Assuntos
Síndrome do QT Longo/genética , Mutação , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio/genética , Feminino , Frequência do Gene , Marcadores Genéticos , Heterozigoto , Humanos , Masculino , Repetições de Microssatélites , Linhagem , Polimorfismo Genético , Canais de Potássio/metabolismoRESUMO
The effect of the uterotonic pituitary hormone oxytocin on the regulation of intracellular calcium concentration ([Ca2+]i) was studied in single cells of a smooth muscle cell line derived from human non-pregnant myometrium. [Ca2+]i was measured with fluorescence microscopy, and by recording the activity of Ca(2+)-activated potassium currents (IK(Ca)) on the whole cell and single channel level. Oxytocin induced a rapid and transient increase in [Ca2+]i that was paralleled by a significant increase in IK(Ca) activity. After removal of extracellular Ca2+, repetitive stimulation with oxytocin did not alter the [Ca2+]i transients initially; however, their amplitude became progressively smaller and the response was eventually abolished completely, indicating that oxytocin increased [Ca2+]i by release of Ca2+ from intracellular stores. Nifedipine did not alter the oxytocin-induced [Ca2+]i-transients suggesting that oxytocin failed to activate Ca2+ entry through voltage-operated Ca2+ channels. Thapsigargin abolished the oxytocin-induced [Ca2+]i transient. Caffeine alone had no effect on [Ca2+]i, however it diminished the oxytocin-induced [Ca2+]i transients. Ryanodine did not affect the oxytocin response indicating that these cells lack release of Ca2+ from the ryanodine receptor release channel. These results demonstrate that oxytocin elicited [Ca2+]i transients predominantly through Ca2+ release from thapsigargin-sensitive stores, presumably by activating an inositol 1,4,5-trisphosphate dependent pathway.
Assuntos
Cálcio/metabolismo , Músculo Liso/citologia , Miométrio/citologia , Ocitocina/farmacologia , Transporte Biológico/efeitos dos fármacos , Cafeína/farmacologia , ATPases Transportadoras de Cálcio/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Feminino , Humanos , Nifedipino/farmacologia , Rianodina/farmacologia , Tapsigargina/farmacologiaRESUMO
Damage to fallopian tubes caused by hydrosalpinges is a major reason for persisting infertility. We describe the mucosal changes that occur following induction of experimentally induced hydrosalpinges in 25 rabbits. Ampullary biopsies obtained at varying intervals were examined with light microscopy and scanning and transmission electron microscopy. Mucosal folds showed progressive atrophy. The epithelium was flattened. The population of ciliated cells decreased and, finally, deciliation became generalized. The morphology of nonciliated cells changed from an initial high content of granules of secretion to a final stage of vacuolated cytoplasm. Cell desquamation was common.
Assuntos
Doenças das Tubas Uterinas/etiologia , Tubas Uterinas/fisiopatologia , Animais , Constrição , Epitélio/ultraestrutura , Tubas Uterinas/patologia , Tubas Uterinas/ultraestrutura , Feminino , CoelhosAssuntos
Anormalidades Múltiplas/genética , Cardiopatias Congênitas/patologia , Deformidades Congênitas dos Membros/patologia , Proteínas com Domínio T/genética , Células 3T3 , Anormalidades Múltiplas/patologia , Transporte Ativo do Núcleo Celular/genética , Animais , Fator Natriurético Atrial/genética , Sequência de Bases , Linhagem Celular , Núcleo Celular/metabolismo , Códon sem Sentido , DNA/química , DNA/genética , DNA/metabolismo , Análise Mutacional de DNA , Saúde da Família , Feminino , Humanos , Luciferases/genética , Luciferases/metabolismo , Masculino , Camundongos , Dados de Sequência Molecular , Mutação , Linhagem , Regiões Promotoras Genéticas , Ligação Proteica/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Deleção de Sequência , Síndrome , Proteínas com Domínio T/metabolismoAssuntos
Fígado/efeitos dos fármacos , Oxigenases de Função Mista/metabolismo , Oxirredutases/metabolismo , Tiadiazóis/farmacologia , Animais , Monóxido de Carbono/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Indução Enzimática/efeitos dos fármacos , Feminino , Hexobarbital/metabolismo , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Fenilalanina/metabolismo , Fatores de TempoAssuntos
Anticoncepcionais Orais/administração & dosagem , Tubas Uterinas/efeitos dos fármacos , Fertilidade/efeitos dos fármacos , Gônadas/efeitos dos fármacos , Norpregnanos/administração & dosagem , Pregnadienodiois/administração & dosagem , Pregnenodionas/administração & dosagem , Progestinas/administração & dosagem , Adulto , Anticoncepcionais Orais/farmacologia , Corpo Lúteo/citologia , Corpo Lúteo/efeitos dos fármacos , Relação Dose-Resposta a Droga , Endométrio/citologia , Células Epiteliais , Epitélio/efeitos dos fármacos , Tubas Uterinas/citologia , Tubas Uterinas/fisiologia , Feminino , Gônadas/citologia , Histocitoquímica , Humanos , Hidroxiesteroides/administração & dosagem , Cetosteroides/administração & dosagem , Microscopia Eletrônica , Folículo Ovariano/citologia , Folículo Ovariano/efeitos dos fármacos , Ovário/citologia , Ovulação/efeitos dos fármacos , Oxirredutases/metabolismo , Pregnatrienos/administração & dosagem , Pregnenodionas/farmacologia , Progestinas/farmacologia , Esteroides Clorados/administração & dosagem , Sulfetos/administração & dosagemAssuntos
Anticoncepcionais Orais/administração & dosagem , Progestinas/administração & dosagem , Adolescente , Adulto , Muco do Colo Uterino/efeitos dos fármacos , Ensaios Clínicos como Assunto , Coito , Anticoncepcionais Orais/farmacologia , Relação Dose-Resposta a Droga , Endométrio/efeitos dos fármacos , Diacetato de Etinodiol/administração & dosagem , Diacetato de Etinodiol/farmacologia , Feminino , Hormônio Foliculoestimulante/metabolismo , Humanos , Hidroxiesteroides/administração & dosagem , Cetosteroides/administração & dosagem , Hormônio Luteinizante/metabolismo , Menstruação/efeitos dos fármacos , Norpregnanos/administração & dosagem , Norpregnanos/farmacologia , Ovário/efeitos dos fármacos , Pregnadienodiois/administração & dosagem , Pregnanodiol/urina , Pregnatrienos/administração & dosagem , Pregnenodionas/administração & dosagem , Progestinas/efeitos adversos , Progestinas/farmacologia , Esteroides Clorados/administração & dosagem , Sulfetos/administração & dosagem , Vagina/citologia , Vagina/efeitos dos fármacosAssuntos
Camelídeos Americanos , Transplante das Ilhotas Pancreáticas , Modelos Biológicos , Animais , Glicemia/metabolismo , Camelídeos Americanos/anatomia & histologia , Camelídeos Americanos/sangue , Camelídeos Americanos/líquido cefalorraquidiano , Cultura em Câmaras de Difusão , Cães , Sobrevivência de Enxerto , Hexoquinase/genética , Insulina/metabolismo , Secreção de Insulina , Transplante das Ilhotas Pancreáticas/imunologia , Transplante das Ilhotas Pancreáticas/patologia , Transplante das Ilhotas Pancreáticas/fisiologia , Camundongos , Camundongos Transgênicos , Ratos , Fatores de Tempo , Transplante HeterólogoRESUMO
The human amnion was examined by means of light microscopy and scanning and transmission electron microscopy. The surface shows apart from microvilli a particular structure, called "blebs"; the intercellular junction is formed by desmosomes and a labyrinthine channel system, and at the base pedicels extend into the basement membrane. Cell shedding uncovers the basement membrane which seems to play a primordial role in preserving the intact amniotic cavity. These data underline the complex structure and the multiple role the amnion performs during gestation.
Assuntos
Âmnio/citologia , Gravidez , Âmnio/ultraestrutura , Desmossomos/ultraestrutura , Células Epiteliais , Feminino , Humanos , Microscopia Eletrônica , Microscopia Eletrônica de Varredura , Microvilosidades/ultraestrutura , Placenta/ultraestrutura , Cordão Umbilical/ultraestruturaRESUMO
The properties of Ca(2+)-activated K+ currents and channels were characterized in pregnant rat myometrium in whole cell and cell-attached patches and in lipid bilayers. Membrane depolarization of cultured myometrial cells from a holding potential of -50 to +70 mV in 10-mV steps under voltage-clamp conditions (whole cell mode) activated K+ outward currents (IK). At +70 mV, in the presence of 0.2 mM external Ca2+, the amplitude and activation time constant of IK were 15.0 +/- 2.1 microA/microF and 1.5 +/- 0.2 ms, respectively. Addition of 1 microM A23187 to the external solution increased the current from a control value of 16.0 +/- 2.0 to 67.9 +/- 9.1 microA/microF. Charybdotoxin, a blocker of Ca(2+)-activated K (KCa) channels, and a low concentration of tetraethylammonium chloride (TEA; 1 mM) decreased the amplitude of IK by 47 and 62%, respectively. In cell-attached patches from these cells, 1 microM A23187 increased the open time probability of a 143 +/- 6.0 pS K+ channel. Incorporation of plasma membrane vesicles from pregnant myometrium into lipid bilayers resulted in one predominant type of K+ channel. The unitary conductance of the K+ channel was 326 +/- 9.0 pS in symmetrical 450 mM KCl. The channel activation was both voltage and Ca2+ dependent. TEA inhibited the channel activity with a dissociation constant (Kd) of 378 +/- 10 microM at -60 mV or 1,477 +/- 80 microM at +60 mV. The whole cell currents were found to be stimulated by isoproterenol, a beta-adrenergic agent.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Agonistas Adrenérgicos beta/farmacologia , Cálcio/farmacologia , Miométrio/metabolismo , Canais de Potássio/metabolismo , Prenhez/metabolismo , Animais , Eletrofisiologia , Feminino , Bicamadas Lipídicas , Canais de Potássio/fisiologia , Gravidez , Ratos , Ratos Sprague-Dawley , Tetraetilamônio , Compostos de Tetraetilamônio/farmacologiaRESUMO
Lens fiber cells contain two gap junction proteins (Cx56 and Cx45.6 in the chicken). Biochemical studies have suggested that these two proteins can form heteromeric connexons. To investigate the biophysical properties of heteromeric lens connexons, Cx56 was co-expressed with Cx45.6 (or its mouse counterpart, Cx50) in Xenopus oocytes. Whole-cell and single-channel currents were measured in single oocytes by conventional two-microelectrode voltage-clamp and patch clamp techniques, respectively. Injection of Cx56 cRNA induced a slowly activating, nonselective cation current that activated on depolarization to potentials higher than -10 mV. In contrast, little or no hemichannel current was induced by injection of Cx50 or Cx45.6 cRNA. Co-expression of Cx56 with Cx45.6 or Cx50 led to a shift in the threshold for activation to -40 or -70 mV, respectively. It also slowed the rate of deactivation of the hemichannel currents. Moreover, an increase in the unitary conductance, steady state probability of hemichannel opening and mean open times at negative potentials, was observed in (Cx56 + Cx45.6) cRNA-injected oocytes compared with Cx56 cRNA-injected oocytes. These results indicate that co-expression of lens fiber connexins gives rise to novel channels that may be explained by the formation of heteromeric hemichannels that contain both connexins.
Assuntos
Conexinas/química , Conexinas/metabolismo , Junções Comunicantes/química , Junções Comunicantes/metabolismo , Canais Iônicos/química , Canais Iônicos/metabolismo , Cristalino/química , Cristalino/metabolismo , Animais , Fenômenos Biofísicos , Biofísica , Cálcio/farmacologia , Galinhas , Conexinas/genética , Proteínas do Olho/química , Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Feminino , Junções Comunicantes/efeitos dos fármacos , Expressão Gênica , Técnicas In Vitro , Ativação do Canal Iônico , Canais Iônicos/efeitos dos fármacos , Cinética , Potenciais da Membrana , Camundongos , Oócitos/metabolismo , Técnicas de Patch-Clamp , Xenopus laevisRESUMO
Relaxin, a hormone that is elevated during pregnancy, can suppress myometrial contractile activity. Ca(2+)-activated K+ channels (KCa) play a role in the modulation of uterine contractions and myometrial Ca2+ homeostasis and have been implicated in the control of smooth muscle excitability. We now show that relaxin stimulates KCa channels in cell-attached patches in a cell line derived from term pregnant human myometrium. This effect was prevented by the protein kinase A (PKA) antagonist, the Rp diastereomer of adenosine 3',5'-cyclic monophosphothioate (Rp-cAMPS). After patch excision, the channel was activated by PKA and inhibited by alkaline phosphatase. These data suggest that relaxin may promote myometrial quiescence in part by stimulation of KCa channels via a PKA-mediated mechanism.
Assuntos
Cálcio/fisiologia , Proteínas Quinases Dependentes de AMP Cíclico/fisiologia , Miométrio/metabolismo , Canais de Potássio/metabolismo , Relaxina/farmacologia , AMP Cíclico/análogos & derivados , AMP Cíclico/farmacologia , Proteínas Quinases Dependentes de AMP Cíclico/antagonistas & inibidores , Condutividade Elétrica , Feminino , Humanos , Miométrio/citologia , Canais de Potássio/efeitos dos fármacos , Gravidez , Relaxina/antagonistas & inibidores , Tionucleotídeos/farmacologiaRESUMO
The role of Ca(2+)-activated potassium (KCa) channels in the regulation of membrane potential, intracellular free calcium ([Ca2+]i) and contraction was investigated in uterine smooth muscle and myometrial cells. In an immortalized human myometrial cell line, oxytocin increased [Ca2+]i and [3H]inositol phosphate formation. Relaxin attenuated the oxytocin-induced increase in [Ca2+]i. In cell-attached patches, membrane depolarization activated a large-conductance KCa channel (179 +/- 4 pS). Iberiotoxin (IbTX), a potent blocker of "maxi" KCa channels (A. Galvez, G. Gimenez-Gallego, J. P. Reuben, L. Roy-Contanciin, P. Feigenbaum, G. J. Kaczorowski, and M. L. Garcia. J. Biol. Chem. 265: 11083-11090, 1990) produced long closed events (approximately 6 min) in these channels. In agreement with this blockage, IbTX depolarized the cells by 9.8 +/- 2.8 mV and caused a dose-dependent increase in [Ca2+]i with a half-maximal effective concentration of 0.79 nM. IbTX also caused phasic contractions in human myometrial strips and increased both the frequency and force of spontaneous contractions in estrogen-primed rat myometrial strips. Moreover, myometrial contractility was also affected by 1 mM tetraethylammonium, a concentration that blocks uterine smooth muscle KCa channels when applied to the extracellular side (G. J. Perez, L. Toro, S. D. Erulkar, and E. Stefani. Am. J. Obstet. Gynecol. 168: 652-660, 1993). These results strongly suggest that the large conductance KCa channels may actively participate in the control of human myometrial cell membrane potential and [Ca2+].
Assuntos
Cálcio/fisiologia , Canais de Potássio/fisiologia , Contração Uterina/fisiologia , Cálcio/metabolismo , Linhagem Celular , Eletrofisiologia , Feminino , Humanos , Fosfatos de Inositol/metabolismo , Membranas Intracelulares/metabolismo , Potenciais da Membrana/efeitos dos fármacos , Miométrio/citologia , Miométrio/metabolismo , Concentração Osmolar , Peptídeos/farmacologia , Bloqueadores dos Canais de Potássio , Contração Uterina/efeitos dos fármacosRESUMO
Long QT syndrome (LQTS) is the prototype of the cardiac ion channelopathies which cause syncope and sudden death. LQT1, due to mutations of KCNQ1 (KVLQT1), is the most common form. This study describes the genotype-phenotype characteristics in 10 families with mutations of KCNQ1, including 5 novel mutations. One hundred and two families with a history of lethal cardiac events, 55 LQTS, 9 Brugada syndrome, 18 idiopathic ventricular fibrillation (IVF), and 20 acquired LQTS, were studied by single-strand conformational polymorphism (SSCP) and DNA sequence analyzes. Families found to have KCNQ1 mutations were phenotyped using ECG parameters and cardiac event history, and genotype-phenotype correlation was performed. No mutations were found in Brugada syndrome, IVF, or acquired LQTS families. Ten out of 55 LQTS families had KCNQ1 mutations and 62 carriers were identified. Mutations included G269S in domain S5; W305X, G314C, Y315C, and D317N in the pore region; A341E and Q357R in domain S6; and 1338insC, G568A and T587M mutations in the C-terminus. W305X, G314C, Q357R, 1338insC, and G568A, appeared to be novel mutations. Gene carriers were 26 +/- 19 years (32 females). Baseline QTc was 0.47 +/- 0.03 s (range 0.40-0.57 s) and 40% had normal to borderline QTc (< or = 0.46 s). Typical LQT1 T wave patterns were present in at least one affected member of each family, and in 73% of all affected members. A history of cardiac events was present in 19/62 (31%), 18 with syncope, 2 with aborted cardiac arrest (ACA) and six with sudden death (SD). Two out of 6 SDs (33%) occurred as the first symptom. No difference in phenotype was evident in pore vs. non-pore mutations. KCNQ1 mutations were limited to LQTS families. All five novel mutations produced a typical LQT1 phenotype. Findings emphasize (1) reduced penetrance of QTc and symptoms, resulting in diagnostic challenges, (2) the problem of sudden death as the first symptom (33% of those who died), and (3) genetic testing is important for identification of gene carriers with reduced penetrance, in order to provide treatment and to prevent lethal cardiac arrhythmias and sudden death.