker, 12.5-50 microg/paw), but not 1H-(1,2,4)-oxadiazolo(4,2-a)quinoxalin-1-one (ODQ, inhibitor of guanylyl cyclase, 12.5-50 microg/paw) or saline, significantly diminished in a dose-dependent manner sildenafil-induced local peripheral antinociception. Given alone, local peripheral administration of inhibitors did not modify formalin-induced nociceptive behavior. Results suggest that sildenafil produces its local peripheral antinociceptive effect via activation of the cyclic GMP-PKG-K+ channel pathway.
Assuntos
Analgésicos/farmacologia , Dor/prevenção & controle , Piperazinas/farmacologia , Animais , Apamina/farmacologia , Charibdotoxina/farmacologia , Proteínas Quinases Dependentes de GMP Cíclico/antagonistas & inibidores , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Feminino , Formaldeído , Guanilato Ciclase/antagonistas & inibidores , Injeções Subcutâneas , Oxidiazóis/farmacologia , Dor/induzido quimicamente , Medição da Dor/métodos , Bloqueadores dos Canais de Potássio/farmacologia , Purinas , Quinoxalinas/farmacologia , Ratos , Ratos Wistar , Citrato de Sildenafila , Sulfonas , Tetraetilamônio/farmacologia , Fatores de Tempo , Tolbutamida/farmacologiaRESUMO
There is evidence that local peripheral administration of morphine produces antinociception through the activation of the nitric oxide (NO)-cyclic GMP-K(+) channels pathway. Therefore we evaluated the possible participation of this pathway in the antinociceptive action produced by codeine in the rat 5% formalin test. Local peripheral injection of codeine produced a dose-dependent antinociception during the first and second phases of the test. Local pretreatment of the paws with the NO synthase inhibitor N(G)-L-nitro-arginine methyl ester (L-NAME), the soluble guanylyl cyclase inhibitor methylene blue, the ATP-sensitive K(+) channel inhibitors glibenclamide and tolbutamide, the non-selective voltage-gated K(+) channel inhibitors 4-aminopyridine (4-AP) and tetraethylammonium (TEA) and the opioid receptor blocker naloxone prevented codeine-induced antinociception in both phases of the test. L-NAME, methylene blue, K(+) channel blockers and naloxone by themselves did not modify formalin-induced nociceptive behavior. Our data suggest that codeine could activate the opioid receptor-NO-cyclic GMP-K(+) channels pathway in order to produce its peripheral antinociceptive effect in the formalin test.
Assuntos
Analgésicos Opioides/farmacologia , Codeína/farmacologia , GMP Cíclico/metabolismo , Óxido Nítrico/metabolismo , Canais de Potássio/metabolismo , Receptores Opioides/metabolismo , 4-Aminopiridina/farmacologia , Animais , Feminino , Glibureto/farmacologia , Azul de Metileno/farmacologia , NG-Nitroarginina Metil Éster/farmacologia , Naloxona/farmacologia , Ratos , Ratos Wistar , Tetraetilamônio/farmacologia , Tolbutamida/farmacologiaRESUMO
Testosterone at physiological intratesticular concentrations induces a dose-dependent depolarisation and an increase in input resistance together with an increment of 45Ca2+ uptake in the Sertoli cells from seminiferous tubules of immature rat. Previous studies have implicated K(+)ATP channels in these testosterone actions. This study demonstrates that testosterone and sulphonylureas (glibenclamide and tolbutamide) depolarise the membrane potential, augment resistance and 45Ca2+ uptake in the Sertoli cells of seminiferous tubules from 10-15 day-old rats. These actions were nullified by the presence of the K(+)ATP channel opener diazoxide. The depolarisation was also observed with the impermeant bovine serum albumin-bound testosterone. Testosterone actions were blocked by both pertussis toxin and the phospholipase C (PLC) inhibitor U73122 implying the involvement of PLC - phosphatidylinositol 4-5 bisphosphate (PIP2) hydrolysis via G protein in testosterone actions. Polycations, including spermine and LaCl3, depolarised the membrane potential and increased the resistance. Hyperpolarisation caused by EGTA was reversed by LaCl3 and by the presence of testosterone. This last effect was nullified by the presence of U73122. All of the above results indicate that the action of testosterone on the Sertoli cell membrane is exercised on the K(+)ATP channels through PLC-PIP2 hydrolysis that closes the channel, depolarises the membrane, and stimulates 45Ca2+ uptake.
Assuntos
Trifosfato de Adenosina/metabolismo , Androgênios/farmacologia , Proteínas de Membrana/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Canais de Potássio/metabolismo , Células de Sertoli/metabolismo , Testosterona/farmacologia , Fosfolipases Tipo C/metabolismo , Animais , Cálcio/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio/efeitos dos fármacos , Impedância Elétrica , Glibureto/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Toxina Pertussis/farmacologia , Ratos , Ratos Wistar , Eletricidade Estática , Tolbutamida/farmacologiaRESUMO
The involvement of the nitric oxide (NO)/cyclic GMP pathway in the molecular mechanisms of antinociceptive drugs like morphine has been previously shown by our group. Additionally, it is known that the desensitisation of nociceptors by K(+) channel opening should be the final target for several analgesic drugs including nitric oxide donors and exogenous micro-opioid receptor agonists. In our previous study, we demonstrated that bremazocine, a kappa-opioid receptor agonist, induces peripheral antinociception by activating nitric oxide/cyclic GMP pathway. In the current study, we assessed whether bremazocine is capable to activate K(+) channels eliciting antinociception. Bremazocine (20, 40 and 50 microg) dose-dependently reversed the hyperalgesia induced in the rat paw by local injection of carrageenan (250 microg) or prostaglandin E(2) (2 microg), measured by the paw pressure test. Using the selective kappa-opioid receptor antagonist nor-binaltorphimine (Nor-BNI, 200 microg/paw), it was confirmed that bremazocine (50 microg/paw) acts specifically on the kappa-opioid receptors present at peripheral sites. Prior treatment with the ATP-sensitive K(+) channel blockers glibenclamide (40, 80 and 160 microg) and tolbutamide (40, 80 and 160 microg) did not antagonise the antinociceptive effect of bremazocine (50 microg). The same results were obtained when we used prostaglandin E(2) (2 microg) as the hyperalgesic stimulus. The supposed participation of other types of K(+) channels was tested using the Ca(2+)-activated K(+) channel blockers dequalinium (12.5, 25 and 50 microg) and charybdotoxin (0.5, 1 and 2 microg) and different types of the non-selective K(+) channel blockers tetraethylammonium (25, 50 and 100 microg) and 4-aminopyridine (10, 25 and 50 microg). None of the K(+) channel blockers reversed the antinociceptive effect of bremazocine. On the basis of these results, we suggest that K(+) channels are not involved in the peripheral antinociceptive effect of bremazocine, although this opioid receptor agonist induces nitric oxide/cGMP pathway activation.
Assuntos
Analgésicos/farmacologia , Benzomorfanos/farmacologia , Naltrexona/análogos & derivados , Sistema Nervoso Periférico/efeitos dos fármacos , Canais de Potássio/efeitos dos fármacos , Receptores Opioides kappa/agonistas , 4-Aminopiridina/farmacologia , Transportadores de Cassetes de Ligação de ATP , Analgésicos/antagonistas & inibidores , Animais , Benzomorfanos/antagonistas & inibidores , Charibdotoxina/farmacologia , GMP Cíclico/fisiologia , Dequalínio/farmacologia , Glibureto/farmacologia , Hiperalgesia/induzido quimicamente , Hiperalgesia/fisiopatologia , Canais KATP , Masculino , Naltrexona/farmacologia , Óxido Nítrico/fisiologia , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio Corretores do Fluxo de Internalização , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Tetraetilamônio/farmacologia , Tolbutamida/farmacologiaRESUMO
In order to investigate to the contribution of K+ channels on the peripheral antinociception induced by diclofenac, we evaluated the effect of several K+ channel blockers, using the rat paw pressure test, in which sensitivity is increased by intraplantar injection (2 microg) of prostaglandin E2. Diclofenac administered locally into the right hindpaw (25, 50, 100 and 200 microg) elicited a dose-dependent antinociceptive effect which was demonstrated to be local, since only higher doses produced an effect when injected in the contralateral paw. This blockade of PGE2 mechanical hyperalgesia induced by diclofenac (100 microg/paw) was antagonized in a dose-dependent manner by intraplantar administration of the sulphonylureas glibenclamide (40, 80 and 160 microg) and tolbutamide (80, 160 and 320 microg), specific blockers of ATP-sensitive K+ channels, and it was observed even when the hyperalgesic agent used was carrageenin, while the antinociceptive action of indomethacin (200 microg/paw), a typical cyclo-oxygenase inhibitor, over carrageenin-induced hyperalgesia was not affected by this treatment. Charybdotoxin (2 microg/paw), a blocker of large conductance Ca2+-activated K+ channels and dequalinium (50 microg/paw), a selective blocker of small conductance Ca2+-activated K+ channels, did not modify the effect of diclofenac. This effect was also unaffected by intraplantar administration of non-specific voltage-dependent K+ channel blockers tetraethylammonium (1700 microg) and 4-aminopyridine (100 microg) or cesium (500 microg), a non-specific K+ channel blocker. The peripheral antinociceptive effect induced by diclofenac was antagonized by NG-Nitro L-arginine (NOarg, 50 microg/paw), a NO synthase inhibitor and methylene blue (MB, 500 microg/paw), a guanylate cyclase inhibitor, and this antagonism was reversed by diazoxide (300 microg/paw), an ATP-sensitive K+ channel opener. We also suggest that an endogenous opioid system may not be involved since naloxone (50 microg/paw) did not affect diclofenac-induced antinociception in the PGE2-induced hyperalgesia model. This study provides evidence that the peripheral antinociceptive effect of diclofenac may result from activation of ATP-sensitive K+ channels, possible involving stimulation of L-arginine/NO/cGMP pathway, while Ca2+-activated K+ channels, voltage-dependent K+ channels as well as endogenous opioids appear not to be involved in the process.