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Sample records for gap junction resistance

  1. Gap Junctions

    PubMed Central

    Nielsen, Morten Schak; Axelsen, Lene Nygaard; Sorgen, Paul L.; Verma, Vandana; Delmar, Mario; Holstein-Rathlou, Niels-Henrik

    2013-01-01

    Gap junctions are essential to the function of multicellular animals, which require a high degree of coordination between cells. In vertebrates, gap junctions comprise connexins and currently 21 connexins are known in humans. The functions of gap junctions are highly diverse and include exchange of metabolites and electrical signals between cells, as well as functions, which are apparently unrelated to intercellular communication. Given the diversity of gap junction physiology, regulation of gap junction activity is complex. The structure of the various connexins is known to some extent; and structural rearrangements and intramolecular interactions are important for regulation of channel function. Intercellular coupling is further regulated by the number and activity of channels present in gap junctional plaques. The number of connexins in cell-cell channels is regulated by controlling transcription, translation, trafficking, and degradation; and all of these processes are under strict control. Once in the membrane, channel activity is determined by the conductive properties of the connexin involved, which can be regulated by voltage and chemical gating, as well as a large number of posttranslational modifications. The aim of the present article is to review our current knowledge on the structure, regulation, function, and pharmacology of gap junctions. This will be supported by examples of how different connexins and their regulation act in concert to achieve appropriate physiological control, and how disturbances of connexin function can lead to disease. © 2012 American Physiological Society. Compr Physiol 2:1981-2035, 2012. PMID:23723031

  2. Gap state charge induced spin-dependent negative differential resistance in tunnel junctions

    NASA Astrophysics Data System (ADS)

    Jiang, Jun; Zhang, X.-G.; Han, X. F.

    2016-04-01

    We propose and demonstrate through first-principles calculation a new spin-dependent negative differential resistance (NDR) mechanism in magnetic tunnel junctions (MTJ) with cubic cation disordered crystals (CCDC) AlO x or Mg1‑x Al x O as barrier materials. The CCDC is a class of insulators whose band gap can be changed by cation doping. The gap becomes arched in an ultrathin layer due to the space charge formed from metal-induced gap states. With an appropriate combination of an arched gap and a bias voltage, NDR can be produced in either spin channel. This mechanism is applicable to 2D and 3D ultrathin junctions with a sufficiently small band gap that forms a large space charge. It provides a new way of controlling the spin-dependent transport in spintronic devices by an electric field. A generalized Simmons formula for tunneling current through junction with an arched gap is derived to show the general conditions under which ultrathin junctions may exhibit NDR.

  3. Gap-junctional communication of bone marrow stromal cells is resistant to irradiation in vitro

    SciTech Connect

    Umezawa, A.; Harigaya, K.; Abe, H.; Watanabe, Y. )

    1990-10-01

    Bone marrow is one of the most radiosensitive organs. Irradiation causes a marked decrease in the total number of hematopoietic cells in the bone marrow. The reticular meshwork structure of marrow stromal cells, however, is relatively resistant to irradiation. Unimpaired stromal cell structure has been thought to be a prerequisite for the repopulation of hematopoietic cells during recovery from the effects of irradiation. The reticular framework is maintained by cell adhesion apparatuses such as gap junctions. The in vitro radiobiologic survival values of a cloned stromal cell line, H-1/A, were studied (n = 1.8, D0 = 138 cGy). Radiation doses of up to 4000 cGy had no detectable effects on the production of colony-stimulating factor 1. H-1/A cells communicate with each other via gap junctions as determined by the sensitive dye-transfer method. Gap-junctional communication between H-1/A cells was resistant to different levels of irradiation (500 to 10,000 cGy), but it was lost during adipocyte differentiation of H-1/A cells. Marrow stromal cells, which are important in the recovery of hematopoiesis, seemed capable of coordination with each other through gap junctions even when exposed to radiation.

  4. [Gap junction and diabetic foot].

    PubMed

    Zou, Xiao-rong; Tao, Jian; Wang, Yun-kai

    2015-11-01

    Gap junctions play a critical role in electrical synchronization and exchange of small molecules between neighboring cells; connexins are a family of structurally related transmembrane proteins that assemble to form vertebrate gap junctions. Hyperglycemia changes the structure gap junction proteins and their expression, resulting in obstruction of neural regeneration, vascular function and wound healing, and also promoting vascular atherosclerosis. These pathogenic factors would cause diabetic foot ulcers. This article reviews the involvement of connexins in pathogenesis of diabetic foot. PMID:26822053

  5. GAP JUNCTION FUNCTION AND CANCER

    EPA Science Inventory

    Gap Junctions (GJs) provide cell-to-cell communication (GJIC) of essential metabolites and ions. Js allow tissues to average responses, clear waste products, and minimize the effects of xenobiotics by dilution and allowing steady-state catabolism. any chemicals can adversely affe...

  6. 1.00 MeV proton radiation resistance studies of single-junction and single gap dual-junction amorphous-silicon alloy solar cells

    NASA Technical Reports Server (NTRS)

    Abdulaziz, Salman; Payson, J. S.; Li, Yang; Woodyard, James R.

    1990-01-01

    A comparative study of the radiation resistance of a-Si:H and a-SiGe:H single-junction and a-Si:H dual-junction solar cells was conducted. The cells were irradiated with 1.00-MeV protons with fluences of 1.0 x 10 to the 14th, 5.0 x 10 to the 14th and 1.0 x 10 to the 15th/sq cm and characterized using I-V and quantum efficiency measurements. The radiation resistance of single-junction cells cannot be used to explain the behavior of dual-junction cells at a fluence of 1.0 x 10 to the 15th/sq cm. The a-Si H single-junction cells degraded the least of the three cells; a-SiGe:H single-junction cells showed the largest reduction in short-circuit current, while a-Si:H dual-junction cells exhibited the largest degradation in the open-circuit voltage. The quantum efficiency of the cells degraded more in the red part of the spectrum; the bottom junction degrades first in dual-junction cells.

  7. Gap junctions as electrical synapses.

    PubMed

    Bennett, M V

    1997-06-01

    Gap junctions are the morphological substrate of one class of electrical synapse. The history of the debate on electrical vs. chemical transmission is instructive. One lesson is that Occam's razor sometimes cuts too deep; the nervous system does its operations in a number of different ways and a unitarian approach can lead one astray. Electrical synapses can do many things that chemical synapses can do, and do them just as slowly. More intriguing are the modulatory actions that chemical synapses can have on electrical synapses. Voltage dependence provides an important window on structure function relations of the connexins, even where the dependence may have no physiological role. The new molecular approaches will greatly advance our knowledge of where gap junctions occur and permit experimental manipulation with high specificity. PMID:9278865

  8. Communication Through Gap Junctions in the Endothelium.

    PubMed

    Schmidt, K; Windler, R; de Wit, C

    2016-01-01

    A swarm of fish displays a collective behavior (swarm behavior) and moves "en masse" despite the huge number of individual animals. In analogy, organ function is supported by a huge number of cells that act in an orchestrated fashion and this applies also to vascular cells along the vessel length. It is obvious that communication is required to achieve this vital goal. Gap junctions with their modular bricks, connexins (Cxs), provide channels that interlink the cytosol of adjacent cells by a pore sealed against the extracellular space. This allows the transfer of ions and charge and thereby the travel of membrane potential changes along the vascular wall. The endothelium provides a low-resistance pathway that depends crucially on connexin40 which is required for long-distance conduction of dilator signals in the microcirculation. The experimental evidence for membrane potential changes synchronizing vascular behavior is manifold but the functional verification of a physiologic role is still open. Other molecules may also be exchanged that possibly contribute to the synchronization (eg, Ca(2+)). Recent data suggest that vascular Cxs have more functions than just facilitating communication. As pharmacological tools to modulate gap junctions are lacking, Cx-deficient mice provide currently the standard to unravel their vascular functions. These include arteriolar dilation during functional hyperemia, hypoxic pulmonary vasoconstriction, vascular collateralization after ischemia, and feedback inhibition on renin secretion in the kidney. PMID:27451099

  9. Adrenocortical Gap Junctions and Their Functions

    PubMed Central

    Bell, Cheryl L.; Murray, Sandra A.

    2016-01-01

    Adrenal cortical steroidogenesis and proliferation are thought to be modulated by gap junction-mediated direct cell–cell communication of regulatory molecules between cells. Such communication is regulated by the number of gap junction channels between contacting cells, the rate at which information flows between these channels, and the rate of channel turnover. Knowledge of the factors regulating gap junction-mediated communication and the turnover process are critical to an understanding of adrenal cortical cell functions, including development, hormonal response to adrenocorticotropin, and neoplastic dedifferentiation. Here, we review what is known about gap junctions in the adrenal gland, with particular attention to their role in adrenocortical cell steroidogenesis and proliferation. Information and insight gained from electrophysiological, molecular biological, and imaging (immunocytochemical, freeze fracture, transmission electron microscopic, and live cell) techniques will be provided. PMID:27445985

  10. Oxidative Stress, Lens Gap Junctions, and Cataracts

    PubMed Central

    Beyer, Eric C.

    2009-01-01

    Abstract The eye lens is constantly subjected to oxidative stress from radiation and other sources. The lens has several mechanisms to protect its components from oxidative stress and to maintain its redox state, including enzymatic pathways and high concentrations of ascorbate and reduced glutathione. With aging, accumulation of oxidized lens components and decreased efficiency of repair mechanisms can contribute to the development of lens opacities or cataracts. Maintenance of transparency and homeostasis of the avascular lens depend on an extensive network of gap junctions. Communication through gap junction channels allows intercellular passage of molecules (up to 1 kDa) including antioxidants. Lens gap junctions and their constituent proteins, connexins (Cx43, Cx46, and Cx50), are also subject to the effects of oxidative stress. These observations suggest that oxidative stress-induced damage to connexins (and consequent altered intercellular communication) may contribute to cataract formation. Antioxid. Redox Signal. 11, 339–353. PMID:18831679

  11. Clathrin and Cx43 gap junction plaque endoexocytosis

    SciTech Connect

    Nickel, Beth M.; DeFranco, B. Hewa; Gay, Vernon L.; Murray, Sandra A.

    2008-10-03

    In earlier transmission electron microscopic studies, we have described pentilaminar gap junctional membrane invaginations and annular gap junction vesicles coated with short, electron-dense bristles. The similarity between these electron-dense bristles and the material surrounding clathrin-coated pits led us to suggest that the dense bristles associated with gap junction structures might be clathrin. To confirm that clathrin is indeed associated with annular gap junction vesicles and gap junction plaques, quantum dot immuno-electron microscopic techniques were used. We report here that clathrin associates with both connexin 43 (Cx43) gap junction plaques and pentilaminar gap junction vesicles. An important finding was the preferential localization of clathrin to the cytoplasmic surface of the annular or of the gap junction plaque membrane of one of the two contacting cells. This is consistent with the possibility that the direction of gap junction plaque internalization into one of two contacting cells is regulated by clathrin.

  12. Magnesium gating of cardiac gap junction channels.

    PubMed

    Matsuda, Hiroyuki; Kurata, Yasutaka; Oka, Chiaki; Matsuoka, Satoshi; Noma, Akinori

    2010-09-01

    We aimed to study kinetics of modulation by intracellular Mg(2+) of cardiac gap junction (Mg(2+) gate). Paired myocytes of guinea-pig ventricle were superfused with solutions containing various concentrations of Mg(2+). In order to rapidly apply Mg(2+) to one aspect of the gap junction, the non-junctional membrane of one of the pair was perforated at nearly the connecting site by pulses of nitrogen laser beam. The gap junction conductance (G(j)) was measured by clamping the membrane potential of the other cell using two-electrode voltage clamp method. The laser perforation immediately increased G(j), followed by slow G(j) change with time constant of 3.5 s at 10 mM Mg(2+). Mg(2+) more than 1.0 mM attenuated dose-dependently the gap junction conductance and lower Mg(2+) (0.6 mM) increased G(j) with a Hill coefficient of 3.4 and a half-maximum effective concentration of 0.6 mM. The time course of G(j) changes was fitted by single exponential function, and the relationship between the reciprocal of time constant and Mg(2+) concentration was almost linear. Based on the experimental data, a mathematical model of Mg(2+) gate with one open state and three closed states well reproduced experimental results. One-dimensional cable model of thirty ventricular myocytes connected to the Mg(2+) gate model suggested a pivotal role of the Mg(2+) gate of gap junction under pathological conditions. PMID:20553744

  13. Lycopene oxidation product enhances gap junctional communication.

    PubMed

    Aust, O; Ale-Agha, N; Zhang, L; Wollersen, H; Sies, H; Stahl, W

    2003-10-01

    Carotenoids as well as their metabolites and oxidation products stimulate gap junctional communication (GJC) between cells, which is thought to be one of the protective mechanisms related to cancer-preventive activities of these compounds. Increased intake of lycopene by consumption of tomatoes or tomato products has been epidemiologically associated with a diminished risk of prostate cancer. Here, we report a stimulatory effect of a lycopene oxidation product on GJC in rat liver epithelial WB-F344 cells. The active compound was obtained by complete in vitro oxidation of lycopene with hydrogen peroxide/osmium tetroxide. For structural analysis high performance liquid chromatography, gas chromatography coupled with mass spectrometry, ultraviolet/visible-, and infrared spectrophotometry were applied. The biologically active oxidation product was identified as 2,7,11-trimethyl-tetradecahexaene-1,14-dial. The present data indicate a potential role of lycopene degradation products in cell signaling enhancing cell-to-cell communication via gap junctions. PMID:12909274

  14. GAP junctional communication in brain secondary organizers.

    PubMed

    Bosone, Camilla; Andreu, Abraham; Echevarria, Diego

    2016-06-01

    Gap junctions (GJs) are integral membrane proteins that enable the direct cytoplasmic exchange of ions and low molecular weight metabolites between adjacent cells. They are formed by the apposition of two connexons belonging to adjacent cells. Each connexon is formed by six proteins, named connexins (Cxs). Current evidence suggests that gap junctions play an important part in ensuring normal embryo development. Mutations in connexin genes have been linked to a variety of human diseases, although the precise role and the cell biological mechanisms of their action remain almost unknown. Among the big family of Cxs, several are expressed in nervous tissue but just a few are expressed in the anterior neural tube of vertebrates. Many efforts have been made to elucidate the molecular bases of Cxs cell biology and how they influence the morphogenetic signal activity produced by brain signaling centers. These centers, orchestrated by transcription factors and morphogenes determine the axial patterning of the mammalian brain during its specification and regionalization. The present review revisits the findings of GJ composed by Cx43 and Cx36 in neural tube patterning and discuss Cx43 putative enrollment in the control of Fgf8 signal activity coming from the well known secondary organizer, the isthmic organizer. PMID:27273333

  15. Thalamic Modulation of Cingulate Seizure Activity Via the Regulation of Gap Junctions in Mice Thalamocingulate Slice

    PubMed Central

    Chang, Wei-Pang; Wu, José Jiun-Shian; Shyu, Bai-Chuang

    2013-01-01

    The thalamus is an important target for deep brain stimulation in the treatment of seizures. However, whether the modulatory effect of thalamic inputs on cortical seizures occurs through the modulation of gap junctions has not been previously studied. Therefore, we tested the effects of different gap junction blockers and couplers in a drug-resistant seizure model and studied the role of gap junctions in the thalamic modulation on cortical seizures. Multielectrode array and calcium imaging were used to record the cortical seizures induced by 4-aminopyridine (250 µM) and bicuculline (5–50 µM) in a novel thalamocingulate slice preparation. Seizure-like activity was significantly attenuated by the pan-gap junction blockers carbenoxolone and octanol and specific neuronal gap junction blocker mefloquine. The gap junction coupler trimethylamine significantly enhanced seizure-like activity. Gap junction blockers did not influence the initial phase of seizure-like activity, but they significantly decreased the amplitude and duration of the maintenance phase. The development of seizures is regulated by extracellular potassium concentration. Carbenoxolone partially restored the amplitude and duration after removing the thalamic inputs. A two-dimensional current source density analysis showed that the sink and source signals shifted to deeper layers after removing the thalamic inputs during the clonic phase. These results indicate that the regulatory mechanism of deep brain stimulation in the thalamus occurs partially though gap junctions. PMID:23690968

  16. Terbinafine inhibits gap junctional intercellular communication.

    PubMed

    Lee, Ju Yeun; Yoon, Sei Mee; Choi, Eun Ju; Lee, Jinu

    2016-09-15

    Terbinafine is an antifungal agent that selectively inhibits fungal sterol synthesis by blocking squalene epoxidase. We evaluated the effect of terbinafine on gap junctional intercellular communication (GJIC). Fluorescence recovery after photobleaching (FRAP) and I-YFP GJIC assays revealed that terbinafine inhibits GJIC in a reversible and dose-dependent manner in FRT-Cx43 and LN215 cells. Treatment with terbinafine did not affect Cx43 phosphorylation status or intracellular Ca(2+) concentration, well-known action mechanisms of various GJIC blockers. While a structurally related chemical, naftifine, attenuated GJIC, epigallocatechin gallate, another potent squalene epoxidase inhibitor with a different structure, did not. These results suggest that terbinafine inhibits GJIC with a so far unknown mechanism of action. PMID:27487578

  17. Electromechanical resistive switching via back-to-back Schottky junctions

    SciTech Connect

    Li, Lijie

    2015-09-15

    The physics of the electromechanical resistive switching is uncovered using the theory of back-to-back Schottky junctions combined with the quantum domain space charge transport. A theoretical model of the basic element of resistive switching devices realized by the metal-ZnO nanowires-metal structure has been created and analyzed. Simulation results show that the reverse biased Schottky junction and the air gap impedance dominate the current-voltage relation at higher external voltages; thereby electromechanically varying the air gap thickness causes the device exhibit resistive tuning characteristics. As the device dimension is in nanometre scale, investigation of the model based on quantum mechanics has also been conducted.

  18. TEMPORAL CHANGE IN GAP JUNCTION FUNCTION IN PRIMARY HEPATOCYTES

    EPA Science Inventory

    TEMPORAL CHANGES IN GAP JUNCTION FUNCTION IN PRIMARY *

    The objective of this study was to examine the reduction in gap junction communication (GJC) in primary hepatocytes due to coincident melatonin and magnetic field treatments to determine if these conditions could prov...

  19. Differences between liver gap junction protein and lens MIP 26 from rat: implications for tissue specificity of gap junctions.

    PubMed

    Nicholson, B J; Takemoto, L J; Hunkapiller, M W; Hood, L E; Revel, J P

    1983-03-01

    Liver gap junctions and gap-junction-like structures from eye lenses are each comprised of a single major protein (Mr 28,000 and 26,000, respectively). These proteins display different two-dimensional peptide fingerprints, distinct amino acid compositions, nonhomologous N-terminal amino acid sequences and different sensitivities to proteases when part of the intact junction. However, the junctional protein of each tissue is well conserved between species, as demonstrated previously for lens and now for liver in several mammalian species. The possiblity of tissue-specific gap junction proteins is discussed in the light of data suggesting that rat heart gap junctions are comprised of yet a third protein. PMID:6299583

  20. Heterotypic gap junctions at glutamatergic mixed synapses are abundant in goldfish brain.

    PubMed

    Rash, J E; Kamasawa, N; Vanderpool, K G; Yasumura, T; O'Brien, J; Nannapaneni, S; Pereda, A E; Nagy, J I

    2015-01-29

    Gap junctions provide for direct intercellular electrical and metabolic coupling. The abundance of gap junctions at "large myelinated club ending (LMCE)" synapses on Mauthner cells (M-cells) of the teleost brain provided a convenient model to correlate anatomical and physiological properties of electrical synapses. There, presynaptic action potentials were found to evoke short-latency electrical "pre-potentials" immediately preceding their accompanying glutamate-induced depolarizations, making these the first unambiguously identified "mixed" (i.e., chemical plus electrical) synapses in the vertebrate CNS. We recently showed that gap junctions at these synapses exhibit asymmetric electrical resistance (i.e., electrical rectification), which we correlated with total molecular asymmetry of connexin composition in their apposing gap junction hemiplaques, with connexin35 (Cx35) restricted to axon terminal hemiplaques and connexin34.7 (Cx34.7) restricted to apposing M-cell plasma membranes. We now show that similarly heterotypic neuronal gap junctions are abundant throughout goldfish brain, with labeling exclusively for Cx35 in presynaptic hemiplaques and exclusively for Cx34.7 in postsynaptic hemiplaques. Moreover, the vast majority of these asymmetric gap junctions occur at glutamatergic axon terminals. The widespread distribution of heterotypic gap junctions at glutamatergic mixed synapses throughout goldfish brain and spinal cord implies that pre- vs. postsynaptic asymmetry at electrical synapses evolved early in the chordate lineage. We propose that the advantages of the molecular and functional asymmetry of connexins at electrical synapses that are so prominently expressed in the teleost CNS are unlikely to have been abandoned in higher vertebrates. However, to create asymmetric coupling in mammals, where most gap junctions are composed of connexin36 (Cx36) on both sides, would require some other mechanism, such as differential phosphorylation of connexins on

  1. Heterotypic gap junctions at glutamatergic mixed synapses are abundant in goldfish brain

    PubMed Central

    Rash, John E.; Kamasawa, Naomi; Vanderpool, Kimberly G.; Yasumura, Thomas; O'Brien, John; Nannapaneni, Srikant; Pereda, Alberto E.; Nagy, James I.

    2014-01-01

    Gap junctions provide for direct intercellular electrical and metabolic coupling. The abundance of gap junctions at “large myelinated club ending” synapses on Mauthner cells of the teleost brain provided a convenient model to correlate anatomical and physiological properties of electrical synapses. There, presynaptic action potentials were found to evoke short-latency electrical “pre-potentials” immediately preceding their accompanying glutamate-induced depolarizations, making these the first unambiguously identified “mixed” (i.e., chemical plus electrical) synapses in the vertebrate CNS. We recently showed that gap junctions at these synapses exhibit asymmetric electrical resistance (i.e., electrical rectification), which we correlated with total molecular asymmetry of connexin composition in their apposing gap junction hemiplaques, with Cx35 restricted to axon terminal hemiplaques and Cx34.7 restricted to apposing Mauthner cell plasma membranes. We now show that similarly heterotypic neuronal gap junctions are abundant throughout goldfish brain, with labeling exclusively for Cx35 in presynaptic hemiplaques and exclusively for Cx34.7 in postsynaptic hemiplaques. Moreover, the vast majority of these asymmetric gap junctions occur at glutamatergic axon terminals. The widespread distribution of heterotypic gap junctions at glutamatergic mixed synapses throughout goldfish brain and spinal cord implies that pre- vs. postsynaptic asymmetry at electrical synapses evolved early in the chordate lineage. We propose that the advantages of the molecular and functional asymmetry of connexins at electrical synapses that are so prominently expressed in the teleost CNS are unlikely to have been abandoned in higher vertebrates. However, to create asymmetric coupling in mammals, where most gap junctions are composed of Cx36 on both sides, would require some other mechanism, such as differential phosphorylation of connexins on opposite sides of the same gap junction or

  2. Pallidal gap junctions-triggers of synchrony in Parkinson's disease?

    PubMed Central

    Schwab, Bettina C; Heida, Tjitske; Zhao, Yan; van Gils, Stephan A; van Wezel, Richard J A

    2014-01-01

    Although increased synchrony of the neural activity in the basal ganglia may underlie the motor deficiencies exhibited in Parkinson's disease (PD), how this synchrony arises, propagates through the basal ganglia, and changes under dopamine replacement remains unknown. Gap junctions could play a major role in modifying this synchrony, because they show functional plasticity under the influence of dopamine and after neural injury. In this study, confocal imaging was used to detect connexin-36, the major neural gap junction protein, in postmortem tissues of PD patients and control subjects in the putamen, subthalamic nucleus (STN), and external and internal globus pallidus (GPe and GPi, respectively). Moreover, we quantified how gap junctions affect synchrony in an existing computational model of the basal ganglia. We detected connexin-36 in the human putamen, GPe, and GPi, but not in the STN. Furthermore, we found that the number of connexin-36 spots in PD tissues increased by 50% in the putamen, 43% in the GPe, and 109% in the GPi compared with controls. In the computational model, gap junctions in the GPe and GPi strongly influenced synchrony. The basal ganglia became especially susceptible to synchronize with input from the cortex when gap junctions were numerous and high in conductance. In conclusion, connexin-36 expression in the human GPe and GPi suggests that gap junctional coupling exists within these nuclei. In PD, neural injury and dopamine depletion could increase this coupling. Therefore, we propose that gap junctions act as a powerful modulator of synchrony in the basal ganglia. PMID:25124148

  3. Pallidal gap junctions-triggers of synchrony in Parkinson's disease?

    PubMed

    Schwab, Bettina C; Heida, Tjitske; Zhao, Yan; van Gils, Stephan A; van Wezel, Richard J A

    2014-10-01

    Although increased synchrony of the neural activity in the basal ganglia may underlie the motor deficiencies exhibited in Parkinson's disease (PD), how this synchrony arises, propagates through the basal ganglia, and changes under dopamine replacement remains unknown. Gap junctions could play a major role in modifying this synchrony, because they show functional plasticity under the influence of dopamine and after neural injury. In this study, confocal imaging was used to detect connexin-36, the major neural gap junction protein, in postmortem tissues of PD patients and control subjects in the putamen, subthalamic nucleus (STN), and external and internal globus pallidus (GPe and GPi, respectively). Moreover, we quantified how gap junctions affect synchrony in an existing computational model of the basal ganglia. We detected connexin-36 in the human putamen, GPe, and GPi, but not in the STN. Furthermore, we found that the number of connexin-36 spots in PD tissues increased by 50% in the putamen, 43% in the GPe, and 109% in the GPi compared with controls. In the computational model, gap junctions in the GPe and GPi strongly influenced synchrony. The basal ganglia became especially susceptible to synchronize with input from the cortex when gap junctions were numerous and high in conductance. In conclusion, connexin-36 expression in the human GPe and GPi suggests that gap junctional coupling exists within these nuclei. In PD, neural injury and dopamine depletion could increase this coupling. Therefore, we propose that gap junctions act as a powerful modulator of synchrony in the basal ganglia. PMID:25124148

  4. Distal gap junctions and active dendrites can tune network dynamics.

    PubMed

    Saraga, Fernanda; Ng, Leo; Skinner, Frances K

    2006-03-01

    Gap junctions allow direct electrical communication between CNS neurons. From theoretical and modeling studies, it is well known that although gap junctions can act to synchronize network output, they can also give rise to many other dynamic patterns including antiphase and other phase-locked states. The particular network pattern that arises depends on cellular, intrinsic properties that affect firing frequencies as well as the strength and location of the gap junctions. Interneurons or GABAergic neurons in hippocampus are diverse in their cellular characteristics and have been shown to have active dendrites. Furthermore, parvalbumin-positive GABAergic neurons, also known as basket cells, can contact one another via gap junctions on their distal dendrites. Using two-cell network models, we explore how distal electrical connections affect network output. We build multi-compartment models of hippocampal basket cells using NEURON and endow them with varying amounts of active dendrites. Two-cell networks of these model cells as well as reduced versions are explored. The relationship between intrinsic frequency and the level of active dendrites allows us to define three regions based on what sort of network dynamics occur with distal gap junction coupling. Weak coupling theory is used to predict the delineation of these regions as well as examination of phase response curves and distal dendritic polarization levels. We find that a nonmonotonic dependence of network dynamic characteristics (phase lags) on gap junction conductance occurs. This suggests that distal electrical coupling and active dendrite levels can control how sensitive network dynamics are to gap junction modulation. With the extended geometry, gap junctions located at more distal locations must have larger conductances for pure synchrony to occur. Furthermore, based on simulations with heterogeneous networks, it may be that one requires active dendrites if phase-locking is to occur in networks formed

  5. Ischemic preconditioning protects against gap junctional uncoupling in cardiac myofibroblasts.

    PubMed

    Sundset, Rune; Cooper, Marie; Mikalsen, Svein-Ole; Ytrehus, Kirsti

    2004-01-01

    Ischemic preconditioning increases the heart's tolerance to a subsequent longer ischemic period. The purpose of this study was to investigate the role of gap junction communication in simulated preconditioning in cultured neonatal rat cardiac myofibroblasts. Gap junctional intercellular communication was assessed by Lucifer yellow dye transfer. Preconditioning preserved intercellular coupling after prolonged ischemia. An initial reduction in coupling in response to the preconditioning stimulus was also observed. This may protect neighboring cells from damaging substances produced during subsequent regional ischemia in vivo, and may preserve gap junctional communication required for enhanced functional recovery during subsequent reperfusion. PMID:16247851

  6. Study of Gap Junctions in Human Embryonic Stem Cells.

    PubMed

    Pébay, Alice; Wong, Raymond C B

    2016-01-01

    Gap junctional intercellular communication (GJIC) has been described in different cell types including stem cells and has been involved in different biological events. GJIC is required for mouse embryonic stem cell maintenance and proliferation and various studies suggest that functional GJIC is a common characteristic of human embryonic stem cells (hESC) maintained in different culture conditions. This chapter introduces methods to study gap junctions in hESC, from expression of gap junction proteins to functional study of GJIC in hESC proliferation, apoptosis, colony growth, and pluripotency. PMID:24859928

  7. Gap junction modulation and its implications for heart function

    PubMed Central

    Kurtenbach, Stefan; Kurtenbach, Sarah; Zoidl, Georg

    2014-01-01

    Gap junction communication (GJC) mediated by connexins is critical for heart function. To gain insight into the causal relationship of molecular mechanisms of disease pathology, it is important to understand which mechanisms contribute to impairment of gap junctional communication. Here, we present an update on the known modulators of connexins, including various interaction partners, kinases, and signaling cascades. This gap junction network (GJN) can serve as a blueprint for data mining approaches exploring the growing number of publicly available data sets from experimental and clinical studies. PMID:24578694

  8. Measurement of tunnel junction resistance during formation

    SciTech Connect

    Barber, W.C.; Johnson, R.T.; Lee, J.S.; Laws, K.E.; Bland, R.W. )

    1993-11-01

    The authors have measured the characteristics of aluminum tunnel junctions during and immediately after the formation of the junction. This has permitted us to observe changes in the oxide barrier, in vacuum and in air. By observing the barrier resistance during sputtering, they were able to diagnose and correct problems due to plasma discharges which were damaging the junctions. They report preliminary results from junctions passivated with a silicon nitride cap layer.

  9. Fixed-Gap Tunnel Junction for Reading DNA Nucleotides

    PubMed Central

    2015-01-01

    Previous measurements of the electronic conductance of DNA nucleotides or amino acids have used tunnel junctions in which the gap is mechanically adjusted, such as scanning tunneling microscopes or mechanically controllable break junctions. Fixed-junction devices have, at best, detected the passage of whole DNA molecules without yielding chemical information. Here, we report on a layered tunnel junction in which the tunnel gap is defined by a dielectric layer, deposited by atomic layer deposition. Reactive ion etching is used to drill a hole through the layers so that the tunnel junction can be exposed to molecules in solution. When the metal electrodes are functionalized with recognition molecules that capture DNA nucleotides via hydrogen bonds, the identities of the individual nucleotides are revealed by characteristic features of the fluctuating tunnel current associated with single-molecule binding events. PMID:25380505

  10. Reduction of Gap Junctional Conductance by Microinjection of Antibodies against the 27-kDa Liver Gap Junction Polypeptide

    NASA Astrophysics Data System (ADS)

    Hertzberg, E. L.; Spray, D. C.; Bennett, M. V. L.

    1985-04-01

    Antibody raised against isolated rat liver gap junctions was microinjected into coupled cells in culture to assess its influence on gap junctional conductance. A rapid inhibition of fluorescent dye transfer and electrical coupling was produced in pairs of freshly dissociated adult rat hepatocytes and myocardial cells as well as in pairs of superior cervical ganglion neurons from neonatal rats cultured under conditions in which electrotonic synapses form. The antibodies have been shown by indirect immunofluorescence to bind to punctate regions of the plasma membrane in liver. By immunoreplica analysis of rat liver homogenates, plasma membranes, and isolated gap junctions resolved on NaDodSO4/polyacrylamide gels, binding was shown to be specific for the 27-kDa major polypeptide of gap junctions. This and similar antibodies should provide a tool for further investigation of the role of cell-cell communication mediated by gap junctions and indicate that immunologically similar polypeptides comprise gap junctions in adult mammalian cells derived from all three germ layers.

  11. Reduction of gap junctional conductance by microinjection of antibodies against the 27-kDa liver gap junction polypeptide.

    PubMed Central

    Hertzberg, E L; Spray, D C; Bennett, M V

    1985-01-01

    Antibody raised against isolated rat liver gap junctions was microinjected into coupled cells in culture to assess its influence on gap junctional conductance. A rapid inhibition of fluorescent dye transfer and electrical coupling was produced in pairs of freshly dissociated adult rat hepatocytes and myocardial cells as well as in pairs of superior cervical ganglion neurons from neonatal rats cultured under conditions in which electrotonic synapses form. The antibodies have been shown by indirect immunofluorescence to bind to punctate regions of the plasma membrane in liver. By immunoreplica analysis of rat liver homogenates, plasma membranes, and isolated gap junctions resolved on NaDodSO4/polyacrylamide gels, binding was shown to be specific for the 27-kDa major polypeptide of gap junctions. This and similar antibodies should provide a tool for further investigation of the role of cell-cell communication mediated by gap junctions and indicate that immunologically similar polypeptides comprise gap junctions in adult mammalian cells derived from all three germ layers. Images PMID:2986116

  12. Dynamic gap junctional communication: a delimiting model for tissue responses.

    PubMed Central

    Christ, G J; Brink, P R; Ramanan, S V

    1994-01-01

    Gap junctions are aqueous intercellular channels formed by a diverse class of membrane-spanning proteins, known as connexins. These aqueous pores provide partial cytoplasmic continuity between cells in most tissues, and are freely permeable to a host of physiologically relevant second messenger molecules/ionic species (e.g., Ca2+, IP3, cAMP, cGMP). Despite the fact that these second messenger molecules/ionic species have been shown to alter junctional patency, there is no clear basis for understanding how dynamic and transient changes in the intracellular concentration of second messenger molecules might modulate the extent of intercellular communication among coupled cells. Thus, we have modified the tissue monolayer model of Ramanan and Brink (1990) to account for both the up-regulatory and down-regulatory effects on junctions by second messenger molecules that diffuse through gap junctions. We have chosen the vascular wall as our morphological correlate because of its anisotropy and large investment of gap junctions. The model allows us to illustrate the putative behavior of gap junctions under a variety of physiologically relevant conditions. The modeling studies demonstrated that transient alterations in intracellular second messenger concentrations are capable of producing 50-125% changes in the number of cells recruited into a functional syncytial unit, after activation of a single cell. Moreover, the model conditions required to demonstrate such physiologically relevant changes in intercellular diffusion among coupled cells are commonly observed in intact tissues and cultured cells. Images FIGURE 2 PMID:7811948

  13. Gap junctions in developing thalamic and neocortical neuronal networks.

    PubMed

    Niculescu, Dragos; Lohmann, Christian

    2014-12-01

    The presence of direct, cytoplasmatic, communication between neurons in the brain of vertebrates has been demonstrated a long time ago. These gap junctions have been characterized in many brain areas in terms of subunit composition, biophysical properties, neuronal connectivity patterns, and developmental regulation. Although interesting findings emerged, showing that different subunits are specifically regulated during development, or that excitatory and inhibitory neuronal networks exhibit various electrical connectivity patterns, gap junctions did not receive much further interest. Originally, it was believed that gap junctions represent simple passageways for electrical and biochemical coordination early in development. Today, we know that gap junction connectivity is tightly regulated, following independent developmental patterns for excitatory and inhibitory networks. Electrical connections are important for many specific functions of neurons, and are, for example, required for the development of neuronal stimulus tuning in the visual system. Here, we integrate the available data on neuronal connectivity and gap junction properties, as well as the most recent findings concerning the functional implications of electrical connections in the developing thalamus and neocortex. PMID:23843439

  14. Functional Properties of Dendritic Gap Junctions in Cerebellar Golgi Cells.

    PubMed

    Szoboszlay, Miklos; Lőrincz, Andrea; Lanore, Frederic; Vervaeke, Koen; Silver, R Angus; Nusser, Zoltan

    2016-06-01

    The strength and variability of electrical synaptic connections between GABAergic interneurons are key determinants of spike synchrony within neuronal networks. However, little is known about how electrical coupling strength is determined due to the inaccessibility of gap junctions on the dendritic tree. We investigated the properties of gap junctions in cerebellar interneurons by combining paired somato-somatic and somato-dendritic recordings, anatomical reconstructions, immunohistochemistry, electron microscopy, and modeling. By fitting detailed compartmental models of Golgi cells to their somato-dendritic voltage responses, we determined their passive electrical properties and the mean gap junction conductance (0.9 nS). Connexin36 immunofluorescence and freeze-fracture replica immunogold labeling revealed a large variability in gap junction size and that only 18% of the 340 channels are open in each plaque. Our results establish that the number of gap junctions per connection is the main determinant of both the strength and variability in electrical coupling between Golgi cells. PMID:27133465

  15. Role of astroglial connexin30 in hippocampal gap junction coupling.

    PubMed

    Gosejacob, Dominic; Dublin, Pavel; Bedner, Peter; Hüttmann, Kerstin; Zhang, Jiong; Tress, Oliver; Willecke, Klaus; Pfrieger, Frank; Steinhäuser, Christian; Theis, Martin

    2011-03-01

    The impact of connexin30 (Cx30) on interastrocytic gap junction coupling in the normal hippocampus is matter of debate; reporter gene analyses indicated a weak expression of Cx30 in the mouse hippocampus. In contrast, mice lacking connexin43 (Cx43) in astrocytes exhibited only 50% reduction in coupling. Complete uncoupling of hippocampal astrocytes in mice lacking both Cx30 and Cx43 suggested that Cx30 participates in interastrocytic gap junction coupling in the hippocampus. With comparative reporter gene assays, immunodetection, and cre/loxP-based reporter approaches we demonstrate that Cx30 is more abundant than previously thought. The specific role of Cx30 in interastrocytic coupling has never been investigated. Employing tracer coupling analyses in acute slices of Cx30 deficient mice here we show that Cx30 makes a substantial contribution to interastrocytic gap junctional communication in the mouse hippocampus. PMID:21264956

  16. Gap junctions in olfactory neurons modulate olfactory sensitivity

    PubMed Central

    2010-01-01

    Background One of the fundamental questions in olfaction is whether olfactory receptor neurons (ORNs) behave as independent entities within the olfactory epithelium. On the basis that mature ORNs express multiple connexins, I postulated that gap junctional communication modulates olfactory responses in the periphery and that disruption of gap junctions in ORNs reduces olfactory sensitivity. The data collected from characterizing connexin 43 (Cx43) dominant negative transgenic mice OlfDNCX, and from calcium imaging of wild type mice (WT) support my hypothesis. Results I generated OlfDNCX mice that express a dominant negative Cx43 protein, Cx43/β-gal, in mature ORNs to inactivate gap junctions and hemichannels composed of Cx43 or other structurally related connexins. Characterization of OlfDNCX revealed that Cx43/β-gal was exclusively expressed in areas where mature ORNs resided. Real time quantitative PCR indicated that cellular machineries of OlfDNCX were normal in comparison to WT. Electroolfactogram recordings showed decreased olfactory responses to octaldehyde, heptaldehyde and acetyl acetate in OlfDNCX compared to WT. Octaldehyde-elicited glomerular activity in the olfactory bulb, measured according to odor-elicited c-fos mRNA upregulation in juxtaglomerular cells, was confined to smaller areas of the glomerular layer in OlfDNCX compared to WT. In WT mice, octaldehyde sensitive neurons exhibited reduced response magnitudes after application of gap junction uncoupling reagents and the effects were specific to subsets of neurons. Conclusions My study has demonstrated that altered assembly of Cx43 or structurally related connexins in ORNs modulates olfactory responses and changes olfactory activation maps in the olfactory bulb. Furthermore, pharmacologically uncoupling of gap junctions reduces olfactory activity in subsets of ORNs. These data suggest that gap junctional communication or hemichannel activity plays a critical role in maintaining olfactory

  17. Roles of gap junctions, connexins, and pannexins in epilepsy

    PubMed Central

    Mylvaganam, Shanthini; Ramani, Meera; Krawczyk, Michal; Carlen, Peter L.

    2014-01-01

    Enhanced gap junctional communication (GJC) between neurons is considered a major factor underlying the neuronal synchrony driving seizure activity. In addition, the hippocampal sharp wave ripple complexes, associated with learning and seizures, are diminished by GJC blocking agents. Although gap junctional blocking drugs inhibit experimental seizures, they all have other non-specific actions. Besides interneuronal GJC between dendrites, inter-axonal and inter-glial GJC is also considered important for seizure generation. Interestingly, in most studies of cerebral tissue from animal seizure models and from human patients with epilepsy, there is up-regulation of glial, but not neuronal gap junctional mRNA and protein. Significant changes in the expression and post-translational modification of the astrocytic connexin Cx43, and Panx1 were observed in an in vitro Co++ seizure model, further supporting a role for glia in seizure-genesis, although the reasons for this remain unclear. Further suggesting an involvement of astrocytic GJC in epilepsy, is the fact that the expression of astrocytic Cx mRNAs (Cxs 30 and 43) is several fold higher than that of neuronal Cx mRNAs (Cxs 36 and 45), and the number of glial cells outnumber neuronal cells in mammalian hippocampal and cortical tissue. Pannexin expression is also increased in both animal and human epileptic tissues. Specific Cx43 mimetic peptides, Gap 27 and SLS, inhibit the docking of astrocytic connexin Cx43 proteins from forming intercellular gap junctions (GJs), diminishing spontaneous seizures. Besides GJs, Cx membrane hemichannels in glia and Panx membrane channels in neurons and glia are also inhibited by traditional gap junctional pharmacological blockers. Although there is no doubt that connexin-based GJs and hemichannels, and pannexin-based membrane channels are related to epilepsy, the specific details of how they are involved and how we can modulate their function for therapeutic purposes remain to be

  18. Gap junctions and inhibitory synapses modulate inspiratory motoneuron synchronization.

    PubMed

    Bou-Flores, C; Berger, A J

    2001-04-01

    Interneuronal electrical coupling via gap junctions and chemical synaptic inhibitory transmission are known to have roles in the generation and synchronization of activity in neuronal networks. Uncertainty exists regarding the roles of these two modes of interneuronal communication in the central respiratory rhythm-generating system. To assess their roles, we performed studies on both the neonatal mouse medullary slice and en bloc brain stem-spinal cord preparations where rhythmic inspiratory motor activity can readily be recorded from both hypoglossal and phrenic nerve roots. The rhythmic inspiratory activity observed had two temporal characteristics: the basic respiratory frequency occurring on a long time scale and the synchronous neuronal discharge within the inspiratory burst occurring on a short time scale. In both preparations, we observed that bath application of gap-junction blockers, including 18 alpha-glycyrrhetinic acid, 18 beta-glycyrrhetinic acid, and carbenoxolone, all caused a reduction in respiratory frequency. In contrast, peak integrated phrenic and hypoglossal inspiratory activity was not significantly changed by gap-junction blockade. On a short-time-scale, gap-junction blockade increased the degree of synchronization within an inspiratory burst observed in both nerves. In contrast, opposite results were observed with blockade of GABA(A) and glycine receptors. We found that respiratory frequency increased with receptor blockade, and simultaneous blockade of both receptors consistently resulted in a reduction in short-time-scale synchronized activity observed in phrenic and hypoglossal inspiratory bursts. These results support the concept that the central respiratory system has two components: a rhythm generator responsible for the production of respiratory cycle timing and an inspiratory pattern generator that is involved in short-time-scale synchronization. In the neonatal rodent, properties of both components can be regulated by interneuronal

  19. Gap junctions in the heart of the adult Protopterus aethiopicus.

    PubMed

    Scheuermann, D W; de Maziere, A

    1984-07-01

    In thin sections and in freeze-fracture replicas small and sparse gap junctions appear to be developed on the longitudinal plasma membrane of Protopterus cardiac cells near a macula or fascia adhaerens. By thin-section electron microscopy, they had septalaminar profiles with a length between 0.042 and 0.260 micron. In freeze-fracture images they appear on the P-fracture face as maculate particle aggregations with complementary pits on the E-fracture face. Particles with a central intercellular channel could be observed. The average center-to-center distance between neighbouring particles or pits is 10.05 +/- 1.87 nm (N = 2429). The diameter of the junctional maculae in replicas lies between 0.037 and 0.229 nm. The particle packing density increases in larger maculate aggregations, while particle-free areas emerge which could be related to the degradation or reformation of gap junctions Atypical configurations of gap junctions observed in the myocardium of lower vertebrates are rarely encountered in this primitive vertebrate. PMID:6485893

  20. Length and energy gap dependences of thermoelectricity in nanostructured junctions.

    PubMed

    Asai, Yoshihiro

    2013-04-17

    The possibilities of an enhanced thermoelectric figure of merit value, ZT, in a nanostructured junction are examined for a wide range of parameter values in a theoretical model. Our research shows that the figure of merit can take a very large maximum, which depends both on the length and the energy gap values. The maximum of ZT is achieved when the Fermi level of the electrodes is aligned to the edge of the electronic transmission function of the junction, where both the conductance and the Seebeck constant are significantly enhanced. On the basis of our results, we conclude that nanowires and molecular junctions form a special class of systems where a large ZT can be expected in some cases. PMID:23528878

  1. Gap Junctions Mediate Human Immunodeficiency Virus-Bystander Killing in Astrocytes

    PubMed Central

    Eugenin, Eliseo A.; Berman, Joan W.

    2007-01-01

    Human immunodeficiency virus (HIV) entry into the CNS is an early event after infection, resulting in neurological dysfunction in a significant number of individuals. As people with acquired immunodeficiency syndrome (AIDS) live longer, the prevalence of cognitive impairment is increasing, despite antiretroviral therapy. The mechanisms that mediate CNS dysfunction are still not completely understood, and include inflammation, viral presence, and/or replication. In this report, we characterize a novel role of gap junctions in transmitting and thereby amplifying toxic signals originating from HIV-infected astrocytes that trigger cell death in uninfected astrocytes. HIV-infected astrocytes were resistant to apoptosis; however, uninfected astrocytes forming gap junctions with infected astrocytes were apoptotic. Gap junction blockers abolished apoptosis in uninfected astrocytes, supporting the role of these channels in amplifying cell death. Our findings describe a novel mechanism of toxicity within the brain, triggered by low numbers of HIV-infected astrocytes and amplified by gap junctions, contributing to the pathogenesis of NeuroAIDS. PMID:18032656

  2. Human Articular Chondrocytes Express Multiple Gap Junction Proteins

    PubMed Central

    Mayan, Maria D.; Carpintero-Fernandez, Paula; Gago-Fuentes, Raquel; Martinez-de-Ilarduya, Oskar; Wang, Hong-Zhang; Valiunas, Virginijus; Brink, Peter; Blanco, Francisco J.

    2014-01-01

    Osteoarthritis (OA) is the most common joint disease and involves progressive degeneration of articular cartilage. The aim of this study was to investigate if chondrocytes from human articular cartilage express gap junction proteins called connexins (Cxs). We show that human chondrocytes in tissue express Cx43, Cx45, Cx32, and Cx46. We also find that primary chondrocytes from adults retain the capacity to form functional voltage-dependent gap junctions. Immunohistochemistry experiments in cartilage from OA patients revealed significantly elevated levels of Cx43 and Cx45 in the superficial zone and down through the next approximately 1000 μm of tissue. These zones corresponded with regions damaged in OA that also had high levels of proliferative cell nuclear antigen. An increased number of Cxs may help explain the increased proliferation of cells in clusters that finally lead to tissue homeostasis loss. Conversely, high levels of Cxs in OA cartilage reflect the increased number of adjacent cells in clusters that are able to interact directly by gap junctions as compared with hemichannels on single cells in normal cartilage. Our data provide strong evidence that OA patients have a loss of the usual ordered distribution of Cxs in the damaged zones and that the reductions in Cx43 levels are accompanied by the loss of correct Cx localization in the nondamaged areas. PMID:23416160

  3. Robert Feulgen Prize Lecture. Distribution and role of gap junctions in normal myocardium and human ischaemic heart disease.

    PubMed

    Green, C R; Severs, N J

    1993-02-01

    In the heart, individual cardiac muscle cells are linked by gap junctions. These junctions form low resistance pathways along which the electrical impulse flows rapidly and repeatedly between all the cells of the myocardium, ensuring their synchronous contraction. To obtain probes for mapping the distribution of gap junctions in cardiac tissue, polyclonal antisera were raised to three synthetic peptides, each matching different cytoplasmically exposed portions of the sequence of connexin43, the major gap-junctional protein reported in the heart. The specificity of each antiserum for the peptide to which it was raised was established by dot blotting. New methods were developed for isolating enriched fractions of gap junctions from whole heart and from dissociated adult myocytes, in which detergent-treatment and raising the temperature (potentially damaging steps in previously described techniques) are avoided. Analysis of these fractions by SDS-polyacrylamide gel electrophoresis revealed major bands at 43 kDa (matching the molecular mass of connexin43) and at 70 kDa. Western blot experiments using our antisera indicated that both the 43-kDa and the 70-kDa bands represent cardiac gap-junctional proteins. Pre-embedding immunogold labelling of isolated gap junctions and post-embedding immunogold labelling of Lowicryl-embedded whole tissue demonstrated the specific binding of the antibodies to ultrastructurally defined gap junctions. One antiserum (raised to residues 131-142) was found to be particularly effective for cytochemical labelling. Using this antiserum for immunofluorescence labelling in combination with confocal scanning laser microscopy enabled highly sensitive detection and three-dimensional mapping of gap junctions through thick slices of cardiac tissue. By means of the serial optical sectioning ability of the confocal microscope, images of the entire gap junction population of complete en face-viewed disks were reconstructed. These reconstructions reveal

  4. Polyamine signal through gap junctions: A key regulator of proliferation and gap-junction organization in mammalian tissues?

    PubMed

    Hamon, Loic; Savarin, Philippe; Pastré, David

    2016-06-01

    We propose that interaction rules derived from polyamine exchange in connected cells may explain the spatio-temporal organization of gap junctions observed during tissue regeneration and tumorigenesis. We also hypothesize that polyamine exchange can be considered as signal that allows cells to sense the proliferation status of their neighbors. Polyamines (putrescine, spermidine, and spermine) are indeed small aliphatic polycations that serve as fuels to sustain elevated proliferation rates of the order observed in cancer cells. Based on recent reports, we consider here that polyamines can be exchanged through gap junction channels between mammalian cells. Such intercellular exchange of polyamines has critical consequences on the local control of growth. In line with this hypothesis, the complex protein network that keeps polyamine levels finely tuned in mammalian cells can translate polyamine efflux or influx into integrated signals controlling transcription, translation, and cell communications. PMID:27125471

  5. The connexin43 mimetic peptide Gap19 inhibits hemichannels without altering gap junctional communication in astrocytes

    PubMed Central

    Abudara, Verónica; Bechberger, John; Freitas-Andrade, Moises; De Bock, Marijke; Wang, Nan; Bultynck, Geert; Naus, Christian C.; Leybaert, Luc; Giaume, Christian

    2014-01-01

    In the brain, astrocytes represent the cellular population that expresses the highest amount of connexins (Cxs). This family of membrane proteins is the molecular constituent of gap junction channels and hemichannels that provide pathways for direct cytoplasm-to-cytoplasm and inside-out exchange, respectively. Both types of Cx channels are permeable to ions and small signaling molecules allowing astrocytes to establish dynamic interactions with neurons. So far, most pharmacological approaches currently available do not distinguish between these two channel functions, stressing the need to develop new specific molecular tools. In astrocytes two major Cxs are expressed, Cx43 and Cx30, and there is now evidence indicating that at least Cx43 operates as a gap junction channel as well as a hemichannel in these cells. Based on studies in primary cultures as well as in acute hippocampal slices, we report here that Gap19, a nonapeptide derived from the cytoplasmic loop of Cx43, inhibits astroglial Cx43 hemichannels in a dose-dependent manner, without affecting gap junction channels. This peptide, which not only selectively inhibits hemichannels but is also specific for Cx43, can be delivered in vivo in mice as TAT-Gap19, and displays penetration into the brain parenchyma. As a result, Gap19 combined with other tools opens up new avenues to decipher the role of Cx43 hemichannels in interactions between astrocytes and neurons in physiological as well as pathological situations. PMID:25374505

  6. Gap junctions modulate glioma invasion by direct transfer of microRNA.

    PubMed

    Hong, Xiaoting; Sin, Wun Chey; Harris, Andrew L; Naus, Christian C

    2015-06-20

    The invasiveness of high-grade glioma is the primary reason for poor survival following treatment. Interaction between glioma cells and surrounding astrocytes are crucial to invasion. We investigated the role of gap junction mediated miRNA transfer in this context. By manipulating gap junctions with a gap junction inhibitor, siRNAs, and a dominant negative connexin mutant, we showed that functional glioma-glioma gap junctions suppress glioma invasion while glioma-astrocyte and astrocyte-astrocyte gap junctions promote it in an in vitro transwell invasion assay. After demonstrating that glioma-astrocyte gap junctions are permeable to microRNA, we compared the microRNA profiles of astrocytes before and after co-culture with glioma cells, identifying specific microRNAs as candidates for transfer through gap junctions from glioma cells to astrocytes. Further analysis showed that transfer of miR-5096 from glioma cells to astrocytes is through gap junctions; this transfer is responsible, in part, for the pro-invasive effect. Our results establish a role for glioma-astrocyte gap junction mediated microRNA signaling in modulation of glioma invasive behavior, and that gap junction coupling among astrocytes magnifies the pro-invasive signaling. Our findings reveal the potential for therapeutic interventions based on abolishing alteration of stromal cells by tumor cells via manipulation of microRNA and gap junction channel activity. PMID:25978028

  7. Models and methods for in vitro testing of hepatic gap junctional communication

    PubMed Central

    Willebrords, Joost; Vinken, Mathieu

    2015-01-01

    Inherent to their pivotal roles in controlling all aspects of the liver cell life cycle, hepatocellular gap junctions are frequently disrupted upon impairment of the homeostatic balance, as occurs during liver toxicity. Hepatic gap junctions, which are mainly built up by connexin32, are specifically targeted by tumor promoters and epigenetic carcinogens. This renders inhibition of gap junction functionality a suitable indicator for the in vitro detection of nongenotoxic hepatocarcinogenicity. The establishment of a reliable liver gap junction inhibition assay for routine in vitro testing purposes requires a cellular system in which gap junctions are expressed at an in vivo-like level as well as an appropriate technique to probe gap junction activity. Both these models and methods are discussed in the current paper, thereby focusing on connexin32-based gap junctions. PMID:26420514

  8. Models and methods for in vitro testing of hepatic gap junctional communication.

    PubMed

    Maes, Michaël; Crespo Yanguas, Sara; Willebrords, Joost; Vinken, Mathieu

    2015-12-25

    Inherent to their pivotal roles in controlling all aspects of the liver cell life cycle, hepatocellular gap junctions are frequently disrupted upon impairment of the homeostatic balance, as occurs during liver toxicity. Hepatic gap junctions, which are mainly built up by connexin32, are specifically targeted by tumor promoters and epigenetic carcinogens. This renders inhibition of gap junction functionality a suitable indicator for the in vitro detection of nongenotoxic hepatocarcinogenicity. The establishment of a reliable liver gap junction inhibition assay for routine in vitro testing purposes requires a cellular system in which gap junctions are expressed at an in vivo-like level as well as an appropriate technique to probe gap junction activity. Both these models and methods are discussed in the current paper, thereby focusing on connexin32-based gap junctions. PMID:26420514

  9. CHLORAL HYDRATE DECREASES GAP JUNCTION COMMUNICATION IN RAT LIVER EPITHELIAL CELLS

    EPA Science Inventory

    Chloral hydrate decreases gap junction communication in rat liver epithelial cells

    Gap junction communication (GJC) is involved in controlling cell proliferation and differentiation. Connexins (Cx) that make up these junctions are composed of a closely related group of m...

  10. Molecular mechanisms regulating formation, trafficking and processing of annular gap junctions.

    PubMed

    Falk, Matthias M; Bell, Cheryl L; Kells Andrews, Rachael M; Murray, Sandra A

    2016-01-01

    Internalization of gap junction plaques results in the formation of annular gap junction vesicles. The factors that regulate the coordinated internalization of the gap junction plaques to form annular gap junction vesicles, and the subsequent events involved in annular gap junction processing have only relatively recently been investigated in detail. However it is becoming clear that while annular gap junction vesicles have been demonstrated to be degraded by autophagosomal and endo-lysosomal pathways, they undergo a number of additional processing events. Here, we characterize the morphology of the annular gap junction vesicle and review the current knowledge of the processes involved in their formation, fission, fusion, and degradation. In addition, we address the possibility for connexin protein recycling back to the plasma membrane to contribute to gap junction formation and intercellular communication. Information on gap junction plaque removal from the plasma membrane and the subsequent processing of annular gap junction vesicles is critical to our understanding of cell-cell communication as it relates to events regulating development, cell homeostasis, unstable proliferation of cancer cells, wound healing, changes in the ischemic heart, and many other physiological and pathological cellular phenomena. PMID:27230503

  11. Switch in Gap Junction Protein Expression is Associated with Selective Changes in Junctional Permeability During Keratinocyte Differentiation

    NASA Astrophysics Data System (ADS)

    Brissette, Janice L.; Kumar, Nalin M.; Gilula, Norton B.; Hall, James E.; Dotto, G. Paolo

    1994-07-01

    Gap junctional communication provides a mechanism for regulating multicellular activities by allowing the exchange of small diffusible molecules between neighboring cells. The diversity of gap junction proteins may exist to form channels that have different permeability properties. We report here that induction of terminal differentiation in mouse primary keratinocytes by calcium results in a specific switch in gap junction protein expression. Expression of α_1 (connexin 43) and β_2 (connexin 26) gap junction proteins is down-modulated, whereas that of β_3 (connexin 31) and β_4 (connexin 31.1) proteins is induced. Although both proliferating and differentiating keratinocytes are electrically coupled, there are significant changes in the permeability properties of the junctions to small molecules. In parallel with the changes in gap junction protein expression during differentiation, the intercellular transfer of the small dyes neurobiotin, carboxyfluorescein, and Lucifer yellow is significantly reduced, whereas that of small metabolites, such as nucleotides and amino acids, proceeds unimpeded. Thus, a switch in gap junction protein expression in differentiating keratinocytes is accompanied by selective changes in junctional permeability that may play an important role in the coordinate control of the differentiation process.

  12. Reduction in gap junction intercellular communication promotes glioma migration.

    PubMed

    Aftab, Qurratulain; Sin, Wun-Chey; Naus, Christian C

    2015-05-10

    Glioblastoma Multiforme (GBM), an aggressive form of adult brain tumor, is difficult to treat due to its invasive nature. One of the molecular changes observed in GBM is a decrease in the expression of the gap junction protein Connexin43 (Cx43); however, how a reduction in Cx43 expression contributes to glioma malignancy is still unclear. In this study we examine whether a decrease in Cx43 protein expression has a role in enhanced cell migration, a key feature associated with increased tumorigenicity. We used a 3D spheroid migration model that mimics the in vivo architecture of tumor cells to quantify migration changes. We found that down-regulation of Cx43 expression in the U118 human glioma cell line increased migration by reducing cell-ECM adhesion, and changed the migration pattern from collective to single cell. In addition gap junction intercellular communication (GJIC) played a more prominent role in mediating migration than the cytoplasmic interactions of the C-terminal tail. Live imaging revealed that reducing Cx43 expression enhanced relative migration by increasing the cell speed and affecting the direction of migration. Taken together our findings reveal an unexplored role of GJIC in facilitating collective migration. PMID:25926558

  13. Gap junction proteins and their role in spinal cord injury

    PubMed Central

    Tonkin, Ryan S.; Mao, Yilin; O’Carroll, Simon J.; Nicholson, Louise F. B.; Green, Colin R.; Gorrie, Catherine A.; Moalem-Taylor, Gila

    2015-01-01

    Gap junctions are specialized intercellular communication channels that are formed by two hexameric connexin hemichannels, one provided by each of the two adjacent cells. Gap junctions and hemichannels play an important role in regulating cellular metabolism, signaling, and functions in both normal and pathological conditions. Following spinal cord injury (SCI), there is damage and disturbance to the neuronal elements of the spinal cord including severing of axon tracts and rapid cell death. The initial mechanical disruption is followed by multiple secondary cascades that cause further tissue loss and dysfunction. Recent studies have implicated connexin proteins as playing a critical role in the secondary phase of SCI by propagating death signals through extensive glial networks. In this review, we bring together past and current studies to outline the distribution, changes and roles of various connexins found in neurons and glial cells, before and in response to SCI. We discuss the contribution of pathologically activated connexin proteins, in particular connexin 43, to functional recovery and neuropathic pain, as well as providing an update on potential connexin specific pharmacological agents to treat SCI. PMID:25610368

  14. Gap-Junction Channels Dysfunction in Deafness and Hearing Loss

    PubMed Central

    Acuña, Rodrigo; Figueroa, Vania; Maripillan, Jaime; Nicholson, Bruce

    2009-01-01

    Abstract Gap-junction channels connect the cytoplasm of adjacent cells, allowing the diffusion of ions and small metabolites. They are formed at the appositional plasma membranes by a family of related proteins named connexins. Mutations in connexins 26, 31, 30, 32, and 43 have been associated with nonsyndromic or syndromic deafness. The majority of these mutations are inherited in an autosomal recessive manner, but a few of them have been associated with dominantly inherited hearing loss. Mutations in the connexin26 gene (GJB2) are the most common cause of genetic deafness. This review summarizes the most relevant and recent information about different mutations in connexin genes found in human patients, with emphasis on GJB2. The possible effects of the mutations on channel expression and function are discussed, in addition to their possible physiologic consequences for inner ear physiology. Finally, we propose that connexin channels (gap junctions and hemichannels) may be targets for age-related hearing loss induced by oxidative damage. Antioxid. Redox Signal. 11, 309–322. PMID:18837651

  15. Gap junction networks in mushroom bodies participate in visual learning and memory in Drosophila.

    PubMed

    Liu, Qingqing; Yang, Xing; Tian, Jingsong; Gao, Zhongbao; Wang, Meng; Li, Yan; Guo, Aike

    2016-01-01

    Gap junctions are widely distributed in the brains across species and play essential roles in neural information processing. However, the role of gap junctions in insect cognition remains poorly understood. Using a flight simulator paradigm and genetic tools, we found that gap junctions are present in Drosophila Kenyon cells (KCs), the major neurons of the mushroom bodies (MBs), and showed that they play an important role in visual learning and memory. Using a dye coupling approach, we determined the distribution of gap junctions in KCs. Furthermore, we identified a single pair of MB output neurons (MBONs) that possess a gap junction connection to KCs, and provide strong evidence that this connection is also required for visual learning and memory. Together, our results reveal gap junction networks in KCs and the KC-MBON circuit, and bring new insight into the synaptic network underlying fly's visual learning and memory. PMID:27218450

  16. Carcinoma-astrocyte gap junctions promote brain metastasis by cGAMP transfer.

    PubMed

    Chen, Qing; Boire, Adrienne; Jin, Xin; Valiente, Manuel; Er, Ekrem Emrah; Lopez-Soto, Alejandro; Jacob, Leni S; Patwa, Ruzeen; Shah, Hardik; Xu, Ke; Cross, Justin R; Massagué, Joan

    2016-05-26

    Brain metastasis represents a substantial source of morbidity and mortality in various cancers, and is characterized by high resistance to chemotherapy. Here we define the role of the most abundant cell type in the brain, the astrocyte, in promoting brain metastasis. We show that human and mouse breast and lung cancer cells express protocadherin 7 (PCDH7), which promotes the assembly of carcinoma-astrocyte gap junctions composed of connexin 43 (Cx43). Once engaged with the astrocyte gap-junctional network, brain metastatic cancer cells use these channels to transfer the second messenger cGAMP to astrocytes, activating the STING pathway and production of inflammatory cytokines such as interferon-α (IFNα) and tumour necrosis factor (TNF). As paracrine signals, these factors activate the STAT1 and NF-κB pathways in brain metastatic cells, thereby supporting tumour growth and chemoresistance. The orally bioavailable modulators of gap junctions meclofenamate and tonabersat break this paracrine loop, and we provide proof-of-principle that these drugs could be used to treat established brain metastasis. PMID:27225120

  17. Effects of microgravity on liposome-reconstituted cardiac gap junction channeling activity

    NASA Technical Reports Server (NTRS)

    Claassen, D. E.; Spooner, B. S.

    1989-01-01

    Effects of microgravity on cardiac gap junction channeling activity were investigated aboard NASA zero-gravity aircraft. Liposome-reconstituted gap junctions were assayed for channel function during free-fall, and the data were compared with channeling at 1 g. Control experiments tested for 0 g effects on the structural stability of liposomes, and on the enzyme-substrate signalling system of the assay. The results demonstrate that short periods of microgravity do not perturb reconstituted cardiac gap junction channeling activity.

  18. Temperature Dependence of Gap Related Structures in YBa2Cu3O7-δ Break Junctions

    NASA Astrophysics Data System (ADS)

    Giubileo, F.; Bobba, F.; Cucolo, A. M.; Akimenko, A. I.

    We have applied the break junction technique to highly epitaxial c-axis oriented YBaCuO thin films with Tc(ρ=0)=91 K deposited on (001) SrTiO3 or LaAlO3 substrates by a high oxygen pressure d.c. sputtering technique. The film thickness was about 1500 Å and a photolithographic process was used to reduce to 100 μm the junction width across the fracture. By this procedure, tunable resistance break junctions with tunneling current favored along the ab-planes have been realized. The junctions were formed at low temperatures with freshly fractured surfaces and an inert tunnel barrier was created by helium gas or liquid. A good stability was obtained with the normal-state resistance RN changing about 15% in the temperature range between 4.2 K and 100 K. We have measured the temperature dependence of the conductance maxima that are related to superconducting energy gap at the Fermi level.

  19. A rapid and sensitive assay of intercellular coupling by voltage imaging of gap junction networks

    PubMed Central

    2013-01-01

    Background A variety of mechanisms that govern connexin channel gating and permeability regulate coupling in gap junction networks. Mutations in connexin genes have been linked to several pathologies, including cardiovascular anomalies, peripheral neuropathy, skin disorders, cataracts and deafness. Gap junction coupling and its patho–physiological alterations are commonly assayed by microinjection experiments with fluorescent tracers, which typically require several minutes to allow dye transfer to a limited number of cells. Comparable or longer time intervals are required by fluorescence recovery after photobleaching experiments. Paired electrophysiological recordings have excellent time resolution but provide extremely limited spatial information regarding network connectivity. Results Here, we developed a rapid and sensitive method to assay gap junction communication using a combination of single cell electrophysiology, large–scale optical recordings and a digital phase–sensitive detector to extract signals with a known frequency from Vf2.1.Cl, a novel fluorescent sensor of plasma membrane potential. Tests performed in HeLa cell cultures confirmed that suitably encoded Vf2.1.Cl signals remained confined within the network of cells visibly interconnected by fluorescently tagged gap junction channels. We used this method to visualize instantly intercellular connectivity over the whole field of view (hundreds of cells) in cochlear organotypic cultures from postnatal mice. A simple resistive network model reproduced accurately the spatial dependence of the electrical signals throughout the cellular network. Our data suggest that each pair of cochlear non−sensory cells of the lesser epithelial ridge is coupled by ~1500 gap junction channels, on average. Junctional conductance was reduced by 14% in cochlear cultures harboring the T5M mutation of connexin30, which induces a moderate hearing loss in connexin30T5M/T5M knock–in mice, and by 91% in cultures from

  20. Establishment of the Dual Whole Cell Recording Patch Clamp Configuration for the Measurement of Gap Junction Conductance.

    PubMed

    Veenstra, Richard D

    2016-01-01

    The development of the patch clamp technique has enabled investigators to directly measure gap junction conductance between isolated pairs of small cells with resolution to the single channel level. The dual patch clamp recording technique requires specialized equipment and the acquired skill to reliably establish gigaohm seals and the whole cell recording configuration with high efficiency. This chapter describes the equipment needed and methods required to achieve accurate measurement of macroscopic and single gap junction channel conductances. Inherent limitations with the dual whole cell recording technique and methods to correct for series access resistance errors are defined as well as basic procedures to determine the essential electrical parameters necessary to evaluate the accuracy of gap junction conductance measurements using this approach. PMID:27207298

  1. Chlorpromazine reduces the intercellular communication via gap junctions in mammalian cells

    SciTech Connect

    Orellana, Juan A.; Palacios-Prado, Nicolas; Saez, Juan C. . E-mail: jsaez@bio.puc.cl

    2006-06-15

    In the work presented herein, we evaluated the effect of chlorpromazine (CPZ) on gap junctions expressed by two mammalian cell types; Gn-11 cells (cell line derived from mouse LHRH neurons) and rat cortical astrocytes maintained in culture. We also attempted to elucidate possible mechanisms of action of CPZ effects on gap junctions. CPZ, in concentrations comparable with doses used to treat human diseases, was found to reduce the intercellular communication via gap junctions as evaluated with measurements of dye coupling (Lucifer yellow). In both cell types, maximal inhibition of functional gap junctions was reached within about 1 h of treatment with CPZ, an recovery was almost complete at about 5 h after CPZ wash out. In both cell types, CPZ treatment increased the phosphorylation state of connexin43 (Cx43), a gap junction protein subunit. Moreover, CPZ reduced the reactivity of Cx43 (immunofluorescence) at cell interfaces and concomitantly increased its reactivity in intracellular vesicles, suggesting an increased retrieval from and/or reduced insertion into the plasma membrane. CPZ also caused cellular retraction reducing cell-cell contacts in a reversible manner. The reduction in contact area might destabilize existing gap junctions and abrogate formation of new ones. Moreover, the CPZ-induced reduction in gap junctional communication may depend on the connexins (Cxs) forming the junctions. If Cx43 were the only connexin expressed, MAPK-dependent phosphorylation of this connexin would induce closure of gap junction channels.

  2. Gap junction-mediated electrical transmission: regulatory mechanisms and plasticity

    PubMed Central

    Pereda, Alberto E.; Curti, Sebastian; Hoge, Gregory; Cachope, Roger; Flores, Carmen E.; Rash, John E.

    2012-01-01

    The term synapse applies to cellular specializations that articulate the processing of information within neural circuits by providing a mechanism for the transfer of information between two different neurons. There are two main modalities of synaptic transmission: chemical and electrical. While most efforts have been dedicated to the understanding of the properties and modifiability of chemical transmission, less is still known regarding the plastic properties of electrical synapses, whose structural correlate is the gap junction. A wealth of data indicates that, rather than passive intercellular channels, electrical synapses are more dynamic and modifiable than was generally perceived. This article will discuss the factors determining the strength of electrical transmission and review current evidence demonstrating its dynamic properties. Like their chemical counterparts, electrical synapses can also be plastic and modifiable. PMID:22659675

  3. Microinjection Technique for Assessment of Gap Junction Function.

    PubMed

    Fridman, Michael D; Liu, Jun; Sun, Yu; Hamilton, Robert M

    2016-01-01

    Gap junctions are essential for the proper function of many native mammalian tissues including neurons, cardiomyocytes, embryonic tissues, and muscle. Assessing these channels is therefore fundamental to understanding disease pathophysiology, developing therapies for a multitude of acquired and genetic conditions, and providing novel approaches to drug delivery and cellular communication. Microinjection is a robust, albeit difficult, technique, which provides considerable information that is superior to many of the simpler techniques due to its ability to isolate cells, quantify kinetics, and allow cross-comparison of multiple cell lines. Despite its user-dependent nature, the strengths of the technique are considerable and with the advent of new, automation technologies may improve further. This text describes the basic technique of microinjection and briefly discusses modern automation advances that can improve the success rates of this technique. PMID:27207292

  4. Synchronization transition in gap-junction-coupled leech neurons

    NASA Astrophysics Data System (ADS)

    Wang, Qingyun; Duan, Zhisheng; Feng, Zhaosheng; Chen, Guanrong; Lu, Qishao

    2008-07-01

    Real neurons can exhibit various types of firings including tonic spiking, bursting as well as silent state, which are frequently observed in neuronal electrophysiological experiments. More interestingly, it is found that neurons can demonstrate the co-existing mode of stable tonic spiking and bursting, which depends on initial conditions. In this paper, synchronization in gap-junction-coupled neurons with co-existing attractors of spiking and bursting firings is investigated as the coupling strength gets increased. Synchronization transitions can be identified by means of the bifurcation diagram and the correlation coefficient. It is illustrated that the coupled neurons can exhibit different types of synchronization transitions between spiking and bursting when the coupling strength increases. In the course of synchronization transitions, an intermittent synchronization can be observed. These results may be instructive to understand synchronization transitions in neuronal systems.

  5. Methamphetamine compromises gap junctional communication in astrocytes and neurons.

    PubMed

    Castellano, Paul; Nwagbo, Chisom; Martinez, Luis R; Eugenin, Eliseo A

    2016-05-01

    Methamphetamine (meth) is a central nervous system (CNS) stimulant that results in psychological and physical dependency. The long-term effects of meth within the CNS include neuronal plasticity changes, blood-brain barrier compromise, inflammation, electrical dysfunction, neuronal/glial toxicity, and an increased risk to infectious diseases including HIV. Most of the reported meth effects in the CNS are related to dysregulation of chemical synapses by altering the release and uptake of neurotransmitters, especially dopamine, norepinephrine, and epinephrine. However, little is known about the effects of meth on connexin (Cx) containing channels, such as gap junctions (GJ) and hemichannels (HC). We examined the effects of meth on Cx expression, function, and its role in NeuroAIDS. We found that meth altered Cx expression and localization, decreased GJ communication between neurons and astrocytes, and induced the opening of Cx43/Cx36 HC. Furthermore, we found that these changes in GJ and HC induced by meth treatment were mediated by activation of dopamine receptors, suggesting that dysregulation of dopamine signaling induced by meth is essential for GJ and HC compromise. Meth-induced changes in GJ and HC contributed to amplified CNS toxicity by dysregulating glutamate metabolism and increasing the susceptibility of neurons and astrocytes to bystander apoptosis induced by HIV. Together, our results indicate that connexin containing channels, GJ and HC, are essential in the pathogenesis of meth and increase the sensitivity of the CNS to HIV CNS disease. Methamphetamine (meth) is an extremely addictive central nervous system stimulant. Meth reduced gap junctional (GJ) communication by inducing internalization of connexin-43 (Cx43) in astrocytes and reducing expression of Cx36 in neurons by a mechanism involving activation of dopamine receptors (see cartoon). Meth-induced changes in Cx containing channels increased extracellular levels of glutamate and resulted in higher

  6. Gap junction dysfunction in the prefrontal cortex induces depressive-like behaviors in rats.

    PubMed

    Sun, Jian-Dong; Liu, Yan; Yuan, Yu-He; Li, Jing; Chen, Nai-Hong

    2012-04-01

    Growing evidence has implicated glial anomalies in the pathophysiology of major depression disorder (MDD). Gap junctional communication is a main determinant of astrocytic function. However, it is unclear whether gap junction dysfunction is involved in MDD development. This study investigates changes in the function of astrocyte gap junction occurring in the rat prefrontal cortex (PFC) after chronic unpredictable stress (CUS), a rodent model of depression. Animals exposed to CUS and showing behavioral deficits in sucrose preference test (SPT) and novelty suppressed feeding test (NSFT) exhibited significant decreases in diffusion of gap junction channel-permeable dye and expression of connexin 43 (Cx43), a major component of astrocyte gap junction, and abnormal gap junctional ultrastructure in the PFC. Furthermore, we analyzed the effects of typical antidepressants fluoxetine and duloxetine and glucocorticoid receptor (GR) antagonist mifepristone on CUS-induced gap junctional dysfunction and depressive-like behaviors. The cellular and behavioral alterations induced by CUS were reversed and/or blocked by treatment with typical antidepressants or mifepristone, indicating that the mechanism of their antidepressant action may involve the amelioration of gap junction dysfunction and the cellular changes may be related to GR activation. We then investigated the effects of pharmacological gap junction blockade in the PFC on depressive-like behaviors. The results demonstrate that carbenoxolone (CBX) infusions induced anhedonia in SPT, and anxiety in NSFT, and Cx43 mimetic peptides Gap27 and Gap26 also induced anhedonia, a core symptom of depression. Together, this study supports the hypothesis that gap junction dysfunction contributes to the pathophysiology of depression. PMID:22189291

  7. Benzalkonium Chloride Suppresses Rabbit Corneal Endothelium Intercellular Gap Junction Communication

    PubMed Central

    Zhang, Zhenhao; Huang, Yue; Xie, Hui; Pan, Juxin; Liu, Fanfei; Li, Xuezhi; Chen, Wensheng; Hu, Jiaoyue; Liu, Zuguo

    2014-01-01

    Purpose Gap junction intercellular communication (GJIC) plays a critical role in the maintenance of corneal endothelium homeostasis. We determined if benzalkonium chloride (BAK) alters GJIC activity in the rabbit corneal endothelium since it is commonly used as a drug preservative in ocular eyedrop preparations even though it can have cytotoxic effects. Methods Thirty-six adult New Zealand albino rabbits were randomly divided into three groups. BAK at 0.01%, 0.05%, and 0.1% was applied twice daily to one eye of each of the rabbits in one of the three groups for seven days. The contralateral untreated eyes were used as controls. Corneal endothelial morphological features were observed by in vivo confocal microscopy (IVCM). Immunofluorescent staining resolved changes in gap junction integrity and localization. Western blot analysis and RT-PCR evaluated changes in levels of connexin43 (Cx43) and tight junction zonula occludens-1 (ZO-1) gene and protein expression, respectively. Cx43 and ZO-1 physical interaction was detected by immunoprecipitation (IP). Primary rabbit corneal endothelial cells were cultured in Dulbecco's Modified Eagle Medium (DMEM) containing BAK for 24 hours. The scrape-loading dye transfer technique (SLDT) was used to assess GJIC activity. Results Topical administration of BAK (0.05%, 0.1%) dose dependently disrupted corneal endothelial cell morphology, altered Cx43 and ZO-1 distribution and reduced Cx43 expression. BAK also markedly induced increases in Cx43 phosphorylation status concomitant with decreases in the Cx43-ZO-1 protein-protein interaction. These changes were associated with marked declines in GJIC activity. Conclusions The dose dependent declines in rabbit corneal endothelial GJIC activity induced by BAK are associated with less Cx43-ZO-1 interaction possibly arising from increases in Cx43 phosphorylation and declines in its protein expression. These novel changes provide additional evidence that BAK containing eyedrop preparations

  8. Regulation of Gap Junction Dynamics by UNC-44/ankyrin and UNC-33/CRMP through VAB-8 in C. elegans Neurons

    PubMed Central

    Yan, Dong

    2016-01-01

    Gap junctions are present in both vertebrates and invertebrates from nematodes to mammals. Although the importance of gap junctions has been documented in many biological processes, the molecular mechanisms underlying gap junction dynamics remain unclear. Here, using the C. elegans PLM neurons as a model, we show that UNC-44/ankyrin acts upstream of UNC-33/CRMP in regulation of a potential kinesin VAB-8 to control gap junction dynamics, and loss-of-function in the UNC-44/UNC-33/VAB-8 pathway suppresses the turnover of gap junction channels. Therefore, we first show a signal pathway including ankyrin, CRMP, and kinesin in regulating gap junctions. PMID:27015090

  9. Regulation of Gap Junction Dynamics by UNC-44/ankyrin and UNC-33/CRMP through VAB-8 in C. elegans Neurons.

    PubMed

    Meng, Lingfeng; Chen, Chia-Hui; Yan, Dong

    2016-03-01

    Gap junctions are present in both vertebrates and invertebrates from nematodes to mammals. Although the importance of gap junctions has been documented in many biological processes, the molecular mechanisms underlying gap junction dynamics remain unclear. Here, using the C. elegans PLM neurons as a model, we show that UNC-44/ankyrin acts upstream of UNC-33/CRMP in regulation of a potential kinesin VAB-8 to control gap junction dynamics, and loss-of-function in the UNC-44/UNC-33/VAB-8 pathway suppresses the turnover of gap junction channels. Therefore, we first show a signal pathway including ankyrin, CRMP, and kinesin in regulating gap junctions. PMID:27015090

  10. GAP JUNCTION COMMUNICATON IN A TRANSFECTED HUMAN CELL LINE: ACTION OF MELATONIN AND MAGNETIC FIELDS

    EPA Science Inventory

    GAP JUNCTION COMMUNICTION IN TRANSFECTED HUMAN CELL LINE: ACTION OF MELATONIN AND MAGNETIC FIELDS.

    OBJECTIVE: We previously showed that functional gap junction communication (GJC), as monitored by dye transfer (DT), could be enhanced in mouse C3H 10T112 cells and in mouse...

  11. ORIENTATION REQUIREMENT TO DETECT MAGNETIC FIELD-INDUCTED ALTERATION OF GAP JUNCTION COMMUNICATION IN EPITHELIAL CELLS

    EPA Science Inventory

    ORIENTATION REQUIREMENT TO DETECT MAGNETIC FIELD-INDUCED ALTERATION OF GAP JUNCTION COMMUNICATION IN EPITHELIAL CELLS.
    OBJECTIVE: We have shown that functional gap junction communication as measured by Lucifer yellow dye transfer (DT) in Clone-9 rat liver epithelial cells, c...

  12. Robustness effect of gap junctions between Golgi cells on cerebellar cortex oscillations

    PubMed Central

    2011-01-01

    Background Previous one-dimensional network modeling of the cerebellar granular layer has been successfully linked with a range of cerebellar cortex oscillations observed in vivo. However, the recent discovery of gap junctions between Golgi cells (GoCs), which may cause oscillations by themselves, has raised the question of how gap-junction coupling affects GoC and granular-layer oscillations. To investigate this question, we developed a novel two-dimensional computational model of the GoC-granule cell (GC) circuit with and without gap junctions between GoCs. Results Isolated GoCs coupled by gap junctions had a strong tendency to generate spontaneous oscillations without affecting their mean firing frequencies in response to distributed mossy fiber input. Conversely, when GoCs were synaptically connected in the granular layer, gap junctions increased the power of the oscillations, but the oscillations were primarily driven by the synaptic feedback loop between GoCs and GCs, and the gap junctions did not change oscillation frequency or the mean firing rate of either GoCs or GCs. Conclusion Our modeling results suggest that gap junctions between GoCs increase the robustness of cerebellar cortex oscillations that are primarily driven by the feedback loop between GoCs and GCs. The robustness effect of gap junctions on synaptically driven oscillations observed in our model may be a general mechanism, also present in other regions of the brain. PMID:22330240

  13. Functionally Active Gap Junctions between Connexin 43-Positive Mesenchymal Stem Cells and Glioma Cells.

    PubMed

    Gabashvili, A N; Baklaushev, V P; Grinenko, N F; Levinskii, A B; Mel'nikov, P A; Cherepanov, S A; Chekhonin, V P

    2015-05-01

    The formation of functional gap junctions between mesenchymal stem cells and cells of low-grade rat glioma C6 cells was studied in in vitro experiments. Immunocytochemical analysis with antibodies to connexin 43 extracellular loop 2 showed that mesenchymal stem cells as well as C6 glioma cells express the main astroglial gap junction protein connexin 43. Analysis of migration activity showed that mesenchymal stem cells actively migrate towards C6 glioma cells. During co-culturing, mesenchymal stem cells and glioma C6 form functionally active gap junctions mediating the transport of cytoplasmic dye from glioma cells to mesenchymal stem cells in the opposite direction. Fluorometry showed that the intensity of transport of low-molecular substances through heterologous gap junctions between mesenchymal stem cells and glioma cells is similar to that through homologous gap junctions between glioma cells. This phenomenon can be used for the development of new methods of cell therapy of high-grade gliomas. PMID:26033611

  14. Emergent Central Pattern Generator Behavior in Gap-Junction-Coupled Hodgkin-Huxley Style Neuron Model

    PubMed Central

    Memelli, Heraldo; Solomon, Irene C.

    2012-01-01

    Most models of central pattern generators (CPGs) involve two distinct nuclei mutually inhibiting one another via synapses. Here, we present a single-nucleus model of biologically realistic Hodgkin-Huxley neurons with random gap junction coupling. Despite no explicit division of neurons into two groups, we observe a spontaneous division of neurons into two distinct firing groups. In addition, we also demonstrate this phenomenon in a simplified version of the model, highlighting the importance of afterhyperpolarization currents (IAHP) to CPGs utilizing gap junction coupling. The properties of these CPGs also appear sensitive to gap junction conductance, probability of gap junction coupling between cells, topology of gap junction coupling, and, to a lesser extent, input current into our simulated nucleus. PMID:23365558

  15. Propofol depresses cisplatin cytotoxicity via the inhibition of gap junctions.

    PubMed

    Zhang, Yuan; Wang, Xiyan; Wang, Qin; Ge, Hui; Tao, Liang

    2016-06-01

    The general anesthetic, propofol, affects chemotherapeutic activity, however, the mechanism underlying its effects remains to be fully elucidated. Our previous study showed that tramadol and flurbiprofen depressed the cytotoxicity of cisplatin via the inhibition of gap junction (GJ) intercellular communication (GJIC) in connexin (Cx)32 HeLa cells. The present study investigated whether the effects of propofol on the cytotoxicity of cisplatin were mediated by GJ in U87 glioma cells and Cx26‑transfected HeLa cells. Standard colony formation assay was used to determine the cytotoxicity of cisplatin. Parachute dye coupling assay was used to measure GJ function, and western blot analysis was used to determine the expression levels of Cx32. The results revealed that exposure of the U87 glioma cells and the Cx26-transfected HeLa cells to cisplatin for 1 h reduced clonogenic survival in low density cultures (without GJs) and high density cultures (with GJs). However, the toxic effect was higher in the high density culture. In addition, pretreatment of the cells with propofol significantly reduced cisplatin‑induced cytotoxicity, but only in the presence of functional GJs. Furthermore, propofol significantly inhibited dye coupling through junctional channels, and a long duration of exposure of the cells to propofol downregulated the expression levels of Cx43 and Cx26. These results demonstrated that the inhibition of GJIC by propofol affected the therapeutic efficacy of chemotherapeutic drugs. The present study provides evidence of a novel mechanism underlying the effects of analgesics in counteracting chemotherapeutic efficiency. PMID:27082707

  16. Abundance of gap junctions at glutamatergic mixed synapses in adult Mosquitofish spinal cord neurons

    PubMed Central

    Serrano-Velez, Jose L.; Rodriguez-Alvarado, Melanie; Torres-Vazquez, Irma I.; Fraser, Scott E.; Yasumura, Thomas; Vanderpool, Kimberly G.; Rash, John E.; Rosa-Molinar, Eduardo

    2014-01-01

    “Dye-coupling”, whole-mount immunohistochemistry for gap junction channel protein connexin 35 (Cx35), and freeze-fracture replica immunogold labeling (FRIL) reveal an abundance of electrical synapses/gap junctions at glutamatergic mixed synapses in the 14th spinal segment that innervates the adult male gonopodium of Western Mosquitofish, Gambusia affinis (Mosquitofish). To study gap junctions’ role in fast motor behavior, we used a minimally-invasive neural-tract-tracing technique to introduce gap junction-permeant or -impermeant dyes into deep muscles controlling the gonopodium of the adult male Mosquitofish, a teleost fish that rapidly transfers (complete in <20 mS) spermatozeugmata into the female reproductive tract. Dye-coupling in the 14th spinal segment controlling the gonopodium reveals coupling between motor neurons and a commissural primary ascending interneuron (CoPA IN) and shows that the 14th segment has an extensive and elaborate dendritic arbor and more gap junctions than do other segments. Whole-mount immunohistochemistry for Cx35 results confirm dye-coupling and show it occurs via gap junctions. Finally, FRIL shows that gap junctions are at mixed synapses and reveals that >50 of the 62 gap junctions at mixed synapses are in the 14th spinal segment. Our results support and extend studies showing gap junctions at mixed synapses in spinal cord segments involved in control of genital reflexes in rodents, and they suggest a link between mixed synapses and fast motor behavior. The findings provide a basis for studies of specific roles of spinal neurons in the generation/regulation of sex-specific behavior and for studies of gap junctions’ role in regulating fast motor behavior. Finally, the CoPA IN provides a novel candidate neuron for future studies of gap junctions and neural control of fast motor behaviors. PMID:25018700

  17. Single-junction solar cells with the optimum band gap for terrestrial concentrator applications

    DOEpatents

    Wanlass, Mark W.

    1994-01-01

    A single-junction solar cell having the ideal band gap for terrestrial concentrator applications. Computer modeling studies of single-junction solar cells have shown that the presence of absorption bands in the direct spectrum has the effect of "pinning" the optimum band gap for a wide range of operating conditions at a value of 1.14.+-.0.02 eV. Efficiencies exceeding 30% may be possible at high concentration ratios for devices with the ideal band gap.

  18. Protective effect of gap junction uncouplers given during hypoxia against reoxygenation injury in isolated rat hearts.

    PubMed

    Rodríguez-Sinovas, Antonio; García-Dorado, David; Ruiz-Meana, Marisol; Soler-Soler, Jordi

    2006-02-01

    It has been shown that cell-to-cell chemical coupling may persist during severe myocardial hypoxia or ischemia. We aimed to analyze the effects of different, chemically unrelated gap junction uncouplers on the progression of ischemic injury in hypoxic myocardium. First, we analyzed the effects of heptanol, 18alpha-glycyrrhetinic acid, and palmitoleic acid on intracellular Ca2+ concentration during simulated hypoxia (2 mM NaCN) in isolated cardiomyocytes. Next, we analyzed their effects on developed and diastolic tension and electrical impedance in 47 isolated rat hearts submitted to 40 min of hypoxia and reoxygenation. All treatments were applied only during the hypoxic period. Cell injury was determined by lactate dehydrogenase (LDH) release. Heptanol, but not 18alpha-glycyrrhetinic acid nor palmitoleic acid, attenuated the increase in cytosolic Ca2+ concentration induced by simulated ischemia in cardiomyocytes and delayed rigor development (rigor onset at 7.31 +/- 0.71 min in controls vs. 14.76 +/- 1.44 in heptanol-treated hearts, P < 0.001) and the onset of the marked changes in electrical impedance (tissue resistivity: 4.02 +/- 0.29 vs. 7.75 +/- 1.84 min, P = 0.016) in hypoxic rat hearts. LDH release from hypoxic hearts was minimal and was not significantly modified by drugs. However, all gap junction uncouplers, given during hypoxia, attenuated LDH release during subsequent reoxygenation. Dose-response analysis showed that increasing heptanol concentration beyond the level associated with maximal effects on cell coupling resulted in further protection against hypoxic injury. In conclusion, gap junction uncoupling during hypoxia has a protective effect on cell death occurring upon subsequent reoxygenation, and heptanol has, in addition, a marked protective effect independent of its uncoupling actions. PMID:16183732

  19. Gap Junctions in the Ventral Hippocampal-Medial Prefrontal Pathway Are Involved in Anxiety Regulation

    PubMed Central

    Schoenfeld, Timothy J.; Kloth, Alexander D.; Hsueh, Brian; Runkle, Matthew B.; Kane, Gary A.; Wang, Samuel S.-H.

    2014-01-01

    Anxiety disorders are highly prevalent but little is known about their underlying mechanisms. Gap junctions exist in brain regions important for anxiety regulation, such as the ventral hippocampus (vHIP) and mPFC, but their functions in these areas have not been investigated. Using pharmacological blockade of neuronal gap junctions combined with electrophysiological recordings, we found that gap junctions play a role in theta rhythm in the vHIP and mPFC of adult mice. Bilateral infusion of neuronal gap junction blockers into the vHIP decreased anxiety-like behavior on the elevated plus maze and open field. Similar anxiolytic effects were observed with unilateral infusion of these drugs into the vHIP combined with contralateral infusion into the mPFC. No change in anxious behavior was observed with gap junction blockade in the unilateral vHIP alone or in the bilateral dorsal HIP. Since physical exercise is known to reduce anxiety, we examined the effects of long-term running on the expression of the neuronal gap junction protein connexin-36 among inhibitory interneurons and found a reduction in the vHIP. Despite this change, we observed no alteration in theta frequency or power in long-term runners. Collectively, these findings suggest that neuronal gap junctions in the vHIP–mPFC pathway are important for theta rhythm and anxiety regulation under sedentary conditions but that additional mechanisms are likely involved in running-induced reduction in anxiety. PMID:25411496

  20. Bioglass promotes wound healing by affecting gap junction connexin 43 mediated endothelial cell behavior.

    PubMed

    Li, Haiyan; He, Jin; Yu, Hongfei; Green, Colin R; Chang, Jiang

    2016-04-01

    It is well known that gap junctions play an important role in wound healing, and bioactive glass (BG) has been shown to help healing when applied as a wound dressing. However, the effects of BG on gap junctional communication between cells involved in wound healing is not well understood. We hypothesized that BG may be able to affect gap junction mediated cell behavior to enhance wound healing. Therefore, we set out to investigate the effects of BG on gap junction related behavior of endothelial cells in order to elucidate the mechanisms through which BG is operating. In in vitro studies, BG ion extracts prevented death of human umbilical vein endothelial cells (HUVEC) following hypoxia in a dose dependent manner, possibly through connexin hemichannel modulation. In addition, BG showed stimulatory effects on gap junction communication between HUVECs and upregulated connexin43 (Cx43) expression. Furthermore, BG prompted expression of vascular endothelial growth factor and basic fibroblast growth factor as well as their receptors, and vascular endothelial cadherin in HUVECs, all of which are beneficial for vascularization. In vivo wound healing results showed that the wound closure of full-thickness excisional wounds of rats was accelerated by BG with reduced inflammation during initial stages of healing and stimulated angiogenesis during the proliferation stage. Therefore, BG can stimulate wound healing through affecting gap junctions and gap junction related endothelial cell behaviors, including prevention of endothelial cell death following hypoxia, stimulation of gap junction communication and upregulation of critical vascular growth factors, which contributes to the enhancement of angiogenesis in the wound bed and finally to accelerate wound healing. Although many studies have reported that BG stimulates angiogenesis and wound healing, this work reveals the relationship between BG and gap junction connexin 43 mediated endothelial cell behavior and elucidates

  1. Minireview: Regulation of Gap Junction Dynamics by Nuclear Hormone Receptors and Their Ligands

    PubMed Central

    Kapadia, Bhumika J.

    2012-01-01

    Gap junctions are plasma membrane channels comprising connexin proteins that mediate intercellular permeability and communication. The presence, composition, and function of gap junctions can be regulated by diverse sets of physiological signals. Evidence from many hormone-responsive tissues has shown that connexin expression, modification, stability, and localization can be targeted by nuclear hormone receptors and their ligands through both transcriptional and nontranscriptional mechanisms. The focus of this review is to discuss molecular, cellular, and physiological studies that directly link receptor- and ligand-triggered signaling pathways to the regulation of gap junction dynamics. PMID:22935924

  2. Gap distance and interactions in a molecular tunnel junction.

    PubMed

    Chang, Shuai; He, Jin; Zhang, Peiming; Gyarfas, Brett; Lindsay, Stuart

    2011-09-14

    The distance between electrodes in a tunnel junction cannot be determined from the external movement applied to the electrodes because of interfacial forces that distort the electrode geometry at the nanoscale. These distortions become particularly complex when molecules are present in the junction, as demonstrated here by measurements of the AC response of a molecular junction over a range of conductivities from microsiemens to picosiemens. Specific chemical interactions within the junction lead to distinct features in break-junction data, and these have been used to determine the electrode separation in a junction functionalized with 4(5)-(2-mercaptoethyl)-1H-imidazole-2-carboxamide, a reagent developed for reading DNA sequences. PMID:21838292

  3. Changes in gap junction expression and function following ischemic injury of spinal cord white matter

    PubMed Central

    Goncharenko, Karina; Eftekharpour, Eftekhar; Velumian, Alexander A.; Carlen, Peter L.

    2014-01-01

    Gap junctions are widely present in spinal cord white matter; however, their role in modulating the dynamics of axonal dysfunction remains largely unexplored. We hypothesized that inhibition of gap junctions reduces the loss of axonal function during oxygen and glucose deprivation (OGD). The functional role of gap junctions was assessed by electrophysiological recordings of compound action potentials (CAPs) in Wistar rat spinal cord slices with the sucrose gap technique. The in vitro slices were subjected to 30-min OGD. Gap junction connexin (Cx) mRNA expression was determined by qPCR and normalized to β-actin. A 30-min OGD resulted in reduction of CAPs to 14.8 ± 4.6% of their pre-OGD amplitude (n = 5). In the presence of gap junction blockers carbenoxolone (Cbx; 100 μM) and 1-octanol (Oct; 300 μM), the CAP reduction in OGD was to only 35.7 ± 5.7% of pre-OGD amplitude in Cbx (n = 9) and to 37.4 ± 8.9% of pre-OGD amplitude in Oct (n = 10). Both drugs also noticeably prolonged the half-decline time of CAP amplitudes in OGD from 6.0 min in no-drug conditions to 9.6 min in the presence of Cbx and to 7.7 min in the presence of Oct, suggesting that blocking gap junctions reduces conduction loss during OGD. With application of Cbx and Oct in the setting of OGD, expression of Cx30 and Cx43 mRNA was downregulated. Our data provide new insights into the role of gap junctions in white matter ischemia and reveal the necessity of a cautious approach in determining detrimental or beneficial effects of gap junction blockade in white matter ischemia. PMID:25080569

  4. Molecular Transport Junctions Created By Self-Contacting Gapped Nanowires.

    PubMed

    Lim, Jong Kuk; Lee, One-Sun; Jang, Jae-Won; Petrosko, Sarah Hurst; Schatz, George C; Mirkin, Chad A

    2016-08-01

    Molecular transport junctions (MTJs) are important components in molecular electronic devices. However, the synthesis of MTJs remains a significant challenge, as the dimensions of the junction must be tailored for each experiment, based on the molecular lengths. A novel methodology is reported for forming MTJs, taking advantage of capillary and van der Waals forces. PMID:27364594

  5. Osmotic forces and gap junctions in spreading depression: a computational model

    NASA Technical Reports Server (NTRS)

    Shapiro, B. E.

    2001-01-01

    In a computational model of spreading depression (SD), ionic movement through a neuronal syncytium of cells connected by gap junctions is described electrodiffusively. Simulations predict that SD will not occur unless cells are allowed to expand in response to osmotic pressure gradients and K+ is allowed to move through gap junctions. SD waves of [K+]out approximately 25 to approximately 60 mM moving at approximately 2 to approximately 18 mm/min are predicted over the range of parametric values reported in gray matter, with extracellular space decreasing up to approximately 50%. Predicted waveform shape is qualitatively similar to laboratory reports. The delayed-rectifier, NMDA, BK, and Na+ currents are predicted to facilitate SD, while SK and A-type K+ currents and glial activity impede SD. These predictions are consonant with recent findings that gap junction poisons block SD and support the theories that cytosolic diffusion via gap junctions and osmotic forces are important mechanisms underlying SD.

  6. Chloral hydrate decreases gap junction communications in rat liver epithelial cells

    EPA Science Inventory

    Gap junction communication (GJC) is involved in controlling cell proliferation and differentiation. Alterations in GJC are associated with carcinogenesis, but the mechanisms involvedareunknown.Chloralhydrate(CH), a by-productofchlorinedisinfection ofwater,is carcinogenic in mice,...

  7. Preparation of Gap Junctions in Membrane Microdomains for Immunoprecipitation and Mass Spectrometry Interactome Analysis.

    PubMed

    Fowler, Stephanie; Akins, Mark; Bennett, Steffany A L

    2016-01-01

    Protein interaction networks at gap junction plaques are increasingly implicated in a variety of intracellular signaling cascades. Identifying protein interactions of integral membrane proteins is a valuable tool for determining channel function. However, several technical challenges exist. Subcellular fractionation of the bait protein matrix is usually required to identify less abundant proteins in complex homogenates. Sufficient solvation of the lipid environment without perturbation of the protein interactome must also be achieved. The present chapter describes the flotation of light and heavy liver tissue membrane microdomains to facilitate the identification and analysis of endogenous gap junction proteins and includes technical notes for translation to other integral membrane proteins, tissues, or cell culture models. These procedures are valuable tools for the enrichment of gap junction membrane compartments and for the identification of gap junction signaling interactomes. PMID:27207290

  8. Innexin gap junctions in nerve cells coordinate spontaneous contractile behavior in Hydra polyps

    NASA Astrophysics Data System (ADS)

    Takaku, Yasuharu; Hwang, Jung Shan; Wolf, Alexander; Böttger, Angelika; Shimizu, Hiroshi; David, Charles N.; Gojobori, Takashi

    2014-01-01

    Nerve cells and spontaneous coordinated behavior first appeared near the base of animal evolution in the common ancestor of cnidarians and bilaterians. Experiments on the cnidarian Hydra have demonstrated that nerve cells are essential for this behavior, although nerve cells in Hydra are organized in a diffuse network and do not form ganglia. Here we show that the gap junction protein innexin-2 is expressed in a small group of nerve cells in the lower body column of Hydra and that an anti-innexin-2 antibody binds to gap junctions in the same region. Treatment of live animals with innexin-2 antibody eliminates gap junction staining and reduces spontaneous body column contractions. We conclude that a small subset of nerve cells, connected by gap junctions and capable of synchronous firing, act as a pacemaker to coordinate the contraction of the body column in the absence of ganglia.

  9. Innexin gap junctions in nerve cells coordinate spontaneous contractile behavior in Hydra polyps

    PubMed Central

    Takaku, Yasuharu; Hwang, Jung Shan; Wolf, Alexander; Böttger, Angelika; Shimizu, Hiroshi; David, Charles N.; Gojobori, Takashi

    2014-01-01

    Nerve cells and spontaneous coordinated behavior first appeared near the base of animal evolution in the common ancestor of cnidarians and bilaterians. Experiments on the cnidarian Hydra have demonstrated that nerve cells are essential for this behavior, although nerve cells in Hydra are organized in a diffuse network and do not form ganglia. Here we show that the gap junction protein innexin-2 is expressed in a small group of nerve cells in the lower body column of Hydra and that an anti-innexin-2 antibody binds to gap junctions in the same region. Treatment of live animals with innexin-2 antibody eliminates gap junction staining and reduces spontaneous body column contractions. We conclude that a small subset of nerve cells, connected by gap junctions and capable of synchronous firing, act as a pacemaker to coordinate the contraction of the body column in the absence of ganglia. PMID:24394722

  10. Innexin gap junctions in nerve cells coordinate spontaneous contractile behavior in Hydra polyps.

    PubMed

    Takaku, Yasuharu; Hwang, Jung Shan; Wolf, Alexander; Böttger, Angelika; Shimizu, Hiroshi; David, Charles N; Gojobori, Takashi

    2014-01-01

    Nerve cells and spontaneous coordinated behavior first appeared near the base of animal evolution in the common ancestor of cnidarians and bilaterians. Experiments on the cnidarian Hydra have demonstrated that nerve cells are essential for this behavior, although nerve cells in Hydra are organized in a diffuse network and do not form ganglia. Here we show that the gap junction protein innexin-2 is expressed in a small group of nerve cells in the lower body column of Hydra and that an anti-innexin-2 antibody binds to gap junctions in the same region. Treatment of live animals with innexin-2 antibody eliminates gap junction staining and reduces spontaneous body column contractions. We conclude that a small subset of nerve cells, connected by gap junctions and capable of synchronous firing, act as a pacemaker to coordinate the contraction of the body column in the absence of ganglia. PMID:24394722

  11. Wind-Resistant Filler for Tile Gaps

    NASA Technical Reports Server (NTRS)

    Bellavia, J.; Quigley, I. A.; Callahan, T. S.

    1982-01-01

    Filler developed for gaps between insulating tiles on Space Shuttle finds application in industries that use tiles for thermal or environmental protection. Filler consists of tight-fitting ceramic tubes and fibrous alumina. Combination resists high wind loads while providing requisite heat protection. Quartz-thread stitching holds envelope together.

  12. Death of Neurons following Injury Requires Conductive Neuronal Gap Junction Channels but Not a Specific Connexin.

    PubMed

    Fontes, Joseph D; Ramsey, Jon; Polk, Jeremy M; Koop, Andre; Denisova, Janna V; Belousov, Andrei B

    2015-01-01

    Pharmacological blockade or genetic knockout of neuronal connexin 36 (Cx36)-containing gap junctions reduces neuronal death caused by ischemia, traumatic brain injury and NMDA receptor (NMDAR)-mediated excitotoxicity. However, whether Cx36 gap junctions contribute to neuronal death via channel-dependent or channel-independent mechanism remains an open question. To address this, we manipulated connexin protein expression via lentiviral transduction of mouse neuronal cortical cultures and analyzed neuronal death twenty-four hours following administration of NMDA (a model of NMDAR excitotoxicity) or oxygen-glucose deprivation (a model of ischemic injury). In cultures prepared from wild-type mice, over-expression and knockdown of Cx36-containing gap junctions augmented and prevented, respectively, neuronal death from NMDAR-mediated excitotoxicity and ischemia. In cultures obtained form from Cx36 knockout mice, re-expression of functional gap junction channels, containing either neuronal Cx36 or non-neuronal Cx43 or Cx31, resulted in increased neuronal death following insult. In contrast, the expression of communication-deficient gap junctions (containing mutated connexins) did not have this effect. Finally, the absence of ethidium bromide uptake in non-transduced wild-type neurons two hours following NMDAR excitotoxicity or ischemia suggested the absence of active endogenous hemichannels in those neurons. Taken together, these results suggest a role for neuronal gap junctions in cell death via a connexin type-independent mechanism that likely relies on channel activities of gap junctional complexes among neurons. A possible contribution of gap junction channel-permeable death signals in neuronal death is discussed. PMID:26017008

  13. Death of Neurons following Injury Requires Conductive Neuronal Gap Junction Channels but Not a Specific Connexin

    PubMed Central

    Fontes, Joseph D.; Ramsey, Jon; Polk, Jeremy M; Koop, Andre; Denisova, Janna V.; Belousov, Andrei B.

    2015-01-01

    Pharmacological blockade or genetic knockout of neuronal connexin 36 (Cx36)-containing gap junctions reduces neuronal death caused by ischemia, traumatic brain injury and NMDA receptor (NMDAR)-mediated excitotoxicity. However, whether Cx36 gap junctions contribute to neuronal death via channel-dependent or channel-independent mechanism remains an open question. To address this, we manipulated connexin protein expression via lentiviral transduction of mouse neuronal cortical cultures and analyzed neuronal death twenty-four hours following administration of NMDA (a model of NMDAR excitotoxicity) or oxygen-glucose deprivation (a model of ischemic injury). In cultures prepared from wild-type mice, over-expression and knockdown of Cx36-containing gap junctions augmented and prevented, respectively, neuronal death from NMDAR-mediated excitotoxicity and ischemia. In cultures obtained form from Cx36 knockout mice, re-expression of functional gap junction channels, containing either neuronal Cx36 or non-neuronal Cx43 or Cx31, resulted in increased neuronal death following insult. In contrast, the expression of communication-deficient gap junctions (containing mutated connexins) did not have this effect. Finally, the absence of ethidium bromide uptake in non-transduced wild-type neurons two hours following NMDAR excitotoxicity or ischemia suggested the absence of active endogenous hemichannels in those neurons. Taken together, these results suggest a role for neuronal gap junctions in cell death via a connexin type-independent mechanism that likely relies on channel activities of gap junctional complexes among neurons. A possible contribution of gap junction channel-permeable death signals in neuronal death is discussed. PMID:26017008

  14. Kinase programs spatiotemporally regulate gap junction assembly and disassembly: Effects on wound repair.

    PubMed

    Solan, Joell L; Lampe, Paul D

    2016-02-01

    Gap junctions are highly ordered plasma membrane domains that are constantly assembled, remodeled and turned over due to the short half-life of connexins, the integral membrane proteins that form gap junctions. Connexin 43 (Cx43), by far the most widely expressed connexin, is phosphorylated at multiple serine residues in the cytoplasmic, C-terminal region allowing for exquisite cellular control over gap junctional communication. This is evident during epidermal wounding where spatiotemporal changes in connexin expression occur as cells are instructed whether to die, proliferate or migrate to promote repair. Early gap junctional communication is required for initiation of keratinocyte migration, but accelerated Cx43 turnover is also critical for proper wound healing at later stages. These events are controlled via a "kinase program" where sequential phosphorylation of Cx43 leads to reductions in Cx43's half-life and significant depletion of gap junctions from the plasma membrane within several hours. The complex regulation of gap junction assembly and turnover affords several steps where intervention might speed wound healing. PMID:26706150

  15. Inhibition of connexin43 gap junction channels by the endocrine disruptor ioxynil

    SciTech Connect

    Leithe, Edward; Kjenseth, Ane; Bruun, Jarle; Sirnes, Solveig; Rivedal, Edgar

    2010-08-15

    Gap junctions are intercellular plasma membrane domains containing channels that mediate transport of ions, metabolites and small signaling molecules between adjacent cells. Gap junctions play important roles in a variety of cellular processes, including regulation of cell growth and differentiation, maintenance of tissue homeostasis and embryogenesis. The constituents of gap junction channels are a family of trans-membrane proteins called connexins, of which the best-studied is connexin43. Connexin43 functions as a tumor suppressor protein in various tissue types and is frequently dysregulated in human cancers. The pesticide ioxynil has previously been shown to act as an endocrine disrupting chemical and has multiple effects on the thyroid axis. Furthermore, both ioxynil and its derivative ioxynil octanoate have been reported to induce tumors in animal bioassays. However, the molecular mechanisms underlying the possible tumorigenic effects of these compounds are unknown. In the present study we show that ioxynil and ioxynil octanoate are strong inhibitors of connexin43 gap junction channels. Both compounds induced rapid loss of connexin43 gap junctions at the plasma membrane and increased connexin43 degradation. Ioxynil octanoate, but not ioxynil, was found to be a strong activator of ERK1/2. The compounds also had different effects on the phosphorylation status of connexin43. Taken together, the data show that ioxynil and ioxynil octanoate are potent inhibitors of intercellular communication via gap junctions.

  16. Neuronal gap junctions play a role in the secondary neuronal death following controlled cortical impact.

    PubMed

    Belousov, Andrei B; Wang, Yongfu; Song, Ji-Hoon; Denisova, Janna V; Berman, Nancy E; Fontes, Joseph D

    2012-08-22

    In the mammalian CNS, excessive release of glutamate and overactivation of glutamate receptors are responsible for the secondary (delayed) neuronal death following neuronal injury, including ischemia, traumatic brain injury (TBI) and epilepsy. Recent studies in mice showed a critical role for neuronal gap junctions in NMDA receptor-mediated excitotoxicity and ischemia-mediated neuronal death. Here, using controlled cortical impact (CCI) in adult mice, as a model of TBI, and Fluoro-Jade B staining for analysis of neuronal death, we set to determine whether neuronal gap junctions play a role in the CCI-mediated secondary neuronal death. We report that 24h post-CCI, substantial neuronal death is detected in a number of brain regions outside the injury core, including the striatum. The striatal neuronal death is reduced both in wild-type mice by systemic administration of mefloquine (a relatively selective blocker of neuronal gap junctions) and in knockout mice lacking connexin 36 (neuronal gap junction protein). It is also reduced by inactivation of group II metabotropic glutamate receptors (with LY341495) which, as reported previously, control the rapid increase in neuronal gap junction coupling following different types of neuronal injury. The results suggest that neuronal gap junctions play a critical role in the CCI-induced secondary neuronal death. PMID:22781494

  17. Gap Junctions, Dendrites and Resonances: A Recipe for Tuning Network Dynamics

    PubMed Central

    2013-01-01

    Gap junctions, also referred to as electrical synapses, are expressed along the entire central nervous system and are important in mediating various brain rhythms in both normal and pathological states. These connections can form between the dendritic trees of individual cells. Many dendrites express membrane channels that confer on them a form of sub-threshold resonant dynamics. To obtain insight into the modulatory role of gap junctions in tuning networks of resonant dendritic trees, we generalise the “sum-over-trips” formalism for calculating the response function of a single branching dendrite to a gap junctionally coupled network. Each cell in the network is modelled by a soma connected to an arbitrary structure of dendrites with resonant membrane. The network is treated as a single extended tree structure with dendro-dendritic gap junction coupling. We present the generalised “sum-over-trips” rules for constructing the network response function in terms of a set of coefficients defined at special branching, somatic and gap-junctional nodes. Applying this framework to a two-cell network, we construct compact closed form solutions for the network response function in the Laplace (frequency) domain and study how a preferred frequency in each soma depends on the location and strength of the gap junction. PMID:23945377

  18. Intracellular spermine prevents acid-induced uncoupling of Cx43 gap junction channels.

    PubMed

    Skatchkov, Serguei N; Bukauskas, Feliksas F; Benedikt, Jan; Inyushin, Mikhail; Kucheryavykh, Yuriy V

    2015-06-17

    Polyamines (PAs), such as spermine and spermidine, modulate the activity of numerous receptors and channels in the central nervous system (CNS) and are stored in glial cells; however, little attention has been paid to their role in the regulation of connexin (Cx)-based gap junction channels. We have previously shown that PAs facilitate diffusion of Lucifer Yellow through astrocytic gap junctions in acute brain slices; therefore, we hypothesized that spermine can regulate Cx43-mediated (as the most abundant Cx in astrocytes) gap junctional communication. We used electrophysiological patch-clamp recording from paired Novikoff cells endogenously expressing Cx43 and HeLaCx43-EGFP transfectants to study pH-dependent modulation of cell-cell coupling in the presence or absence of PAs. Our results showed (i) a higher increase in gap junctional communication at higher concentrations of cytoplasmic spermine, and (ii) that spermine prevented uncoupling of gap junctions at low intracellular pH. Taken together, we conclude that spermine enhances Cx43-mediated gap junctional communication and may preserve neuronal excitability during ischemia and trauma when pH in the brain acidifies. We, therefore, suggest a new role of spermine in the regulation of a Cx43-based network under (patho)physiological conditions. PMID:26011388

  19. Bi-directional gap junction-mediated soma-germline communication is essential for spermatogenesis.

    PubMed

    Smendziuk, Christopher M; Messenberg, Anat; Vogl, A Wayne; Tanentzapf, Guy

    2015-08-01

    Soma-germline interactions play conserved essential roles in regulating cell proliferation, differentiation, patterning and homeostasis in the gonad. In the Drosophila testis, secreted signalling molecules of the JAK-STAT, Hedgehog, BMP and EGF pathways are used to mediate soma-germline communication. Here, we demonstrate that gap junctions may also mediate direct, bi-directional signalling between the soma and germ line. When gap junctions between the soma and germ line are disrupted, germline differentiation is blocked and germline stem cells are not maintained. In the soma, gap junctions are required to regulate proliferation and differentiation. Localization and RNAi-mediated knockdown studies reveal that gap junctions in the fly testis are heterotypic channels containing Zpg (Inx4) and Inx2 on the germ line and the soma side, respectively. Overall, our results show that bi-directional gap junction-mediated signalling is essential to coordinate the soma and germ line to ensure proper spermatogenesis in Drosophila. Moreover, we show that stem cell maintenance and differentiation in the testis are directed by gap junction-derived cues. PMID:26116660

  20. Intracellular spermine prevents acid-induced uncoupling of Cx43 gap junction channels

    PubMed Central

    Skatchkov, Serguei N.; Bukauskas, Feliksas F.; Benedikt, Jan; Inyushin, Mikhail

    2015-01-01

    Polyamines (PAs), such as spermine and spermidine, modulate the activity of numerous receptors and channels in the central nervous system (CNS) and are stored in glial cells; however, little attention has been paid to their role in the regulation of connexin (Cx)-based gap junction channels. We have previously shown that PAs facilitate diffusion of Lucifer Yellow through astrocytic gap junctions in acute brain slices; therefore, we hypothesized that spermine can regulate Cx43-mediated (as the most abundant Cx in astrocytes) gap junctional communication. We used electrophysiological patch-clamp recording from paired Novikoff cells endogenously expressing Cx43 and HeLaCx43-EGFP transfectants to study pH-dependent modulation of cell–cell coupling in the presence or absence of PAs. Our results showed (i) a higher increase in gap junctional communication at higher concentrations of cytoplasmic spermine, and (ii) that spermine prevented uncoupling of gap junctions at low intracellular pH. Taken together, we conclude that spermine enhances Cx43-mediated gap junctional communication and may preserve neuronal excitability during ischemia and trauma when pH in the brain acidifies. We, therefore, suggest a new role of spermine in the regulation of a Cx43-based network under (patho)physiological conditions. PMID:26011388

  1. Gap junction networks in mushroom bodies participate in visual learning and memory in Drosophila

    PubMed Central

    Liu, Qingqing; Yang, Xing; Tian, Jingsong; Gao, Zhongbao; Wang, Meng; Li, Yan; Guo, Aike

    2016-01-01

    Gap junctions are widely distributed in the brains across species and play essential roles in neural information processing. However, the role of gap junctions in insect cognition remains poorly understood. Using a flight simulator paradigm and genetic tools, we found that gap junctions are present in Drosophila Kenyon cells (KCs), the major neurons of the mushroom bodies (MBs), and showed that they play an important role in visual learning and memory. Using a dye coupling approach, we determined the distribution of gap junctions in KCs. Furthermore, we identified a single pair of MB output neurons (MBONs) that possess a gap junction connection to KCs, and provide strong evidence that this connection is also required for visual learning and memory. Together, our results reveal gap junction networks in KCs and the KC-MBON circuit, and bring new insight into the synaptic network underlying fly’s visual learning and memory. DOI: http://dx.doi.org/10.7554/eLife.13238.001 PMID:27218450

  2. Low-bias negative differential resistance effect in armchair graphene nanoribbon junctions

    SciTech Connect

    Li, Suchun; Gan, Chee Kwan; Son, Young-Woo; Feng, Yuan Ping; Quek, Su Ying

    2015-01-05

    Graphene nanoribbons with armchair edges (AGNRs) have bandgaps that can be flexibly tuned via the ribbon width. A junction made of a narrower AGNR sandwiched between two wider AGNR leads was recently reported to possess two perfect transmission channels close to the Fermi level. Here, we report that by using a bias voltage to drive these transmission channels into the gap of the wider AGNR lead, we can obtain a negative differential resistance (NDR) effect. Owing to the intrinsic properties of the AGNR junctions, the on-set bias reaches as low as ∼0.2 V and the valley current almost vanishes. We further show that such NDR effect is robust against details of the atomic structure of the junction, substrate, and whether the junction is made by etching or by hydrogenation.

  3. Role of gap junctions in the propagation of the cardiac action potential.

    PubMed

    Rohr, Stephan

    2004-05-01

    Gap junctions play a pivotal role for the velocity and the safety of impulse propagation in cardiac tissue. Under physiologic conditions, the specific subcellular distribution of gap junctions together with the tight packaging of the rod-shaped cardiomyocytes underlies anisotropic conduction, which is continuous at the macroscopic scale. However, when breaking down the three-dimensional network of cells into linear single cell chains, gap junctions can be shown to limit axial current flow and to induce 'saltatory' conduction at unchanged overall conduction velocities. In two- and three-dimensional tissue, these discontinuities disappear due to lateral averaging of depolarizing current flow at the activation wavefront. During gap junctional uncoupling, discontinuities reappear and are accompanied by slowed and meandering conduction. Critical gap junctional uncoupling reduces conduction velocities to a much larger extent than does a reduction of excitability, which suggests that the safety for conduction is higher at any given conduction velocity for gap junctional uncoupling. In uniformly structured tissue, gap junctional uncoupling is accompanied by a parallel decrease in conduction velocity. However, this is not necessarily the case for non-uniform structures like tissue expansion where partial uncoupling paradoxically increases conduction velocity and has the capacity to remove unidirectional conduction blocks. Whereas the impact of gap junctions on impulse conduction is generally assessed from the point of view of cell coupling among cardiomyocytes, it is possible that other cell types within the myocardium might be coupled to cardiomyocytes as well. In this context, it has been shown that fibroblasts establish successful conduction between sheets of cardiomyocytes over distances as long as 300 microm. This might not only explain electrical synchronization of heart transplants but might be of importance for cardiac diseases involving fibrosis. Finally, the

  4. Single-junction solar cells with the optimum band gap for terrestrial concentrator applications

    DOEpatents

    Wanlass, M.W.

    1994-12-27

    A single-junction solar cell is described having the ideal band gap for terrestrial concentrator applications. Computer modeling studies of single-junction solar cells have shown that the presence of absorption bands in the direct spectrum has the effect of ''pinning'' the optimum band gap for a wide range of operating conditions at a value of 1.14[+-]0.02 eV. Efficiencies exceeding 30% may be possible at high concentration ratios for devices with the ideal band gap. 7 figures.

  5. LRP6 acts as a scaffold protein in cardiac gap junction assembly

    PubMed Central

    Li, Jun; Li, Changming; Liang, Dandan; Lv, Fei; Yuan, Tianyou; The, Erlinda; Ma, Xiue; Wu, Yahan; Zhen, Lixiao; Xie, Duanyang; Wang, Shiyi; Liu, Yuan; Huang, Jian; Shi, Jingyi; Liu, Yi; Shi, Dan; Xu, Liang; Lin, Li; Peng, Luying; Cui, Jianmin; Zhu, Weidong; Chen, Yi-Han

    2016-01-01

    Low-density lipoprotein receptor-related protein 6 (LRP6) is a Wnt co-receptor in the canonical Wnt/β-catenin signalling. Here, we report the scaffold function of LRP6 in gap junction formation of cardiomyocytes. Cardiac LRP6 is spatially restricted to intercalated discs and binds to gap junction protein connexin 43 (Cx43). A deficiency in LRP6 disrupts Cx43 gap junction formation and thereby impairs the cell-to-cell coupling, which is independent of Wnt/β-catenin signalling. The defect in Cx43 gap junction resulting from LRP6 reduction is attributable to the defective traffic of de novo Cx43 proteins from the endoplasmic reticulum to the Golgi apparatus, leading to the lysosomal degradation of Cx43 proteins. Accordingly, the hearts of conditional cardiac-specific Lrp6-knockout mice consistently exhibit overt reduction of Cx43 gap junction plaques without any abnormality in Wnt signalling and are predisposed to lethal arrhythmias. These findings uncover a distinct role of LRP6 as a platform for intracellular protein trafficking. PMID:27250245

  6. Netrin and Frazzled regulate presynaptic gap junctions at a Drosophila giant synapse.

    PubMed

    Orr, Brian O; Borgen, Melissa A; Caruccio, Phyllis M; Murphey, Rodney K

    2014-04-16

    Netrin and its receptor, Frazzled, dictate the strength of synaptic connections in the giant fiber system (GFS) of Drosophila melanogaster by regulating gap junction localization in the presynaptic terminal. In Netrin mutant animals, the synaptic coupling between a giant interneuron and the "jump" motor neuron was weakened and dye coupling between these two neurons was severely compromised or absent. In cases in which Netrin mutants displayed apparently normal synaptic anatomy, half of the specimens exhibited physiologically defective synapses and dye coupling between the giant fiber (GF) and the motor neuron was reduced or eliminated, suggesting that gap junctions were disrupted in the Netrin mutants. When we examined the gap junctions with antibodies to Shaking-B (ShakB) Innexin, they were significantly decreased or absent in the presynaptic terminal of the mutant GF. Frazzled loss of function mutants exhibited similar defects in synaptic transmission, dye coupling, and gap junction localization. These data are the first to show that Netrin and Frazzled regulate the placement of gap junctions presynaptically at a synapse. PMID:24741033

  7. Mix and match: Investigating heteromeric and heterotypic gap junction channels in model systems and native tissues

    PubMed Central

    Koval, Michael; Molina, Samuel A.; Burt, Janis M.

    2014-01-01

    This review is based in part on a roundtable discussion session: “Physiological roles for heterotypic/heteromeric channels” at the 2013 International Gap Junction Conference (IGJC 2013) in Charleston, South Carolina. It is well recognized that multiple connexins can specifically co-assemble to form mixed gap junction channels with unique properties as a means to regulate intercellular communication. Compatibility determinants for both heteromeric and heterotypic gap junction channel formation have been identified and associated with specific connexin amino acid motifs. Hetero-oligomerization is also a regulated process; differences in connexin quality control and monomer stability are likely to play integral roles to control interactions between compatible connexins. Gap junctions in oligodendrocyte:astrocyte communication and in the cardiovascular system have emerged as key systems where heterotypic and heteromeric channels have unique physiologic roles. There are several methodologies to study heteromeric and heterotypic channels that are best applied to either heterologous expression systems, native tissues or both. There remains a need to use and develop different experimental approaches in order to understand the prevalence and roles for mixed gap junction channels in human physiology. PMID:24561196

  8. Modelling the effect of gap junctions on tissue-level cardiac electrophysiology.

    PubMed

    Bruce, Doug; Pathmanathan, Pras; Whiteley, Jonathan P

    2014-02-01

    When modelling tissue-level cardiac electrophysiology, a continuum approximation to the discrete cell-level equations, known as the bidomain equations, is often used to maintain computational tractability. Analysing the derivation of the bidomain equations allows us to investigate how microstructure, in particular gap junctions that electrically connect cells, affect tissue-level conductivity properties. Using a one-dimensional cable model, we derive a modified form of the bidomain equations that take gap junctions into account, and compare results of simulations using both the discrete and continuum models, finding that the underlying conduction velocity of the action potential ceases to match up between models when gap junctions are introduced at physiologically realistic coupling levels. We show that this effect is magnified by: (i) modelling gap junctions with reduced conductivity; (ii) increasing the conductance of the fast sodium channel; and (iii) an increase in myocyte length. From this, we conclude that the conduction velocity arising from the bidomain equations may not be an accurate representation of the underlying discrete system. In particular, the bidomain equations are less likely to be valid when modelling certain diseased states whose symptoms include a reduction in gap junction coupling or an increase in myocyte length. PMID:24338526

  9. Gap junctions between CA3 pyramidal cells contribute to network synchronization in neonatal hippocampus.

    PubMed

    Molchanova, Svetlana M; Huupponen, Johanna; Lauri, Sari E; Taira, Tomi

    2016-08-01

    Direct electrical coupling between neurons through gap junctions is prominent during development, when synaptic connectivity is scarce, providing the additional intercellular connectivity. However, functional studies of gap junctions are hampered by the unspecificity of pharmacological tools available. Here we have investigated gap-junctional coupling between CA3 pyramidal cells in neonatal hippocampus and its contribution to early network activity. Four different gap junction inhibitors, including the general blocker carbenoxolone, decreased the frequency of network activity bursts in CA3 area of hippocampus of P3-6 rats, suggesting the involvement of electrical connections in the generation of spontaneous network activity. In CA3 pyramidal cells, spikelets evoked by local stimulation of stratum oriens, were inhibited by carbenoxolone, but not by inhibitors of glutamatergic and GABAergic synaptic transmission, signifying the presence of electrical connectivity through axo-axonic gap junctions. Carbenoxolone also decreased the success rate of firing antidromic action potentials in response to stimulation, and changed the pattern of spontaneous action potential firing of CA3 pyramidal cells. Altogether, these data suggest that electrical coupling of CA3 pyramidal cells contribute to the generation of the early network events in neonatal hippocampus by modulating their firing pattern and synchronization. PMID:26926429

  10. LRP6 acts as a scaffold protein in cardiac gap junction assembly.

    PubMed

    Li, Jun; Li, Changming; Liang, Dandan; Lv, Fei; Yuan, Tianyou; The, Erlinda; Ma, Xiue; Wu, Yahan; Zhen, Lixiao; Xie, Duanyang; Wang, Shiyi; Liu, Yuan; Huang, Jian; Shi, Jingyi; Liu, Yi; Shi, Dan; Xu, Liang; Lin, Li; Peng, Luying; Cui, Jianmin; Zhu, Weidong; Chen, Yi-Han

    2016-01-01

    Low-density lipoprotein receptor-related protein 6 (LRP6) is a Wnt co-receptor in the canonical Wnt/β-catenin signalling. Here, we report the scaffold function of LRP6 in gap junction formation of cardiomyocytes. Cardiac LRP6 is spatially restricted to intercalated discs and binds to gap junction protein connexin 43 (Cx43). A deficiency in LRP6 disrupts Cx43 gap junction formation and thereby impairs the cell-to-cell coupling, which is independent of Wnt/β-catenin signalling. The defect in Cx43 gap junction resulting from LRP6 reduction is attributable to the defective traffic of de novo Cx43 proteins from the endoplasmic reticulum to the Golgi apparatus, leading to the lysosomal degradation of Cx43 proteins. Accordingly, the hearts of conditional cardiac-specific Lrp6-knockout mice consistently exhibit overt reduction of Cx43 gap junction plaques without any abnormality in Wnt signalling and are predisposed to lethal arrhythmias. These findings uncover a distinct role of LRP6 as a platform for intracellular protein trafficking. PMID:27250245

  11. Cross-talk between pulmonary injury, oxidant stress, and gap junctional communication.

    PubMed

    Johnson, Latoya N; Koval, Michael

    2009-02-01

    Gap junction channels interconnect several different types of cells in the lung, ranging from the alveolar epithelium to the pulmonary vasculature, each of which expresses a unique subset of gap junction proteins (connexins). Major lung functions regulated by gap junctional communication include coordination of ciliary beat frequency and inflammation. Gap junctions help enable the alveolus to regulate surfactant secretion as an integrated system, in which type I cells act as mechanical sensors that transmit calcium transients to type II cells. Thus, disruption of epithelial gap junctional communication, particularly during acute lung injury, can interfere with these processes and increase the severity of injury. Consistent with this, connexin expression is altered during lung injury, and connexin-deficiency has a negative impact on the injury response and lung-growth control. It has recently been shown that alcohol abuse is a significant risk factor associated with acute respiratory distress syndrome. Oxidant stress and hormone-signaling cascades in the lung induced by prolonged alcohol ingestion are discussed, as well as the effects of these pathways on connexin expression and function. PMID:18816185

  12. Josephson Current in a Gapped Graphene Superconductor/Barrier/Superconductor Junction: Case of Massive Electrons

    NASA Astrophysics Data System (ADS)

    Suwannasit, Tatnatchai; Tang, I.-Ming; Hoonsawat, Rassmidara; Soodchomshom, Bumned

    2011-10-01

    The Josephson effect in a gapped graphene-based superconductor/barrier/superconductor junction is studied. The superconductivity in gapped graphene may be achieved by depositing conventional superconductor on the top of the gapped graphene such as graphene grown on SiC substrate. In gapped graphene system, the carriers exhibit massive Dirac fermions. We focus on the effect of pseudo-Dirac-like mass on the supercurrent. In contrast to that in the gapless graphene superconductor/barrier/superconductor junction, we find that the supercurrent exhibits dependency of the Fermi energy. Also, the massive supercurrent anomalously oscillates as a function of the gate potential. This novel behavior is due to the effect of electrons acquiring mass in gapped graphene.

  13. Pharmacological and Genetic Evidence for Gap Junctions as Potential New Insecticide Targets in the Yellow Fever Mosquito, Aedes aegypti

    PubMed Central

    Calkins, Travis L.; Piermarini, Peter M.

    2015-01-01

    The yellow fever mosquito Aedes aegypti is an important vector of viral diseases that impact global health. Insecticides are typically used to manage mosquito populations, but the evolution of insecticide resistance is limiting their effectiveness. Thus, identifying new molecular and physiological targets in mosquitoes is needed to facilitate insecticide discovery and development. Here we test the hypothesis that gap junctions are valid molecular and physiological targets for new insecticides. Gap junctions are intercellular channels that mediate direct communication between neighboring cells and consist of evolutionarily distinct proteins in vertebrate (connexins) and invertebrate (innexins) animals. We show that the injection of pharmacological inhibitors of gap junctions (i.e., carbenoxolone, meclofenamic acid, or mefloquine) into the hemolymph of adult female mosquitoes elicits dose-dependent toxic effects, with mefloquine showing the greatest potency. In contrast, when applied topically to the cuticle, carbenoxolone was the only inhibitor to exhibit full efficacy. In vivo urine excretion assays demonstrate that both carbenoxolone and mefloquine inhibit the diuretic output of adult female mosquitoes, suggesting inhibition of excretory functions as part of their mechanism of action. When added to the rearing water of 1st instar larvae, carbenoxolone and meclofenamic acid both elicit dose-dependent toxic effects, with meclofenamic acid showing the greatest potency. Injecting a double-stranded RNA cocktail against innexins into the hemolymph of adult female mosquitoes knock down whole-animal innexin mRNA expression and decreases survival of the mosquitoes. Taken together these data indicate that gap junctions may provide novel molecular and physiological targets for the development of insecticides. PMID:26325403

  14. High throughput assay of diffusion through Cx43 gap junction channels with a microfluidic chip.

    PubMed

    Bathany, Cédric; Beahm, Derek; Felske, James D; Sachs, Frederick; Hua, Susan Z

    2011-02-01

    This paper describes a microfluidic-based assay capable of measuring gap-junction mediated dye diffusion in cultured cells. The technique exploits multistream laminar flow to selectively expose cells to different environments, enabling continuous loading of cells in one compartment while monitoring, in real time, dye diffusion into cells of a neighboring compartment. A simple one-dimensional diffusion model fit to the data extracted the diffusion coefficient of four different dyes, 5-(6)-carboxyfluorescein, 5-chloromethylfluorescein, Oregon green 488 carboxylic acid, and calcein. Different inhibitors were assayed for their ability to reduce dye coupling. The chip can screen multiple inhibitors in parallel in the same cell preparation, demonstrating its potential for high throughput. The technique provides a convenient method to measure gap junction mediated diffusion and a screen for drugs that affect gap junction communication. PMID:21182279

  15. Synchronization of Ca(2+)-signals within insulin-secreting pseudoislets: effects of gap-junctional uncouplers.

    PubMed

    Squires, P E; Hauge-Evans, A C; Persaud, S J; Jones, P M

    2000-05-01

    The secretory response of the intact islet is greater than the response of individual beta-cells in isolation, and functional coupling between cells is critical in insulin release. The changes in intracellular Ca(2+)([Ca(2+)](i)) which initiate insulin secretory responses are synchronized between groups of cells within the islet, and gap-junctions are thought to play a central role in coordinating signalling events. We have used the MIN6 insulin-secreting cell line, to examine whether uncoupling gap-junctions alters the synchronicity of nutrient- and non-nutrient-evoked Ca(2+)oscillations, or affects insulin secretion. MIN6 cells express mRNA species that can be amplified using PCR primers for connexin 36. A commonly used gap-junctional inhibitor, heptanol, inhibited glucose- and tolbutamide-induced Ca(2+)-oscillations to basal levels in MIN6 cell clusters at concentrations of 0.5 mM and greater, and it had similar effects in pseudoislets when used at 2.5 mM. Lower heptanol concentrations altered the frequency of Ca(2+)transients without affecting their synchronicity, in both monolayers and pseudoislets. Heptanol also had effects on insulin secretion from MIN6 pseudoislets such that 1 mM enhanced secretion while 2.5 mM was inhibitory. These data suggest that heptanol has multiple effects in pancreatic beta-cells, none of which appears to be related to uncoupling of synchronicity of Ca(2+)signalling between cells. A second gap-junction uncoupler, 18 alpha-glycyrrhetinic acid, also failed to uncouple synchronized Ca(2+)-oscillations, and it had no effect on insulin secretion. These data provide evidence that Ca(2+)signalling events occur simultaneously across the bulk mass of the pseudoislet, and suggest that gap-junctions are not required to coordinate the synchronicity of these events, nor is communication via gap junctions essential for integrated insulin secretory responses. PMID:10859595

  16. Astrocytic gap junctional networks suppress cellular damage in an in vitro model of ischemia

    SciTech Connect

    Shinotsuka, Takanori; Yasui, Masato; Nuriya, Mutsuo

    2014-02-07

    Highlights: • Astrocytes exhibit characteristic changes in [Ca{sup 2+}]{sub i} under OGD. • Astrocytic [Ca{sup 2+}]{sub i} increase is synchronized with a neuronal anoxic depolarization. • Gap junctional couplings protect neurons as well as astrocytes during OGD. - Abstract: Astrocytes play pivotal roles in both the physiology and the pathophysiology of the brain. They communicate with each other via extracellular messengers as well as through gap junctions, which may exacerbate or protect against pathological processes in the brain. However, their roles during the acute phase of ischemia and the underlying cellular mechanisms remain largely unknown. To address this issue, we imaged changes in the intracellular calcium concentration ([Ca{sup 2+}]{sub i}) in astrocytes in mouse cortical slices under oxygen/glucose deprivation (OGD) condition using two-photon microscopy. Under OGD, astrocytes showed [Ca{sup 2+}]{sub i} oscillations followed by larger and sustained [Ca{sup 2+}]{sub i} increases. While the pharmacological blockades of astrocytic receptors for glutamate and ATP had no effect, the inhibitions of gap junctional intercellular coupling between astrocytes significantly advanced the onset of the sustained [Ca{sup 2+}]{sub i} increase after OGD exposure. Interestingly, the simultaneous recording of the neuronal membrane potential revealed that the onset of the sustained [Ca{sup 2+}]{sub i} increase in astrocytes was synchronized with the appearance of neuronal anoxic depolarization. Furthermore, the blockade of gap junctional coupling resulted in a concurrent faster appearance of neuronal depolarizations, which remain synchronized with the sustained [Ca{sup 2+}]{sub i} increase in astrocytes. These results indicate that astrocytes delay the appearance of the pathological responses of astrocytes and neurons through their gap junction-mediated intercellular network under OGD. Thus, astrocytic gap junctional networks provide protection against tissue damage

  17. The Mr 28,000 gap junction proteins from rat heart and liver are different but related.

    PubMed

    Nicholson, B J; Gros, D B; Kent, S B; Hood, L E; Revel, J P

    1985-06-10

    The sequence of the amino-terminal 32 residues of the rat heart Mr 28,000 gap junction protein presented here allows, for the first time, a sequence comparison of gap junctional proteins from different tissues (heart and liver). Comparison of the rat heart gap junction protein sequence and that available from rat liver reveals 43% sequence identity and conservative changes at an additional 25% of the positions. Both proteins exhibit a hydrophobic domain which could represent a transmembrane span of the junction. This result unequivocally demonstrates the existence of at least two forms of the gap junction protein. As yet, no homology is evident between the gap junctional proteins of either heart or liver and main intrinsic protein from rat eye lens. PMID:2987225

  18. Relating specific connexin co-expression ratio to connexon composition and gap junction function.

    PubMed

    Desplantez, T; Grikscheit, K; Thomas, N M; Peters, N S; Severs, N J; Dupont, E

    2015-12-01

    Cardiac connexin 43 (Cx43), Cx40 and Cx45 are co-expressed at distinct ratios in myocytes. This pattern is considered a key factor in regulating the gap junction channels composition, properties and function and remains poorly understood. This work aims to correlate gap junction function with the connexin composition of the channels at accurate ratios Cx43:Cx40 and Cx43:Cx45. Rat liver epithelial cells that endogenously express Cx43 were stably transfected to induce expression of accurate levels of Cx40 or Cx45 that may be present in various areas of the heart (e.g. atria and ventricular conduction system). Induction of Cx40 does not increase the amounts of junctional connexins (Cx43 and Cx40), whereas induction of Cx45 increases the amounts of junctional connexins (Cx43 and Cx45). Interestingly, the non-junctional fraction of Cx43 remains unaffected upon induction of Cx40 and Cx45. Co-immunoprecipitation studies show low level of Cx40/Cx43 heteromerisation and undetectable Cx45/Cx43 heteromerisation. Functional characterisation shows that induction of Cx40 and Cx45 decreases Lucifer Yellow transfer. Electrical coupling is decreased by Cx45 induction, whereas it is decreased at low induction of Cx40 and increased at high induction. These data indicate a fine regulation of the gap junction channel make-up in function of the type and the ratio of co-expressed Cxs that specifically regulates chemical and electrical coupling. This reflects specific gap junction function in regulating impulse propagation in the healthy heart, and a pro-arrhythmic potential of connexin remodelling in the diseased heart. PMID:26550940

  19. Connexin26 regulates assembly and maintenance of cochlear gap junction macromolecular complex for normal hearing

    NASA Astrophysics Data System (ADS)

    Kamiya, Kazusaku; Fukunaga, Ichiro; Hatakeyama, Kaori; Ikeda, Katsuhisa

    2015-12-01

    Hereditary deafness affects about 1 in 2000 children and GJB2 gene mutation is most frequent cause for this disease in the world. GJB2 encodes connexin26 (Cx26), a component in cochlear gap junction. Recently, we found macromolecular change of gap junction plaques with two different types of Cx26 mutation as major classification of clinical case, one is a model of dominant negative type, Cx26R75W+ and the other is conditional gene deficient mouse, Cx26f/fP0Cre as a model for insufficiency of gap junction protein [6]. Gap junction composed mainly of Cx26 and Cx30 in wild type mice formed large planar gap junction plaques (GJP). In contrast, Cx26R75W+ and Cx26f/fP0Cre showed fragmented small round GJPs around the cell border. In Cx26f/fP0Cre, some of the cells with Cx26 expression due to their cellular mosaicism showed normal large GJP with Cx26 and Cx30 only at the cell junction site between two Cx26 positive cells. These indicate that bilateral Cx26 expressions from both adjacent cells are essential for the formation of the cochlear linear GJP, and it is not compensated by other cochlear Connexins such as Connexin30. In the present study, we demonstrated a new molecular pathology in most common hereditary deafness with different types of Connexin26 mutations, and this machinery can be a new target for drag design of hereditary deafness.

  20. Low resistance contacts for shallow junction semiconductors

    NASA Technical Reports Server (NTRS)

    Fatemi, Navid S. (Inventor); Weizer, Victor G. (Inventor)

    1994-01-01

    A method of enhancing the specific contact resistivity in InP semiconductor devices and improved devices produced thereby are disclosed. Low resistivity values are obtained by using gold ohmic contacts that contain small amounts of gallium or indium and by depositing a thin gold phosphide interlayer between the surface of the InP device and the ohmic contact. When both the thin interlayer and the gold-gallium or gold-indium contact metallizations are used, ultra low specific contact resistivities are achieved. Thermal stability with good contact resistivity is achieved by depositing a layer of refractory metal over the gold phosphide interlayer.

  1. Unconventional resistive switching behavior in ferroelectric tunnel junctions.

    PubMed

    Mao, H J; Song, C; Xiao, L R; Gao, S; Cui, B; Peng, J J; Li, F; Pan, F

    2015-04-21

    We investigate an unconventional resistive switching (RS) behavior in La0.67Sr0.33MnO3/BaTiO3/metal (LSMO/BTO) ferroelectric tunnel junctions (FTJs), which is dominated by the variation of the barrier potential profile modulated by the migration of oxygen vacancies in the p-LSMO/n-BTO junction. The LSMO/BTO/Co junction exhibits a remarkable self-rectifying effect ascribed to the high-density interface state at the BTO/Co interface, in contrast to the symmetric conductivity when the top metal electrode is inert Pt. The effects of ferroelectric polarization on the RS behavior are also emphasized. Our work builds a bridge between FTJs and resistive random access memory devices. PMID:25789877

  2. Multi-junction, monolithic solar cell using low-band-gap materials lattice matched to GaAs or Ge

    DOEpatents

    Olson, Jerry M.; Kurtz, Sarah R.; Friedman, Daniel J.

    2001-01-01

    A multi-junction, monolithic, photovoltaic solar cell device is provided for converting solar radiation to photocurrent and photovoltage with improved efficiency. The solar cell device comprises a plurality of semiconductor cells, i.e., active p/n junctions, connected in tandem and deposited on a substrate fabricated from GaAs or Ge. To increase efficiency, each semiconductor cell is fabricated from a crystalline material with a lattice constant substantially equivalent to the lattice constant of the substrate material. Additionally, the semiconductor cells are selected with appropriate band gaps to efficiently create photovoltage from a larger portion of the solar spectrum. In this regard, one semiconductor cell in each embodiment of the solar cell device has a band gap between that of Ge and GaAs. To achieve desired band gaps and lattice constants, the semiconductor cells may be fabricated from a number of materials including Ge, GaInP, GaAs, GaInAsP, GaInAsN, GaAsGe, BGaInAs, (GaAs)Ge, CuInSSe, CuAsSSe, and GaInAsNP. To further increase efficiency, the thickness of each semiconductor cell is controlled to match the photocurrent generated in each cell. To facilitate photocurrent flow, a plurality of tunnel junctions of low-resistivity material are included between each adjacent semiconductor cell. The conductivity or direction of photocurrent in the solar cell device may be selected by controlling the specific p-type or n-type characteristics for each active junction.

  3. Connexins: a myriad of functions extending beyond assembly of gap junction channels

    PubMed Central

    Dbouk, Hashem A; Mroue, Rana M; El-Sabban, Marwan E; Talhouk, Rabih S

    2009-01-01

    Connexins constitute a large family of trans-membrane proteins that allow intercellular communication and the transfer of ions and small signaling molecules between cells. Recent studies have revealed complex translational and post-translational mechanisms that regulate connexin synthesis, maturation, membrane transport and degradation that in turn modulate gap junction intercellular communication. With the growing myriad of connexin interacting proteins, including cytoskeletal elements, junctional proteins, and enzymes, gap junctions are now perceived, not only as channels between neighboring cells, but as signaling complexes that regulate cell function and transformation. Connexins have also been shown to form functional hemichannels and have roles altogether independent of channel functions, where they exert their effects on proliferation and other aspects of life and death of the cell through mostly-undefined mechanisms. This review provides an updated overview of current knowledge of connexins and their interacting proteins, and it describes connexin modulation in disease and tumorigenesis. PMID:19284610

  4. Gap junction networks can generate both ripple-like and fast ripple-like oscillations

    PubMed Central

    Simon, Anna; Traub, Roger D.; Vladimirov, Nikita; Jenkins, Alistair; Nicholson, Claire; Whittaker, Roger G.; Schofield, Ian; Clowry, Gavin J.; Cunningham, Mark O.; Whittington, Miles A.

    2014-01-01

    Fast ripples (FRs) are network oscillations, defined variously as having frequencies of > 150 to > 250 Hz, with a controversial mechanism. FRs appear to indicate a propensity of cortical tissue to originate seizures. Here, we demonstrate field oscillations, at up to 400 Hz, in spontaneously epileptic human cortical tissue in vitro, and present a network model that could explain FRs themselves, and their relation to ‘ordinary’ (slower) ripples. We performed network simulations with model pyramidal neurons, having axons electrically coupled. Ripples (< 250 Hz) were favored when conduction of action potentials, axon to axon, was reliable. Whereas ripple population activity was periodic, firing of individual axons varied in relative phase. A switch from ripples to FRs took place when an ectopic spike occurred in a cell coupled to another cell, itself multiply coupled to others. Propagation could then start in one direction only, a condition suitable for re-entry. The resulting oscillations were > 250 Hz, were sustained or interrupted, and had little jitter in the firing of individual axons. The form of model FR was similar to spontaneously occurring FRs in excised human epileptic tissue. In vitro, FRs were suppressed by a gap junction blocker. Our data suggest that a given network can produce ripples, FRs, or both, via gap junctions, and that FRs are favored by clusters of axonal gap junctions. If axonal gap junctions indeed occur in epileptic tissue, and are mediated by connexin 26 (recently shown to mediate coupling between immature neocortical pyramidal cells), then this prediction is testable. PMID:24118191

  5. A unified framework for spiking and gap-junction interactions in distributed neuronal network simulations.

    PubMed

    Hahne, Jan; Helias, Moritz; Kunkel, Susanne; Igarashi, Jun; Bolten, Matthias; Frommer, Andreas; Diesmann, Markus

    2015-01-01

    Contemporary simulators for networks of point and few-compartment model neurons come with a plethora of ready-to-use neuron and synapse models and support complex network topologies. Recent technological advancements have broadened the spectrum of application further to the efficient simulation of brain-scale networks on supercomputers. In distributed network simulations the amount of spike data that accrues per millisecond and process is typically low, such that a common optimization strategy is to communicate spikes at relatively long intervals, where the upper limit is given by the shortest synaptic transmission delay in the network. This approach is well-suited for simulations that employ only chemical synapses but it has so far impeded the incorporation of gap-junction models, which require instantaneous neuronal interactions. Here, we present a numerical algorithm based on a waveform-relaxation technique which allows for network simulations with gap junctions in a way that is compatible with the delayed communication strategy. Using a reference implementation in the NEST simulator, we demonstrate that the algorithm and the required data structures can be smoothly integrated with existing code such that they complement the infrastructure for spiking connections. To show that the unified framework for gap-junction and spiking interactions achieves high performance and delivers high accuracy in the presence of gap junctions, we present benchmarks for workstations, clusters, and supercomputers. Finally, we discuss limitations of the novel technology. PMID:26441628

  6. Impact of Gap Junctional Intercellular Communication on MLO-Y4 Sclerostin and Soluble Factor Expression.

    PubMed

    York, S L; Sethu, P; Saunders, M M

    2016-04-01

    Bone remodeling is a continual process in which old bone is resorbed by osteoclasts and new bone is formed by osteoblasts, providing a mechanism for bones' ability to adapt to changes in its mechanical environment. While the role of osteoblasts and osteoclasts in bone remodeling is well understood, the cellular regulation of bone remodeling is unclear. One theory is that osteocytes, found within bone, play an important role in controlling the bone remodeling response. Osteocytes possess gap junctions, narrow channels that extend between nearby cells and allow communication between cells via the transfer of small molecules and ions. This work investigated the potential role of gap junctional intercellular communication in bone remodeling by exposing osteocyte-like MLO-Y4 cells to mechanical strains and quantifying the expression of soluble factors, including sclerostin, a protein closely associated with bone remodeling. The soluble factors and sclerostin expression were further examined after inhibiting gap junctional intercellular communication to study the impact of the communication. At supraphysiologic strains, the inhibition of gap junctional intercellular communication led to increases in sclerostin expression relative to cells in which communication was present, indicating that the communication may play a significant role in regulating bone remodeling. PMID:26154422

  7. Impaired Astrocytic Gap Junction Coupling and Potassium Buffering in a Mouse Model of Tuberous Sclerosis Complex

    PubMed Central

    Xu, Lin; Zeng, Ling-Hui; Wong, Michael

    2009-01-01

    Abnormalities in astrocytes occur in the brains of patients with Tuberous Sclerosis Complex (TSC) and may contribute to the pathogenesis of neurological dysfunction in this disease. Here, we report that knock-out mice with Tsc1 gene inactivation in glia (Tsc1GFAPCKO mice) exhibit decreased expression of the astrocytic connexin protein, Cx43, and an associated impairment in gap junction coupling between astrocytes. Correspondingly, hippocampal slices from Tsc1GFAPCKO mice have increased extracellular potassium concentration in response to stimulation. This impaired potassium buffering can be attributed to abnormal gap junction coupling, as a gap junction inhibitor elicits an additional increase in potassium concentration in control, but not Tsc1GFAPCKO slices. Furthermore, treatment with a mammalian target of rapamycin inhibitor reverses the deficient Cx43 expression and impaired potassium buffering. These findings suggest that Tsc1 inactivation in astrocytes causes defects in astrocytic gap junction coupling and potassium clearance, which may contribute to epilepsy in Tsc1GFAPCKO mice. PMID:19385061

  8. LIMITATIONS IN THE USE OF MAGNETIC FIELDS TO EXAMINE GAP JUNCTION COMMUNICATION

    EPA Science Inventory

    OBJECTIVE: We have previously shown that gap junction communication (GJC) in mouse primary hepatocytes can be enhanced by treatment with physiological levels of melatonin, and that 45-Hz magnetic fields can eliminate this enhancement in a time-dependent manner. The objective of t...

  9. Increased phosphorylation of Cx36 gap junctions in the AII amacrine cells of RD retina

    PubMed Central

    Ivanova, Elena; Yee, Christopher W.; Sagdullaev, Botir T.

    2015-01-01

    Retinal degeneration (RD) encompasses a family of diseases that lead to photoreceptor death and visual impairment. Visual decline due to photoreceptor cell loss is further compromised by emerging spontaneous hyperactivity in inner retinal cells. This aberrant activity acts as a barrier to signals from the remaining photoreceptors, hindering therapeutic strategies to restore light sensitivity in RD. Gap junctions, particularly those expressed in AII amacrine cells, have been shown to be integral to the generation of aberrant activity. It is unclear whether gap junction expression and coupling are altered in RD. To test this, we evaluated the expression and phosphorylation state of connexin36 (Cx36), the gap junction subunit predominantly expressed in AII amacrine cells, in two mouse models of RD, rd10 (slow degeneration) and rd1 (fast degeneration). Using Ser293-P antibody, which recognizes a phosphorylated form of connexin36, we found that phosphorylation of connexin36 in both slow and fast RD models was significantly greater than in wildtype controls. This elevated phosphorylation may underlie the increased gap junction coupling of AII amacrine cells exhibited by RD retina. PMID:26483638

  10. In vivo analysis of undocked connexin43 gap junction hemichannels in ovarian granulosa cells.

    PubMed

    Tong, Dan; Li, Tony Y; Naus, Kathryn E; Bai, Donglin; Kidder, Gerald M

    2007-11-15

    Connexin43 (Cx43, encoded by Gja1) is required for ovarian follicle development in the mouse. It is strongly expressed in granulosa cells, in which it forms intercellular gap junction channels that couple the cells metabolically. However, recent evidence indicates that undocked gap junction hemichannels can also have physiological roles such as mediating the release of small messenger molecules, including ATP. In this study, the presence of undocked Cx43 hemichannels in granulosa cells was revealed by dye uptake induced either by mechanical stimulation or by the reduction of extracellular divalent cations, both of which are known triggers for hemichannel opening. ATP release was also detected, and could be abolished by connexin-channel blockers. None of these putative hemichannel-mediated activities were detected in Cx43-deficient granulosa cells. Therefore, we hypothesized that hemichannels account for the essential role of Cx43 in folliculogenesis. To test this, a Cx43 mutant lacking the conserved cysteines on the extracellular loops (cys-less Cx43), reported to form hemichannels but not intercellular channels, was retrovirally expressed in Cx43-deficient granulosa cells. The infected cells were then combined with wild-type oocytes to make reaggregated ovaries, which were grafted into host kidneys. Although re-introduction of wild-type Cx43 rescued folliculogenesis, introduction of cys-less Cx43 did not. Therefore, although Cx43 gap junction hemichannels might play a role in ovarian folliculogenesis, their contribution does not supplant the need for intercellular gap junction channels. PMID:17971414

  11. A unified framework for spiking and gap-junction interactions in distributed neuronal network simulations

    PubMed Central

    Hahne, Jan; Helias, Moritz; Kunkel, Susanne; Igarashi, Jun; Bolten, Matthias; Frommer, Andreas; Diesmann, Markus

    2015-01-01

    Contemporary simulators for networks of point and few-compartment model neurons come with a plethora of ready-to-use neuron and synapse models and support complex network topologies. Recent technological advancements have broadened the spectrum of application further to the efficient simulation of brain-scale networks on supercomputers. In distributed network simulations the amount of spike data that accrues per millisecond and process is typically low, such that a common optimization strategy is to communicate spikes at relatively long intervals, where the upper limit is given by the shortest synaptic transmission delay in the network. This approach is well-suited for simulations that employ only chemical synapses but it has so far impeded the incorporation of gap-junction models, which require instantaneous neuronal interactions. Here, we present a numerical algorithm based on a waveform-relaxation technique which allows for network simulations with gap junctions in a way that is compatible with the delayed communication strategy. Using a reference implementation in the NEST simulator, we demonstrate that the algorithm and the required data structures can be smoothly integrated with existing code such that they complement the infrastructure for spiking connections. To show that the unified framework for gap-junction and spiking interactions achieves high performance and delivers high accuracy in the presence of gap junctions, we present benchmarks for workstations, clusters, and supercomputers. Finally, we discuss limitations of the novel technology. PMID:26441628

  12. INFLUENCE OF SODIUM ARSENITE ON GAP JUNCTION COMMUNICATION IN RAT LIVER EPITHELIAL CELLS

    EPA Science Inventory

    Influence of sodium arsenite on gap junction communication in rat-Iiver epitheiial cells.

    Arsenic is known to cause certain types of cancers, hepatitis, cirrhosis and neurological disorders as well as cardiovascular and reproductive effects and skin lesions. The mechanism...

  13. A functional assay for gap junctional examination; electroporation of adherent cells on indium-tin oxide.

    PubMed

    Geletu, Mulu; Guy, Stephanie; Firth, Kevin; Raptis, Leda

    2014-01-01

    In this technique, cells are cultured on a glass slide that is partly coated with indium-tin oxide (ITO), a transparent, electrically conductive material. A variety of molecules, such as peptides or oligonucleotides can be introduced into essentially 100% of the cells in a non-traumatic manner. Here, we describe how it can be used to study intercellular, gap junctional communication. Lucifer yellow penetrates into the cells when an electric pulse, applied to the conductive surface on which they are growing, causes pores to form through the cell membrane. This is electroporation. Cells growing on the nonconductive glass surface immediately adjacent to the electroporated region do not take up Lucifer yellow by electroporation but do acquire the fluorescent dye as it is passed to them via gap junctions that link them to the electroporated cells. The results of the transfer of dye from cell to cell can be observed microscopically under fluorescence illumination. This technique allows for precise quantitation of gap junctional communication. In addition, it can be used for the introduction of peptides or other non-permeant molecules, and the transfer of small electroporated peptides via gap junctions to inhibit the signal in the adjacent, non-electroporated cells is a powerful demonstration of signal inhibition. PMID:25350637

  14. Electrical signal transmission in a bone cell network: the influence of a discrete gap junction

    NASA Technical Reports Server (NTRS)

    Zhang, D.; Weinbaum, S.; Cowin, S. C.

    1998-01-01

    A refined electrical cable model is formulated to investigate the role of a discrete gap junction in the intracellular transmission of electrical signals in an electrically coupled system of osteocytes and osteoblasts in an osteon. The model also examines the influence of the ratio q between the membrane's electrical time constant and the characteristic time of pore fluid pressure, the circular, cylindrical geometry of the osteon, and key simplifying assumptions in our earlier continuous cable model (see Zhang, D., S. C. Cowin, and S. Weinbaum. Electrical signal transmission and gap junction regulation in a bone cell network: A cable model for an osteon. Ann. Biomed. Eng. 25:379-396, 1997). Using this refined model, it is shown that (1) the intracellular potential amplitude at the osteoblastic end of the osteonal cable retains the character of a combination of a low-pass and a high-pass filter as the corner frequency varies in the physiological range; (2) the presence of a discrete gap junction near a resting osteoblast can lead to significant modulation of the intracellular potential and current in the osteoblast for measured values of the gap junction coupling strength; and (3) the circular, cylindrical geometry of the osteon is well simulated by the beam analogy used in Zhang et al.

  15. Oxaliplatin enhances gap junction-mediated coupling in cell cultures of mouse trigeminal ganglia.

    PubMed

    Poulsen, Jeppe Nørgaard; Warwick, Rebekah; Duroux, Meg; Hanani, Menachem; Gazerani, Parisa

    2015-08-01

    Communications between satellite glial cells and neighboring neurons within sensory ganglia may contribute to neuropathic and inflammatory pain. To elucidate the role of satellite glial cells in chemotherapy-induced pain, we examined the effects of oxaliplatin on the gap junction-mediated coupling between these cells. We also examined whether the gap junction blocker, carbenoxolone, can reverse the coupling. Primary cultures of mice trigeminal ganglia, 24-48h after cell isolation, were used. Satellite glial cells were injected with Lucifer yellow in the presence or absence of oxaliplatin (60 μM). In addition, the effect of carbenoxolone (100 μM) on coupling, and the expression of connexin 43 proteins were evaluated. Dye coupling between adjacent satellite glial cells was significantly increased (2.3-fold, P<0.05) following a 2h incubation with oxaliplatin. Adding carbenoxolone to the oxaliplatin-treated cultures reversed oxaliplatin-evoked coupling to baseline (P<0.05). Immunostaining showed no difference between expression of connexin 43 in control and oxaliplatin-treated cultures. Our findings indicated that oxaliplatin-increased gap junction-mediated coupling between satellite glial cells in primary cultures of mouse trigeminal ganglia, and carbenoxolone reversed this effect. Hence, it is proposed that increased gap junction-mediated coupling was seen between satellite glial cells in TG. This observation together with our previous data obtained from a behavioral study suggests that this phenomenon might contribute to chemotherapy-induced nociception following oxaliplatin treatment. PMID:25999145

  16. FREQUENCY-DEPENDENT CHANGES IN GAP JUNCTION FUNCTION IN PRIMARY HEPATOCYTES

    EPA Science Inventory

    FREQUENCY-DEPENDENT CHANGES IN GAP JUNCTION FUNCTION IN PRIMARY HEPATOCYTES. X. Wang1 *, D.E. Housel *, J. Page2, C.F. Blackmanl. 1 National Health and Environmental Effects Research Laboratory, USEPA, Research Triangle Park, North Carolina 27711 USA, 2Oakland, California USA
    ...

  17. INTEGRIN-MEDIATED CELL ATTACHMENT SHOWS TIME-DEPENDENT UPREGULATION OF GAP JUNCTION COMMUNICATION.

    EPA Science Inventory


    Integrin-mediated Cell Attachment Shows Time-Dependent Upregulation of Gap Junction
    Communication

    Rachel Grindstaff and Carl Blackman, National Health & Environmental Effects Research
    Laboratory, Office of Research and Development, US EPA, Research Triang...

  18. Gap-junctional coupling of mammalian rod photoreceptors and its effect on visual detection

    PubMed Central

    Li, Peter H.; Verweij, Jan; Long, James H.; Schnapf, Julie L.

    2012-01-01

    The presence of gap junctions between rods in mammalian retina suggests a role for rod-rod coupling in human vision. Rod coupling is known to reduce response variability, but because junctional conductances are not known, the downstream effects on visual performance are uncertain. Here we assessed rod coupling in guinea pig retina by measuring: 1) the variability in responses to dim flashes, 2) Neurobiotin tracer coupling, and 3) junctional conductances. Results were consolidated into an electrical network model and a model of human psychophysical detection. Guinea pig rods form tracer pools of 1 to ~20 rods, with junctional conductances averaging ~350 pS. We calculate that coupling will reduce human dark-adapted sensitivity ~10% by impairing the noise filtering of the synapse between rods and rod bipolar cells. However, coupling also mitigates synaptic saturation and is thus calculated to improve sensitivity when stimuli are spatially restricted or are superimposed over background illumination. PMID:22399777

  19. Biophysical properties of gap junctions between freshly dispersed pairs of mouse pancreatic beta cells.

    PubMed Central

    Pérez-Armendariz, M; Roy, C; Spray, D C; Bennett, M V

    1991-01-01

    Coupling between beta cells through gap junctions has been postulated as a principal mechanism of electrical synchronization of glucose-induced activity throughout the islet of Langerhans. We characterized junctional conductance between isolated pairs of mouse pancreatic beta cells by whole-cell recording with two independent patch-clamp circuits. Most pairs were coupled (67%, n = 155), although the mean junctional conductance (gj) (215 +/- 110 pS) was lower than reported in other tissues. Coupling could be recorded for long periods, up to 40 min. Voltage imposed across the junctional or nonjunctional membranes had no effect on gj. Up to several hours of treatment to increase intracellular cAMP levels did not affect gj. Electrically coupled pairs did not show transfer of the dye Lucifer yellow. Octanol (2 mM) reversibly decreased gj. Lower concentrations of octanol (0.5 mM) and heptanol (0.5 mM) than required to uncouple beta cells decreased voltage-dependent K+ and Ca2+ currents in nonjunctional membranes. Although gj recorded in these experiments would be expected to be provided by current flowing through only a few channels of the unitary conductance previously reported for other gap junctions, no unitary junctional currents were observed even during reversible suppression of gj by octanol. This result suggests either that the single channel conductance of gap junction channels between beta cells is smaller than in other tissues (less than 20 pS) or that the small mean conductance is due to transitions between open and closed states that are too rapid or too slow to be resolved. Images FIGURE 1 FIGURE 5 PMID:2015391

  20. A methodology for experimentally based determination of gap shrinkage and effective lifetimes in the emitter and base of p-n junction solar cells and other p-n junction devices

    NASA Technical Reports Server (NTRS)

    Lindholm, F. A.; Neugroschel, A.; Sah, C.-T.; Godlewski, M. P.; Brandhorst, H. W., Jr.

    1977-01-01

    An experimentally based methodology that determines the effective gap shrinkage and lifetime in the emitter of a p-n junction solar cell is described which provides an experimental means for assessing the importance of gap shrinkage relative to that of large recombination rates in the highly doped emitter. The base lifetime is also determined. The methodology pertains to a solar cell after the junction is formed, so that each material parameter determined includes the effects of the processing used in junction fabrication. The methodology consists of strategy and procedures for designing experiments and interpreting data consistently with the physical mechanisms governing device behavior. This careful linking to the device physics uncover the material parameters concealed in the data. To illustrate the procedures, they are applied to an n(+)-p solar cell having substrate resistivity of about 0.1 ohm-cm.

  1. Regulation of gap junction channels and hemichannels by phosphorylation and redox changes: a revision.

    PubMed

    Pogoda, Kristin; Kameritsch, Petra; Retamal, Mauricio A; Vega, José L

    2016-01-01

    Post-translational modifications of connexins play an important role in the regulation of gap junction and hemichannel permeability. The prerequisite for the formation of functional gap junction channels is the assembly of connexin proteins into hemichannels and their insertion into the membrane. Hemichannels can affect cellular processes by enabling the passage of signaling molecules between the intracellular and extracellular space. For the intercellular communication hemichannels from one cell have to dock to its counterparts on the opposing membrane of an adjacent cell to allow the transmission of signals via gap junctions from one cell to the other. The controlled opening of hemichannels and gating properties of complete gap junctions can be regulated via post-translational modifications of connexins. Not only channel gating, but also connexin trafficking and assembly into hemichannels can be affected by post-translational changes. Recent investigations have shown that connexins can be modified by phosphorylation/dephosphorylation, redox-related changes including effects of nitric oxide (NO), hydrogen sulfide (H2S) or carbon monoxide (CO), acetylation, methylation or ubiquitination. Most of the connexin isoforms are known to be phosphorylated, e.g. Cx43, one of the most studied connexin at all, has 21 reported phosphorylation sites. In this review, we provide an overview about the current knowledge and relevant research of responsible kinases, connexin phosphorylation sites and reported effects on gap junction and hemichannel regulation. Regarding the effects of oxidants we discuss the role of NO in different cell types and tissues and recent studies about modifications of connexins by CO and H2S. PMID:27229925

  2. Regulation of gap junctional intercellular communication by TCDD in HMEC and MCF-7 breast cancer cells

    SciTech Connect

    Gakhar, Gunjan Schrempp, Diane Nguyen, Thu Annelise

    2009-03-01

    Previous studies suggest that many neoplastic tissues exhibit a decrease in gap junctional intercellular communication (GJIC). Many hydrocarbons and organochlorine compounds are environmental pollutants known to be carcinogenic. The effect of an organochlorine compound, TCDD, on GJIC in human breast cell lines has not been established. In the present study, we showed that TCDD causes an inhibition in the gap junctional activity in MCF-7 (breast cancer cells). In MCF-7 cells, an increase in the phosphorylated form of gap junctional protein, connexin 43 (Cx43), and PKC {alpha} was seen in the presence of TCDD. Gap junctional plaque formation was significantly decreased in MCF-7 cells in the presence of TCDD. Immunoprecipitation studies of PKC {alpha} showed that TCDD caused a significant 40% increase in the phosphorylated Cx43 in MCF-7 cells. TCDD also modulated the translocation of PKC {alpha} from the cytosol to the membrane and caused a 2-fold increase in the PKC {alpha} activity at 50 nM TCDD in MCF-7 cells. Calphostin C, an inhibitor of PKC {alpha}, showed a significant inhibition of PKC {alpha} activity in the presence of TCDD. Furthermore, TCDD also caused a decrease in the gap junctional activity and Cx43 protein in human mammary epithelial cells (HMEC). However, we observed a shift in the Cx43 plaques towards the perinuclear membrane in the presence of TCDD by confocal microscopy and Western blot. Overall, these results conclude that TCDD decreases GJIC by phosphorylating Cx43 via PKC {alpha} signaling pathway in MCF-7 cells; however, TCDD decreases the GJIC by affecting the localization of Cx43 in HMEC. These new findings elucidate the differential mode of effect of TCDD in the downregulation of GJIC in HMEC and MCF-7 cells.

  3. High temperature coefficient of resistance for a ferroelectric tunnel junction

    NASA Astrophysics Data System (ADS)

    Zhao, Xiaolin; Wang, Jianlu; Tian, Bobo; Liu, Bolu; Wang, Xudong; Huang, Hai; Zou, Yuhong; Sun, Shuo; Lin, Tie; Han, Li; Sun, Jinglan; Meng, Xiangjian; Chu, Junhao

    2015-08-01

    An infrared detector is proposed that is based on a ferroelectric tunnel junction (FTJ) working under bolometer-like principles. Electron tunneling, either direct or indirect, through the ferroelectric barrier depends on the temperature of the devices. During tunneling, infrared radiation alters the polarization of the ferroelectric film via pyroelectricity, resulting in a change in the barrier height of the tunnel junction. A high temperature coefficient of resistance of up to -3.86% was observed at room temperature. These results show that the FTJ structure has potential to be adapted for use in uncooled infrared detectors.

  4. High temperature coefficient of resistance for a ferroelectric tunnel junction

    SciTech Connect

    Zhao, Xiaolin; Tian, Bobo; Liu, Bolu; Wang, Xudong; Huang, Hai; Wang, Jianlu E-mail: xjmeng@mail.sitp.ac.cn; Zou, Yuhong; Sun, Shuo; Lin, Tie; Han, Li; Sun, Jinglan; Meng, Xiangjian E-mail: xjmeng@mail.sitp.ac.cn; Chu, Junhao

    2015-08-10

    An infrared detector is proposed that is based on a ferroelectric tunnel junction (FTJ) working under bolometer-like principles. Electron tunneling, either direct or indirect, through the ferroelectric barrier depends on the temperature of the devices. During tunneling, infrared radiation alters the polarization of the ferroelectric film via pyroelectricity, resulting in a change in the barrier height of the tunnel junction. A high temperature coefficient of resistance of up to −3.86% was observed at room temperature. These results show that the FTJ structure has potential to be adapted for use in uncooled infrared detectors.

  5. Experimental Study of Resistive Bistability in Metal Oxide Junctions

    SciTech Connect

    Su, D.; Tan, Z.; Patel, V.; Likharev, K.K.; Zhu, Y.

    2011-05-01

    We have studied resistive bistability (memory) effects in junctions based on metal oxides, with a focus on sample-to-sample reproducibility, which is necessary for the use of such junctions as crosspoint devices of hybrid CMOS/nanoelectronic circuits. Few-nm-thick layers of NbO{sub x}, CuO{sub x} and TiO{sub x} have been formed by thermal and plasma oxidation, at various deposition and oxidation conditions, both with and without rapid thermal post-annealing. The resistive bistability effect has been observed for all these materials, with particularly high endurance (over 10{sup 3} switching cycles) obtained for single-layer TiO{sub 2} junctions, and the best reproducibility reached for multi-layer junctions of the same material. Fabrication optimization has allowed us to improve the OFF/ON resistance ratio to about 10{sup 3}, but the sample-to-sample reproducibility is so far lower than that required for large-scale integration.

  6. Baicalein increases the cytotoxicity of cisplatin by enhancing gap junction intercellular communication.

    PubMed

    Wang, Yiwen; Wang, Qin; Zhang, Suzhi; Zhang, Yuan; Tao, Liang

    2014-07-01

    Drug resistance limits the clinical application of cisplatin, a widely used chemotherapeutic agent. Gap junction (GJ) is a channel that enhances cytotoxicity of certain chemotherapeutic agents. Baicalein is well known for its antitumor activity. This study investigated the effect of baicalein on cisplatin cytotoxicity and the relationship between this effect and the modulation of the GJ function in connexin 26 (Cx26)‑transfected HeLa cells. The sulforhodamine B (SRB) assay was used to examine the effect of baicalein on cell viability. A 'parachute' assay was used to investigate the effect of baicalein on GJ function. The effects of baicalein on cisplatin cytotoxicity and GJ function were assayed by standard colony‑forming assays. The expression of Cx26 was monitored by western blotting. It was observed that exposure of Cx26‑transfected cells to cisplatin reduced the number of colonies formed in low‑density cultures (no GJ formation) and in high‑density cultures (GJ formation), but the toxic effect was greater when cells were seeded at a high density. In the absence of connexin expression or with blockage of connexin channels however, cell density had no effect on cisplatin toxicity. Baicalein significantly enhanced cisplatin cytotoxicity, but this effect required the presence of functional GJs between the cells. In conclusion, the dependence of cisplatin toxicity on cell density is mediated by GJs. Baicalein increases cisplatin cytotoxicity through enhancing GJ intercellular communication. PMID:24736991

  7. High band gap 2-6 and 3-5 tunneling junctions for silicon multijunction solar cells

    NASA Technical Reports Server (NTRS)

    Daud, Taher (Inventor); Kachare, Akaram H. (Inventor)

    1986-01-01

    A multijunction silicon solar cell of high efficiency is provided by providing a tunnel junction between the solar cell junctions to connect them in series. The tunnel junction is comprised of p+ and n+ layers of high band gap 3-5 or 2-6 semiconductor materials that match the lattice structure of silicon, such as GaP (band gap 2.24 eV) or ZnS (band gap 3.6 eV). Each of which has a perfect lattice match with silicon to avoid defects normally associated with lattice mismatch.

  8. A fluorescence photobleaching assay of gap junction-mediated communication between human cells.

    PubMed

    Wade, M H; Trosko, J E; Schindler, M

    1986-04-25

    Gap junction-mediated communication between contiguous cells has been implicated in the regulation of cell proliferation and differentiation. This report describes a new technique to measure cell-cell communication, gap fluorescence redistribution after photobleaching, which is based on the diffusion-dependent return of 6-carboxyfluorescein-mediated fluorescence in a photobleached cell that is in contact with other fluorescently labeled cells. Fluorescence recovery rates are interpreted as dye transport across gap junctions. Results of experiments on normal human fibroblasts and human teratocarcinoma cells show that this technique can measure rapid dye transfer and detect inhibition of communication (between teratocarcinoma cells) by the tumor promoters 12-O-tetradecanoyl-phorbol-13-acetate and the pesticide dieldrin. PMID:3961495

  9. An electrostatic mechanism for Ca(2+)-mediated regulation of gap junction channels.

    PubMed

    Bennett, Brad C; Purdy, Michael D; Baker, Kent A; Acharya, Chayan; McIntire, William E; Stevens, Raymond C; Zhang, Qinghai; Harris, Andrew L; Abagyan, Ruben; Yeager, Mark

    2016-01-01

    Gap junction channels mediate intercellular signalling that is crucial in tissue development, homeostasis and pathologic states such as cardiac arrhythmias, cancer and trauma. To explore the mechanism by which Ca(2+) blocks intercellular communication during tissue injury, we determined the X-ray crystal structures of the human Cx26 gap junction channel with and without bound Ca(2+). The two structures were nearly identical, ruling out both a large-scale structural change and a local steric constriction of the pore. Ca(2+) coordination sites reside at the interfaces between adjacent subunits, near the entrance to the extracellular gap, where local, side chain conformational rearrangements enable Ca(2+)chelation. Computational analysis revealed that Ca(2+)-binding generates a positive electrostatic barrier that substantially inhibits permeation of cations such as K(+) into the pore. Our results provide structural evidence for a unique mechanism of channel regulation: ionic conduction block via an electrostatic barrier rather than steric occlusion of the channel pore. PMID:26753910

  10. An electrostatic mechanism for Ca2+-mediated regulation of gap junction channels

    PubMed Central

    Bennett, Brad C.; Purdy, Michael D.; Baker, Kent A.; Acharya, Chayan; McIntire, William E.; Stevens, Raymond C.; Zhang, Qinghai; Harris, Andrew L.; Abagyan, Ruben; Yeager, Mark

    2016-01-01

    Gap junction channels mediate intercellular signalling that is crucial in tissue development, homeostasis and pathologic states such as cardiac arrhythmias, cancer and trauma. To explore the mechanism by which Ca2+ blocks intercellular communication during tissue injury, we determined the X-ray crystal structures of the human Cx26 gap junction channel with and without bound Ca2+. The two structures were nearly identical, ruling out both a large-scale structural change and a local steric constriction of the pore. Ca2+ coordination sites reside at the interfaces between adjacent subunits, near the entrance to the extracellular gap, where local, side chain conformational rearrangements enable Ca2+chelation. Computational analysis revealed that Ca2+-binding generates a positive electrostatic barrier that substantially inhibits permeation of cations such as K+ into the pore. Our results provide structural evidence for a unique mechanism of channel regulation: ionic conduction block via an electrostatic barrier rather than steric occlusion of the channel pore. PMID:26753910

  11. TLR-Activated Gap Junction Channels Protect Mice against Bacterial Infection through Extracellular UDP Release.

    PubMed

    Qin, Juliang; Zhang, Guangxu; Zhang, Xiaoyu; Tan, Binghe; Lv, Zhangsheng; Liu, Mingyao; Ren, Hua; Qian, Min; Du, Bing

    2016-02-15

    Extracellular UDP (eUDP), released as a danger signal by stressed or apoptotic cells, plays an important role in a series of physiological processes. Although the mechanism of eUDP release in apoptotic cells has been well defined, how the eUDP is released in innate immune responses remains unknown. In this study, we demonstrated that UDP was released in both Escherichia coli-infected mice and LPS- or Pam3CSK4-treated macrophages. Also, LPS-induced UDP release could be significantly blocked by selective TLR4 inhibitor Atractylenolide I and selective gap junction inhibitors carbenoxolone and flufenamic acid (FFA), suggesting the key role of TLR signaling and gap junction channels in this process. Meanwhile, eUDP protected mice from peritonitis by reducing invaded bacteria that could be rescued by MRS2578 (selective P2Y6 receptor inhibitor) and FFA. Then, connexin 43, as one of the gap junction proteins, was found to be clearly increased by LPS in a dose- and time-dependent manner. Furthermore, if we blocked LPS-induced ERK signaling by U0126, the expression of connexin 43 and UDP release was also inhibited dramatically. In addition, UDP-induced MCP-1 secretion was significantly reduced by MRS2578, FFA, and P2Y6 mutation. Accordingly, pretreating mice with U0126 and Gap26 increased invaded bacteria and aggravated mice death. Taken together, our study reveals an internal relationship between danger signals and TLR signaling in innate immune responses, which suggests a potential therapeutic significance of gap junction channel-mediated UDP release in infectious diseases. PMID:26783339

  12. Oligodendrocytes in mouse corpus callosum are coupled via gap junction channels formed by connexin47 and connexin32.

    PubMed

    Maglione, Marta; Tress, Oliver; Haas, Brigitte; Karram, Khalad; Trotter, Jacqueline; Willecke, Klaus; Kettenmann, Helmut

    2010-07-01

    According to previously published ultrastructural studies, oligodendrocytes in white matter exhibit gap junctions with astrocytes, but not among each other, while in vitro oligodendrocytes form functional gap junctions. We have studied functional coupling among oligodendrocytes in acute slices of postnatal mouse corpus callosum. By whole-cell patch clamp we dialyzed oligodendrocytes with biocytin, a gap junction-permeable tracer. On average 61 cells were positive for biocytin detected by labeling with streptavidin-Cy3. About 77% of the coupled cells stained positively for the oligodendrocyte marker protein CNPase, 9% for the astrocyte marker GFAP and 14% were negative for both CNPase and GFAP. In the latter population, the majority expressed Olig2 and some NG2, markers for oligodendrocyte precursors. Oligodendrocytes are known to express Cx47, Cx32 and Cx29, astrocytes Cx43 and Cx30. In Cx47-deficient mice, the number of coupled cells was reduced by 80%. Deletion of Cx32 or Cx29 alone did not significantly reduce the number of coupled cells, but coupling was absent in Cx32/Cx47-double-deficient mice. Cx47-ablation completely abolished coupling of oligodendrocytes to astrocytes. In Cx43-deficient animals, oligodendrocyte-astrocyte coupling was still present, but coupling to oligodendrocyte precursors was not observed. In Cx43/Cx30-double deficient mice, oligodendrocyte-to-astrocyte coupling was almost absent. Uncoupled oligodendrocytes showed a higher input resistance. We conclude that oligodendrocytes in white matter form a functional syncytium predominantly among each other dependent on Cx47 and Cx32 expression, while astrocytic connexins expression can promote the size of this network. PMID:20468052

  13. Protein Kinase Cδ-mediated Phosphorylation of Connexin43 Gap Junction Channels Causes Movement within Gap Junctions followed by Vesicle Internalization and Protein Degradation*

    PubMed Central

    Cone, Angela C.; Cavin, Gabriel; Ambrosi, Cinzia; Hakozaki, Hiroyuki; Wu-Zhang, Alyssa X.; Kunkel, Maya T.; Newton, Alexandra C.; Sosinsky, Gina E.

    2014-01-01

    Phosphorylation of gap junction proteins, connexins, plays a role in global signaling events involving kinases. Connexin43 (Cx43), a ubiquitous and important connexin, has several phosphorylation sites for specific kinases. We appended an imaging reporter tag for the activity of the δ isoform of protein kinase C (PKCδ) to the carboxyl terminus of Cx43. The FRET signal of this reporter is inversely related to the phosphorylation of serine 368 of Cx43. By activating PKC with the phorbol ester phorbol 12,13-dibutyrate (PDBu) or a natural stimulant, UTP, time lapse live cell imaging movies indicated phosphorylated Ser-368 Cx43 separated into discrete domains within gap junctions and was internalized in small vesicles, after which it was degraded by lysosomes and proteasomes. Mutation of Ser-368 to an Ala eliminated the response to PDBu and changes in phosphorylation of the reporter. A phosphatase inhibitor, calyculin A, does not change this pattern, indicating PKC phosphorylation causes degradation of Cx43 without dephosphorylation, which is in accordance with current hypotheses that cells control their intercellular communication by a fast and constant turnover of connexins, using phosphorylation as part of this mechanism. PMID:24500718

  14. Gap Junctions Contribute to the Regulation of Walking-Like Activity in the Adult Mudpuppy (Necturus Maculatus)

    PubMed Central

    Lavrov, Igor; Fox, Lyle; Shen, Jun; Han, Yingchun; Cheng, Jianguo

    2016-01-01

    Although gap junctions are widely expressed in the developing central nervous system, the role of electrical coupling of neurons and glial cells via gap junctions in the spinal cord in adults is largely unknown. We investigated whether gap junctions are expressed in the mature spinal cord of the mudpuppy and tested the effects of applying gap junction blocker on the walking-like activity induced by NMDA or glutamate in an in vitro mudpuppy preparation. We found that glial and neural cells in the mudpuppy spinal cord expressed different types of connexins that include connexin 32 (Cx32), connexin 36 (Cx36), connexin 37 (Cx37), and connexin 43 (Cx43). Application of a battery of gap junction blockers from three different structural classes (carbenexolone, flufenamic acid, and long chain alcohols) substantially and consistently altered the locomotor-like activity in a dose-dependent manner. In contrast, these blockers did not significantly change the amplitude of the dorsal root reflex, indicating that gap junction blockers did not inhibit neuronal excitability nonselectively in the spinal cord. Taken together, these results suggest that gap junctions play a significant modulatory role in the spinal neural networks responsible for the generation of walking-like activity in the adult mudpuppy. PMID:27023006

  15. Gap Junctions Contribute to the Regulation of Walking-Like Activity in the Adult Mudpuppy (Necturus Maculatus).

    PubMed

    Lavrov, Igor; Fox, Lyle; Shen, Jun; Han, Yingchun; Cheng, Jianguo

    2016-01-01

    Although gap junctions are widely expressed in the developing central nervous system, the role of electrical coupling of neurons and glial cells via gap junctions in the spinal cord in adults is largely unknown. We investigated whether gap junctions are expressed in the mature spinal cord of the mudpuppy and tested the effects of applying gap junction blocker on the walking-like activity induced by NMDA or glutamate in an in vitro mudpuppy preparation. We found that glial and neural cells in the mudpuppy spinal cord expressed different types of connexins that include connexin 32 (Cx32), connexin 36 (Cx36), connexin 37 (Cx37), and connexin 43 (Cx43). Application of a battery of gap junction blockers from three different structural classes (carbenexolone, flufenamic acid, and long chain alcohols) substantially and consistently altered the locomotor-like activity in a dose-dependent manner. In contrast, these blockers did not significantly change the amplitude of the dorsal root reflex, indicating that gap junction blockers did not inhibit neuronal excitability nonselectively in the spinal cord. Taken together, these results suggest that gap junctions play a significant modulatory role in the spinal neural networks responsible for the generation of walking-like activity in the adult mudpuppy. PMID:27023006

  16. A versatile optical junction using photonic band-gap guidance and self collimation

    SciTech Connect

    Gupta, Man Mohan; Medhekar, Sarang

    2014-09-29

    We show that it is possible to design two photonic crystal (PC) structures such that an optical beam of desired wavelength gets guided within the line defect of the first structure (photonic band gap guidance) and the same beam gets guided in the second structure by self-collimation. Using two dimensional simulation of a design made of the combination of these two structures, we propose an optical junction that allows for crossing of two optical signals of same wavelength and same polarization with very low crosstalk. Moreover, the junction can be operated at number of frequencies in a wide range. Crossing of multiple beams with very low cross talk is also possible. The proposed junction should be important in future integrated photonic circuits.

  17. Neural progenitor cells isolated from the subventricular zone present hemichannel activity and form functional gap junctions with glial cells

    PubMed Central

    Talaverón, Rocío; Fernández, Paola; Escamilla, Rosalba; Pastor, Angel M.; Matarredona, Esperanza R.; Sáez, Juan C.

    2015-01-01

    The postnatal subventricular zone (SVZ) lining the walls of the lateral ventricles contains neural progenitor cells (NPCs) that generate new olfactory bulb interneurons. Communication via gap junctions between cells in the SVZ is involved in NPC proliferation and in neuroblast migration towards the olfactory bulb. SVZ NPCs can be expanded in vitro in the form of neurospheres that can be used for transplantation purposes after brain injury. We have previously reported that neurosphere-derived NPCs form heterocellular gap junctions with host glial cells when they are implanted after mechanical injury. To analyze functionality of NPC-glial cell gap junctions we performed dye coupling experiments in co-cultures of SVZ NPCs with astrocytes or microglia. Neurosphere-derived cells expressed mRNA for at least the hemichannel/gap junction channel proteins connexin 26 (Cx26), Cx43, Cx45 and pannexin 1 (Panx1). Dye coupling experiments revealed that gap junctional communication occurred among neurosphere cells (incidence of coupling: 100%). Moreover, hemichannel activity was also detected in neurosphere cells as evaluated in time-lapse measurements of ethidium bromide uptake. Heterocellular coupling between NPCs and glial cells was evidenced in co-cultures of neurospheres with astrocytes (incidence of coupling: 91.0 ± 4.7%) or with microglia (incidence of coupling: 71.9 ± 6.7%). Dye coupling in neurospheres and in co-cultures was inhibited by octanol, a gap junction blocker. Altogether, these results suggest the existence of functional hemichannels and gap junction channels in postnatal SVZ neurospheres. In addition, they demonstrate that SVZ-derived NPCs can establish functional gap junctions with astrocytes or microglia. Therefore, cell-cell communication via gap junctions and hemichannels with host glial cells might subserve a role in the functional integration of NPCs after implantation in the damaged brain. PMID:26528139

  18. Ionic blockade of the rat connexin40 gap junction channel by large tetraalkylammonium ions.

    PubMed

    Musa, H; Gough, J D; Lees, W J; Veenstra, R D

    2001-12-01

    The rat connexin40 gap junction channel is permeable to monovalent cations including tetramethylammonium and tetraethylammonium ions. Larger tetraalkyammonium (TAA(+)) ions beginning with tetrabutylammonium (TBA(+)) reduced KCl junctional currents disproportionately. Ionic blockade by tetrapentylammonium (TPeA(+)) and tetrahexylammonium (THxA(+)) ions were concentration- and voltage-dependent and occurred only when TAA(+) ions were on the same side as net K(+) efflux across the junction, indicative of block of the ionic permeation pathway. The voltage-dependent dissociation constants (K(m)(V(j))) were lower for THxA(+) than TPeA(+), consistent with steric effects within the pore. The K(m)-V(j) relationships for TPeA(+) and THxA(+) were fit with different reaction rate models for a symmetrical (homotypic) connexin gap junction channel and were described by either a one- or two-site model that assumed each ion traversed the entire V(j) field. Bilateral addition of TPeA(+) ions confirmed a common site of interaction within the pore that possessed identical K(m)(V(j)) values for cis-trans concentrations of TPeA(+) ions as indicated by the modeled I-V relations and rapid channel block that precluded unitary current measurements. The TAA(+) block of K(+) currents and bilateral TPeA(+) interactions did not alter V(j)-gating of Cx40 gap junctions. N-octyl-tributylammonium and -triethylammonium also blocked rCx40 channels with higher affinity and faster kinetics than TBA(+) or TPeA(+), indicative of a hydrophobic site within the pore near the site of block. PMID:11720990

  19. Regulation of gap junction channels by infectious agents and inflammation in the CNS

    PubMed Central

    Castellano, Paul; Eugenin, Eliseo A.

    2014-01-01

    Gap junctions (GJs) are conglomerates of intercellular channels that connect the cytoplasm of two or more cells, and facilitate the transfer of ions and small molecules, including second messengers, resulting in metabolic and electrical coordination. In general, loss of gap junctional communication (GJC) has been associated with cellular damage and inflammation resulting in compromise of physiological functions. Recently, it has become evident that GJ channels also play a critical role in the pathogenesis of infectious diseases and associated inflammation. Several pathogens use the transfer of intracellular signals through GJ channels to spread infection and toxic signals that amplify inflammation to neighboring cells. Thus, identification of the mechanisms by which several infectious agents alter GJC could result in new potential therapeutic approaches to reduce inflammation and their pathogenesis. PMID:24847208

  20. Tonabersat, a novel gap-junction modulator for the prevention of migraine.

    PubMed

    Silberstein, S D

    2009-11-01

    Migraine is a common, recurrent, primary headache disorder associated with significant morbidity as well as high direct and indirect costs. Despite its impact, only a proportion of migraineurs who meet criteria for prophylactic treatment take preventive medication. Antiepileptic drugs and beta-blockers are among the most used preventive therapies, but their exact mechanisms of action in migraine prophylaxis are unknown. Recent research has pointed to the role of cortical spreading depression in the genesis of migraine aura and pain, with neuronal-glial gap junctions playing a prominent part in cortical spreading depression. Tonabersat is a unique compound with demonstrated activity as a gap-junction inhibitor in animal studies. In preclinical and clinical trials, tonabersat was well tolerated, with no cardiovascular effects; the pharmacokinetic profile suggested its usefulness in the prophylaxis of migraine. PMID:19723123

  1. Effect of sound on gap-junction-based intercellular signaling: Calcium waves under acoustic irradiation

    NASA Astrophysics Data System (ADS)

    Deymier, P. A.; Swinteck, N.; Runge, K.; Deymier-Black, A.; Hoying, J. B.

    2015-11-01

    We present a previously unrecognized effect of sound waves on gap-junction-based intercellular signaling such as in biological tissues composed of endothelial cells. We suggest that sound irradiation may, through temporal and spatial modulation of cell-to-cell conductance, create intercellular calcium waves with unidirectional signal propagation associated with nonconventional topologies. Nonreciprocity in calcium wave propagation induced by sound wave irradiation is demonstrated in the case of a linear and a nonlinear reaction-diffusion model. This demonstration should be applicable to other types of gap-junction-based intercellular signals, and it is thought that it should be of help in interpreting a broad range of biological phenomena associated with the beneficial therapeutic effects of sound irradiation and possibly the harmful effects of sound waves on health.

  2. Spontaneous calcium signals induced by gap junctions in a network model of astrocytes

    NASA Astrophysics Data System (ADS)

    Kazantsev, V. B.

    2009-01-01

    The dynamics of a network model of astrocytes coupled by gap junctions is investigated. Calcium dynamics of the single cell is described by the biophysical model comprising the set of three nonlinear differential equations. Intercellular dynamics is provided by the diffusion of inositol 1,4,5-trisphosphate (IP3) through gap junctions between neighboring astrocytes. It is found that the diffusion induces the appearance of spontaneous activity patterns in the network. Stability of the network steady state is analyzed. It is proved that the increase of the diffusion coefficient above a certain critical value yields the generation of low-amplitude subthreshold oscillatory signals in a certain frequency range. It is shown that such spontaneous oscillations can facilitate calcium pulse generation and provide a certain time scale in astrocyte signaling.

  3. Closing the proximity gap in a metallic Josephson junction between three superconductors

    NASA Astrophysics Data System (ADS)

    Padurariu, C.; Jonckheere, T.; Rech, J.; Mélin, R.; Feinberg, D.; Martin, T.; Nazarov, Yu. V.

    2015-11-01

    We describe the proximity effect in a short disordered metallic junction between three superconducting leads. Andreev bound states in the multiterminal junction may cross the Fermi level. We reveal that for a quasicontinuous metallic density of states, crossings at the Fermi level manifest as a closing of the proximity-induced gap. We calculate the local density of states for a wide range of transport parameters using quantum circuit theory. The gap closes inside an area of the space spanned by the superconducting phase differences. We derive an approximate analytic expression for the boundary of the area and compare it to the full numerical solution. The size of the area increases with the transparency of the junction and is sensitive to asymmetry. The finite density of states at zero energy is unaffected by the electron-hole decoherence present in the junction, although decoherence is important at higher energies. Our predictions can be tested using tunneling transport spectroscopy. To encourage experiments, we calculate the current-voltage characteristic in a typical measurement setup. We show how the structure of the local density of states can be mapped out from the measurement.

  4. Antofine-induced connexin43 gap junction disassembly in rat astrocytes involves protein kinase Cβ.

    PubMed

    Huang, Yu-Fang; Liao, Chih-Kai; Lin, Jau-Chen; Jow, Guey-Mei; Wang, Hwai-Shi; Wu, Jiahn-Chun

    2013-03-01

    Antofine, a phenanthroindolizidine alkaloid derived from Cryptocaryachinensis and Ficusseptica in the Asclepiadaceae milkweed family, is cytotoxic for various cancer cell lines. In this study, we demonstrated that treatment of rat primary astrocytes with antofine induced dose-dependent inhibition of gap junction intercellular communication (GJIC), as assessed by scrape-loading 6-carboxyfluorescein dye transfer. Levels of Cx43 protein were also decreased in a dose- and time-dependent manner following antofine treatment. Double-labeling immunofluorescence microscopy showed that antofine (10ng/ml) induced endocytosis of surface gap junctions into the cytoplasm, where Cx43 was co-localized with the early endosome marker EEA1. Inhibition of lysosomes or proteasomes by co-treatment with antofine and their respective specific inhibitors, NH4Cl or MG132, partially inhibited the antofine-induced decrease in Cx43 protein levels, but did not inhibit the antofine-induced inhibition of GJIC. After 30min of treatment, antofine induced a rapid increase in the intracellular Ca(2+) concentration and activation of protein kinase C (PKC)α/βII, which was maintained for at least 6h. Co-treatment of astrocytes with antofine and the intracellular Ca(2+) chelator BAPTA-AM prevented downregulation of Cx43 and inhibition of GJIC. Moreover, co-treatment with antofine and a specific PKCβ inhibitor prevented endocytosis of gap junctions, downregulation of Cx43, and inhibition of GJIC. Taken together, these findings indicate that antofine induces Cx43 gap junction disassembly by the PKCβ signaling pathway. Inhibition of GJIC by antofine may undermine the neuroprotective effect of astrocytes in CNS. PMID:23403203

  5. SLO BK Potassium Channels Couple Gap Junctions to Inhibition of Calcium Signaling in Olfactory Neuron Diversification

    PubMed Central

    Schumacher, Jennifer A.; Wang, Xiaohong; Merrill, Sean A.; Millington, Grethel; Bayne, Brittany; Jorgensen, Erik M.; Chuang, Chiou-Fen

    2016-01-01

    The C. elegans AWC olfactory neuron pair communicates to specify asymmetric subtypes AWCOFF and AWCON in a stochastic manner. Intercellular communication between AWC and other neurons in a transient NSY-5 gap junction network antagonizes voltage-activated calcium channels, UNC-2 (CaV2) and EGL-19 (CaV1), in the AWCON cell, but how calcium signaling is downregulated by NSY-5 is only partly understood. Here, we show that voltage- and calcium-activated SLO BK potassium channels mediate gap junction signaling to inhibit calcium pathways for asymmetric AWC differentiation. Activation of vertebrate SLO-1 channels causes transient membrane hyperpolarization, which makes it an important negative feedback system for calcium entry through voltage-activated calcium channels. Consistent with the physiological roles of SLO-1, our genetic results suggest that slo-1 BK channels act downstream of NSY-5 gap junctions to inhibit calcium channel-mediated signaling in the specification of AWCON. We also show for the first time that slo-2 BK channels are important for AWC asymmetry and act redundantly with slo-1 to inhibit calcium signaling. In addition, nsy-5-dependent asymmetric expression of slo-1 and slo-2 in the AWCON neuron is necessary and sufficient for AWC asymmetry. SLO-1 and SLO-2 localize close to UNC-2 and EGL-19 in AWC, suggesting a role of possible functional coupling between SLO BK channels and voltage-activated calcium channels in AWC asymmetry. Furthermore, slo-1 and slo-2 regulate the localization of synaptic markers, UNC-2 and RAB-3, in AWC neurons to control AWC asymmetry. We also identify the requirement of bkip-1, which encodes a previously identified auxiliary subunit of SLO-1, for slo-1 and slo-2 function in AWC asymmetry. Together, these results provide an unprecedented molecular link between gap junctions and calcium pathways for terminal differentiation of olfactory neurons. PMID:26771544

  6. Mathematical modeling of gap junction coupling and electrical activity in human β-cells

    NASA Astrophysics Data System (ADS)

    Loppini, Alessandro; Braun, Matthias; Filippi, Simonetta; Gram Pedersen, Morten

    2015-12-01

    Coordinated insulin secretion is controlled by electrical coupling of pancreatic β-cells due to connexin-36 gap junctions. Gap junction coupling not only synchronizes the heterogeneous β-cell population, but can also modify the electrical behavior of the cells. These phenomena have been widely studied with mathematical models based on data from mouse β-cells. However, it is now known that human β-cell electrophysiology shows important differences to its rodent counterpart, and although human pancreatic islets express connexin-36 and show evidence of β-cell coupling, these aspects have been little investigated in human β-cells. Here we investigate theoretically, the gap junction coupling strength required for synchronizing electrical activity in a small cluster of cells simulated with a recent mathematical model of human β-cell electrophysiology. We find a lower limit for the coupling strength of approximately 20 pS (i.e., normalized to cell size, ˜2 pS pF-1) below which spiking electrical activity is asynchronous. To confront this theoretical lower bound with data, we use our model to estimate from an experimental patch clamp recording that the coupling strength is approximately 100-200 pS (10-20 pS pF-1), similar to previous estimates in mouse β-cells. We then investigate the role of gap junction coupling in synchronizing and modifying other forms of electrical activity in human β-cell clusters. We find that electrical coupling can prolong the period of rapid bursting electrical activity, and synchronize metabolically driven slow bursting, in particular when the metabolic oscillators are in phase. Our results show that realistic coupling conductances are sufficient to promote synchrony in small clusters of human β-cells as observed experimentally, and provide motivation for further detailed studies of electrical coupling in human pancreatic islets.

  7. Pharmacological blockade of gap junctions induces repetitive surging of extracellular potassium within the locust CNS.

    PubMed

    Spong, Kristin E; Robertson, R Meldrum

    2013-10-01

    The maintenance of cellular ion homeostasis is crucial for optimal neural function and thus it is of great importance to understand its regulation. Glial cells are extensively coupled by gap junctions forming a network that is suggested to serve as a spatial buffer for potassium (K(+)) ions. We have investigated the role of glial spatial buffering in the regulation of extracellular K(+) concentration ([K(+)]o) within the locust metathoracic ganglion by pharmacologically inhibiting gap junctions. Using K(+)-sensitive microelectrodes, we measured [K(+)]o near the ventilatory neuropile while simultaneously recording the ventilatory rhythm as a model of neural circuit function. We found that blockade of gap junctions with either carbenoxolone (CBX), 18β-glycyrrhetinic acid (18β-GA) or meclofenamic acid (MFA) reliably induced repetitive [K(+)]o surges and caused a progressive impairment in the ability to maintain baseline [K(+)]o levels throughout the treatment period. We also show that a low dose of CBX that did not induce surging activity increased the vulnerability of locust neural tissue to spreading depression (SD) induced by Na(+)/K(+)-ATPase inhibition with ouabain. CBX pre-treatment increased the number of SD events induced by ouabain and hindered the recovery of [K(+)]o back to baseline levels between events. Our results suggest that glial spatial buffering through gap junctions plays an essential role in the regulation of [K(+)]o under normal conditions and also contributes to a component of [K(+)]o clearance following physiologically elevated levels of [K(+)]o. PMID:23916994

  8. SLO BK Potassium Channels Couple Gap Junctions to Inhibition of Calcium Signaling in Olfactory Neuron Diversification.

    PubMed

    Alqadah, Amel; Hsieh, Yi-Wen; Schumacher, Jennifer A; Wang, Xiaohong; Merrill, Sean A; Millington, Grethel; Bayne, Brittany; Jorgensen, Erik M; Chuang, Chiou-Fen

    2016-01-01

    The C. elegans AWC olfactory neuron pair communicates to specify asymmetric subtypes AWCOFF and AWCON in a stochastic manner. Intercellular communication between AWC and other neurons in a transient NSY-5 gap junction network antagonizes voltage-activated calcium channels, UNC-2 (CaV2) and EGL-19 (CaV1), in the AWCON cell, but how calcium signaling is downregulated by NSY-5 is only partly understood. Here, we show that voltage- and calcium-activated SLO BK potassium channels mediate gap junction signaling to inhibit calcium pathways for asymmetric AWC differentiation. Activation of vertebrate SLO-1 channels causes transient membrane hyperpolarization, which makes it an important negative feedback system for calcium entry through voltage-activated calcium channels. Consistent with the physiological roles of SLO-1, our genetic results suggest that slo-1 BK channels act downstream of NSY-5 gap junctions to inhibit calcium channel-mediated signaling in the specification of AWCON. We also show for the first time that slo-2 BK channels are important for AWC asymmetry and act redundantly with slo-1 to inhibit calcium signaling. In addition, nsy-5-dependent asymmetric expression of slo-1 and slo-2 in the AWCON neuron is necessary and sufficient for AWC asymmetry. SLO-1 and SLO-2 localize close to UNC-2 and EGL-19 in AWC, suggesting a role of possible functional coupling between SLO BK channels and voltage-activated calcium channels in AWC asymmetry. Furthermore, slo-1 and slo-2 regulate the localization of synaptic markers, UNC-2 and RAB-3, in AWC neurons to control AWC asymmetry. We also identify the requirement of bkip-1, which encodes a previously identified auxiliary subunit of SLO-1, for slo-1 and slo-2 function in AWC asymmetry. Together, these results provide an unprecedented molecular link between gap junctions and calcium pathways for terminal differentiation of olfactory neurons. PMID:26771544

  9. Lateral interactions among membrane proteins. Implications for the organization of gap junctions.

    PubMed Central

    Abney, J R; Braun, J; Owicki, J C

    1987-01-01

    We have studied the relationship between interprotein forces and the lateral distribution of proteins in disordered mouse liver gap junctions. Data on protein positions are obtained from freeze-fracture electron micrographs. Short-ranged correlations in observed positions are characteristic of interacting particles in a fluid state. An analysis derived from statistical mechanics allows the determination of the magnitude and functional form of interprotein forces. We find that jap junction proteins are mutually repulsive, in a manner consistent with electrostatics and excluded volume. This dictates that long-ranged protein aggregation into jap junction plaques cannot arise solely from interparticle interactions. An alternative is the balance of lateral pressures between the junction and the surrounding glycocalyx. This idea is quantified into a model. Junctional pressure arises from protein-protein interactions and is computed from a pressure equation based on the force and a radial distribution function describing order. The pressure from the glycocalyx is assumed to arise from mixing, electrostatic, and elastic interactions of sugar residues, and is described with terms from Flory-Krigbaum and McMillan-Mayer theories. The results of this modeling are in reasonable agreement with available experimental data. Images FIGURE 1 PMID:3651562

  10. Osteocytes up-regulate the terminal differentiation of pre-osteoblasts via gap junctions.

    PubMed

    Nishikawa, Yoichi; Akiyama, Yuko; Yamamoto, Kiyofumi; Kobayashi, Masayuki; Watanabe, Eri; Watanabe, Nobukazu; Shimizu, Noriyoshi; Mikami, Yoshikazu; Komiyama, Kazuo

    2015-01-01

    We examined cell-to-cell interaction between pre-osteoblasts and osteocytes using MC3T3-E1 and MLO-Y4, respectively. First, GFP expressing MC3T3-E1 (E1-GFP) cells were generated to isolate the cells from co-culture with MLO-Y4. No changes were observed in the expression of osteogenic transcription factors Runx2, Osterix, Dlx5 and Msx2, but expression of alkaline phosphatase (ALP) and bone sialoprotein (BSP) in E1-GFP co-cultured with MLO-Y4 was 300-400-fold greater than that in mono-cultured E1-GFP. In addition, mineralized nodule formation was drastically increased in co-cultured E1-GFP cells compared to mono-cultured cells. Patch clamp assay showed the presence of gap junctions between E1-GFP and MLO-Y4. Furthermore, when the gap junction inhibitor carbenoxolone (CBX) was added to the culture, increased expression of ALP and BSP in E1-GFP co-cultured with MLO-Y4 was suppressed. These results suggest that gap junction detected between pre-osteoblasts and osteocytes plays an important role on the terminal differentiation of pre-osteoblasts. PMID:25450679

  11. Expression and role of connexin-based gap junctions in pulmonary inflammatory diseases.

    PubMed

    Freund-Michel, Véronique; Muller, Bernard; Marthan, Roger; Savineau, Jean-Pierre; Guibert, Christelle

    2016-08-01

    Connexins are transmembrane proteins that can generate intercellular communication channels known as gap junctions. They contribute to the direct movement of ions and larger cytoplasmic solutes between various cell types. In the lung, connexins participate in a variety of physiological functions, such as tissue homeostasis and host defence. In addition, emerging evidence supports a role for connexins in various pulmonary inflammatory diseases, such as asthma, pulmonary hypertension, acute lung injury, lung fibrosis or cystic fibrosis. In these diseases, the altered expression of connexins leads to disruption of normal intercellular communication pathways, thus contributing to various pathophysiological aspects, such as inflammation or tissue altered reactivity and remodeling. The present review describes connexin structure and organization in gap junctions. It focuses on connexins in the lung, including pulmonary bronchial and arterial beds, by looking at their expression, regulation and physiological functions. This work also addresses the issue of connexin expression alteration in various pulmonary inflammatory diseases and describes how targeting connexin-based gap junctions with pharmacological tools, synthetic blocking peptides or genetic approaches, may open new therapeutic perspectives in the treatment of these diseases. PMID:27126473

  12. Structure of the extracellular surface of the gap junction by atomic force microscopy.

    PubMed Central

    Hoh, J H; Sosinsky, G E; Revel, J P; Hansma, P K

    1993-01-01

    The extracellular surface of the gap junction cell-to-cell channels was imaged in phosphate-buffered saline with an atomic force microscope. The fully hydrated isolated gap junction membranes adsorbed to mica were irregular sheets approximately 1-2 microns across and 13.2 (+/- 1.3) nm thick. The top bilayer of the gap junction was dissected by increasing the force applied to the tip or sometimes by increasing the scan rate at moderate forces. The exposed extracellular surface revealed a hexagonal array with a center-to-center spacing of 9.4 (+/- 0.9) nm between individual channels (connexons). Images of individual connexons with a lateral resolution of < 3.5 nm, and in the best case approximately 2.5 nm, were reliably and reproducibly obtained with high-quality tips. These membrane channels protruded 1.4 (+/- 0.4) nm from the extracellular surface of the lipid membrane, and the atomic force microscope tip reached up to 0.7 nm into the pore, which opened up to a diameter of 3.8 (+/- 0.6) nm on the extracellular side. Images FIGURE 2 FIGURE 1 FIGURE 3 FIGURE 4 FIGURE 5 FIGURE 6 FIGURE 7 FIGURE 8 FIGURE 9 FIGURE 10 PMID:8396452

  13. Phylogenetic and bioinformatic analysis of gap junction-related proteins, innexins, pannexins and connexins.

    PubMed

    Fushiki, Daisuke; Hamada, Yasuo; Yoshimura, Ryoichi; Endo, Yasuhisa

    2010-04-01

    All multi-cellular animals, including hydra, insects and vertebrates, develop gap junctions, which communicate directly with neighboring cells. Gap junctions consist of protein families called connexins in vertebrates and innexins in invertebrates. Connexins and innexins have no homology in their amino acid sequence, but both are thought to have some similar characteristics, such as a tetra-membrane-spanning structure, formation of a channel by hexamer, and transmission of small molecules (e.g. ions) to neighboring cells. Pannexins were recently identified as a homolog of innexins in vertebrate genomes. Although pannexins are thought to share the function of intercellular communication with connexins and innexins, there is little information about the relationship among these three protein families of gap junctions. We phylgenetically and bioinformatically examined these protein families and other tetra-membrane-spanning proteins using a database and three analytical softwares. The clades formed by pannexin families do not belong to the species classification but do to paralogs of each member of pannexins. Amino acid sequences of pannexins are closely related to those of innexins but less to those of connexins. These data suggest that innexins and pannexins have a common origin, but the relationship between innexins/pannexins and connexins is as slight as that of other tetra-membrane-spanning members. PMID:20460741

  14. Connexin 43 mediated gap junctional communication enhances breast tumor cell diapedesis in culture

    PubMed Central

    Pollmann, Mary-Ann; Shao, Qing; Laird, Dale W; Sandig, Martin

    2005-01-01

    Introduction Metastasis involves the emigration of tumor cells through the vascular endothelium, a process also known as diapedesis. The molecular mechanisms regulating tumor cell diapedesis are poorly understood, but may involve heterocellular gap junctional intercellular communication (GJIC) between tumor cells and endothelial cells. Method To test this hypothesis we expressed connexin 43 (Cx43) in GJIC-deficient mammary epithelial tumor cells (HBL100) and examined their ability to form gap junctions, establish heterocellular GJIC and migrate through monolayers of human microvascular endothelial cells (HMVEC) grown on matrigel-coated coverslips. Results HBL100 cells expressing Cx43 formed functional heterocellular gap junctions with HMVEC monolayers within 30 minutes. In addition, immunocytochemistry revealed Cx43 localized to contact sites between Cx43 expressing tumor cells and endothelial cells. Quantitative analysis of diapedesis revealed a two-fold increase in diapedesis of Cx43 expressing cells compared to empty vector control cells. The expression of a functionally inactive Cx43 chimeric protein in HBL100 cells failed to increase migration efficiency, suggesting that the observed up-regulation of diapedesis in Cx43 expressing cells required heterocellular GJIC. This finding is further supported by the observation that blocking homocellular and heterocellular GJIC with carbenoxolone in co-cultures also reduced diapedesis of Cx43 expressing HBL100 tumor cells. Conclusion Collectively, our results suggest that heterocellular GJIC between breast tumor cells and endothelial cells may be an important regulatory step during metastasis. PMID:15987459

  15. Modulation of human cell responses to space radiation by gap-junction communication

    NASA Astrophysics Data System (ADS)

    Autsavapromporn, Narongchai; de Toledo, Sonia M.; Buonanno, Manuela; Yang, Zhi; Harris, Andrew; Jay-Gerin, Jean-Paul; Azzam, Edouard

    Understanding the biological effects of space radiation and their underlying mechanism is critical to estimating the health risk associated with human exploration of space. A coordinated interaction of multiple cellular processes is likely involved in the sensing and processing of stressful effects induced by different types of space radiation. Here, we focused on the role of gap-junction intercellular communication (GJIC) in responses of human cells exposed to 1 GeV/n protons or 56 Fe-ions. We compared the results with data obtained in human cells exposed, in parallel, to γ-rays or α-particles. As expected, a higher level of cell killing and DNA damage, per unit dose, was induced in confluent, density-inhibited cells (98% in G0 /G1 ) exposed to α-particles or energetic 56 Fe-ions than γ-rays or protons. Strikingly, greatly attenuated effects occurred when sub-confluent cultures, synchronized in G0 /G1 ,were exposed to 56 Fe-ions. These data suggest that direct intercellular communication is involved in the effects of high linear energy transfer (LET) 56 Fe-ions. To examine the role of gap-junctions in propagating stressful effect, confluent cultures were exposed to 56 Fe-ions or α-particles and incubated for various time periods at 37° C in the presence or absence of the gap-junction inhibitor α-glycyrrhetinic acid (AGA). No repair of potentially lethal radiation damage occurred in cells incubated in the absence of AGA. In contrast, inhibition of functional GJIC significantly enhanced clonogenic survival of irradiated cells. To test the role of junctional channel permeability in the observed effects, we used human adenocarcinoma (HeLa) cells in which specific connexins (Cx) can be expressed in the absence of endogenous connexins. Whereas HeLa cells with selective inducible expression of Cx26 gap-junctions promoted radiation toxic effects, expression of Cx32 junctional channels in HeLa cells promoted pro-survival effects. Experiments are in progress to

  16. Enhanced effect of gap junction uncouplers on macroscopic electrical properties of reperfused myocardium.

    PubMed

    Rodriguez-Sinovas, Antonio; García-Dorado, David; Ruiz-Meana, Marisol; Soler-Soler, Jordi

    2004-08-15

    Transient inhibition of gap junction (GJ)-mediated communication with heptanol during myocardial reperfusion limits infarct size. However, inhibition of cell coupling in normal myocardium may be arrhythmogenic. The purpose of this study was to test the hypothesis that the consequences of GJ inhibition may be magnified in reperfused myocardium compared with normal tissue, thus allowing the inhibition of GJs in reperfused tissue while only minimally modifying overall macroscopic cell coupling in normal myocardium. Concentration-response curves were defined for the effects of heptanol, 18alpha-glycyrrhetinic acid, halothane, and palmitoleic acid on conduction velocity, tissue electrical impedance, developed tension and lactate dehydrogenase (LDH) release in normoxically perfused rat hearts (n= 17). Concentrations lacking significant effects on tissue impedance were added during the initial 15 min of reperfusion in hearts submitted to 60 min (n= 43) or 30 min (n= 35) of ischaemia. These concentrations markedly increased myocardial electrical impedance (resistivity and phase angle) in myocardium reperfused after either 30 or 60 min of ischaemia, and reduced reperfusion-induced LDH release after 1 h of ischaemia by 83.6, 57.9, 51.7 and 52.5% for heptanol, 18alpha-glycyrrhetinic acid, halothane and palmitoleic acid, respectively. LDH release was minimal in hearts submitted to 30 min of ischaemia, independently of group allocation. In conclusion, the present results strongly support the hypothesis that intercellular communication in postischaemic myocardium may be effectively reduced by concentrations of GJ inhibitors affecting only minimally overall electrical impedance in normal myocardium. Reduction of cell coupling during initial reperfusion was consistently associated with attenuated lethal reperfusion injury. PMID:15218064

  17. Enhanced effect of gap junction uncouplers on macroscopic electrical properties of reperfused myocardium

    PubMed Central

    Rodriguez-Sinovas, Antonio; García-Dorado, David; Ruiz-Meana, Marisol; Soler-Soler, Jordi

    2004-01-01

    Transient inhibition of gap junction (GJ)-mediated communication with heptanol during myocardial reperfusion limits infarct size. However, inhibition of cell coupling in normal myocardium may be arrhythmogenic. The purpose of this study was to test the hypothesis that the consequences of GJ inhibition may be magnified in reperfused myocardium compared with normal tissue, thus allowing the inhibition of GJs in reperfused tissue while only minimally modifying overall macroscopic cell coupling in normal myocardium. Concentration–response curves were defined for the effects of heptanol, 18α-glycyrrhetinic acid, halothane, and palmitoleic acid on conduction velocity, tissue electrical impedance, developed tension and lactate dehydrogenase (LDH) release in normoxically perfused rat hearts (n = 17). Concentrations lacking significant effects on tissue impedance were added during the initial 15 min of reperfusion in hearts submitted to 60 min (n = 43) or 30 min (n = 35) of ischaemia. These concentrations markedly increased myocardial electrical impedance (resistivity and phase angle) in myocardium reperfused after either 30 or 60 min of ischaemia, and reduced reperfusion-induced LDH release after 1 h of ischaemia by 83.6, 57.9, 51.7 and 52.5% for heptanol, 18α-glycyrrhetinic acid, halothane and palmitoleic acid, respectively. LDH release was minimal in hearts submitted to 30 min of ischaemia, independently of group allocation. In conclusion, the present results strongly support the hypothesis that intercellular communication in postischaemic myocardium may be effectively reduced by concentrations of GJ inhibitors affecting only minimally overall electrical impedance in normal myocardium. Reduction of cell coupling during initial reperfusion was consistently associated with attenuated lethal reperfusion injury. PMID:15218064

  18. Cell communication across gap junctions: a historical perspective and current developments.

    PubMed

    Evans, W Howard

    2015-06-01

    Collaborative communication lies at the centre of multicellular life. Gap junctions (GJs) are surface membrane structures that allow direct communication between cells. They were discovered in the 1960s following the convergence of the detection of low-resistance electrical interactions between cells and anatomical studies of intercellular contact points. GJs purified from liver plasma membranes contained a 27 kDa protein constituent; it was later named Cx32 (connexin 32) after its full sequence was determined by recombinant technology. Identification of Cx43 in heart and later by a further GJ protein, Cx26 followed. Cxs have a tetraspan organization in the membrane and oligomerize during intracellular transit to the plasma membrane; these were shown to be hexameric hemichannels (connexons) that could interact end-to-end to generate GJs at areas of cell-to-cell contact. The structure of the GJ was confirmed and refined by a combination of biochemical and structural approaches. Progress continues towards obtaining higher atomic 3D resolution of the GJ channel. Today, there are 20 and 21 highly conserved members of the Cx family in the human and mouse genomes respectively. Model organisms such as Xenopus oocytes and zebra fish are increasingly used to relate structure to function. Proteins that form similar large pore membrane channels in cells called pannexins have also been identified in chordates. Innexins form GJs in prechordates; these two other proteins, although functionally similar, are very different in amino acid sequence to the Cxs. A time line tracing the historical progression of wide ranging research in GJ biology over 60 years is mapped out. The molecular basis of channel dysfunctions in disease is becoming evident and progress towards addressing Cx channel-dependent pathologies, especially in ischaemia and tissue repair, continues. PMID:26009190

  19. Combined electric field and gap junctions on propagation of action potentials in cardiac muscle and smooth muscle in PSpice simulation.

    PubMed

    Sperelakis, Nicholas

    2003-10-01

    Propagation of action potentials in cardiac muscle and smooth muscle were simulated using the PSpice program. Excitation was transmitted from cell to cell along a strand of 6 cells (cardiac muscle) or 10 cells (smooth muscle) either not connected (control) or connected by low-resistance tunnels (gap-junction connexons). A significant negative cleft potential (V(jv) ) develops in the narrow junctional cleft when the pre-JM fires. V(jc) depolarizes the postjunctional membrane (post-JM) to threshold by a patch-clamp action. With few connecting tunnels, cell-to-cell transmission by the EF mechanism was facilitated. With many tunnels, propagation was dominated by the low-resistance mechanism, and propagation velocity (theta) became very fast and nonphysiological. In conclusion, when the 2 mechanisms for cell-to-cell transfer of excitation were combined, the two mechanisms facilitated each other in a synergistic manner. When there were many connecting tunnels, the tunnel mechanism was dominant. PMID:14661164

  20. The Carboxyl Tail of Connexin32 Regulates Gap Junction Assembly in Human Prostate and Pancreatic Cancer Cells*

    PubMed Central

    Katoch, Parul; Mitra, Shalini; Ray, Anuttoma; Kelsey, Linda; Roberts, Brett J.; Wahl, James K.; Johnson, Keith R.; Mehta, Parmender P.

    2015-01-01

    Connexins, the constituent proteins of gap junctions, are transmembrane proteins. A connexin (Cx) traverses the membrane four times and has one intracellular and two extracellular loops with the amino and carboxyl termini facing the cytoplasm. The transmembrane and the extracellular loop domains are highly conserved among different Cxs, whereas the carboxyl termini, often called the cytoplasmic tails, are highly divergent. We have explored the role of the cytoplasmic tail of Cx32, a Cx expressed in polarized and differentiated cells, in regulating gap junction assembly. Our results demonstrate that compared with the full-length Cx32, the cytoplasmic tail-deleted Cx32 is assembled into small gap junctions in human pancreatic and prostatic cancer cells. Our results further document that the expression of the full-length Cx32 in cells, which express the tail-deleted Cx32, increases the size of gap junctions, whereas the expression of the tail-deleted Cx32 in cells, which express the full-length Cx32, has the opposite effect. Moreover, we show that the tail is required for the clustering of cell-cell channels and that in cells expressing the tail-deleted Cx32, the expression of cell surface-targeted cytoplasmic tail alone is sufficient to enhance the size of gap junctions. Our live-cell imaging data further demonstrate that gap junctions formed of the tail-deleted Cx32 are highly mobile compared with those formed of full-length Cx32. Our results suggest that the cytoplasmic tail of Cx32 is not required to initiate the assembly of gap junctions but for their subsequent growth and stability. Our findings suggest that the cytoplasmic tail of Cx32 may be involved in regulating the permeability of gap junctions by regulating their size. PMID:25548281

  1. Gap junction blockage promotes cadmium-induced apoptosis in BRL 3A derived from Buffalo rat liver cells

    PubMed Central

    Hu, Di; Zou, Hui; Han, Tao; Xie, Junze; Dai, Nannan; Zhuo, Liling; Gu, Jianhong; Bian, Jianchun; Yuan, Yan; Liu, Xuezhong

    2016-01-01

    Gap junctions mediate direct communication between cells; however, toxicological cascade triggered by nonessential metals can abrogate cellular signaling mediated by gap junctions. Although cadmium (Cd) is known to induce apoptosis in organs and tissues, the mechanisms that underlie gap junction activity in Cd-induced apoptosis in BRL 3A rat liver cells has yet to be established. In this study, we showed that Cd treatment decreased the cell index (a measure of cellular electrical impedance) in BRL 3A cells. Mechanistically, we found that Cd exposure decreased expression of connexin 43 (Cx43), increased expression of p-Cx43 and elevated intracellular free Ca2+ concentration, corresponding to a decrease in gap junctional intercellular communication. Gap junction blockage pretreatment with 18β-glycyrrhizic acid (GA) promoted Cd-induced apoptosis, involving changes in expression of Bax, Bcl-2, caspase-3 and the mitochondrial transmembrane electrical potential (Δψm). Additionally, GA was found to enhance ERK and p38 activation during Cd-induced activation of mitogen-activated protein kinases, but had no significant effect on JNK activation. Our results indicated the apoptosis-related proteins and the ERK and p38 signaling pathways may participate in gap junction blockage promoting Cd-induced apoptosis in BRL 3A cells. PMID:27051341

  2. Gap junctions on hippocampal mossy fiber axons demonstrated by thin-section electron microscopy and freeze fracture replica immunogold labeling.

    PubMed

    Hamzei-Sichani, Farid; Kamasawa, Naomi; Janssen, William G M; Yasumura, Thomas; Davidson, Kimberly G V; Hof, Patrick R; Wearne, Susan L; Stewart, Mark G; Young, Steven R; Whittington, Miles A; Rash, John E; Traub, Roger D

    2007-07-24

    Gap junctions have been postulated to exist between the axons of excitatory cortical neurons based on electrophysiological, modeling, and dye-coupling data. Here, we provide ultrastructural evidence for axoaxonic gap junctions in dentate granule cells. Using combined confocal laser scanning microscopy, thin-section transmission electron microscopy, and grid-mapped freeze-fracture replica immunogold labeling, 10 close appositions revealing axoaxonic gap junctions ( approximately 30-70 nm in diameter) were found between pairs of mossy fiber axons ( approximately 100-200 nm in diameter) in the stratum lucidum of the CA3b field of the rat ventral hippocampus, and one axonal gap junction ( approximately 100 connexons) was found on a mossy fiber axon in the CA3c field of the rat dorsal hippocampus. Immunogold labeling with two sizes of gold beads revealed that connexin36 was present in that axonal gap junction. These ultrastructural data support computer modeling and in vitro electrophysiological data suggesting that axoaxonic gap junctions play an important role in the generation of very fast (>70 Hz) network oscillations and in the hypersynchronous electrical activity of epilepsy. PMID:17640909

  3. Analysis of longitudinal vibration band gaps in periodic carbon nanotube intramolecular junctions using finite element method

    NASA Astrophysics Data System (ADS)

    Li, Jiaqian; Shen, Haijun

    2015-12-01

    The longitudinal vibration band gaps in periodic (n, 0)-(2n, 0) single-walled carbon nanotube(SWCNT) intramolecular junctions(IMJs) are investigated based on the finite element calculation. The frequency ranges of band gaps in frequency response functions(FRF) simulated by finite element method (FEM) show good agreement with those in band structure obtained by simple spring-mass model. Moreover, a comprehensive parametric study is also conducted to highlight the influences of the geometrical parameters such as the size of unit cell, component ratios of the IMJs and diameters of the CNT segments as well as geometric imperfections on the first band gap. The results show that the frequency ranges and the bandwidth of the gap strongly depend on the geometrical parameters. Furthermore, the influences of geometrical parameters on gaps are nuanced in IMJs with different topological defects. The existence of vibration band gaps in periodic IMJs lends a new insight into the development of CNT-based nano-devices in application of vibration isolation.

  4. Electric-field-induced magnetization switching in CoFeB/MgO magnetic tunnel junctions with high junction resistance

    NASA Astrophysics Data System (ADS)

    Kanai, S.; Matsukura, F.; Ohno, H.

    2016-05-01

    We show the electric-field induced magnetization switching for CoFeB/MgO magnetic tunnel junctions with thick MgO barrier layer of 2.8 nm, whose resistance-area product is 176 kΩ μm2, and achieve the small switching energy of 6.3 fJ/bit. The increase of the junction resistance is expected to suppress the energy consumption due to the Joule heating during the switching; however, the energy is still dominated by the Joule energy rather than the charging energy. This is because the junction resistance decreases more rapidly for junctions with thicker MgO as bias voltage increases.

  5. Entrainment, retention, and transport of freely swimming fish in junction gaps between commercial barges operating on the Illinois Waterway

    USGS Publications Warehouse

    Davis, Jeremiah J.; Jackson, Patrick; Engel, Frank; LeRoy, Jessica Z.; Neeley, Rebecca N.; Finney, Samuel T.; Murphy, Elizabeth

    2016-01-01

    Large Electric Dispersal Barriers were constructed in the Chicago Sanitary and Ship Canal (CSSC) to prevent the transfer of invasive fish species between the Mississippi River Basin and the Great Lakes Basin while simultaneously allowing the passage of commercial barge traffic. We investigated the potential for entrainment, retention, and transport of freely swimming fish within large gaps (> 50 m3) created at junction points between barges. Modified mark and capture trials were employed to assess fish entrainment, retention, and transport by barge tows. A multi-beam sonar system enabled estimation of fish abundance within barge junction gaps. Barges were also instrumented with acoustic Doppler velocity meters to map the velocity distribution in the water surrounding the barge and in the gap formed at the junction of two barges. Results indicate that the water inside the gap can move upstream with a barge tow at speeds near the barge tow travel speed. Water within 1 m to the side of the barge junction gaps was observed to move upstream with the barge tow. Observed transverse and vertical water velocities suggest pathways by which fish may potentially be entrained into barge junction gaps. Results of mark and capture trials provide direct evidence that small fish can become entrained by barges, retained within junction gaps, and transported over distances of at least 15.5 km. Fish entrained within the barge junction gap were retained in that space as the barge tow transited through locks and the Electric Dispersal Barriers, which would be expected to impede fish movement upstream.

  6. Sub-harmonic gap structure and Magneto-transport in suspended graphene -Superconductor ballistic junctions

    NASA Astrophysics Data System (ADS)

    Kumaravadivel, Piranavan; Du, Xu

    2015-03-01

    Inducing superconductivity in graphene via the proximity effect enables to study the rich transport of the massless Dirac fermions at the Superconductor(S) - Graphene (G) interface. Some of the predictions are pseudo diffusive transport in Ballistic SGS junctions at low carrier densities and the unique specular and retro Andreev reflections in graphene. One of the challenges in observing these experimentally is to fabricate highly transparent ballistic SGS junctions that can be probed at low carrier densities near the Dirac point. In this talk we will present our recent results on suspended graphene- Niobium Josephson weak links. Our devices exhibit a mobility of ~ 350000 cm2V-1s-1 with a carrier density as low as 109 cm-2. Below the Superconducting transition temperature (Tc) ~ 9K, the devices show supercurrent and sub-harmonic gap structure due to Multiple Andreev reflections. In the vicinity of the Dirac point, the sub-harmonic gap structure becomes more pronounced, which as predicated, is indicative of pseudo-diffusive transport. With a fine scanning of gate voltage close to Dirac point we see emergence of some unusual sub- gap structures. We also report on our study of these samples below the upper critical field of Nb (~ 3.5T), where superconducting proximity effect coexists with Quantum Hall effect.

  7. Connexin 33: a rodent-specific member of the gap junction protein family?

    PubMed

    Fischer, Petra; Brehm, Ralph; Konrad, Lutz; Hartmann, Sonja; Kliesch, Sabine; Bohle, Rainer M; Bergmann, Martin

    2005-01-01

    Gap junctional intercellular communication between Sertoli cells and between Sertoli cells and spermatogonia is considered to play a key role in the regulation of both proliferation and differentiation of germ cells. A member of the gap junction protein family, Connexin 33 (cx33), probably has an inhibitory effect on the formation of gap junctions and so far it is the only cx that has been exclusively found in rat and mouse testes. Thus, this connexin seems to be a special member of the cx family. Using immunohistochemistry, Western blot analysis, polymerase chain reaction, and reverse transcription (RT)-PCR (tissue homogenate and microdissected cells), we studied the possible occurrence of cx33 at the protein, the DNA, and the RNA level in human testis. Whereas immunohistochemistry using the only commercially available anti-cx33 antibody showed similar labeling to the rat within the seminiferous epithelium, we could not find any further evidence for the existence of cx33 using Western blot analysis, PCR, and RT-PCR in human testis. Based on the demonstration of the staining pattern of mitochondria in human germ cells and on preabsorption studies, we could demonstrate anti-cx33 antibody cross-reacting with mitochondrial ferritin, a protein localized in the mitochondria of human testicular spermatids. Therefore, we were not able to abide by the suspicion that cx33 is present in human testis. Additionally, it was not possible to demonstrate cx33 via PCR and immunohistochemistry in the testis of different mammals (dog, cattle, pig, horse, and marmoset monkey) with normal spermatogenesis. These data indicate that cx33 seems to be the first rodent-specific testicular cx. PMID:15611570

  8. Simvastatin Sodium Salt and Fluvastatin Interact with Human Gap Junction Gamma-3 Protein.

    PubMed

    Marsh, Andrew; Casey-Green, Katherine; Probert, Fay; Withall, David; Mitchell, Daniel A; Dilly, Suzanne J; James, Sean; Dimitri, Wade; Ladwa, Sweta R; Taylor, Paul C; Singer, Donald R J

    2016-01-01

    Finding pleiomorphic targets for drugs allows new indications or warnings for treatment to be identified. As test of concept, we applied a new chemical genomics approach to uncover additional targets for the widely prescribed lipid-lowering pro-drug simvastatin. We used mRNA extracted from internal mammary artery from patients undergoing coronary artery surgery to prepare a viral cardiovascular protein library, using T7 bacteriophage. We then studied interactions of clones of the bacteriophage, each expressing a different cardiovascular polypeptide, with surface-bound simvastatin in 96-well plates. To maximise likelihood of identifying meaningful interactions between simvastatin and vascular peptides, we used a validated photo-immobilisation method to apply a series of different chemical linkers to bind simvastatin so as to present multiple orientations of its constituent components to potential targets. Three rounds of biopanning identified consistent interaction with the clone expressing part of the gene GJC3, which maps to Homo sapiens chromosome 7, and codes for gap junction gamma-3 protein, also known as connexin 30.2/31.3 (mouse connexin Cx29). Further analysis indicated the binding site to be for the N-terminal domain putatively 'regulating' connexin hemichannel and gap junction pores. Using immunohistochemistry we found connexin 30.2/31.3 to be present in samples of artery similar to those used to prepare the bacteriophage library. Surface plasmon resonance revealed that a 25 amino acid synthetic peptide representing the discovered N-terminus did not interact with simvastatin lactone, but did bind to the hydrolysed HMG CoA inhibitor, simvastatin acid. This interaction was also seen for fluvastatin. The gap junction blockers carbenoxolone and flufenamic acid also interacted with the same peptide providing insight into potential site of binding. These findings raise key questions about the functional significance of GJC3 transcripts in the vasculature and

  9. Regulation of gap junction function and Connexin 43 expression by cytochrome P450 oxidoreductase (CYPOR)

    SciTech Connect

    Polusani, Srikanth R.; Kar, Rekha; Riquelme, Manuel A.; Masters, Bettie Sue; Panda, Satya P.

    2011-08-05

    Highlights: {yields} Humans with severe forms of cytochrome P450 oxidoreductase (CYPOR) mutations show bone defects as observed in Antley-Bixler Syndrome. {yields} First report showing knockdown of CYPOR in osteoblasts decreased Connexin 43 (Cx43) protein levels. Cx43 is known to play an important role in bone modeling. {yields} Knockdown of CYPOR decreased Gap Junctional Intercellular Communication and hemichannel activity. {yields} Knockdown of CYPOR decreased Cx43 in mouse primary calvarial osteoblasts. {yields} Decreased Cx43 expression was observed at the transcriptional level. -- Abstract: Cytochrome P450 oxidoreductase (CYPOR) is a microsomal electron-transferring enzyme containing both FAD and FMN as co-factors, which provides the reducing equivalents to various redox partners, such as cytochromes P450 (CYPs), heme oxygenase (HO), cytochrome b{sub 5} and squalene monooxygenase. Human patients with severe forms of CYPOR mutation show bone defects such as cranio- and humeroradial synostoses and long bone fractures, known as Antley-Bixler-like Syndrome (ABS). To elucidate the role of CYPOR in bone, we knocked-down CYPOR in multiple osteoblast cell lines using RNAi technology. In this study, knock-down of CYPOR decreased the expression of Connexin 43 (Cx43), known to play a critical role in bone formation, modeling, and remodeling. Knock-down of CYPOR also decreased Gap Junction Intercellular Communication (GJIC) and hemichannel activity. Promoter luciferase assays revealed that the decrease in expression of Cx43 in CYPOR knock-down cells was due to transcriptional repression. Primary osteoblasts isolated from bone specific Por knock-down mice calvariae confirmed the findings in the cell lines. Taken together, our study provides novel insights into the regulation of gap junction function by CYPOR and suggests that Cx43 may play an important role(s) in CYPOR-mediated bone defects seen in patients.

  10. Octanol, a gap junction uncoupling agent, changes intracellular [H+] in rat astrocytes.

    PubMed

    Pappas, C A; Rioult, M G; Ransom, B R

    1996-01-01

    Octanol rapidly closes gap junction channels but its mechanism of action is not known. Because intracellular [H+], pHi, also affects the conductance of gap junctions, we studied octanol's effects on pHi in cultured rat astrocytes, which are highly coupled cells. Octanol (1 mM) caused an acid shift in the pHi of 90% of rat hippocampal astrocytes which averaged -0.19 +/- 0.09 pH units in magnitude. In 58% of the cells tested, a biphasic change in pHi was seen; octanol produced an initial acidification lasting approximately 10 min that was followed by a persistent alkalinization. The related gap junction uncoupling agent, heptanol, had similar effects on pHi. Octanol-induced changes in pHi were similar in nominally HCO(3-)-free and HCO(3-)-containing solutions, although the rate of initial acidification was significantly greater in the presence of HCO3-. The initial acidification was inhibited in the presence of the stilbene DIDS, an inhibitor of Na+/HCO3- cotransport, indicating that octanol caused acidification by blocking this powerful acid extruder. The alkalinization was inhibited by amiloride which blocks the Na+/H+ exchanger (NHE), an acid extruder, suggesting that the alkaline shift induced by octanol was caused by stimulation of NHE. As expected, octanol's effects on astrocytic pHi were prevented by removal of external Na+, which blocks both Na+/HCO3- cotransport and NHE. Octanol had only small effects on intracellular Ca2+ (Ca2+i) in astrocytes. Hepatocytes which, like astrocytes, are strongly coupled to one another, showed no change in pHi with octanol application. Fluorescence recovery after photobleaching (FRAP) was used to study the effect of changes in astrocyte pHi on degree of coupling in hippocampal astrocytes. Coupling was decreased by intracellular acid shifts approximately -0.2 pH units in size. Octanol's effects on astrocyte pHi were complex but a prompt initial acidification was nearly always seen and could contribute to the uncoupling action of

  11. Voltage-dependent gating of single gap junction channels in an insect cell line.

    PubMed Central

    Bukauskas, F F; Weingart, R

    1994-01-01

    De novo formation of cell pairs was used to examine the gating properties of single gap junction channels. Two separate cells of an insect cell line (clone C6/36, derived from the mosquito Aedes albopictus) were pushed against each other to provoke formation of gap junction channels. A dual voltage-clamp method was used to control the voltage gradient between the cells (Vj) and measure the intercellular current (Ij). The first sign of channel activity was apparent 4.7 min after cell contact. Steady-state coupling reached after 30 min revealed a conductance of 8.7 nS. Channel formation involved no leak between the intra- and extracellular space. The first opening of a newly formed channel was slow (25-28 ms). Each preparation passed through a phase with only one operational gap junction channel. This period was exploited to examine the single channel properties. We found that single channels exhibit several conductance states with different conductances gamma j; a fully open state (gamma j(main state)), several substates (gamma j(substates)), a residual state (gamma j(residual)) and a closed state (gamma j(closed)). The gamma j(main state) was 375 pS, and gamma j(residual) ranged from 30 to 90 pS. The transitions between adjacent substates were 1/7-1/4 of gamma j(main state). Vj had no effect on gamma j(main state), but slightly affected gamma j (residual). The lj transitions involving gamma j(closed) were slow (15-60 ms), whereas those not involving gamma j(closed) were fast (< 2 ms). An increase in Vj led to a decrease in open channel probability. Depolarization of the membrane potential (Vm) increased the incidence of slow transitions leading to gamma j(closed). We conclude that insect gap junctions possess two gates, a fast gate controlled by Vj and giving rise to gamma j(substates) and gamma j(residual), and a slow gate sensitive to Vm and able to close the channel completely. PMID:7524710

  12. Blockade of Gap Junction Hemichannel Suppresses Disease Progression in Mouse Models of Amyotrophic Lateral Sclerosis and Alzheimer's Disease

    PubMed Central

    Takeuchi, Hideyuki; Mizoguchi, Hiroyuki; Doi, Yukiko; Jin, Shijie; Noda, Mariko; Liang, Jianfeng; Li, Hua; Zhou, Yan; Mori, Rarami; Yasuoka, Satoko; Li, Endong; Parajuli, Bijay; Kawanokuchi, Jun; Sonobe, Yoshifumi; Sato, Jun; Yamanaka, Koji; Sobue, Gen; Mizuno, Tetsuya; Suzumura, Akio

    2011-01-01

    Background Glutamate released by activated microglia induces excitotoxic neuronal death, which likely contributes to non-cell autonomous neuronal death in neurodegenerative diseases, including amyotrophic lateral sclerosis and Alzheimer's disease. Although both blockade of glutamate receptors and inhibition of microglial activation are the therapeutic candidates for these neurodegenerative diseases, glutamate receptor blockers also perturbed physiological and essential glutamate signals, and inhibitors of microglial activation suppressed both neurotoxic/neuroprotective roles of microglia and hardly affected disease progression. We previously demonstrated that activated microglia release a large amount of glutamate specifically through gap junction hemichannel. Hence, blockade of gap junction hemichannel may be potentially beneficial in treatment of neurodegenerative diseases. Methods and Findings In this study, we generated a novel blood-brain barrier permeable gap junction hemichannel blocker based on glycyrrhetinic acid. We found that pharmacologic blockade of gap junction hemichannel inhibited excessive glutamate release from activated microglia in vitro and in vivo without producing notable toxicity. Blocking gap junction hemichannel significantly suppressed neuronal loss of the spinal cord and extended survival in transgenic mice carrying human superoxide dismutase 1 with G93A or G37R mutation as an amyotrophic lateral sclerosis mouse model. Moreover, blockade of gap junction hemichannel also significantly improved memory impairments without altering amyloid β deposition in double transgenic mice expressing human amyloid precursor protein with K595N and M596L mutations and presenilin 1 with A264E mutation as an Alzheimer's disease mouse model. Conclusions Our results suggest that gap junction hemichannel blockers may represent a new therapeutic strategy to target neurotoxic microglia specifically and prevent microglia-mediated neuronal death in various

  13. Gapped graphene-based Josephson junction with d-wave pair coupling

    NASA Astrophysics Data System (ADS)

    Goudarzi, H.; Khezerlou, M.; Dezhaloud, T.

    2013-06-01

    The Josephson current passing through a S/I/S gapped graphene-based junction, where superconductivity in the S region is induced by depositing unconventional d-wave superconductor is investigated. The energy levels of massive Dirac fermions are exactly found for Andreev bound states. We illustrate the effect of characteristic of d-wave pairing symmetry on the Andreev bound states and the Josephson current. It is shown that the Josephson current vanishes for special range of superconductivity phase, φ = φ1 - φ2 and the position of the maximum current depends on the mass gap of graphene. The critical supercurrent varies in an oscillatory manner as function of the barrier strength, so that the period of oscillations does not change by increasing the effective mass of quasiparticles.

  14. Junction-to-Case Thermal Resistance of a Silicon Carbide Bipolar Junction Transistor Measured

    NASA Technical Reports Server (NTRS)

    Niedra, Janis M.

    2006-01-01

    Junction temperature of a prototype SiC-based bipolar junction transistor (BJT) was estimated by using the base-emitter voltage (V(sub BE)) characteristic for thermometry. The V(sub BE) was measured as a function of the base current (I(sub B)) at selected temperatures (T), all at a fixed collector current (I(sub C)) and under very low duty cycle pulse conditions. Under such conditions, the average temperature of the chip was taken to be the same as that of the temperature-controlled case. At increased duty cycle such as to substantially heat the chip, but same I(sub C) pulse height, the chip temperature was identified by matching the V(sub BE) to the thermometry curves. From the measured average power, the chip-to-case thermal resistance could be estimated, giving a reasonable value. A tentative explanation for an observed bunching with increasing temperature of the calibration curves may relate to an increasing dopant atom ionization. A first-cut analysis, however, does not support this.

  15. Tracking Dynamic Gap Junctional Coupling in Live Cells by Local Photoactivation and Fluorescence Imaging.

    PubMed

    Yang, Song; Li, Wen-Hong

    2016-01-01

    Intercellular communication through gap junction channels is crucial for maintaining cell homeostasis and synchronizing physiological functions of tissues and organs. In this chapter, we present a noninvasive fluorescence imaging assay termed LAMP (local activation of a molecular fluorescent probe) that consists of the following steps: loading cells with a caged and cell permeable coumarin probe (NPE-HCCC2/AM), locally photolyzing the caged coumarin in one or a subpopulation of coupled cells, monitoring cell-cell dye transfer by digital fluorescence microscopy, and post-acquisition analysis to quantify the rate of junction dye transfer using Fick's equation. The LAMP assay can be conveniently carried out in fully intact cells to assess the extent and degree of cell coupling, and is compatible with other fluorophores emitting at different wavelengths to allow multicolor imaging. Moreover, by carrying out multiple photo-activations in a coupled cell pair, LAMP assay can track changes in cell coupling strength between coupled cells, hence providing a powerful method for investigating the regulation of junctional coupling by cellular biochemical changes. PMID:27207295

  16. Degradation of endocytosed gap junctions by autophagosomal and endo-/lysosomal pathways: a perspective

    PubMed Central

    Falk, Matthias M.; Fong, John T.; Kells, Rachael M.; O’Laughlin, Michael C.; Kowal, Tia J.; Thévenin, Anastasia F.

    2012-01-01

    Gap junctions (GJs) are composed of tens to many thousands of double-membrane spanning GJ channels that cluster together to form densely packed channel arrays (termed GJ plaques) in apposing plasma membranes of neighboring cells. In addition to providing direct intercellular communication (GJIC, their hallmark function), GJs, based on their characteristic double-membrane-spanning configuration, likely also significantly contribute to physical cell-to-cell adhesion. Clearly, modulation (up-/down-regulation) of GJIC and of physical cell-to-cell adhesion is as vitally important as the basic ability of GJ formation itself. Others and we have previously described that GJs can be removed from the plasma membrane via the internalization of entire GJ plaques (or portions thereof) in a cellular process that resembles clathrin-mediated endocytosis. GJ endocytosis results in the formation of double-membrane vesicles (termed annular gap junctions [AGJs] or connexosomes) in the cytoplasm of one of the coupled cells. Four recent independent studies, consistent with earlier ultrastructural analyses, demonstrate the degradation of endocytosed AGJ vesicles via autophagy. However, in TPA-treated cells others report degradation of AGJs via the endo-/lysosomal degradation pathway. Here we summarize evidence that supports the concept that autophagy serves as the cellular default pathway for the degradation of internalized GJs. Furthermore, we highlight and discuss structural criteria that seem required for an alternate degradation via the endo-/lysosomal pathway. PMID:22825714

  17. Gap junction channel. Its aqueous nature as indicated by deuterium oxide effects

    SciTech Connect

    Verselis, V.; Brink, P.R.

    1986-11-01

    The effects of temperature and solvent substitution with deuterium oxide (D2O) on axoplasmic (ga) and gap junctional (gj) conductances were examined in the earthworm septate median giant axon (MGA). The temperature coefficients (Q10) for ga and gj were 1.4 and 1.5, respectively, between 5 and 15 degrees C. Substitution with D/sub 2/O rapidly reduced both ga and gj by 20% and increased the Q10's to 1.5 and 1.8, respectively. The reduction in ga upon substitution with D/sub 2/O and with cooling in either solvent reflects the changes that occur in solvent viscosity, which indicates that ion mobility in axoplasm, as in free solution, is primarily governed by viscous properties of the solvent. The similar initial reduction observed for gj suggests that solvent occupies the gap junction channel volume and influences transjunctional ion mobility. With time there was a further reduction in gj at 20 degrees C and a larger Q10 in D/sub 2/O. The enhanced effects of D/sub 2/O on gj cannot be accounted for by solvent viscosity alone and may be due to an increased hydration of the channels and/or the transport ions and by isotope effects of hydrogen-deuterium exchange on the channel protein that reduce gj.

  18. The gap junctional protein INX-14 functions in oocyte precursors to promote C. elegans sperm guidance

    PubMed Central

    Edmonds, Johnathan W.; McKinney, Shauna L.; Prasain, Jeevan K.; Miller, Michael A.

    2011-01-01

    Innexins are the subunits of invertebrate gap junctions. Here we show that the innexin INX-14 promotes sperm guidance to the fertilization site in the C. elegans hermaphrodite reproductive tract. inx-14 loss causes cell nonautonomous defects in sperm migration velocity and directional velocity. Results from genetic and immunocytochemical analyses provide strong evidence that INX-14 acts in transcriptionally active oocyte precursors in the distal gonad, not in transcriptionally inactive oocytes that synthesize prostaglandin sperm-attracting cues. Somatic gonadal sheath cell interaction is necessary for INX-14 function, likely via INX-8 and INX-9 expressed in sheath cells. However, electron microscopy has not identified gap junctions in oocyte precursors, suggesting that INX-14 acts in a channel-independent manner or INX-14 channels are difficult to document. INX-14 promotes prostaglandin signaling to sperm at a step after F-series prostaglandin synthesis in oocytes. Taken together, our results support the model that INX-14 functions in a somatic gonad/germ cell signaling mechanism essential for sperm function. We propose that this mechanism regulates the transcription of a factor(s) that modulates prostaglandin metabolism, transport, or activity in the reproductive tract. PMID:21889935

  19. Modulatory Effects of Connexin-43 Expression on Gap Junction Intercellular Communications with Mast Cells and Fibroblasts

    PubMed Central

    Pistorio, Ashey L.; Ehrlich, H. Paul

    2011-01-01

    The influence of mast cells upon aberrant wound repair and excessive fibrosis has supportive evidence, but the mechanism for these mast cell activities is unclear. It is proposed that heterocellular gap junctional intercellular communication (GJIC) between fibroblasts and mast cells directs some fibroblast activities. An in vitro model was used employing a rodent derived peritoneal mast cell line (RMC-1) and human dermal derived fibroblasts. The influence of the expression of the gap junction channel structural protein, connexin 43 (Cx-43) on heterocellular GJIC, the expression of microtubule β-tubulin and microfilament α smooth muscle actin (SMA) were investigated. The knockdown of Cx-43 by siRNA in RMC-1 cells completely blocked GJIC between RMC-1 cells. SiRNA knockdown of Cx-43 within fibroblasts only dampened GJIC between fibroblasts. It appears Cx-43 is the only expressed connexin in RMC-1 cells. Fibroblasts express other connexins that participate in GJIC between fibroblasts in the absence of Cx-43 expression. Heterocellular GJIC between RMC-1 cells and fibroblasts transformed fibroblasts into myofibroblasts, expressing α SMA within cytoplasmic stress fibers. The knockdown of Cx-43 in RMC-1 cells increased β-tubulin expression, but its knockdown in fibroblasts reduced β-tubulin expression. Knocking down the expression of Cx-43 in fibroblasts limited α SMA expression. Cx-43 participation is critical for heterocellular GJIC between mast cells and fibroblasts, which may herald a novel direction for controlling fibrosis. PMID:21328609

  20. Spatial properties of astrocyte gap junction coupling in the rat hippocampus

    PubMed Central

    Anders, Stefanie; Minge, Daniel; Griemsmann, Stephanie; Herde, Michel K.; Steinhäuser, Christian; Henneberger, Christian

    2014-01-01

    Gap junction coupling enables astrocytes to form large networks. Its strength determines how easily a signalling molecule diffuses through the network and how far a locally initiated signal can spread. Changes of coupling strength are well-documented during development and in response to various stimuli. Precise quantification of coupling is needed for studying such modifications and their functional consequences. We therefore explored spatial properties of astrocyte coupling in a model simulating dye loading of single astrocytes. Dye spread into the astrocyte network could be characterized by a coupling length constant and coupling anisotropy. In experiments, the fluorescent marker Alexa Fluor 594 was used to measure these parameters in CA1 and dentate gyrus of the rat hippocampus. Coupling did not differ between regions but showed a temperature-dependence, partially owing to changes of intracellular diffusivity, detected by measuring coupling length constants but not the more variable cell counts of dye-coupled astrocytes. We further found that coupling is anisotropic depending on distance to the pyramidal cell layer, which correlated with regional differences of astrocyte morphology. This demonstrates that applying these new analytical approaches provides useful quantitative information on gap junction coupling and its heterogeneity. PMID:25225094

  1. Spatial properties of astrocyte gap junction coupling in the rat hippocampus.

    PubMed

    Anders, Stefanie; Minge, Daniel; Griemsmann, Stephanie; Herde, Michel K; Steinhäuser, Christian; Henneberger, Christian

    2014-10-19

    Gap junction coupling enables astrocytes to form large networks. Its strength determines how easily a signalling molecule diffuses through the network and how far a locally initiated signal can spread. Changes of coupling strength are well-documented during development and in response to various stimuli. Precise quantification of coupling is needed for studying such modifications and their functional consequences. We therefore explored spatial properties of astrocyte coupling in a model simulating dye loading of single astrocytes. Dye spread into the astrocyte network could be characterized by a coupling length constant and coupling anisotropy. In experiments, the fluorescent marker Alexa Fluor 594 was used to measure these parameters in CA1 and dentate gyrus of the rat hippocampus. Coupling did not differ between regions but showed a temperature-dependence, partially owing to changes of intracellular diffusivity, detected by measuring coupling length constants but not the more variable cell counts of dye-coupled astrocytes. We further found that coupling is anisotropic depending on distance to the pyramidal cell layer, which correlated with regional differences of astrocyte morphology. This demonstrates that applying these new analytical approaches provides useful quantitative information on gap junction coupling and its heterogeneity. PMID:25225094

  2. Hereditary mucoepithelial dysplasia: a disease apparently of desmosome and gap junction formation.

    PubMed Central

    Witkop, C J; White, J G; King, R A; Dahl, M V; Young, W G; Sauk, J J

    1979-01-01

    A previously unrecognized autosomal dominant syndrome affecting oral, nasal, vaginal, urethral, anal, bladder, and conjunctival mucosa with cataracts, follicular keratosis, nonscarring alopecia, and terminal lung disease is described in a four-generation kindred of German extraction. Severe photophobia, tearing, and nystagmus in infancy heralds the development of keratitis, corneal vascularization, and lens cataracts. Repeated corneal transplants have failed. Red, periorificial mucosal lesions involving the above structures are noted by 1 year of age and may persist throughout life. Chronic rhinorrhea and repeated upper respiratory infections frequently progress to bilateral pneumonia accompanied by loss of hair, diarrhea, occasional melena, enuresis, pyuria, and hematuria. Spontaneous pneumothorax is frequent, terminating in fibrocystic-type lung disease and cor pulmonale. Women have had repeated abnormal vaginal PAP smears. Histologically the mucosal epithelium shows dyshesion, thinning of the epithelial layer, and dyskeratosis. Mucosal PAP smears show lack of epithelial maturation, cytoplasmic vacuoles and inclusions, and individual cell dyskeratosis. Histochemically there is a lack of cornification and keratinization. Ultrastructural studies show lack of keratohyalin granules, a paucity of desmosomes, intercellular accumulations, cytoplasmic vacuolization, and formation of bands and aggregates of filamentous fibers and structures in the cytoplasm resembling desmosomes and gap junctions. The condition is probably a panepithelial cell defect of desmosomal and gap junction structure most prominently affecting mucosal epithelia associated with an increased susceptibility to a variety of adventitious organisms. Images Fig. 2-5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 PMID:484550

  3. Experimental febrile seizures impair interastrocytic gap junction coupling in juvenile mice.

    PubMed

    Khan, Dilaware; Dupper, Alexander; Deshpande, Tushar; Graan, Pierre N E De; Steinhäuser, Christian; Bedner, Peter

    2016-09-01

    Prolonged and focal febrile seizures (FSs) have been associated with the development of temporal lobe epilepsy (TLE), although the underlying mechanism and the contribution of predisposing risk factors are still poorly understood. Using a kainate model of TLE, we previously provided strong evidence that interruption of astrocyte gap junction-mediated intercellular communication represents a crucial event in epileptogenesis. To elucidate this aspect further, we induced seizures in immature mice by hyperthermia (HT) to study the consequences of FSs on the hippocampal astrocytic network. Changes in interastrocytic coupling were assessed by tracer diffusion studies in acute slices from mice 5 days after experimental FS induction. The results reveal that HT-induced FSs cause a pronounced reduction of astrocyte gap junctional coupling in the hippocampus by more than 50%. Western blot analysis indicated that reduced connexin43 protein expression and/or changes in the phosphorylation status account for this astrocyte dysfunction. Remarkably, uncoupling occurred in the absence of neuronal death and reactive gliosis. These data provide a mechanistic link between FSs and the subsequent development of TLE and further strengthen the emerging view that astrocytes have a central role in the pathogenesis of this disorder. © 2016 Wiley Periodicals, Inc. PMID:26931373

  4. Spin-dependent transport properties through gapless graphene-based ferromagnet and gapped graphene-based superconductor junction

    NASA Astrophysics Data System (ADS)

    Hajati, Y.; Zargar Shoushtari, M.; Rashedi, G.

    2012-07-01

    By depositing a superconductor on gapped graphene (graphene grown on SiC substrate), the motion of quasiparticles in this superconductor is explained by the massive Dirac equation. In this paper, we study the spin dependent transport properties of graphene-based ferromagnetic/insulator/superconductor (FIS) junction and graphene-based ferromagnetic/ferromagnetic barrier/superconductor (FFBS) junction in which only the superconducting region is deposited on the gapped graphene and the other graphene regions are gapless. We found that in the graphene-based FIS junction and in the thin barrier approximation, by opening the energy gap in the superconducting region, the charge conductance is an oscillatory function of barrier strength (χG), despite the large Fermi energy mismatch between ferromagnetic and superconductor regions. As an important result, we analytically obtained that for the normal incident of charge carriers, this junction is not totally transparent. This means that the second characteristic of Klein tunneling is not satisfied due to the massive Dirac fermions carrying the current in the superconductor region. For the graphene-based FFBS junction, opening the energy gap causes a phase shift as large as π/2 to appear between the peaks of charge conductance for parallel and antiparallel configuration versus χG. Interestingly, we obtained that by increasing the energy gap in this junction, the magnetoresistance increases and by approaching the energy gap to the Fermi energy of the superconductor, it reaches its maximum value (more than -150%). This characteristic shows that this junction can be a suitable candidate for application in the graphene-based spintronics devices.

  5. Gap junction communication between cells aggregated on a cellulose-coated polystyrene: influence of connexin 43 phosphorylation.

    PubMed

    Faucheux, N; Zahm, J M; Bonnet, N; Legeay, G; Nagel, M D

    2004-06-01

    The appropriate functioning of tissues and organ systems depends on intercellular communication such as gap junctions formed by connexin (Cx) protein channels between adjacent cells. We have previously shown that Swiss 3T3 cells aggregated on hydrophilic cellulose substratum Cuprophan (CU) establish short linear gap junctions composed of Cx 43 in cell surface plaques. This phenomenon seems to depend on the high intracellular cyclic AMP (cAMP) concentration triggered by attachment of the cells to CU. We have now used a cellulose-coated polystyrene inducing the same cell behaviour to analyse the gap junction communication between aggregated cells. The transfer of the dye Lucifer Yellow (LY) between cells showed that cells aggregated on cellulose substratum rapidly (within 90 min) establish functional gap junctions. Inhibitors of cAMP protein kinase (PKI) or protein kinase C (GF109203X) both inhibited the diffusion of LY between neighbouring cells. Western blot analysis showed that this change in permeability was correlated with a decrease in Cx 43 phosphorylation. Thus, cellulose substrata seem to induce cell-cell communication through Cx 43 phosphorylation modulated by PKA and PKC. To understand the mechanisms by which a substratum regulates gap junctional communication is critically important for the emerging fields of tissue engineering and biohybrid devices. PMID:14751734

  6. Virally-expressed connexin26 restores gap junction function in the cochlea of conditional Gjb2 knockout mice

    PubMed Central

    Yu, Qing; Wang, Yunfeng; Chang, Qing; Wang, Jianjun; Gong, Shushen; Li, Huawei; Lin, Xi

    2013-01-01

    Mutations in GJB2, which codes for the gap junction protein connexin26, are the most common causes of human nonsyndromic hereditary deafness. We inoculated modified adeno-associated viral vectors into the scala media of early postnatal conditional Gjb2 knockout mice to drive exogenous connexin26 expression. We found extensive virally-expressed connexin26 in cells lining the scala media, and intercellular gap junction network was re-established in the organ of Corti of mutant mouse cochlea. Widespread ectopic connexin26 expression neither formed ectopic gap junctions nor affected normal hearing thresholds in wild type mice, suggesting that autonomous cellular mechanisms regulate proper membrane trafficking of exogenously-expressed connexin26 and govern the functional manifestation of them. Functional recovery of gap-junction-mediated coupling among the supporting cells was observed. We found that both cell death in the organ of Corti and degeneration of spiral ganglion neurons in the cochlea of mutant mice were substantially reduced, although auditory brainstem responses did not show significant hearing improvement. This is the first report demonstrating that virally-mediated gene therapy restored extensive gap junction intercellular network among cochlear non-sensory cells in vivo. Such a treatment performed at early postnatal stages resulted in a partial rescue of disease phenotypes in the cochlea of the mutant mice. PMID:24225640

  7. Increased incidence of gap junctional coupling between spinal motoneurones following transient blockade of NMDA receptors in neonatal rats

    PubMed Central

    Mentis, George Z; Díaz, Eugenia; Moran, Linda B; Navarrete, Roberto

    2002-01-01

    Neonatal rat motoneurones are electrically coupled via gap junctions and the incidence of this coupling declines during postnatal development. The mechanisms involved in this developmental regulation of gap junctional communication are largely unknown. Here we have studied the role of NMDA receptor-mediated glutamatergic synaptic activity in the regulation of motoneurone coupling. Gap junctional coupling was demonstrated by the presence of graded, short latency depolarising potentials following ventral root stimulation, and by the transfer of the low molecular weight tracer Neurobiotin to neighbouring motoneurones. Sites of close apposition between the somata and/or dendrites of the dye-coupled motoneurones were identified as potential sites of gap junctional coupling. Early postnatal blockade of the NMDA subtype of glutamate receptors using the non-competitive antagonist dizocilpine maleate (MK801) arrested the developmental decrease in electrotonic and dye coupling during the first postnatal week. These results suggest that the postnatal increase in glutamatergic synaptic activity associated with the onset of locomotion promote the loss of gap junctional connections between developing motoneurones. PMID:12411521

  8. Connexin-47 and connexin-32 in gap junctions of oligodendrocyte somata, myelin sheaths, paranodal loops and Schmidt-Lanterman incisures: implications for ionic homeostasis and potassium siphoning.

    PubMed

    Kamasawa, N; Sik, A; Morita, M; Yasumura, T; Davidson, K G V; Nagy, J I; Rash, J E

    2005-01-01

    The subcellular distributions and co-associations of the gap junction-forming proteins connexin 47 and connexin 32 were investigated in oligodendrocytes of adult mouse and rat CNS. By confocal immunofluorescence light microscopy, abundant connexin 47 was co-localized with astrocytic connexin 43 on oligodendrocyte somata, and along myelinated fibers, whereas connexin 32 without connexin 47 was co-localized with contactin-associated protein (caspr) in paranodes. By thin-section transmission electron microscopy, connexin 47 immunolabeling was on the oligodendrocyte side of gap junctions between oligodendrocyte somata and astrocytes. By freeze-fracture replica immunogold labeling, large gap junctions between oligodendrocyte somata and astrocyte processes contained much more connexin 47 than connexin 32. Along surfaces of internodal myelin, connexin 47 was several times as abundant as connexin 32, and in the smallest gap junctions, often occurred without connexin 32. In contrast, connexin 32 was localized without connexin 47 in newly-described autologous gap junctions in Schmidt-Lanterman incisures and between paranodal loops bordering nodes of Ranvier. Thus, connexin 47 in adult rodent CNS is the most abundant connexin in most heterologous oligodendrocyte-to-astrocyte gap junctions, whereas connexin 32 is the predominant if not sole connexin in autologous ("reflexive") oligodendrocyte gap junctions. These results clarify the locations and connexin compositions of heterologous and autologous oligodendrocyte gap junctions, identify autologous gap junctions at paranodes as potential sites for modulating paranodal electrical properties, and reveal connexin 47-containing and connexin 32-containing gap junctions as conduits for long-distance intracellular and intercellular movement of ions and associated osmotic water. The autologous gap junctions may regulate paranodal electrical properties during saltatory conduction. Acting in series and in parallel, autologous and

  9. Effective nonlocal spin injection through low-resistance oxide junctions

    NASA Astrophysics Data System (ADS)

    Cai, Yunjiao; Luo, Yongming; Qin, Chuan; Chen, Shuhan; Wu, Yizheng; Ji, Yi

    2016-05-01

    Many (>40) nonlocal spin valves on the same substrate have been characterized at 6 K and 295 K by using a probe station. Low-resistance oxide junctions (0.2-0.8 Ω) are used to inject spin current into mesoscopic Cu channels. Spin signals exceeding 10 mΩ at 6 K have been consistently observed, indicating efficient spin injection and detection. However, complex switching behavior and possible variations between devices pose a challenge to using a standard fitting method to quantify the spin signals. Two methods are used for quantitative analysis. The range of the effective spin polarizations can be estimated by assuming a reasonable range for the Cu spin diffusion lengths. A nonlocal spin polarization is introduced to evaluate the spin current in the Cu channels.

  10. Effect of several uncouplers of cell-to-cell communication on gap junction morphology in mammalian heart.

    PubMed

    Délèze, J; Hervé, J C

    1983-01-01

    Electrical conduction in sheep Purkinje fibers has been blocked by three different procedures: (I) 1 mM 2-4-dinitrophenol, (II) 3.5 mM n-Heptan-1-ol (heptanol), and (III) treatment by a hypotonic (120 mOsmoles) Ca2+-free solution for half an hour, followed by return to normal conditions. The gap junction morphology was analyzed quantitatively in freeze-fracture replicas and compared in electrically conducting and nonconducting fibers. It is found that the three uncouplers of cell-to-cell conduction induce consistent and statistically significant alterations of the gap junction structure. The investigated morphological criteria: (a) P-face junctional particle diameter, control value 8.18 +/- 0.70 nm (mean +/- SD), (b) P-face junctional particles center-to-center spacing, control value 10.23 +/- 1.57 nm, and (c) E-face pits spacing, control value 9.45 +/- 0.98 nm, are, respectively, decreased to 7.46 +/- 0.62 nm, 9.25 +/- 1.34 nm and 8.67 +/- 1.13 nm in Purkinje fibers with complete conduction blocks. All three gap junctional dimensions are seen to decline progressively with time from the onset of an uncoupling treatment towards stable minima reached in half an hour. The observed morphological transitions appear related to the electrical uncoupling for the following reasons: partial electrical uncoupling results in values of the gap junctional dimensions that are intermediate between those measured in electrically coupled and uncoupled preparations, and the three morphological indices are seen to increase again towards control values very soon after electrical conduction has been re-established. It is concluded that the junctional channels closure on electrical uncoupling correlates with a measurable (-0.72 +/- 0.01 nm, difference of the means +/- SE) decrease of the junctional particle diameters. PMID:6887233

  11. Long-range gap junctional signaling controls oncogene-mediated tumorigenesis in Xenopus laevis embryos

    PubMed Central

    Chernet, Brook T.; Fields, Chris; Levin, Michael

    2015-01-01

    In addition to the immediate microenvironment, long-range signaling may be an important component of cancer. Molecular-genetic analyses have implicated gap junctions—key mediators of cell-cell communication—in carcinogenesis. We recently showed that the resting voltage potential of distant cell groups is a key determinant of metastatic transformation and tumor induction. Here, we show in the Xenopus laevis model that gap junctional communication (GJC) is a modulator of the long-range bioelectric signaling that regulates tumor formation. Genetic disruption of GJC taking place within tumors, within remote host tissues, or between the host and tumors significantly lowers the incidence of tumors induced by KRAS mutations. The most pronounced suppression of tumor incidence was observed upon GJC disruption taking place farther away from oncogene-expressing cells, revealing a role for GJC in distant cells in the control of tumor growth. In contrast, enhanced GJC communication through the overexpression of wild-type connexin Cx26 increased tumor incidence. Our data confirm a role for GJC in tumorigenesis, and reveal that this effect is non-local. Based on these results and on published data on movement of ions through GJs, we present a quantitative model linking the GJC coupling and bioelectrical state of cells to the ability of oncogenes to initiate tumorigenesis. When integrated with data on endogenous bioelectric signaling during left-right patterning, the model predicts differential tumor incidence outcomes depending on the spatial configurations of gap junction paths relative to tumor location and major anatomical body axes. Testing these predictions, we found that the strongest influence of GJ modulation on tumor suppression by hyperpolarization occurred along the embryonic left-right axis. Together, these data reveal new, long-range aspects of cancer control by the host's physiological parameters. PMID:25646081

  12. The beneficial effects of cumulus cells and oocyte-cumulus cell gap junctions depends on oocyte maturation and fertilization methods in mice

    PubMed Central

    Zhou, Cheng-Jie; Wu, Sha-Na; Shen, Jiang-Peng; Wang, Dong-Hui; Kong, Xiang-Wei; Lu, Angeleem; Li, Yan-Jiao; Zhou, Hong-Xia; Zhao, Yue-Fang

    2016-01-01

    Cumulus cells are a group of closely associated granulosa cells that surround and nourish oocytes. Previous studies have shown that cumulus cells contribute to oocyte maturation and fertilization through gap junction communication. However, it is not known how this gap junction signaling affects in vivo versus in vitro maturation of oocytes, and their subsequent fertilization and embryonic development following insemination. Therefore, in our study, we performed mouse oocyte maturation and insemination using in vivo- or in vitro-matured oocyte-cumulus complexes (OCCs, which retain gap junctions between the cumulus cells and the oocytes), in vitro-matured, denuded oocytes co-cultured with cumulus cells (DCs, which lack gap junctions between the cumulus cells and the oocytes), and in vitro-matured, denuded oocytes without cumulus cells (DOs). Using these models, we were able to analyze the effects of gap junction signaling on oocyte maturation, fertilization, and early embryo development. We found that gap junctions were necessary for both in vivo and in vitro oocyte maturation. In addition, for oocytes matured in vivo, the presence of cumulus cells during insemination improved fertilization and blastocyst formation, and this improvement was strengthened by gap junctions. Moreover, for oocytes matured in vitro, the presence of cumulus cells during insemination improved fertilization, but not blastocyst formation, and this improvement was independent of gap junctions. Our results demonstrate, for the first time, that the beneficial effect of gap junction signaling from cumulus cells depends on oocyte maturation and fertilization methods. PMID:26966678

  13. Functional assessment of gap junctions in monolayer and three-dimensional cultures of human tendon cells using fluorescence recovery after photobleaching

    NASA Astrophysics Data System (ADS)

    Kuzma-Kuzniarska, Maria; Yapp, Clarence; Pearson-Jones, Thomas W.; Jones, Andrew K.; Hulley, Philippa A.

    2014-01-01

    Gap junction-mediated intercellular communication influences a variety of cellular activities. In tendons, gap junctions modulate collagen production, are involved in strain-induced cell death, and are involved in the response to mechanical stimulation. The aim of the present study was to investigate gap junction-mediated intercellular communication in healthy human tendon-derived cells using fluorescence recovery after photobleaching (FRAP). The FRAP is a noninvasive technique that allows quantitative measurement of gap junction function in living cells. It is based on diffusion-dependent redistribution of a gap junction-permeable fluorescent dye. Using FRAP, we showed that human tenocytes form functional gap junctions in monolayer and three-dimensional (3-D) collagen I culture. Fluorescently labeled tenocytes following photobleaching rapidly reacquired the fluorescent dye from neighboring cells, while HeLa cells, which do not communicate by gap junctions, remained bleached. Furthermore, both 18 β-glycyrrhetinic acid and carbenoxolone, standard inhibitors of gap junction activity, impaired fluorescence recovery in tendon cells. In both monolayer and 3-D cultures, intercellular communication in isolated cells was significantly decreased when compared with cells forming many cell-to-cell contacts. In this study, we used FRAP as a tool to quantify and experimentally manipulate the function of gap junctions in human tenocytes in both two-dimensional (2-D) and 3-D cultures.

  14. Neuronal gap junction coupling is regulated by glutamate and plays critical role in cell death during neuronal injury.

    PubMed

    Wang, Yongfu; Song, Ji-Hoon; Denisova, Janna V; Park, Won-Mee; Fontes, Joseph D; Belousov, Andrei B

    2012-01-11

    In the mammalian CNS, excessive release of glutamate and overactivation of glutamate receptors are responsible for the secondary (delayed) neuronal death following neuronal injury, including ischemia, traumatic brain injury (TBI), and epilepsy. The coupling of neurons by gap junctions (electrical synapses) increases during neuronal injury. We report here that the ischemic increase in neuronal gap junction coupling is regulated by glutamate via group II metabotropic glutamate receptors (mGluRs). Specifically, using electrotonic coupling, Western blots, and siRNA in the mouse somatosensory cortex in vivo and in vitro, we demonstrate that activation of group II mGluRs increases background levels of neuronal gap junction coupling and expression of connexin 36 (Cx36) (neuronal gap junction protein), and inactivation of group II mGluRs prevents the ischemia-mediated increases in the coupling and Cx36 expression. We also show that the regulation is via cAMP/PKA (cAMP-dependent protein kinase)-dependent signaling and posttranscriptional control of Cx36 expression and that other glutamate receptors are not involved in these regulatory mechanisms. Furthermore, using the analysis of neuronal death, we show that inactivation of group II mGluRs or genetic elimination of Cx36 both dramatically reduce ischemia-mediated neuronal death in vitro and in vivo. Similar results are obtained using in vitro models of TBI and epilepsy. Our results indicate that neuronal gap junction coupling is a critical component of glutamate-dependent neuronal death. They also suggest that causal link among group II mGluR function, neuronal gap junction coupling, and neuronal death has a universal character and operates in different types of neuronal injuries. PMID:22238107

  15. Distribution of the gap junction protein connexin 35 in the central nervous system of developing zebrafish larvae

    PubMed Central

    Jabeen, Shaista; Thirumalai, Vatsala

    2013-01-01

    Gap junctions are membrane specializations that allow the passage of ions and small molecules from one cell to another. In vertebrates, connexins are the protein subunits that assemble to form gap junctional plaques. Connexin-35 (Cx35) is the fish ortholog of mammalian Cx36, which is enriched in the retina and the brain and has been shown to form neuronal gap junctions. As a first step toward understanding the role of neuronal gap junctions in central nervous system (CNS) development, we describe here the distribution of Cx35 in the CNS during zebrafish development. Cx35 expression is first seen at 1 day post fertilization (dpf) along cell boundaries throughout the nervous system. At 2 dpf, Cx35 immunoreactivity appears in commissures and fiber tracts throughout the CNS and along the edges of the tectal neuropil. In the rhombencephalon, the Mauthner neurons and fiber tracts show strong Cx35 immunoreactivity. As the larva develops, the commissures and fiber tracts continue to be immunoreactive for Cx35. In addition, the area of the tectal neuropil stained increases vastly and tectal commissures are visible. Furthermore, at 4–5 dpf, Cx35 is seen in the habenulae, cerebellum and in radial glia lining the rhombencephalic ventricle. This pattern of Cx35 immunoreactivity is stable at least until 15 dpf. To test whether the Cx35 immunoreactivity seen corresponds to functional gap junctional coupling, we documented the number of dye-coupled neurons in the hindbrain. We found several dye-coupled neurons within the reticulospinal network indicating functional gap junctional connectivity in the developing zebrafish brain. PMID:23717264

  16. GnRH Episodic Secretion Is Altered by Pharmacological Blockade of Gap Junctions: Possible Involvement of Glial Cells.

    PubMed

    Pinet-Charvet, Caroline; Geller, Sarah; Desroziers, Elodie; Ottogalli, Monique; Lomet, Didier; Georgelin, Christine; Tillet, Yves; Franceschini, Isabelle; Vaudin, Pascal; Duittoz, Anne

    2016-01-01

    Episodic release of GnRH is essential for reproductive function. In vitro studies have established that this episodic release is an endogenous property of GnRH neurons and that GnRH secretory pulses are associated with synchronization of GnRH neuron activity. The cellular mechanisms by which GnRH neurons synchronize remain largely unknown. There is no clear evidence of physical coupling of GnRH neurons through gap junctions to explain episodic synchronization. However, coupling of glial cells through gap junctions has been shown to regulate neuron activity in their microenvironment. The present study investigated whether glial cell communication through gap junctions plays a role in GnRH neuron activity and secretion in the mouse. Our findings show that Glial Fibrillary Acidic Protein-expressing glial cells located in the median eminence in close vicinity to GnRH fibers expressed Gja1 encoding connexin-43. To study the impact of glial-gap junction coupling on GnRH neuron activity, an in vitro model of primary cultures from mouse embryo nasal placodes was used. In this model, GnRH neurons possess a glial microenvironment and were able to release GnRH in an episodic manner. Our findings show that in vitro glial cells forming the microenvironment of GnRH neurons expressed connexin-43 and displayed functional gap junctions. Pharmacological blockade of the gap junctions with 50 μM 18-α-glycyrrhetinic acid decreased GnRH secretion by reducing pulse frequency and amplitude, suppressed neuronal synchronization and drastically reduced spontaneous electrical activity, all these effects were reversed upon 18-α-glycyrrhetinic acid washout. PMID:26562259

  17. Interleukin-1β Increases Gap Junctional Communication among Synovial Fibroblasts via the Extracellular Signal Regulated Kinase Pathway

    PubMed Central

    Niger, Corinne; Howell, Floyd D.; Stains, Joseph P.

    2009-01-01

    Background information The gap junction protein, connexin43, has been implicated in the etiology of osteoarthritis. Work from others has revealed that the size and number of gap junctions increases in synovial biopsies from patients with osteoarthritis. Further, pharmacologic inhibition of connexin43 function has been shown to reduce interleukin-1β induced metalloproteinase production by synovial fibroblasts in vitro. Results In this study, we examine the link between interleukin-1β and connexin43 function. We demonstrate that treatment of a rabbit synovial fibroblast cell line with interleukin-1β markedly increases connexin43 protein in a dose- and time-dependent manner. The impact on connexin43 protein levels appears to occur post-transcriptionally as mRNA levels are unaffected by interleukin-1β administration. Additionally, we show by fluorescence microscopy that interleukin-1β alters the cellular distribution of connexin43 to cell-cell junctions and is concomitant to a striking increase in gap junction communication. Further, we demonstrate that the increase in connexin43 protein and the associated change in protein localization and gap junction communication following interleukin-1β treatment are dependent upon activation of the extracellular signal regulated kinase signaling cascade. Conclusions These data show that interleukin-1β acts through the extracellular signal regulated kinase signaling cascade to alter the expression and function of connexin43 in synovial fibroblasts. PMID:19656083

  18. Simvastatin Sodium Salt and Fluvastatin Interact with Human Gap Junction Gamma-3 Protein

    PubMed Central

    Marsh, Andrew; Casey-Green, Katherine; Probert, Fay; Withall, David; Mitchell, Daniel A.; Dilly, Suzanne J.; James, Sean; Dimitri, Wade; Ladwa, Sweta R.; Taylor, Paul C.; Singer, Donald R. J.

    2016-01-01

    Finding pleiomorphic targets for drugs allows new indications or warnings for treatment to be identified. As test of concept, we applied a new chemical genomics approach to uncover additional targets for the widely prescribed lipid-lowering pro-drug simvastatin. We used mRNA extracted from internal mammary artery from patients undergoing coronary artery surgery to prepare a viral cardiovascular protein library, using T7 bacteriophage. We then studied interactions of clones of the bacteriophage, each expressing a different cardiovascular polypeptide, with surface-bound simvastatin in 96-well plates. To maximise likelihood of identifying meaningful interactions between simvastatin and vascular peptides, we used a validated photo-immobilisation method to apply a series of different chemical linkers to bind simvastatin so as to present multiple orientations of its constituent components to potential targets. Three rounds of biopanning identified consistent interaction with the clone expressing part of the gene GJC3, which maps to Homo sapiens chromosome 7, and codes for gap junction gamma-3 protein, also known as connexin 30.2/31.3 (mouse connexin Cx29). Further analysis indicated the binding site to be for the N-terminal domain putatively ‘regulating’ connexin hemichannel and gap junction pores. Using immunohistochemistry we found connexin 30.2/31.3 to be present in samples of artery similar to those used to prepare the bacteriophage library. Surface plasmon resonance revealed that a 25 amino acid synthetic peptide representing the discovered N-terminus did not interact with simvastatin lactone, but did bind to the hydrolysed HMG CoA inhibitor, simvastatin acid. This interaction was also seen for fluvastatin. The gap junction blockers carbenoxolone and flufenamic acid also interacted with the same peptide providing insight into potential site of binding. These findings raise key questions about the functional significance of GJC3 transcripts in the vasculature and

  19. Expression of gap junctional connexin proteins in ovine fetal ovaries: Effects of maternal diet

    PubMed Central

    Grazul-Bilska, A.T.; Vonnahme, K.A.; Bilski, J.J.; Borowczyk, E.; Soni, D.; Mikkelson, B.; Johnson, M.L.; Reynolds, L.P.; Redmer, D.A.; Caton, J.S.

    2011-01-01

    Gap junctions have been implicated in the regulation of cellular metabolism and the coordination of cellular functions during growth and differentiation of organs and tissues, and gap junctions play a major role in direct cell-cell communication. Gap junctional channels and connexin (Cx) proteins have been detected in adult ovaries in several species. Furthermore, it has been demonstrated that several environmental factors including maternal diet may affect fetal organ growth and function. To determine if maternal diet impacts expression of connexin (Cx) 26, 32, 37 and 43 in fetal ovaries, sheep were fed a maintenance (M) diet with adequate (A) selenium (Se) or high (H) Se levels from 21 days before breeding to day 132 of pregnancy. From day 50 to 132 of pregnancy (tissue collection day), a portion of the ewes from ASe and HSe groups was fed restricted (R; 60% of M) diet. Sections of fetal ovaries were immunostained for the presence of connexins followed by image analysis. All four connexins were detected, but the distribution pattern differed. Cx26 was immunolocalized in the oocytes from primordial, primary, secondary and antral follicles, in granulosa and theca layers of secondary and antral follicles, stroma and blood vessels; Cx32 was in oocytes, granulosa and theca cells in a portion of antral follicles; Cx37 was on the borders between oocyte and granulosa/cumulus cells of primordial to antral follicles, and in endothelium; and Cx43 was on cellular borders in granulosa and theca layers, and between oocyte and granulosa/cumulus cells of primordial to antral follicles. Maternal diet affected Cx26 and Cx43 expression; Cx26 in granulosa layer of antral follicles was decreased (P<0.01) by HSe in M and R diets, and Cx43 in granulosa layer of primary and granulosa and theca of antral follicles was increased (P<0.05) by M diet with HSe. Thus, connexins may be differentially involved in regulation of fetal ovarian function in sheep. These data emphasize the importance

  20. Cx23, a connexin with only four extracellular-loop cysteines, forms functional gap junction channels and hemichannels

    PubMed Central

    Iovine, M. Kathryn; Gumpert, Anna; Falk, Matthias; Mendelson, Tamra C.

    2008-01-01

    Gap junction channels may be comprised of either connexin or pannexin proteins (innexins and pannexins). Membrane topologies of both families are similar, but sequence similarity is lacking. Recently, connexin-like sequences have been identified in mammalian and zebrafish genomes that have only four conserved cysteines in the extracellular domains (Cx23), a feature of the pannexins. Phylogenetic analyses of the non-canonical “C4” connexins reveal that these sequences are indeed connexins. Functional assays reveal that the Cx23 gap junctions are capable of sharing neurobiotin, and further, that Cx23 connexins form hemichannels in vitro. PMID:18068130

  1. Stochastic Model of Gap Junctions Exhibiting Rectification and Multiple Closed States of Slow Gates.

    PubMed

    Snipas, Mindaugas; Kraujalis, Tadas; Paulauskas, Nerijus; Maciunas, Kestutis; Bukauskas, Feliksas F

    2016-03-29

    Gap-junction (GJ) channels formed from connexin (Cx) proteins provide direct pathways for electrical and metabolic cell-cell communication. Earlier, we developed a stochastic 16-state model (S16SM) of voltage gating of the GJ channel containing two pairs of fast and slow gates, each operating between open (o) and closed (c) states. However, experimental data suggest that gates may in fact contain two or more closed states. We developed a model in which the slow gate operates according to a linear reaction scheme, o↔c1↔c2, where c1 and c2 are initial-closed and deep-closed states that both close the channel fully, whereas the fast gate operates between the open state and the closed state and exhibits a residual conductance. Thus, we developed a stochastic 36-state model (S36SM) of GJ channel gating that is sensitive to transjunctional voltage (Vj). To accelerate simulation and eliminate noise in simulated junctional conductance (gj) records, we transformed an S36SM into a Markov chain 36-state model (MC36SM) of GJ channel gating. This model provides an explanation for well-established experimental data, such as delayed gj recovery after Vj gating, hysteresis of gj-Vj dependence, and the low ratio of functional channels to the total number of GJ channels clustered in junctional plaques, and it has the potential to describe chemically mediated gating, which cannot be reflected using an S16SM. The MC36SM, when combined with global optimization algorithms, can be used for automated estimation of gating parameters including probabilities of c1↔c2 transitions from experimental gj-time and gj-Vj dependencies. PMID:27028642

  2. Conduction abnormalities and ventricular arrhythmogenesis: The roles of sodium channels and gap junctions

    PubMed Central

    Tse, Gary; Yeo, Jie Ming

    2015-01-01

    Ventricular arrhythmias arise from disruptions in the normal orderly sequence of electrical activation and recovery of the heart. They can be categorized into disorders affecting predominantly cellular depolarization or repolarization, or those involving action potential (AP) conduction. This article briefly discusses the factors causing conduction abnormalities in the form of unidirectional conduction block and reduced conduction velocity (CV). It then examines the roles that sodium channels and gap junctions play in AP conduction. Finally, it synthesizes experimental results to illustrate molecular mechanisms of how abnormalities in these proteins contribute to such conduction abnormalities and hence ventricular arrhythmogenesis, in acquired pathologies such as acute ischaemia and heart failure, as well as inherited arrhythmic syndromes. PMID:26839915

  3. Effect of reabsorbed recombination radiation on the saturation current of direct gap p-n junctions

    NASA Technical Reports Server (NTRS)

    Von Roos, O.; Mavromatis, H.

    1984-01-01

    The application of the radiative transfer theory for semiconductors to p-n homojunctions subject to low level injection conditions is discussed. By virtue of the interaction of the radiation field with free carriers across the depletion layer, the saturation current density in Shockley's expression for the diode current is reduced at high doping levels. The reduction, due to self-induced photon generation, is noticeable for n-type material owing to the small electron effective mass in direct band-gap III-V compounds. The effect is insignificant in p-type material. At an equilibrium electron concentration of 2 x 10 to the 18th/cu cm in GaAs, a reduction of the saturation current density by 15 percent is predicted. It is concluded that realistic GaAs p-n junctions possess a finite thickness.

  4. Block of gap junctions eliminates aberrant activity and restores light responses during retinal degeneration.

    PubMed

    Toychiev, Abduqodir H; Ivanova, Elena; Yee, Christopher W; Sagdullaev, Botir T

    2013-08-28

    Retinal degeneration leads to progressive photoreceptor cell death, resulting in vision loss. Subsequently, inner retinal neurons develop aberrant synaptic activity, compounding visual impairment. In retinal ganglion cells, light responses driven by surviving photoreceptors are obscured by elevated levels of aberrant spiking activity. Here, we demonstrate in rd10 mice that targeting disruptive neuronal circuitry with a gap junction antagonist can significantly reduce excessive spiking. This treatment increases the sensitivity of the degenerated retina to light stimuli driven by residual photoreceptors. Additionally, this enhances signal transmission from inner retinal neurons to ganglion cells, potentially allowing the retinal network to preserve the fidelity of signals either from prosthetic electronic devices, or from cells optogenetically modified to transduce light. Thus, targeting maladaptive changes to the retina allows for treatments to use existing neuronal tissue to restore light sensitivity, and to augment existing strategies to replace lost photoreceptors. PMID:23986234

  5. Managing the complexity of communication: regulation of gap junctions by post-translational modification

    PubMed Central

    Axelsen, Lene N.; Calloe, Kirstine; Holstein-Rathlou, Niels-Henrik; Nielsen, Morten S.

    2013-01-01

    Gap junctions are comprised of connexins that form cell-to-cell channels which couple neighboring cells to accommodate the exchange of information. The need for communication does, however, change over time and therefore must be tightly controlled. Although the regulation of connexin protein expression by transcription and translation is of great importance, the trafficking, channel activity and degradation are also under tight control. The function of connexins can be regulated by several post translational modifications, which affect numerous parameters; including number of channels, open probability, single channel conductance or selectivity. The most extensively investigated post translational modifications are phosphorylations, which have been documented in all mammalian connexins. Besides phosphorylations, some connexins are known to be ubiquitinated, SUMOylated, nitrosylated, hydroxylated, acetylated, methylated, and γ-carboxyglutamated. The aim of the present review is to summarize our current knowledge of post translational regulation of the connexin family of proteins. PMID:24155720

  6. Interaction between fractional Josephson vortices in multi-gap superconductor tunnel junctions

    NASA Astrophysics Data System (ADS)

    Kim, Ju H.

    In a long Josephson junction (LJJ) with two-band superconductors, fractionalization of Josephson vortices (fluxons) can occur in the broken time reversal symmetry state when spatial phase textures (i-solitons) are excited. Excitation of i-solitons in each superconductor layer of the junction, arising due to the presence of two condensates and the interband Josephson effect, leads to spatial variation of the critical current density between the superconductor layers. Similar to the situation in a YBa2 Cu3O7 - x superconductor film grain boundary, this spatial dependence of the crtitical current density can self-generate magnetic flux in the insulator layer, resulting in fractional fluxons with large and small fraction of flux quantum. Similar to fluxons in one-band superconductor LJJ, these fractional fluxons are found to interact with each other. The interaction between large and small fractional fluxons determines the size of a fluxon which includes two (one large and one small) fractional fluxons. We discuss the nature of interaction between fractional fluxons and suggest that i-soliton excitations in multi-gap superconductor LJJs may be probed by using magnetic flux measurements.

  7. Gap junctions are selectively associated with interlocking ball-and-sockets but not protrusions in the lens

    PubMed Central

    Biswas, Sondip K.; Lee, Jai Eun; Brako, Lawrence; Jiang, Jean X.

    2010-01-01

    Purpose Ball-and-sockets and protrusions are specialized interlocking membrane domains between lens fibers of all species studied. Ball-and-sockets and protrusions are similar in their shape, size, and surface morphology, and are traditionally believed to play a key role in maintaining fiber-to-fiber stability. Here, we evaluate the hypothesis that ball-and-sockets and protrusions possess important structural and functional differences during fiber cell differentiation and maturation. Methods Intact lenses of leghorn chickens (E7 days to P62 weeks old) and rhesus monkeys (1.5–20 years old) were studied with SEM, freeze-fracture TEM, freeze-fracture immunogold labeling (FRIL), and filipin cytochemistry for membrane cholesterol detection. Results SEM showed that ball-and-sockets were distributed along the long and short sides of hexagonal fiber cells, whereas protrusions were located along the cell corners, from superficial to deep cortical regions in both chicken and monkey lenses. Importantly, by freeze-fracture TEM, we discovered the selective association of gap junctions with all ball-and-sockets examined, but not with protrusions, in both species. In the embryonic chicken lens (E18), the abundant distribution of ball-and-socket gap junctions was regularly found in an approximate zone extending at least 300 μm deep from the equatorial surface of the superficial cortical fibers. Many ball-and-socket gap junctions often protruded deeply into neighboring cells. However, in the mature fibers of monkey lenses, several ball-and-sockets exhibited only partial occupancy of gap junctions with disorganized connexons, possibly due to degradation of gap junctions during fiber maturation and aging. FRIL analysis confirmed that both connexin46 (Cx46) and connexin50 (Cx50) antibodies specifically labeled ball-and-socket gap junctions, but not protrusions. Furthermore, filipin cytochemistry revealed that the ball-and-socket gap junctions contained different amounts of

  8. Pannexin-1 channels show distinct morphology and no gap junction characteristics in mammalian cells.

    PubMed

    Beckmann, Anja; Grissmer, Alexander; Krause, Elmar; Tschernig, Thomas; Meier, Carola

    2016-03-01

    Pannexins (Panx) are proteins with a similar membrane topology to connexins, the integral membrane protein of gap junctions. Panx1 channels are generally of major importance in a large number of system and cellular processes and their function has been thoroughly characterized. In contrast, little is known about channel structure and subcellular distribution. We therefore determine the subcellular localization of Panx1 channels in cultured cells and aim at the identification of channel morphology in vitro. Using freeze-fracture replica immunolabeling on EYFP-Panx1-overexpressing HEK 293 cells, large particles were identified in plasma membranes, which were immunogold-labeled using either GFP or Panx1 antibodies. There was no labeling or particles in the nuclear membranes of these cells, pointing to plasma membrane localization of Panx1-EYFP channels. The assembly of particles was irregular, this being in contrast to the regular pattern of gap junctions. The fact that no counterparts were identified on apposing cells, which would have been indicative of intercellular signaling, supported the idea of Panx1 channels within one membrane. Control cells (transfected with EYFP only, non-transfected) were devoid of both particles and immunogold labeling. Altogether, this study provides the first demonstration of Panx1 channel morphology and assembly in intact cells. The identification of Panx1 channels as large particles within the plasma membrane provides the knowledge required to enable recognition of Panx1 channels in tissues in future studies. Thus, these results open up new avenues for the detailed analysis of the subcellular localization of Panx1 and of its nearest neighbors such as purinergic receptors in vivo. PMID:26386583

  9. Modulated gap junctional intercellular communication as a biomarker of PAH epigenetic toxicity: structure-function relationship.

    PubMed Central

    Upham, B L; Weis, L M; Trosko, J E

    1998-01-01

    Cancer is a multistage multimechanism process involving gene and/or chromosomal mutations (genotoxic events), altered gene expression at the transcriptional, translational, and post-translational levels (epigenetic events), and altered cell survival (proliferation and apoptosis or necrosis), resulting in an imbalance of the organism's homeostasis. Maintenance of the organism's homeostasis depends on the intricate coordination of genetic and metabolic events between cells via extracellular and intercellular communication mechanisms. The release of a quiescent cell, whether normal or premalignant, from the suppressing effects of communicating neighbors requires the downregulation of intercellular communication via gap junctions, thereby allowing factors that control intracellular events to exceed a critical mass necessary for the cell to either proliferate or undergo apoptosis. Therefore, determining the role an environmental pollutant must play in the multistage carcinogenic process includes mechanisms of epigenetic toxicity such as the effects of a compound on gap junctional intercellular communication (GJIC). A classic example of a class of compounds in which determination of carcinogenicity focused on genotoxic events and ignored epigenetic events is polycyclic aromatic hydrocarbons (PAHs). The study of structure-activity relationships of PAHs has focused exclusively on the genotoxic and tumor-initiating properties of the compound. We report on the structure-activity relationships of two- to four-ringed PAHs on GJIC in a rat liver epithelial cell line. PAHs containing a bay or baylike region were more potent inhibitors of GJIC than the linear PAHs that do not contain these regions. These are some of the first studies of determine the epigenetic toxicity of PAHs at the epigenetic level. Images Figure 1 PMID:9703481

  10. Radiation induced bystander effect by GAP junction channels in human fibroblast cell

    NASA Astrophysics Data System (ADS)

    Furusawa, Y.; Shao, C.; Aoki, M.; Kobayashi, Y.; Funayama, T.; Ando, K.

    The chemical factor involved in bystander effect and its transfer pathway were investigated in a confluent human fibroblast cell (AG1522) population. Micronuclei (MN) and G1-phase arrest were detected in cells irradiated by carbon (~100 keV/μm) ions at HIMAC. A very low dose irradiation showed a high effectiveness in producing MN, suggesting a bystander effect. This effectiveness was enhanced by 8-Br-cAMP treatment that increases gap junctional intercellular communication (GJIC). On the other hand, the effect was reduced by 5% DMSO treatment, which reduce the reactive oxygen species (ROS), and suppressed by 100 μM lindane treatment, an inhibitor of GJIC. In addition, the radiation-induced G1-phase arrest was also enhanced by cAMP, and reduced or suppressed by DMSO or lindane. A microbeam device (JAERI) was also used for these studies. It was found that exposing one single cell in a confluent cell population to exactly one argon (~1260 keV/μm) or neon (~430 keV/ μm) ion, additional MN could be detected in many other unirradiated cells. The yield of MN increased with the number of irradiated cells. However, there was no significant difference in the MN induction when the cells were irradiated by increasing number of particles. MN induction by bystander effect was partly reduced by DMSO, and effectively suppressed by lindane. Our results obtained from both random irradiation and precise numbered irradiation indicate that both GJIC and ROS contributed to the radiation-induced bystander effect, but the cell gap junction channels likely play an essential role in the release and transfer of radiation-induced chemical factors.

  11. N-cadherin haploinsufficiency affects cardiac gap junctions and arrhythmic susceptibility

    PubMed Central

    Li, Jifen; Levin, Mark D; Xiong, Yanming; Petrenko, Nataliya; Patel, Vickas V.; Radice, Glenn L.

    2008-01-01

    Cardiac-specific deletion of the murine gene (Cdh2) encoding the cell adhesion molecule, N-cadherin, results in disassembly of the intercalated disc (ICD) structure and sudden arrhythmic death. Connexin 43 (Cx43)-containing gap junctions are significantly reduced in the heart after depleting N-cadherin, therefore we hypothesized that animals expressing half the normal levels of N-cadherin would exhibit an intermediate phenotype. We examined the effect of N-cadherin haploinsufficiency on Cx43 expression and susceptibility to induced arrhythmias in mice either wild-type or heterozygous for the Cx43 (Gja1)-null allele. An increase in hypophosphorylated Cx43 accompanied by a modest decrease in total Cx43 protein levels was observed in the N-cadherin heterozygous mice. Consistent with these findings N-cadherin heterozygotes exhibited increased susceptibility to ventricular arrhythmias compared to wild-type mice. Quantitative immunofluorescence microscopy revealed a reduction in size of large Cx43-containing plaques in the N-cadherin heterozygous animals compared to wild-type. Gap junctions were further decreased in number and size in the N-cad/Cx43 compound heterozygous mice with increased arrhythmic susceptibility compared to the single mutants. The scaffold protein, ZO-1, was reduced at the ICD in N-cadherin heterozygous cardiomyocytes providing a possible explanation for the reduction in Cx43 plaque size. These data provide further support for the intimate relationship between N-cadherin and Cx43 in the heart, and suggest that germline mutations in the human N-cadherin (Cdh2) gene may predispose patients to increased risk of cardiac arrhythmias. PMID:18201716

  12. Influence of drugs on gap junctions in glioma cell lines and primary astrocytes in vitro

    PubMed Central

    Moinfar, Zahra; Dambach, Hannes; Faustmann, Pedro M.

    2014-01-01

    Gap junctions (GJs) are hemichannels on cell membrane. Once they are intercellulary connected to the neighboring cells, they build a functional syncytium which allows rapid transfer of ions and molecules between cells. This characteristic makes GJs a potential modulator in proliferation, migration, and development of the cells. So far, several types of GJs are recognized on different brain cells as well as in glioma. Astrocytes, as one of the major cells that maintain neuronal homeostasis, express different types of GJs that let them communicate with neurons, oligodendrocytes, and endothelial cells of the blood brain barrier; however, the main GJ in astrocytes is connexin 43. There are different cerebral diseases in which astrocyte GJs might play a role. Several drugs have been reported to modulate gap junctional communication in the brain which can consequently have beneficial or detrimental effects on the course of treatment in certain diseases. However, the exact cellular mechanism behind those pharmaceutical efficacies on GJs is not well-understood. Accordingly, how specific drugs would affect GJs and what some consequent specific brain diseases would be are the interests of the authors of this chapter. We would focus on pharmaceutical effects on GJs on astrocytes in specific diseases where GJs could possibly play a role including: (1) migraine and a novel therapy for migraine with aura, (2) neuroautoimmune diseases and immunomodulatory drugs in the treatment of demyelinating diseases of the central nervous system such as multiple sclerosis, (3) glioma and antineoplastic and anti-inflammatory agents that are used in treating brain tumors, and (4) epilepsy and anticonvulsants that are widely used for seizures therapy. All of the above-mentioned therapeutic categories can possibly affect GJs expression of astrocytes and the role is discussed in the upcoming chapter. PMID:24904426

  13. Maternal treatment with glucocorticoids modulates gap junction protein expression in the ovine fetal brain.

    PubMed

    Sadowska, G B; Stonestreet, B S

    2014-09-01

    Gap junctions facilitate intercellular communication and are important in brain development. Connexins (Cx) comprise a transmembrane protein family that forms gap junctions. Cx-32 is expressed in oligodendrocytes and neurons, Cx-36 in neurons, and Cx-43 in astrocytes. Although single antenatal steroid courses are recommended for fetal lung maturation, multiple courses can be given to women at recurrent risk for premature delivery. We examined the effects of single and multiple glucocorticoid courses on Cx-32, Cx-36, and Cx-43 protein expressions in the fetal cerebral cortex, cerebellum, and spinal cord, and differences in Cx expression among brain regions under basal conditions. In the single-course groups, the ewes received dexamethasone (6 mg) or placebo as four intramuscular injections every 12h over 48 h. In the multiple-course groups, the ewes received the same treatment, once a week for 5 weeks starting at 76-78 days of gestation. Cx were measured by Western immunoblot on brain samples from 105 to 108-day gestation fetuses. A single dexamethasone course was associated with increases (P<0.05) in cerebral cortical and spinal cord Cx-36 and Cx-43 and multiple courses with increases in cerebellar and spinal cord Cx-36, and cerebral cortical and cerebellar Cx-43. Cx-32 did not change. Cx-32 was higher in the cerebellum than cerebral cortex and spinal cord, Cx-36 higher in the spinal cord than cerebellum, and Cx-43 higher in the cerebellum and spinal cord than cerebral cortex during basal conditions. In conclusion, maternal glucocorticoid therapy increases specific Cx, responses to different maternal courses vary among Cx and brain regions, and Cx expression differs among brain regions under basal conditions. Maternal treatment with glucocorticoids differentially modulates Cx in the fetal brain. PMID:24929069

  14. Phosphorylation on Ser-279 and Ser-282 of connexin43 regulates endocytosis and gap junction assembly in pancreatic cancer cells

    PubMed Central

    Johnson, Kristen E.; Mitra, Shalini; Katoch, Parul; Kelsey, Linda S.; Johnson, Keith R.; Mehta, Parmender P.

    2013-01-01

    The molecular mechanisms regulating the assembly of connexins (Cxs) into gap junctions are poorly understood. Using human pancreatic tumor cell lines BxPC3 and Capan-1, which express Cx26 and Cx43, we show that, upon arrival at the cell surface, the assembly of Cx43 is impaired. Connexin43 fails to assemble, because it is internalized by clathrin-mediated endocytosis. Assembly is restored upon expressing a sorting-motif mutant of Cx43, which does not interact with the AP2 complex, and by expressing mutants that cannot be phosphorylated on Ser-279 and Ser-282. The mutants restore assembly by preventing clathrin-mediated endocytosis of Cx43. Our results also document that the sorting-motif mutant is assembled into gap junctions in cells in which the expression of endogenous Cx43 has been knocked down. Remarkably, Cx43 mutants that cannot be phosphorylated on Ser-279 or Ser-282 are assembled into gap junctions only when connexons are composed of Cx43 forms that can be phosphorylated on these serines and forms in which phosphorylation on these serines is abolished. Based on the subcellular fate of Cx43 in single and contacting cells, our results document that the endocytic itinerary of Cx43 is altered upon cell–cell contact, which causes Cx43 to traffic by EEA1-negative endosomes en route to lysosomes. Our results further show that gap-junctional plaques formed of a sorting motif–deficient mutant of Cx43, which is unable to be internalized by the clathrin-mediated pathway, are predominantly endocytosed in the form of annular junctions. Thus the differential phosphorylation of Cx43 on Ser-279 and Ser-282 is fine-tuned to control Cx43’s endocytosis and assembly into gap junctions. PMID:23363606

  15. Resistance of Single Ag Nanowire Junctions and Their Role in the Conductivity of Nanowire Networks.

    PubMed

    Bellew, Allen T; Manning, Hugh G; Gomes da Rocha, Claudia; Ferreira, Mauro S; Boland, John J

    2015-11-24

    Networks of silver nanowires appear set to replace expensive indium tin oxide as the transparent conducting electrode material in next generation devices. The success of this approach depends on optimizing the material conductivity, which until now has largely focused on minimizing the junction resistance between wires. However, there have been no detailed reports on what the junction resistance is, nor is there a known benchmark for the minimum attainable sheet resistance of an optimized network. In this paper, we present junction resistance measurements of individual silver nanowire junctions, producing for the first time a distribution of junction resistance values and conclusively demonstrating that the junction contribution to the overall resistance can be reduced beyond that of the wires through standard processing techniques. We find that this distribution shows the presence of a small percentage (6%) of high-resistance junctions, and we show how these may impact the performance of network-based materials. Finally, through combining experiment with a rigorous model, we demonstrate the important role played by the network skeleton and the specific connectivity of the network in determining network performance. PMID:26448205

  16. Conductance Measurements of Magnesium Diboride-based Josephson Junctions Below 1 Kelvin: Beyond the 2-Gap Model

    NASA Astrophysics Data System (ADS)

    Carabello, Steven; Lambert, Joseph; Mlack, Jerome; Dai, Wenqing; Li, Qi; Chen, Ke; Cunnane, Daniel; Zhuang, C. G.; Xi, X. X.; Ramos, Roberto

    2014-03-01

    Theoretical and experimental studies have probed the nature of magnesium diboride's two superconducting energy gaps Δπ and Δσ. Several theoretical analyses have predicted fine structures within each energy gap, with recent experiments revealing similar structures. We have performed high-resolution tunneling measurements of low-transparency Josephson junctions using ``terraced,'' ``columnar,'' and c-axis MgB2 films separated by its native oxide from either lead (Pb) or tin (Sn) counter-electrodes. Using high-resolution I-V data at T as low as 23mK, we observe sub-structures within both energy gaps. We also observe sharp peaks in the subgap that identify, to high precision, the energy gap values of the junction counter-electrodes (Pb and Sn). These lead us to conclude that the substructures seen in the gaps are due to MgB2. We then fit the data using simplified two-gap and four-gap models with variable weights and broadening factors. By demonstrating the inadequacy of a simple two-gap model in fitting the data, we illustrate that some distinctions between theoretical models of energy gap substructures are experimentally observable. R.C.R. acknowledges partial support from National Science Foundation Grant # DMR-1206561.

  17. Photoperiod-Dependent Effects of 4-tert-Octylphenol on Adherens and Gap Junction Proteins in Bank Vole Seminiferous Tubules

    PubMed Central

    Kuras, Paulina; Lydka-Zarzycka, Marta; Bilinska, Barbara

    2013-01-01

    In the present study we evaluated in vivo and in vitro effects of 4-tert-octylphenol (OP) on the expression and distribution of adherens and gap junction proteins, N-cadherin, β-catenin, and connexin 43 (Cx43), in testes of seasonally breeding rodents, bank voles. We found that in bank vole testes expression and distribution of N-cadherin, β-catenin, and Cx43 were photoperiod dependent. Long-term treatment with OP (200 mg/kg b.w.) resulted in the reduction of junction proteins expressions (P < 0.05, P < 0.01) and their delocalization in the testes of males kept in long photoperiod, whereas in short-day animals slight increase of Cx43 (P < 0.05), N-cadherin, and β-catenin (statistically nonsignificant) levels was observed. Effects of OP appeared to be independent of FSH and were maintained during in vitro organ culture, indicating that OP acts directly on adherens and gap junction proteins in the testes. An experiment performed using an antiestrogen ICI 182,780 demonstrated that the biological effects of OP on β-catenin and Cx43 involve an estrogen receptor-mediated response. Taken together, in bank vole organization of adherens and gap junctions and their susceptibility to OP are related to the length of photoperiod. Alterations in cadherin/catenin and Cx43-based junction may partially result from activation of estrogen receptor α and/or β signaling pathway. PMID:23737770

  18. Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells

    NASA Technical Reports Server (NTRS)

    Jorgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne; Civitelli, Roberto; Sorensen, Ole Helmer; Steinberg, Thomas H.

    2003-01-01

    The propagation of mechanically induced intercellular calcium waves (ICW) among osteoblastic cells occurs both by activation of P2Y (purinergic) receptors by extracellular nucleotides, resulting in "fast" ICW, and by gap junctional communication in cells that express connexin43 (Cx43), resulting in "slow" ICW. Human osteoblastic cells transmit intercellular calcium signals by both of these mechanisms. In the current studies we have examined the mechanism of slow gap junction-dependent ICW in osteoblastic cells. In ROS rat osteoblastic cells, gap junction-dependent ICW were inhibited by removal of extracellular calcium, plasma membrane depolarization by high extracellular potassium, and the L-type voltage-operated calcium channel inhibitor, nifedipine. In contrast, all these treatments enhanced the spread of P2 receptor-mediated ICW in UMR rat osteoblastic cells. Using UMR cells transfected to express Cx43 (UMR/Cx43) we confirmed that nifedipine sensitivity of ICW required Cx43 expression. In human osteoblastic cells, gap junction-dependent ICW also required activation of L-type calcium channels and influx of extracellular calcium.

  19. Intercellular calcium signaling in a gap junction-coupled cell network establishes asymmetric neuronal fates in C. elegans

    PubMed Central

    Schumacher, Jennifer A.; Hsieh, Yi-Wen; Chen, Shiuhwei; Pirri, Jennifer K.; Alkema, Mark J.; Li, Wen-Hong; Chang, Chieh; Chuang, Chiou-Fen

    2012-01-01

    The C. elegans left and right AWC olfactory neurons specify asymmetric subtypes, one default AWCOFF and one induced AWCON, through a stochastic, coordinated cell signaling event. Intercellular communication between AWCs and non-AWC neurons via a NSY-5 gap junction network coordinates AWC asymmetry. However, the nature of intercellular signaling across the network and how individual non-AWC cells in the network influence AWC asymmetry is not known. Here, we demonstrate that intercellular calcium signaling through the NSY-5 gap junction neural network coordinates a precise 1AWCON/1AWCOFF decision. We show that NSY-5 gap junctions in C. elegans cells mediate small molecule passage. We expressed vertebrate calcium-buffer proteins in groups of cells in the network to reduce intracellular calcium levels, thereby disrupting intercellular communication. We find that calcium in non-AWC cells of the network promotes the AWCON fate, in contrast to the autonomous role of calcium in AWCs to promote the AWCOFF fate. In addition, calcium in specific non-AWCs promotes AWCON side biases through NSY-5 gap junctions. Our results suggest a novel model in which calcium has dual roles within the NSY-5 network: autonomously promoting AWCOFF and non-autonomously promoting AWCON. PMID:23093425

  20. Effect of Mefloquine, a Gap Junction Blocker, on Circadian Period2 Gene Oscillation in the Mouse Suprachiasmatic Nucleus Ex Vivo

    PubMed Central

    Koo, Jinmi; Choe, Han Kyoung; Kim, Hee-Dae; Chun, Sung Kook; Son, Gi Hoon

    2015-01-01

    Background In mammals, the master circadian pacemaker is localized in an area of the ventral hypothalamus known as the suprachiasmatic nucleus (SCN). Previous studies have shown that pacemaker neurons in the SCN are highly coupled to one another, and this coupling is crucial for intrinsic self-sustainability of the SCN central clock, which is distinguished from peripheral oscillators. One plausible mechanism underlying the intercellular communication may involve direct electrical connections mediated by gap junctions. Methods We examined the effect of mefloquine, a neuronal gap junction blocker, on circadian Period 2 (Per2) gene oscillation in SCN slice cultures prepared from Per2::luciferase (PER2::LUC) knock-in mice using a real-time bioluminescence measurement system. Results Administration of mefloquine causes instability in the pulse period and a slight reduction of amplitude in cyclic PER2::LUC expression. Blockade of gap junctions uncouples PER2::LUC-expressing cells, in terms of phase transition, which weakens synchrony among individual cellular rhythms. Conclusion These findings suggest that neuronal gap junctions play an important role in synchronizing the central pacemaker neurons and contribute to the distinct self-sustainability of the SCN master clock. PMID:25491783

  1. Oxidative-Dependent Integration of Signal Transduction with Intercellular Gap Junctional Communication in the Control of Gene Expression

    PubMed Central

    Trosko, James E.

    2009-01-01

    Abstract Research on oxidative stress focused primarily on determining how reactive oxygen species (ROS) damage cells by indiscriminate reactions with their macromolecular machinery, particularly lipids, proteins, and DNA. However, many chronic diseases are not always a consequence of tissue necrosis, DNA, or protein damage, but rather to altered gene expression. Gene expression is highly regulated by the coordination of cell signaling systems that maintain tissue homeostasis. Therefore, much research has shifted to the understanding of how ROS reversibly control gene expression through cell signaling mechanisms. However, most research has focused on redox regulation of signal transduction within a cell, but we introduce a more comprehensive-systems biology approach to understanding oxidative signaling that includes gap junctional intercellular communication, which plays a role in coordinating gene expression between cells of a tissue needed to maintain tissue homeostasis. We propose a hypothesis that gap junctions are critical in modulating the levels of second messengers, such as low molecular weight reactive oxygen, needed in the transduction of an external signal to the nucleus in the expression of genes. Thus, any comprehensive-systems biology approach to understanding oxidative signaling must also include gap junctions, in which aberrant gap junctions have been clearly implicated in many human diseases. Antioxid. Redox Signal. 11, 297–307. PMID:18834329

  2. GAP JUNCTION COMMUNICATION MEDIATES TRANSFORMING GROWTH FACTOR-BETA ACTIVATION AND ENDOTHELIAL-INDUCED MURAL CELL DIFFERENTIATION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During blood vessel assembly, endothelial cells recruit mesenchymal progenitors and induce their differentiation into mural cells via contact-dependent transforming growth factor-beta (TGF-beta) activation. We investigated whether gap junction channels are formed between endothelial cells and recrui...

  3. The B[a]P-increased intercellular communication via translocation of connexin-43 into gap junctions reduces apoptosis

    SciTech Connect

    Tekpli, X.; Rivedal, E.; Gorria, M.; Landvik, N.E.; Rissel, M.; Dimanche-Boitrel, M.-T.; Baffet, G.; Holme, J.A.; Lagadic-Gossmann, D.

    2010-01-15

    Gap junctions are channels in plasma membrane composed of proteins called connexins. These channels are organized in special domains between cells, and provide for direct gap junctional intercellular communication (GJIC), allowing diffusion of signalling molecules < 1 kD. GJIC regulates cell homeostasis and notably the balance between proliferation, cell cycle arrest, cell survival and apoptosis. Here, we have investigated benzo[a]pyrene (B[a]P) effects on GJIC and on the subcellular localization of the major protein of gap junction: connexin-43 (Cx43). Our results showed that B[a]P increased GJIC between mouse hepatoma Hepa1c1c7 cells via translocation of Cx43 from Golgi apparatus and lipid rafts into gap junction plaques. Interestingly, inhibition of GJIC by chlordane or small interference RNA directed against Cx43 enhanced B[a]P-induced apoptosis in Hepa1c1c7 cells. The increased apoptosis caused by inhibition of GJIC appeared to be mediated by ERK/MAPK pathway. It is suggested that B[a]P could induce transfer of cell survival signal or dilute cell death signal via regulation of ERK/MAPK through GJIC.

  4. Gap junctional communication modulates gene transcription by altering the recruitment of Sp1 and Sp3 to connexin-response elements in osteoblast promoters

    NASA Technical Reports Server (NTRS)

    Stains, Joseph P.; Lecanda, Fernando; Screen, Joanne; Towler, Dwight A.; Civitelli, Roberto

    2003-01-01

    Loss-of-function mutations of gap junction proteins, connexins, represent a mechanism of disease in a variety of tissues. We have shown that recessive (gene deletion) or dominant (connexin45 overexpression) disruption of connexin43 function results in osteoblast dysfunction and abnormal expression of osteoblast genes, including down-regulation of osteocalcin transcription. To elucidate the molecular mechanisms of gap junction-sensitive transcriptional regulation, we systematically analyzed the rat osteocalcin promoter for sensitivity to gap junctional intercellular communication. We identified an Sp1/Sp3 containing complex that assembles on a minimal element in the -70 to -57 region of the osteocalcin promoter in a gap junction-dependent manner. This CT-rich connexin-response element is necessary and sufficient to confer gap junction sensitivity to the osteocalcin proximal promoter. Repression of osteocalcin transcription occurs as a result of displacement of the stimulatory Sp1 by the inhibitory Sp3 on the promoter when gap junctional communication is perturbed. Modulation of Sp1/Sp3 recruitment also occurs on the collagen Ialpha1 promoter and translates into gap junction-sensitive transcriptional control of collagen Ialpha1 gene expression. Thus, regulation of Sp1/Sp3 recruitment to the promoter may represent a potential general mechanism for transcriptional control of target genes by signals passing through gap junctions.

  5. Homotypic gap junctional communication associated with metastasis increases suppression increases with PKA kinase activity and is unaffected by P13K inhibition

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Loss of gap junctional intercellular communication (GJIC) between cancer cells is a common characteristic of malignant transformation. This communication is mediated by connexin proteins that make up the functional units of gap junctions. Connexins are highly regulated at the protein level and phosp...

  6. Deficiency of transcription factor Brn4 disrupts cochlear gap junction plaques in a model of DFN3 non-syndromic deafness.

    PubMed

    Kidokoro, Yoshinobu; Karasawa, Keiko; Minowa, Osamu; Sugitani, Yoshinobu; Noda, Tetsuo; Ikeda, Katsuhisa; Kamiya, Kazusaku

    2014-01-01

    Brn4, which encodes a POU transcription factor, is the gene responsible for DFN3, an X chromosome-linked, non-syndromic type of hearing loss. Brn4-deficient mice have a low endocochlear potential (EP), hearing loss, and ultrastructural alterations in spiral ligament fibrocytes, however the molecular pathology through which Brn4 deficiency causes low EP is still unclear. Mutations in the Gjb2 and Gjb6 genes encoding the gap junction proteins connexin26 (Cx26) and connexin30 (Cx30) genes, respectively, which encode gap junction proteins and are expressed in cochlear fibrocytes and non-sensory epithelial cells (i.e., cochlear supporting cells) to maintain the proper EP, are responsible for hereditary sensorineural deafness. It has been hypothesized that the gap junction in the cochlea provides an intercellular passage by which K+ is transported to maintain the EP at the high level necessary for sensory hair cell excitation. Here we analyzed the formation of gap junction plaques in cochlear supporting cells of Brn4-deficient mice at different stages by confocal microscopy and three-dimensional graphic reconstructions. Gap junctions from control mice, which are composed mainly of Cx26 and Cx30, formed linear plaques along the cell-cell junction sites with adjacent cells. These plaques formed pentagonal or hexagonal outlines of the normal inner sulcus cells and border cells. Gap junction plaques in Brn4-deficient mice did not, however, show the normal linear structure but instead formed small spots around the cell-cell junction sites. Gap junction lengths were significantly shorter, and the level of Cx26 and Cx30 was significantly reduced in Brn4-deficient mice compared with littermate controls. Thus the Brn4 mutation affected the assembly and localization of gap junction proteins at the cell borders of cochlear supporting cells, suggesting that Brn4 substantially contributes to cochlear gap junction properties to maintain the proper EP in cochleae, similar to connexin

  7. Engineered Cx40 variants increased docking and function of heterotypic Cx40/Cx43 gap junction channels.

    PubMed

    Jassim, Arjewan; Aoyama, Hiroshi; Ye, Willy G; Chen, Honghong; Bai, Donglin

    2016-01-01

    Gap junction (GJ) channels provide low resistance passages for rapid action potential propagation in the heart. Both connexin40 (Cx40) and Cx43 are abundantly expressed in and frequently co-localized between atrial myocytes, possibly forming heterotypic GJ channels. However, conflicting results have been obtained on the functional status of heterotypic Cx40/Cx43 GJs. Here we provide experimental evidence that the docking and formation of heterotypic Cx40/Cx43 GJs can be substantially increased by designed Cx40 variants on the extracellular domains (E1 and E2). Specifically, Cx40 D55N and P193Q, substantially increased the probability to form GJ plaque-like structures at the cell-cell interfaces with Cx43 in model cells. More importantly the coupling conductance (Gj) of D55N/Cx43 and P193Q/Cx43 GJ channels are significantly increased from the Gj of Cx40/Cx43 in N2A cells. Our homology models indicate the electrostatic interactions and surface structures at the docking interface are key factors preventing Cx40 from docking to Cx43. Improving heterotypic Gj of these atrial connexins might be potentially useful in improving the coupling and synchronization of atrial myocardium. PMID:26625713

  8. Endothelium-Derived Hyperpolarization and Coronary Vasodilation: Diverse and Integrated Roles of Epoxyeicosatrienoic Acids, Hydrogen Peroxide, and Gap Junctions.

    PubMed

    Ellinsworth, David C; Sandow, Shaun L; Shukla, Nilima; Liu, Yanping; Jeremy, Jamie Y; Gutterman, David D

    2016-01-01

    Myocardial perfusion and coronary vascular resistance are regulated by signaling metabolites released from the local myocardium that act either directly on the VSMC or indirectly via stimulation of the endothelium. A prominent mechanism of vasodilation is EDH of the arteriolar smooth muscle, with EETs and H(2)O(2) playing important roles in EDH in the coronary microcirculation. In some cases, EETs and H(2)O(2) are released as transferable hyperpolarizing factors (EDHFs) that act directly on the VSMCs. By contrast, EETs and H(2)O(2) can also promote endothelial KCa activity secondary to the amplification of extracellular Ca(2+) influx and Ca(2+) mobilization from intracellular stores, respectively. The resulting endothelial hyperpolarization may subsequently conduct to the media via myoendothelial gap junctions or potentially lead to the release of a chemically distinct factor(s). Furthermore, in human isolated coronary arterioles dilator signaling involving EETs and H(2)O(2) may be integrated, being either complimentary or inhibitory depending on the stimulus. With an emphasis on the human coronary microcirculation, this review addresses the diverse and integrated mechanisms by which EETs and H(2)O(2) regulate vessel tone and also examines the hypothesis that myoendothelial microdomain signaling facilitates EDH activity in the human heart. PMID:26541094

  9. pH-dependent modulation of connexin-based gap junctional uncouplers

    PubMed Central

    Skeberdis, Vytenis A; Rimkute, Lina; Skeberdyte, Aiste; Paulauskas, Nerijus; Bukauskas, Feliksas F

    2011-01-01

    Abstract Gap junction (GJ) channels formed from connexin (Cx) proteins provide a direct pathway for electrical and metabolic cell–cell communication exhibiting high sensitivity to intracellular pH (pHi). We examined pHi-dependent modulation of junctional conductance (gj) of GJs formed of Cx26, mCx30.2, Cx36, Cx40, Cx43, Cx45, Cx46, Cx47 and Cx50 by reagents representing several distinct groups of uncouplers, such as long carbon chain alkanols (LCCAs), arachidonic acid, carbenoxolone, isoflurane, flufenamic acid and mefloquine. We demonstrate that alkalization by NH4Cl to pH ∼8 increased gj in cells expressing mCx30.2 and Cx45, yet did not affect gj of Cx26, Cx40, Cx46, Cx47 and Cx50 and decreased it in Cx43 and Cx36 GJs. Unexpectedly, cells expressing Cx45, but not other Cxs, exhibited full coupling recovery after alkalization with NH4Cl under the continuous presence of LCCAs, isoflurane and mefloquine. There was no coupling recovery by alkalization in the presence of arachidonic acid, carbenoxolone and flufenamic acid. In cells expressing Cx45, IC50 for octanol was 0.1, 0.25 and 2.68 mm at pHi values of 6.9, 7.2 and 8.1, respectively. Histidine modification of Cx45 protein by N-bromosuccinimide reduced the coupling-promoting effect of NH4Cl as well as the uncoupling effect of octanol. This suggests that LCCAs and some other uncouplers may act through the formation of hydrogen bonds with the as-of-yet unidentified histidine/s of the Cx45 GJ channel protein. PMID:21606109

  10. pH-dependent modulation of connexin-based gap junctional uncouplers.

    PubMed

    Skeberdis, Vytenis A; Rimkute, Lina; Skeberdyte, Aiste; Paulauskas, Nerijus; Bukauskas, Feliksas F

    2011-07-15

    Gap junction (GJ) channels formed from connexin (Cx) proteins provide a direct pathway for electrical and metabolic cell–cell communication exhibiting high sensitivity to intracellular pH (pH(i)). We examined pH(i)-dependent modulation of junctional conductance (g(j)) of GJs formed of Cx26, mCx30.2, Cx36, Cx40, Cx43, Cx45, Cx46, Cx47 and Cx50 by reagents representing several distinct groups of uncouplers, such as long carbon chain alkanols (LCCAs), arachidonic acid, carbenoxolone, isoflurane, flufenamic acid and mefloquine. We demonstrate that alkalization by NH4Cl to pH ∼8 increased g(j) in cells expressing mCx30.2 and Cx45, yet did not affect g(j) of Cx26, Cx40, Cx46, Cx47 and Cx50 and decreased it in Cx43 and Cx36 GJs. Unexpectedly, cells expressing Cx45, but not other Cxs, exhibited full coupling recovery after alkalization with NH4Cl under the continuous presence of LCCAs, isoflurane and mefloquine. There was no coupling recovery by alkalization in the presence of arachidonic acid, carbenoxolone and flufenamic acid. In cells expressing Cx45, IC50 for octanol was 0.1, 0.25 and 2.68 mm at pH(i) values of 6.9, 7.2 and 8.1, respectively. Histidine modification of Cx45 protein by N-bromosuccinimide reduced the coupling-promoting effect of NH4Cl as well as the uncoupling effect of octanol. This suggests that LCCAs and some other uncouplers may act through the formation of hydrogen bonds with the as-of-yet unidentified histidine/s of the Cx45 GJ channel protein. PMID:21606109