RESUMO
Venous valves (VVs) are critical for unidirectional blood flow from superficial and deep veins towards the heart. Congenital valve aplasia or agenesis may, in some cases, be a direct cause of vascular disease, motivating an understanding of the molecular mechanisms underlying the development and maintenance of VVs. Three gap junction proteins (Connexins), Cx37, Cx43, and Cx47, are specifically expressed at VVs in a highly polarized fashion. VVs are absent from adult mice lacking Cx37; however it is not known if Cx37 is required for the initial formation of valves. In addition, the requirement of Cx43 and Cx47 for VV development has not been studied. Here, we provide a detailed description of Cx37, Cx43, and Cx47 expression during mouse vein development and show by gene knockout that each Cx is necessary for normal valve development. The valve phenotypes in the knockout lines exhibit Cx-specific differences, however, including whether peripheral or central VVs are affected by gene inactivation. In addition, we show that a Cx47 null mutation impairs peripheral VV development but does not affect lymphatic valve formation, a finding of significance for understanding how some CX47 mutations cause inherited lymphedema in humans. Finally, we demonstrate a striking segregation of Foxc2 and NFATc1 transcription factor expression between the downstream and upstream faces, respectively, of developing VV leaflets and show that this segregation is closely associated with the highly polarized expression of Cx37, Cx43, and Cx47. The partition of Foxc2 and NFATc1 expression at VV leaflets makes it unlikely that these factors directly cooperate during the leaflet elongation stage of VV development.
Assuntos
Conexina 43/metabolismo , Conexinas/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Fatores de Transcrição NFATC/metabolismo , Válvulas Venosas/metabolismo , Animais , Conexina 43/genética , Conexinas/genética , Fatores de Transcrição Forkhead/genética , Imuno-Histoquímica , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fatores de Transcrição NFATC/genética , Fenótipo , Fatores de Tempo , Válvulas Venosas/embriologia , Válvulas Venosas/crescimento & desenvolvimento , Proteína alfa-4 de Junções ComunicantesRESUMO
BACKGROUND: Very few telemedicine projects in medically underserved areas have been sustained over time. This research furthers understanding of telemedicine service sustainability by examining teleconsultation projects from the perspective of healthcare providers. Drivers influencing healthcare providers' continued participation in teleconsultation projects and how projects can be designed to effectively and efficiently address these drivers is examined. METHODS: Case studies of fourteen teleconsultation projects that were part of two health sciences center (HSC) based telemedicine networks was utilized. Semi-structured interviews of 60 key informants (clinicians, administrators, and IT professionals) involved in teleconsultation projects were the primary data collection method. RESULTS: Two key drivers influenced providers' continued participation. First was severe time constraints. Second was remote site healthcare providers' (RSHCPs) sense of professional isolation. Two design steps to address these were identified. One involved implementing relatively simple technology and process solutions to make participation convenient. The more critical and difficult design step focused on designing teleconsultation projects for collaborative, active learning. This learning empowered participating RSHCPs by leveraging HSC specialists' expertise. CONCLUSIONS: In order to increase sustainability the fundamental purpose of teleconsultation projects needs to be re-conceptualized. Doing so requires HSC specialists and RSHCPs to assume new roles and highlights the importance of trust. By implementing these design steps, healthcare delivery in medically underserved areas can be positively impacted.
Assuntos
Pessoal de Saúde/estatística & dados numéricos , Área Carente de Assistência Médica , Consulta Remota/estatística & dados numéricos , Atitude do Pessoal de Saúde , Atenção à Saúde/estatística & dados numéricos , Pessoal de Saúde/psicologia , Humanos , Prática Profissional/estatística & dados numéricos , Papel Profissional , Consulta Remota/métodos , Saúde da População RuralRESUMO
BACKGROUND: This research analyzes teleconsultation from both a mechanistic and complex adaptive system (CAS) dominant logic in order to further understand the influence of dominant logic on utilization rates of teleconsultation projects. In both dominant logics, the objective of teleconsultation projects is to increase access to and quality of healthcare delivery in a cost efficient manner. A mechanistic dominant logic perceives teleconsultation as closely resembling the traditional service delivery model, while a CAS dominant logic focuses on the system's emergent behavior of learning resulting from the relationships and interactions of participating healthcare providers. METHODS: Qualitative case studies of 17 teleconsultation projects that were part of four health sciences center (HSC) based telemedicine networks was utilized. Data were collected at two points in time approximately 10 years apart. Semi-structured interviews of 85 key informants (clinicians, administrators, and IT professionals) involved in teleconsultation projects were the primary data collection method. RESULTS: The findings indicated that the emergent behavior of effective and sustainable teleconsultation projects differed significantly from what was anticipated in a mechanistic dominant logic. Teleconsultation projects whose emergent behavior focused on continuous learning enabled remote site generalists to manage and treat more complex cases and healthcare problems on their own without having to refer to HSC specialists for assistance. In teleconsultation projects that continued to be effectively utilized, participant roles evolved and were expanded. Further, technology requirements for teleconsultation projects whose emergent behavior was learning did not need to be terribly sophisticated. CONCLUSIONS: When a teleconsultation project is designed with a mechanistic dominant logic, it is less likely to be sustained, whereas a teleconsultation project designed with a CAS dominant logic is more likely to be sustained. Consistent with a CAS dominant logic, teleconsultation projects that continued to be utilized involved participants taking on new roles and continuously learning. This continuous learning enabled remote site generalists to better handle the constantly changing nature of the problems faced. A CAS dominant logic provides a theoretical framework which explains why the teleconsultation literature about the role of technology, which is based on a mechanistic dominate logic, does not have adequate explanatory power.
Assuntos
Modelos Organizacionais , Consulta Remota/estatística & dados numéricos , HumanosRESUMO
Secondary cell death via gap junctions (GJs) plays a role in the propagation of neuronal loss under a number of degenerative disorders. Here, we examined the role of GJs in neuronal death in the retina, which has arguably the most diverse expression of GJs in the CNS. Initially, we induced apoptotic death by injecting single retinal ganglion cells and glia with cytochrome C and found that this resulted in the loss of neighboring cells to which they were coupled via GJs. We next found that pharmacological blockade of GJs eradicated nearly all amacrine cell loss and reduced retinal ganglion cell loss by â¼70% after induction of either excitotoxic or ischemic insult conditions. These data indicate that the GJ-mediated secondary cell death was responsible for the death of most cells. Whereas genetic deletion of the GJ subunit Cx36 increased cell survivability by â¼50% under excitotoxic condition, cell loss in Cx45 knock-out mouse retinas was similar to that seen in wild-type mice. In contrast, ablation of Cx45 reduced neuronal loss by â¼50% under ischemic insult, but ablation of Cx36 offered no protection. Immunolabeling of the connexins showed differential changes in protein expression consistent with their differing roles in propagating death signals under the two insults. These data indicate that secondary cell death is mediated by different cohorts of GJs dependent on the connexins they express and the type of initial insult. Our results suggest that targeting specific connexins offers a novel therapeutic strategy to reduce progressive cell loss under different neurodegenerative conditions.
Assuntos
Apoptose/fisiologia , Conexinas/metabolismo , Junções Comunicantes/fisiologia , Retina/citologia , Células Ganglionares da Retina/fisiologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Toxina da Cólera/metabolismo , Colina O-Acetiltransferase/metabolismo , Conexinas/genética , Inibidores Enzimáticos/farmacologia , Agonistas de Aminoácidos Excitatórios/toxicidade , Feminino , Fluoresceínas , Junções Comunicantes/efeitos dos fármacos , Junções Comunicantes/genética , Proteína Glial Fibrilar Ácida/metabolismo , Ácido Glicirretínico/análogos & derivados , Ácido Glicirretínico/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , N-Metilaspartato/toxicidade , Retina/lesões , Células Ganglionares da Retina/efeitos dos fármacos , Vias Visuais/efeitos dos fármacos , Vias Visuais/metabolismoRESUMO
The existence of functional connexin36 (Cx36) hemichannels in ß-cells was investigated in pancreatic islets of rat and wild-type (Cx36(+/+)), monoallelic (Cx36(+/-)), and biallelic (Cx36(-/-)) knockout mice. Hemichannel opening by KCl depolarization was studied by measuring ATP release and changes of intracellular ATP (ADP). Cx36(+/+) islets lost ATP after depolarization with 70 mM KCl at 5 mM glucose; ATP loss was prevented by 8 and 20 mM glucose or 50 µM mefloquine (connexin inhibitor). ATP content was higher in Cx36(-/-) than Cx36(+/+) islets and was not decreased by KCl depolarization; Cx36(+/-) islets showed values between that of control and homozygous islets. Five minimolar extracellular ATP increased ATP content and ATP/ADP ratio and induced a biphasic insulin secretion in depolarized Cx36(+/+) and Cx36(+/-) but not Cx36(-/-) islets. Cx36 hemichannels expressed in oocytes opened upon depolarization of membrane potential, and their activation was inhibited by mefloquine and glucose (IC50 â¼8 mM). It is postulated that glucose-induced inhibition of Cx36 hemichannels in islet ß-cells might avoid depolarization-induced ATP loss, allowing an optimum increase of the ATP/ADP ratio by sugar metabolism and a biphasic stimulation of insulin secretion. Gradual suppression of glucose-induced insulin release in Cx36(+/-) and Cx36(-/-) islets confirms that Cx36 gap junction channels are necessary for a full secretory stimulation and might account for the glucose intolerance observed in mice with defective Cx36 expression. Mefloquine targeting of Cx36 on both gap junctions and hemichannels also suppresses glucose-stimulated secretion. By contrast, glucose stimulation of insulin secretion requires Cx36 hemichannels' closure but keeping gap junction channels opened.
Assuntos
Glicemia/metabolismo , Conexinas/antagonistas & inibidores , Intolerância à Glucose/metabolismo , Hiperglicemia/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Regulação para Cima , Trifosfato de Adenosina/metabolismo , Animais , Glicemia/análise , Conexinas/genética , Conexinas/metabolismo , Junções Comunicantes/efeitos dos fármacos , Junções Comunicantes/metabolismo , Intolerância à Glucose/sangue , Heterozigoto , Hiperglicemia/etiologia , Secreção de Insulina , Células Secretoras de Insulina/efeitos dos fármacos , Masculino , Potenciais da Membrana/efeitos dos fármacos , Moduladores de Transporte de Membrana/farmacologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ratos , Ratos Wistar , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Técnicas de Cultura de Tecidos , Regulação para Cima/efeitos dos fármacos , Proteína delta-2 de Junções ComunicantesRESUMO
Hypothyroidism, defined as inadequate production of thyroid hormone, can be secondary to various underlying abnormalities in the pediatric population. Most frequently, hypothyroidism is related to structural abnormalities of the gland (dysgenesis), particularly in the neonatal population. However, other etiologies including intrinsic biochemical (dyshormonogenesis) and autoimmune abnormalities, as well as other rare causes, must be considered. Imaging is required to differentiate among the various etiologies of hypothyroidism and can be helpful in guiding therapy. This review aims to present an organized approach to hypothyroidism in the pediatric population, and assist the imager in guiding patient care.
Assuntos
Hipotireoidismo/diagnóstico , Aumento da Imagem/métodos , Posicionamento do Paciente/métodos , Cintilografia/métodos , Glândula Tireoide/diagnóstico por imagem , Ultrassonografia/métodos , Adolescente , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , RadiografiaRESUMO
Bipolar cells are the central neurons of the retina that transmit visual signals from rod and cone photoreceptors to third-order neurons in the inner retina and the brain. A dogma set forth by early anatomical studies is that bipolar cells in mammalian retinas receive segregated rod/cone synaptic inputs (either from rods or from cones), and here, we present evidence that challenges this traditional view. By analyzing light-evoked cation currents from morphologically identified depolarizing bipolar cells (DBCs) in the wild-type and three pathway-specific knockout mice (rod transducin knockout [Tralpha(-/-)], connexin36 knockout [Cx36(-/-)], and transcription factor beta4 knockout [Bhlhb4(-/-)]), we show that a subpopulation of rod DBCs (DBC(R2)s) receives substantial input directly from cones and a subpopulation of cone DBCs (DBC(C1)s) receives substantial input directly from rods. These results provide evidence of the existence of functional rod-DBC(C) and cone-DBC(R) synaptic pathways in the mouse retina as well as the previously proposed rod hyperpolarizing bipolar-cells pathway. This is grounds for revising the mammalian rod/cone bipolar cell dogma.
Assuntos
Células Bipolares da Retina , Células Fotorreceptoras Retinianas Cones/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/fisiologia , Vias Visuais , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Forma Celular , Conexinas/genética , Conexinas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Técnicas de Patch-Clamp , Estimulação Luminosa , Células Bipolares da Retina/citologia , Células Bipolares da Retina/fisiologia , Células Fotorreceptoras Retinianas Cones/citologia , Células Fotorreceptoras Retinianas Bastonetes/citologia , Transmissão Sináptica/fisiologia , Transducina/genética , Transducina/metabolismo , Vias Visuais/anatomia & histologia , Vias Visuais/fisiologia , Proteína delta-2 de Junções ComunicantesRESUMO
Periodic Cheyne-Stokes breathing (CSB) oscillating between apnea and crescendo-decrescendo hyperpnea is the most common central apnea. Currently, there is no proven therapy for CSB, probably because the fundamental pathophysiological question of how the respiratory center generates this form of breathing instability is still unresolved. Therefore, we aimed to determine the respiratory motor pattern of CSB resulting from the interaction of inspiratory and expiratory oscillators and identify the neural mechanism responsible for breathing regularization induced by the supplemental CO2 administration. Analysis of the inspiratory and expiratory motor pattern in a transgenic mouse model lacking connexin-36 electrical synapses, the neonatal (P14) Cx36 knockout male mouse, with a persistent CSB, revealed that the reconfigurations recurrent between apnea and hyperpnea and vice versa result from cyclical turn on/off of active expiration driven by the expiratory oscillator, which acts as a master pacemaker of respiration and entrains the inspiratory oscillator to restore ventilation. The results also showed that the suppression of CSB by supplemental 12% CO2 in inhaled air is due to the stabilization of coupling between expiratory and inspiratory oscillators, which causes the regularization of respiration. CSB rebooted after washout of CO2 excess when the inspiratory activity depressed again profoundly, indicating that the disability of the inspiratory oscillator to sustain ventilation is the triggering factor of CSB. Under these circumstances, the expiratory oscillator activated by the cyclic increase of CO2 behaves as an "anti-apnea" center generating the crescendo-decrescendo hyperpnea and periodic breathing. The neurogenic mechanism of CSB identified highlights the plasticity of the two-oscillator system in the neural control of respiration and provides a rationale base for CO2 therapy.
RESUMO
Mechanosensory neurons innervating the skin underlie our sense of touch. Fast-conducting, rapidly adapting mechanoreceptors innervating glabrous (non-hairy) skin form Meissner corpuscles, while in hairy skin, they associate with hair follicles, forming longitudinal lanceolate endings. How mechanoreceptors develop axonal endings appropriate for their skin targets is unknown. We report that mechanoreceptor morphologies across different skin regions are indistinguishable during early development but diverge post-natally, in parallel with skin maturation. Neurons terminating along the glabrous and hairy skin border exhibit hybrid morphologies, forming both Meissner corpuscles and lanceolate endings. Additionally, molecular profiles of neonatal glabrous and hairy skin-innervating neurons largely overlap. In mouse mutants with ectopic glabrous skin, mechanosensory neurons form end-organs appropriate for the altered skin type. Finally, BMP5 and BMP7 are enriched in glabrous skin, and signaling through type I bone morphogenetic protein (BMP) receptors in neurons is critical for Meissner corpuscle morphology. Thus, mechanoreceptor morphogenesis is flexibly instructed by target tissues.
Assuntos
Mecanorreceptores , Neurônios , Camundongos , Animais , Mecanorreceptores/metabolismo , Pele/inervação , Tato/fisiologia , CabeloRESUMO
Bipolar cells are the central neurons of the retina that convey visual signals from rod and cone photoreceptors in the outer retina to higher-order neurons in the inner retina and the brain. Early anatomical studies have suggested that there are four types of cone hyperpolarizing (OFF) bipolar cells (HBCs) in the mouse retina, but no light responses have been systematically examined. By analysing light-evoked cation and chloride currents (I(C) and I(Cl)) from over 50 morphologically identified HBCs in the dark-adapted wildtype and connexin36 knockout (Cx36(-/-)) mouse retinas, we identified three types of HBCs, each with distinct light responses and morphological characteristics. The HBC(R/MC)s with axon terminals ramifying between 0% and 30% of the inner plexiform layer (IPL) receive mixed inputs from rods and M-cones, the HBC(MC)s with axon terminals ramifying between 10% and 50% of the IPL receive inputs primarily from M-cones, and the HBC(M/SC)s with axon terminals ramifying between 25% and 50% of IPL receive inputs primarily from cones with mixed M- and S-cone pigments. Moreover, we found that HBC(R/MC)s in the Cx36(-/-) mice exhibit light responses very similar to the wildtype HBC(R/MC)s, suggesting that the mixed rod-cone inputs are not mediated by connexin36-dependent rod-cone coupling, but rather by direct synaptic contacts from rods and M-cones. This study constitutes the first systematic investigation that correlates light response characteristics and axonal morphology of HBCs in dark-adapted mouse retina, and contributes to recently emerging evidence that revises the traditional view that mammalian HBCs only contact cone photoreceptors.
Assuntos
Células Fotorreceptoras Retinianas Cones/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/fisiologia , Sinapses/fisiologia , Animais , Conexinas/deficiência , Conexinas/genética , Luz , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína delta-2 de Junções ComunicantesRESUMO
Claudins are tight junction integral membrane proteins that are key regulators of the paracellular pathway. Defects in claudin-16 (CLDN16) and CLDN19 function result in the inherited human renal disorder familial hypomagnesemia with hypercalciuria and nephrocalcinosis (FHHNC). Previous studies showed that siRNA knockdown of CLDN16 in mice results in a mouse model for FHHNC. Here, we show that CLDN19-siRNA mice also developed the FHHNC symptoms of chronic renal wasting of magnesium and calcium together with defective renal salt handling. siRNA knockdown of CLDN19 caused a loss of CLDN16 from tight junctions in the thick ascending limb (TAL) without a decrease in CLDN16 expression level, whereas siRNA knockdown of CLDN16 produced a similar effect on CLDN19. In both mouse lines, CLDN10, CLDN18, occludin, and ZO-1, normal constituents of TAL tight junctions, remained correctly localized. CLDN16- and CLDN19-depleted tight junctions had normal barrier function but defective ion selectivity. These data, together with yeast two-hybrid binding studies, indicate that a heteromeric CLDN16 and CLDN19 interaction was required for assembling them into the tight junction structure and generating cation-selective paracellular channels.
Assuntos
Alça do Néfron/metabolismo , Magnésio/metabolismo , Proteínas de Membrana/metabolismo , Junções Íntimas/metabolismo , Absorção , Animais , Claudinas , Clonagem Molecular , Immunoblotting , Lentivirus , Proteínas de Membrana/genética , Camundongos , Camundongos Transgênicos , Microscopia de Fluorescência , Oligonucleotídeos/genéticaRESUMO
In the retina, signals originating from rod and cone photoreceptors can reach retinal ganglion cells (RGCs)-the output neurons-through different pathways. However, little is known about the exact sensitivities and operating ranges of these pathways. Previously, we created rod- or cone-specific Cx36 knockout (KO) mouse lines. Both lines are deficient in rod/cone electrical coupling and therefore provide a way to selectively remove the secondary rod pathway. We measured the threshold of the primary rod pathway in RGCs of wild-type mice. Under pharmacological blockade of the primary rod pathway, the threshold was elevated. This secondary component was removed in the Cx36 KOs to unmask the threshold of the third rod pathway, still below cone threshold. In turn, the cone threshold was estimated by several independent methods. Our work defines the functionality of the secondary rod pathway and describes an additive contribution of the different pathways to the retinal output.
RESUMO
CNS glia exhibit a variety of gap junctional interactions: between neighboring astrocytes, between neighboring oligodendrocytes, between astrocytes and oligodendrocytes, and as 'reflexive' structures between layers of myelin in oligodendrocytes. Together, these junctions are thought to form a network facilitating absorption and removal of extracellular K(+) released during neuronal activity. In mice, loss of the two major oligodendrocyte connexins causes severe demyelination and early mortality, while loss of the two major astrocyte connexins causes mild dysmyelination and sensorimotor impairment, suggesting that reflexive and/or oligo-oligo coupling may be more important for the maintenance of myelin than other forms. To further explore the functional relationships between glial connexins, we generated double knockout mice lacking one oligodendrocyte and one astrocyte connexin. Cx32-Cx43 dKO animals develop white matter vacuolation without obvious ultrastructural abnormalities in myelin. Progressive loss of astrocytes but not oligodendrocytes or microglia accompanies sensorimotor impairment, seizure activity and early mortality at around 16 weeks of age. Our data reveal an unexpected role for connexins in the survival of white matter astrocytes, requiring the expression of particular isoforms in both oligodendrocytes and astrocytes.
Assuntos
Astrócitos/metabolismo , Conexina 43/deficiência , Conexinas/deficiência , Leucoencefalopatias , Fibras Nervosas Mielinizadas/patologia , Oligodendroglia/metabolismo , Fatores Etários , Animais , Morte Celular , Progressão da Doença , Proteína Glial Fibrilar Ácida/genética , Humanos , Leucoencefalopatias/complicações , Leucoencefalopatias/genética , Leucoencefalopatias/mortalidade , Camundongos , Camundongos Transgênicos , Microscopia Eletrônica de Transmissão , Transtornos dos Movimentos/etiologia , Transtornos dos Movimentos/genética , Transtornos dos Movimentos/patologia , Bainha de Mielina/patologia , Bainha de Mielina/ultraestrutura , Fibras Nervosas Mielinizadas/ultraestrutura , Convulsões/etiologia , Convulsões/genética , Convulsões/patologia , Proteína beta-1 de Junções ComunicantesRESUMO
Human genetic diseases and mouse knockouts illustrate that the maintenance of central nervous system myelin requires connexin expression by both astrocytes and oligodendrocytes. Because these cell types express nonoverlapping sets of connexins, the intercellular channels formed between them must be asymmetric with regard to connexin content, defined as heterotypic. Here, we show that oligodendrocyte Cx47 can form heterotypic channels with astrocyte Cx43 or Cx30 but not Cx26, whereas oligodendrocyte Cx32 can functionally interact with astrocyte Cx30 or Cx26 but not Cx43. Thus, as many as four types of intercellular channels could be formed between astrocytes and oligodendrocytes.
Assuntos
Astrócitos/metabolismo , Comunicação Celular/fisiologia , Conexinas/metabolismo , Junções Comunicantes/metabolismo , Oligodendroglia/metabolismo , Animais , Células Cultivadas , Conexina 26 , Conexinas/genética , Junções Comunicantes/genética , Células HeLa , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Transgênicos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Tight junctions (TJs) play a key role in mediating paracellular ion reabsorption in the kidney. Familial hypomagnesemia with hypercalciuria and nephrocalcinosis (FHHNC) is an inherited disorder caused by mutations in the genes encoding the TJ proteins claudin-16 (CLDN16) and CLDN19; however, the mechanisms underlying the roles of these claudins in mediating paracellular ion reabsorption in the kidney are not understood. Here we showed that in pig kidney epithelial cells, CLDN19 functioned as a Cl(-) blocker, whereas CLDN16 functioned as a Na(+) channel. Mutant forms of CLDN19 that are associated with FHHNC were unable to block Cl(-) permeation. Coexpression of CLDN16 and CLDN19 generated cation selectivity of the TJ in a synergistic manner, and CLDN16 and CLDN19 were observed to interact using several criteria. In addition, disruption of this interaction by introduction of FHHNC-causing mutant forms of either CLDN16 or CLDN19 abolished their synergistic effect. Our data show that CLDN16 interacts with CLDN19 and that their association confers a TJ with cation selectivity, suggesting a mechanism for the role of mutant forms of CLDN16 and CLDN19 in the development of FHHNC.
Assuntos
Hipercalciúria/genética , Magnésio/metabolismo , Proteínas de Membrana/metabolismo , Nefrocalcinose/genética , Canais de Sódio/metabolismo , Junções Íntimas/metabolismo , Animais , Cátions Monovalentes/metabolismo , Células Cultivadas , Cloro/metabolismo , Claudinas , Imunoprecipitação , Transporte de Íons , Proteínas de Membrana/genética , Mutação , Permeabilidade , Canais de Sódio/genética , SuínosRESUMO
Emerging studies indicate that connexins have activities completely unrelated to gap junctions and, conversely, that non-connexin proteins can form gap junction channels.
Assuntos
Conexinas/fisiologia , Junções Comunicantes/fisiologia , Transdução de Sinais/fisiologia , Bactérias/patogenicidade , Transporte Biológico/fisiologia , Cálcio/metabolismo , Proliferação de Células , Modelos BiológicosRESUMO
Neural circuits at the brainstem involved in the central generation of the motor patterns of respiration and cardiorespiratory chemoreflexes organize as cell assemblies connected by chemical and electrical synapses. However, the role played by the electrical connectivity mainly mediated by connexin36 (Cx36), which expression reaches peak value during the postnatal period, is still unknown. To address this issue, we analyzed here the respiratory phenotype of a mouse strain devoid constitutively of Cx36 at P14. Male Cx36-knockout mice at rest showed respiratory instability of variable degree, including a periodic Cheyne-Stokes breathing. Moreover, mice lacking Cx36 exhibited exacerbated chemoreflexes to normoxic and hypoxic hypercapnia characterized by a stronger inspiratory/expiratory coupling due to an increased sensitivity to CO2 . Deletion of Cx36 also impaired the generation of the recurrent episodes of transient bradycardia (ETBs) evoked during hypercapnic chemoreflexes; these EBTs constituted a powerful mechanism of cardiorespiratory coupling capable of improving alveolar gaseous exchange under hypoxic hypercapnia conditions. Approximately half of the homo- and heterozygous Cx36KO, but none WT, mice succumbed by respiratory arrest when submitted to hypoxia-hypercapnia, the principal exogenous stressor causing sudden infant death syndrome (SIDS). The early suppression of EBTs, which worsened arterial O2 saturation, and the generation of a paroxysmal generalized clonic-tonic activity, which provoked the transition from eupneic to gasping respiration, were the critical events causing sudden death in the Cx36KO mice. These results indicate that Cx36 expression plays a pivotal role in respiratory control, cardiorespiratory coordination, and protection against SIDS at the postnatal period.
Assuntos
Conexinas/genética , Respiração , Morte Súbita do Lactente/genética , Animais , Conexinas/metabolismo , Feminino , Deleção de Genes , Humanos , Lactente , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Reflexo , Centro Respiratório/metabolismo , Centro Respiratório/fisiopatologia , Proteína delta-2 de Junções ComunicantesRESUMO
Mouse photoreceptors are electrically coupled via gap junctions, but the relative importance of rod/rod, cone/cone, or rod/cone coupling is unknown. Furthermore, while connexin36 (Cx36) is expressed by cones, the identity of the rod connexin has been controversial. We report that FACS-sorted rods and cones both express Cx36 but no other connexins. We created rod- and cone-specific Cx36 knockout mice to dissect the photoreceptor network. In the wild type, Cx36 plaques at rod/cone contacts accounted for more than 95% of photoreceptor labeling and paired recordings showed the transjunctional conductance between rods and cones was ~300 pS. When Cx36 was eliminated on one side of the gap junction, in either conditional knockout, Cx36 labeling and rod/cone coupling were almost abolished. We could not detect direct rod/rod coupling, and cone/cone coupling was minor. Rod/cone coupling is so prevalent that indirect rod/cone/rod coupling via the network may account for previous reports of rod coupling.
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BACKGROUND: Gap junction protein and extracellular matrix signalling systems act in concert to influence developmental specification of neural stem and progenitor cells. It is not known how these two signalling systems interact. Here, we examined the role of ECM components in regulating connexin expression and function in postnatal hippocampal progenitor cells. RESULTS: We found that Cx26, Cx29, Cx30, Cx37, Cx40, Cx43, Cx45, and Cx47 mRNA and protein but only Cx32 and Cx36 mRNA are detected in distinct neural progenitor cell populations cultured in the absence of exogenous ECM. Multipotential Type 1 cells express Cx26, Cx30, and Cx43 protein. Their Type 2a progeny but not Type 2b and 3 neuronally committed progenitor cells additionally express Cx37, Cx40, and Cx45. Cx29 and Cx47 protein is detected in early oligodendrocyte progenitors and mature oligodendrocytes respectively. Engagement with a laminin substrate markedly increases Cx26 protein expression, decreases Cx40, Cx43, Cx45, and Cx47 protein expression, and alters subcellular localization of Cx30. These changes are associated with decreased neurogenesis. Further, laminin elicits the appearance of Cx32 protein in early oligodendrocyte progenitors and Cx36 protein in immature neurons. These changes impact upon functional connexin-mediated hemichannel activity but not gap junctional intercellular communication. CONCLUSION: Together, these findings demonstrate a new role for extracellular matrix-cell interaction, specifically laminin, in the regulation of intrinsic connexin expression and function in postnatal neural progenitor cells.
Assuntos
Conexinas/metabolismo , Matriz Extracelular/fisiologia , Hipocampo/fisiologia , Neurônios/fisiologia , Células-Tronco/fisiologia , Animais , Western Blotting , Comunicação Celular/fisiologia , Células Cultivadas , Conexinas/genética , Citometria de Fluxo , Imuno-Histoquímica , Laminina/metabolismo , Camundongos , Camundongos Knockout , Neurogênese/fisiologia , Oligodendroglia/fisiologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Primary hyperparathyroidism (PHPT) and familial hypocalciuric hypercalcemia (FHH) have significantly different treatment approaches, so physicians must be careful to differentiate these 2 diseases. Herein, we report a 14-year-old female who presented with symptomatic hypercalcemia (12 mg/dL; reference range, 9.2-10.7 mg/dL), elevated intact parathyroid hormone (iPTH) (236 pg/mL; reference range, 9-69 pg/mL), and vitamin D deficiency (6 ng/mL; reference range, ≥ 20 ng/mL). On numerous occasions, her 24-hour urine calcium was more than 4 mg/kg/day, consistent with PHPT, but her fractional excretion of calcium on 24-hour urine collection was consistently below 1%, in line with FHH. 99mTc-Sestamibi scan failed to detect any abnormalities. However, a 4-dimensional computed tomography scan of the neck revealed a right superior parathyroid adenoma which was excised with a focused parathyroidectomy. Although the patient's calcium and iPTH levels normalized, her nonspecific symptoms persisted. This case illustrates both the challenges of differentiating PHPT from FHH and the limitations of a first-line imaging tool in identifying a parathyroid adenoma.