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Scott C Steffensen - One of the best experts on this subject based on the ideXlab platform.
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Connexin 36 knock out mice have increased threshold for kindled seizures role of gaba inhibition
Biochemical Pharmacology, 2013Co-Authors: Samuel I Shin, Dallin J Andersen, Micah Hansen D, Jordan T Yorgason, Nathan D Schilaty, David D Busath, Scott C SteffensenAbstract:Blockade of gap junctions (GJs) has been shown to reduce seizures in different epilepsy models. Gap junction mediated, electrically-coupled neuronal networks have been implicated in neuronal synchronization, which is the hallmark of seizure activity. To further understand the role of GJs in seizures, in particular hippocampal seizures, we evaluated electrophysiological responses in the dentate gyrus subfield of the hippocampus, including GABA-mediated recurrent inhibition, seizuregenic stimulation, and seizure activity in response to perforant path kindling in Cx36 knockout (KO) mice compared to wild-type (WT) controls. In anesthetized mice, Cx36 KO mice were characterized by enhanced GABA-mediated recurrent inhibition. Stimulation of the perforant path at 10 Hz for 10 sec (i.e., 100 pulses) markedly enhanced population spike (PS) amplitudes and reduced paired-pulse GABA-mediated inhibition during the stimulation, induced an after discharge for approximately 10 sec at 7-10 sec into the stimulation, and suppressed PS amplitudes postictally for 5-10 min in both KO and WT mice. Repeated epochs of 10 Hz for 10 sec stimulation of the perforant path at 20 min intervals resulted in progressive and persistent disinhibition of paired-pulse responses in WT, but not KO mice. In freely-behaving seizure studies, stimulation of the perforant path (10 Hz for 10 sec) once/day resulted in progressive Stage I-IV seizures in both WT and KO mice. Once Stage IV was achieved, another Stage IV seizure could be elicited each day with the same behavioral response (i.e., Stage V). The threshold for kindled seizures was significantly greater in KO mice compared to WT controls for most stages of seizures. The Cx36 antagonist mefloquine (MFQ) and the typical anticonvulsant pentobarbital reduced Stage IV seizures in both WT and KO mice. Knock-out mice were more sensitive to the anti-epileptic effects of the typical anti-convulsant pentobarbital (20 mg/ kg). Taken together, these findings support the emerging view that reduction in GJ-mediated GABA electrical coupling reduces seizures, perhaps through enhancement of GABAergic feedback inhibition, which results from the uncoupling of the GABA recurrent interneurons from the resistive load that is inherent in their electrical connectivity.
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the role of Connexin 36 gap junctions in alcohol intoxication and consumption
Synapse, 2011Co-Authors: Scott C Steffensen, Kathryn Diane Bradley, David M Hansen, Jeffrey D Wilcox, Rebecca S Wilcox, David W Allison, Collin B Merrill, Jeffrey G EdwardsAbstract:Ventral tegmental area (VTA) GABA neurons appear to be critical substrates underlying the acute and chronic effects of ethanol on dopamine (DA) neurotransmission in the mesocorticolimbic system implicated in alcohol reward. The aim of this study was to examine the role of midbrain Connexin-36 (Cx36) gap junctions (GJs) in ethanol's rewarding effects. Using behavioral, molecular and electrophysiological methods we compared the effects of ethanol in mature Cx36 knockout (KO) mice and age-matched wild-type (WT) controls. Cx36 KO mice exhibited significantly more ethanol-induced ataxia in the open field test, but less disruption in motor coordination than their WT controls in the rotarod paradigm. Cx36 KO mice and WT mice treated with the Cx36 antagonist mefloquine (MFQ) consumed significantly less ethanol than their vehicle-treated WT controls in the drink-in-the-dark procedure. The firing rate of VTA GABA neurons in WT mice was inhibited by ethanol with an IC(50) of 0.25 g/kg, while VTA GABA neurons in KO mice were significantly less sensitive to ethanol. Dopamine neuron sIPSC frequency was reduced by ethanol (30 mM) in WT mice, but not affected in KO mice. Cx36 KO mice evinced a significant up-regulation in DAT and D2 receptors in the VTA, as assessed by quantitative RT-PCR. These findings demonstrate the behavioral relevance of Cx36 GJ-mediated electrical coupling between GABA neurons in mature animals, and suggest that loss of coupling between VTA GABA neurons results in disinhibition of DA neurons, a hyper-DAergic state and lowered hedonic valence for ethanol consumption. Synapse, 2010. © 2010 Wiley-Liss, Inc. Language: en
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mefloquine effects on ventral tegmental area dopamine and gaba neuron inhibition a physiologic role for Connexin 36 gap junctions
Synapse, 2011Co-Authors: David W Allison, David M Hansen, Jeffrey D Wilcox, Rebecca S Wilcox, Kyle L Ellefsen, Caitlin E Askew, Stephanie S Sandoval, Dennis L Eggett, Yuchio Yanagawa, Scott C SteffensenAbstract:Connexin-36 (Cx36) gap junctions (GJs) appear to be involved in the synchronization of GABA interneurons in many brain areas. We have previously identified a population of Cx36-connected ventral tegmental area (VTA) GABA neurons that may regulate mesolimbic dopamine (DA) neurotransmission, a system implicated in reward from both natural behaviors and drugs of abuse. The aim of this study was to determine the effect mefloquine (MFQ) has on midbrain DA and GABA neuron inhibition, and the role Cx36 GJs play in regulating midbrain VTA DA neuron activity in mice. In brain slices from adolescent wild-type (WT) mice the Cx36-selective GJ blocker mefloquine (MFQ, 25 μM) increased VTA DA neuron sIPSC frequency sixfold, and mIPSC frequency threefold. However, in Cx36 KO mice, MFQ only increased sIPSC and mIPSC frequency threefold. The nonselective GJ blocker carbenoxolone (CBX, 100 μM) increased DA neuron sIPSC frequency twofold in WT mice, did not affect Cx36 KO mouse sIPSCs, and did not affect mIPSCs in WT or Cx36 KO mice. Interestingly, MFQ had no effect on VTA GABA neuron sIPSC frequency. We also examined MFQ effects on VTA DA neuron firing rate and current-evoked spiking in WT and Cx36 KO mice, and found that MFQ decreased WT DA neuron firing rate and current-evoked spiking, but did not alter these measures in Cx36 KO mice. Taken together these findings suggest that blocking Cx36 GJs increases VTA DA neuron inhibition, and that GJs play in key role in regulating inhibition of VTA DA neurons.
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Connexin 36 gap junctions mediate electrical coupling between ventral tegmental area gaba neurons
Synapse, 2006Co-Authors: David W Allison, Allison J Ohran, Sarah H Stobbs, Manuel Mameli, Fernando C Valenzuela, Sterling N Sudweeks, Steven J Henriksen, Scott C SteffensenAbstract:Communication between neurons in the mammalian brain is primarily through chemical synapses; however, evidence is accumulating in support of electrical synaptic transmission between some neuronal types in the mature nervous system. The authors have recently demonstrated that the gap junction (GJ) blocker quinidine suppresses stimulus-induced and dopamine-evoked coupling of gamma amino butyric acid (GABA) neurons in the ventral tegmental area (VTA) of mature rats (Stobbs et al., 2004). The aim of this study was to evaluate the role of Connexin-36 (Cx36) GJs in mediating electrical coupling between VTA GABA neurons in P50-80 rats in vivo and P25-50 rats in vitro. Single stimulation of the internal capsule (IC) evoked VTA GABA neuron spike couplets in mature rats when activated antidromically, and multiple poststimulus spike discharges (PSDs) when activated with brief high-frequency stimulation of the IC (ICPSDs). The Cx36 GJ blocker mefloquine (30 mg/kg) suppressed VTA GABA neuron ICPSDs in mature freely behaving rats. VTA GABA neurons recorded via whole-cell patch clamp in the midbrain slice preparation of P25-50 rats showed robust expression of Cx36 transcripts when tested with single-cell quantitative reverse transcription polymerase chain reaction. In P50-80 rats, Cx36 protein immunoreactivity was evident in the VTA and surrounding structures. Dye-coupling between VTA neurons was observed following Neurobiotin labeling of VTA GABA neurons, as well as with the fluorochrome Alexa Fluor 488 using real-time video fluorescent microscopy. Thus, mature VTA GABA neurons appear to be connected by electrical synapses via Cx36 GJs, whose coupling is enhanced by corticotegmental input and by dopamine. Synapse 60:20–31, 2006. © 2006 Wiley-Liss, Inc.
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Connexin‐36 gap junctions mediate electrical coupling between ventral tegmental area GABA neurons
Synapse, 2006Co-Authors: David W Allison, Allison J Ohran, Sarah H Stobbs, Manuel Mameli, Sterling N Sudweeks, Steven J Henriksen, C. Fernando Valenzuela, Scott C SteffensenAbstract:Communication between neurons in the mammalian brain is primarily through chemical synapses; however, evidence is accumulating in support of electrical synaptic transmission between some neuronal types in the mature nervous system. The authors have recently demonstrated that the gap junction (GJ) blocker quinidine suppresses stimulus-induced and dopamine-evoked coupling of gamma amino butyric acid (GABA) neurons in the ventral tegmental area (VTA) of mature rats (Stobbs et al., 2004). The aim of this study was to evaluate the role of Connexin-36 (Cx36) GJs in mediating electrical coupling between VTA GABA neurons in P50-80 rats in vivo and P25-50 rats in vitro. Single stimulation of the internal capsule (IC) evoked VTA GABA neuron spike couplets in mature rats when activated antidromically, and multiple poststimulus spike discharges (PSDs) when activated with brief high-frequency stimulation of the IC (ICPSDs). The Cx36 GJ blocker mefloquine (30 mg/kg) suppressed VTA GABA neuron ICPSDs in mature freely behaving rats. VTA GABA neurons recorded via whole-cell patch clamp in the midbrain slice preparation of P25-50 rats showed robust expression of Cx36 transcripts when tested with single-cell quantitative reverse transcription polymerase chain reaction. In P50-80 rats, Cx36 protein immunoreactivity was evident in the VTA and surrounding structures. Dye-coupling between VTA neurons was observed following Neurobiotin labeling of VTA GABA neurons, as well as with the fluorochrome Alexa Fluor 488 using real-time video fluorescent microscopy. Thus, mature VTA GABA neurons appear to be connected by electrical synapses via Cx36 GJs, whose coupling is enhanced by corticotegmental input and by dopamine. Synapse 60:20–31, 2006. © 2006 Wiley-Liss, Inc.
John E Rash - One of the best experts on this subject based on the ideXlab platform.
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mixed electrical chemical synapses in adult rat hippocampus are primarily glutamatergic and coupled by Connexin 36
Frontiers in Neuroanatomy, 2012Co-Authors: Farid Hamzeisichani, Roger D Traub, Kimberly G V Davidson, Thomas Yasumura, William G M Janssen, Susan L Wearne, Rafael Gutierrez, Ole Petter Ottersen, John E RashAbstract:Dendrodendritic electrical signaling via gap junctions is now an accepted feature of neuronal communication in the mammalian brain, whereas axodendritic and axosomatic gap junctions have rarely been described. We present ultrastructural, immunocytochemical, and dye-coupling evidence for “mixed” (electrical/chemical) synapses in adult rat hippocampus on both principal cells and interneurons. Thin-section electron microscopic images of small gap junction-like appositions were found at mossy fiber (MF) terminals on thorny excrescences of CA3 pyramidal neurons (CA3pyr), apparently forming glutamatergic mixed synapses. Lucifer Yellow injected into four weakly-fixed CA3pyr was detected in MF axons that contacted the injected CA3pyr, supporting gap junction-mediated coupling between those two types of principal cells. Freeze-fracture replica immunogold-labeling revealed diverse sizes and morphologies of Connexin36-containing gap junctions throughout hippocampus. Of 20 immunogold-labeled gap junctions, seven were large (328-1140 connexons), three of which were consistent with electrical synapses between interneurons; but nine were at axon terminal synapses, three of which were immediately adjacent to distinctive glutamate receptor-containing postsynaptic densities, forming mixed glutamatergic synapses. Four others were adjacent to small clusters of immunogold-labeled 10-nm E-face intramembrane particles, apparently representing extrasynaptic glutamate receptor particles. Gap junctions also were on spines in stratum lucidum, stratum oriens, dentate gyrus, and hilus, on both interneurons and unidentified neurons. In addition, one putative GABAergic mixed synapse was found in thin section images of a CA3pyr, but none found by immunogold-labeling were at GABAergic mixed synapses, suggesting their rarity. Cx36-containing gap junctions throughout hippocampus suggest the possibility of reciprocal modulation of electrical and chemical signals in diverse hippocampal neurons.
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Mixed Electrical–Chemical Synapses in Adult Rat Hippocampus are Primarily Glutamatergic and Coupled by Connexin-36
Frontiers in Neuroanatomy, 2012Co-Authors: Farid Hamzei-sichani, Roger D Traub, Kimberly G V Davidson, Thomas Yasumura, William G M Janssen, Susan L Wearne, Rafael Gutierrez, Ole Petter Ottersen, John E RashAbstract:Dendrodendritic electrical signaling via gap junctions is now an accepted feature of neuronal communication in the mammalian brain, whereas axodendritic and axosomatic gap junctions have rarely been described. We present ultrastructural, immunocytochemical, and dye-coupling evidence for “mixed” (electrical/chemical) synapses in adult rat hippocampus on both principal cells and interneurons. Thin-section electron microscopic images of small gap junction-like appositions were found at mossy fiber (MF) terminals on thorny excrescences of CA3 pyramidal neurons (CA3pyr), apparently forming glutamatergic mixed synapses. Lucifer Yellow injected into four weakly-fixed CA3pyr was detected in MF axons that contacted the injected CA3pyr, supporting gap junction-mediated coupling between those two types of principal cells. Freeze-fracture replica immunogold-labeling revealed diverse sizes and morphologies of Connexin36-containing gap junctions throughout hippocampus. Of 20 immunogold-labeled gap junctions, seven were large (328-1140 connexons), three of which were consistent with electrical synapses between interneurons; but nine were at axon terminal synapses, three of which were immediately adjacent to distinctive glutamate receptor-containing postsynaptic densities, forming mixed glutamatergic synapses. Four others were adjacent to small clusters of immunogold-labeled 10-nm E-face intramembrane particles, apparently representing extrasynaptic glutamate receptor particles. Gap junctions also were on spines in stratum lucidum, stratum oriens, dentate gyrus, and hilus, on both interneurons and unidentified neurons. In addition, one putative GABAergic mixed synapse was found in thin section images of a CA3pyr, but none found by immunogold-labeling were at GABAergic mixed synapses, suggesting their rarity. Cx36-containing gap junctions throughout hippocampus suggest the possibility of reciprocal modulation of electrical and chemical signals in diverse hippocampal neurons.
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immunogold evidence that neuronal gap junctions in adult rat brain and spinal cord contain Connexin 36 but not Connexin 32 or Connexin 43
Proceedings of the National Academy of Sciences of the United States of America, 2000Co-Authors: John E Rash, Thomas Yasumura, W Staines, D Patel, C S Furman, Gerald L Stelmack, J I NagyAbstract:Physiological and ultrastructural evidence indicates that gap junctions link many classes of neurons in mammalian central nervous system (CNS), allowing direct electrical and metabolic communication. Among at least six gap junction-forming Connexin proteins in adult rat brain, Connexin- (Cx) 32, Cx36, and Cx43 have been reported to occur in neurons. However, no Connexin has been documented at ultrastructurally defined neuronal gap junctions. To address this question directly, freeze-fracture replica immunogold labeling (FRIL) and immunofluorescence (IF) were used to visualize the subcellular and regional localization of Cx36 in rat brain and spinal cord. Three antibodies were generated against different sequences in Cx36. By Western blotting, these antibodies detected protein at 36 and 66 kDa, corresponding to Cx36 monomer and dimer forms, respectively. After double-labeling for Cx36 and Cx43 by FRIL, neuronal gap junctions in inferior olive, spinal cord, and retina were consistently immunogold-labeled for Cx36, but none were labeled for Cx43. Conversely, Cx43 but not Cx36 was detected in astrocyte and ependymocyte gap junctions. In >250 Cx32/Cx43 single- and double-labeled replicas from 10 CNS regions, no neuronal gap junctions were labeled for either Cx32 or Cx43. Instead, Cx32 and Cx43 were restricted to glial gap junctions. By IF, Cx36 labeling was widely distributed in neuropil, including along dendritic processes and within neuronal somata. On the basis of FRIL identification of Cx36 in neuronal gap junctions and IF imaging of Cx36 throughout rat brain and spinal cord, neuronal gap junctions containing Cx36 appear to occur in sufficient density to provide widespread electrical and metabolic coupling in adult CNS.
Andrei B Belousov - One of the best experts on this subject based on the ideXlab platform.
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a potential role for neuronal Connexin 36 in the pathogenesis of amyotrophic lateral sclerosis
Neuroscience Letters, 2018Co-Authors: Andrei B Belousov, Hiroshi Nishimune, Janna V Denisova, Joseph D FontesAbstract:Abstract Neuronal gap junctional protein Connexin 36 (Cx36) contributes to neuronal death following a range of acute brain insults such as ischemia, traumatic brain injury and epilepsy. Whether Cx36 contributes to neuronal death and pathological outcomes in chronic neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS), is not known. We show here that the expression of Cx36 is significantly decreased in lumbar segments of the spinal cord of both human ALS subjects and SOD1G93A mice as compared to healthy human and wild-type mouse controls, respectively. In purified neuronal cultures prepared from the spinal cord of wild-type mice, knockdown of Cx36 reduces neuronal death caused by overexpression of the mutant human SOD1-G93A protein. Taken together, these data suggest a possible contribution of Cx36 to ALS pathogenesis. A perspective for the use of blockers of Cx36 gap junction channels for ALS therapy is discussed.
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Neuronal gap junctions: making and breaking connections during development and injury
Trends in Neurosciences, 2012Co-Authors: Andrei B Belousov, Joseph D FontesAbstract:In the mammalian central nervous system (CNS), coupling of neurons by gap junctions (i.e., electrical synapses) and the expression of the neuronal gap junction protein, Connexin 36 (Cx36), transiently increase during early postnatal development. The levels of both subsequently decline and remain low in the adult, confined to specific subsets of neurons. However, following neuronal injury [such as ischemia, traumatic brain injury (TBI), and epilepsy], the coupling and expression of Cx36 rise. Here we summarize new findings on the mechanisms of regulation of Cx36-containing gap junctions in the developing and mature CNS and following injury. We also review recent studies suggesting various roles for neuronal gap junctions and in particular their role in glutamate-mediated neuronal death.
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regulation of Connexin 36 expression during development
Neuroscience Letters, 2012Co-Authors: Jihoon Song, Yongfu Wang, Joseph D Fontes, Andrei B BelousovAbstract:Abstract In the mammalian CNS, the expression of neuronal gap junction protein, Connexin 36 (Cx36), increases during the first 2 weeks of postnatal development and then decreases during the following 2 weeks. Recently we showed that the developmental increase in Cx36 expression is augmented by chronic (2 weeks) activation of group II metabotropic glutamate receptors (mGluR), prevented by chronic receptor inactivation, and the receptor-dependent increase in Cx36 expression is regulated via transcriptional control of the Cx36 gene activity. We demonstrate here that acute (60 min) activation of group II mGluRs in developing cortical neuronal cultures causes transient increase in Cx36 protein expression with decrease during the following 24 h. However, there is no change in Cx36 mRNA expression. In addition, the data indicate that transient increase in Cx36 expression is due to new protein synthesis. The results suggest that, during development, acute activation of group II mGluRs causes up-regulation of Cx36 via post-transcriptional mechanisms. However, if the receptor activation is sustained, transcriptional activation of the Cx36 gene occurs.
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deletion of neuronal gap junction protein Connexin 36 impairs hippocampal ltp
Neuroscience Letters, 2011Co-Authors: Yongfu Wang, Andrei B BelousovAbstract:In the mammalian CNS, deletion of neuronal gap junction protein, Connexin 36 (Cx36), causes deficiencies in learning and memory. Here we tested whether Cx36 deletion affects the hippocampal long-term potentiation (LTP), which is considered as a cellular model of learning and memory mechanisms. We report that in acute slices of the hippocampal CA1 area, LTP is reduced in Cx36 knockout mice as compared to wild-type mice. Western blot analysis of NMDA receptor subunits indicates a higher NR2A/NR2B ratio in Cx36 knockout mice, indicating that there is shift in the threshold for LTP induction in knockout animals. Data suggest a possibility that learning and memory deficiencies in Cx36 knockout mice are due to deficiencies in LTP mechanisms.
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Neuronal gap junctions are required for NMDA receptor-mediated excitotoxicity: implications in ischemic stroke.
Journal of Neurophysiology, 2010Co-Authors: Yongfu Wang, Joseph D Fontes, Janna V Denisova, Ki Sung Kang, Andrei B BelousovAbstract:N-methyl-d-aspartate receptors (NMDARs) play an important role in cell survival versus cell death decisions during neuronal development, ischemia, trauma, and epilepsy. Coupling of neurons by electrical synapses (gap junctions) is high or increases in neuronal networks during all these conditions. In the developing CNS, neuronal gap junctions are critical for two different types of NMDAR-dependent cell death. However, whether neuronal gap junctions play a role in NMDAR-dependent neuronal death in the mature CNS was not known. Using Fluoro-Jade B staining, we show that a single intraperitoneal administration of NMDA (100 mg/kg) to adult wild-type mice induces neurodegeneration in three forebrain regions, including rostral dentate gyrus. However, the NMDAR-mediated neuronal death is prevented by pharmacological blockade of neuronal gap junctions (with mefloquine, 30 mg/kg) and does not occur in mice lacking neuronal gap junction protein, Connexin 36. Using Western blots, electrophysiology, calcium imaging, and gas chromatography–mass spectrometry in wild-type and Connexin 36 knockout mice, we show that the reduced level of neuronal death in knockout animals is not caused by the reduced expression of NMDARs, activity of NMDARs, or permeability of the blood–brain barrier to NMDA. In wild-type animals, this neuronal death is not caused by upregulation of Connexin 36 by NMDA. Finally, pharmacological and genetic inactivation of neuronal gap junctions in mice also dramatically reduces neuronal death caused by photothrombotic focal cerebral ischemia. The results indicate that neuronal gap junctions are required for NMDAR-dependent excitotoxicity and play a critical role in ischemic neuronal death.
Hannah Monyer - One of the best experts on this subject based on the ideXlab platform.
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Gap Junctions between Interneurons Are Required for Normal Spatial Coding in the Hippocampus and Short-Term Spatial Memory
The Journal of Neuroscience, 2011Co-Authors: Kevin Allen, Elke C Fuchs, Hannah Jaschonek, David M. Bannerman, Hannah MonyerAbstract:Gap junctions containing Connexin 36 electrically couple interneurons in many brain regions and synchronize their activity. We used Connexin-36 knock-out mice ( Cx36 −/−) to study the importance of electrical coupling between interneurons for spatial coding in the hippocampus and for different forms of hippocampus-dependent spatial memory. Recordings in behaving mice revealed that the spatial selectivity of hippocampal pyramidal neurons was reduced and less stable in Cx36 −/− mice. Altered network activity was reflected in slower theta oscillations in the mutants. Temporal coding, assessed by determining the presence and characteristics of theta phase precession, had different dynamics in Cx36 −/− mice compared with controls. At the behavioral level, Cx36 −/− mice displayed impaired short-term spatial memory but normal spatial reference memory. These results highlight the functional role of electrically coupled interneurons for spatial coding and cognition. Moreover, they suggest that the precise spatial selectivity of place cells is not essential for normal performance on spatial tasks assessing associative long-term memory.
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in vivo evidence for the involvement of the carboxy terminal domain in assembling Connexin 36 at the electrical synapse
Molecular and Cellular Neuroscience, 2010Co-Authors: Ingo Helbig, Andrei Rozov, Hannah Monyer, Esther Sammler, Marina Eliava, Alexey P Bolshakov, Roberto Bruzzone, Sheriar G HormuzdiAbstract:Connexin 36 (Cx36)-containing electrical synapses contribute to the timing and amplitude of neural responses in many brain regions. A Cx36-EGFP transgenic was previously generated to facilitate their identification and study. In this study we demonstrate that electrical coupling is normal in transgenic mice expressing Cx36 from the genomic locus and suggest that fluorescent puncta present in brain tissue represent distributed electrical synapses. These qualities emphasize the usefulness of the Cx36-EGFP reporter as a tool for the detailed anatomical characterization of electrical synapses in fixed and living tissue. However, though the fusion protein is able to form gap junctions between Xenopus laevis oocytes it is unable to restore electrical coupling to interneurons in the Cx36-deficient mouse. Further experiments in transgenic tissue and non-neural cell lines reveal impaired transport to the plasma membrane as the possible cause. By analyzing the functional deficits exhibited by the fusion protein in vivo and in vitro, we identify a motif within Cx36 that may interact with other trafficking or scaffold proteins and thereby be responsible for its incorporation into electrical synapses.
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sharp wave like activity in the hippocampus in vitro in mice lacking the gap junction protein Connexin 36
Journal of Neurophysiology, 2003Co-Authors: Isabel Pais, Sheriar G Hormuzdi, Eberhard H Buhl, Miles A Whittington, Hannah Monyer, Roger D Traub, Ian C Wood, Fiona E N LebeauAbstract:Bath application of kainate (100–300 nM) induced a persistent gamma-frequency (30–80 Hz) oscillation that could be recorded in stratum radiatum of the CA3 region in vitro. We have previously described that in knockout mice lacking the gap junction protein Connexin 36 (Cx36KO), γ-frequency oscillations are reduced but still present. We now demonstrate that in the Cx36KO mice, but not in wild-type (WT), large population field excitatory postsynaptic potentials, or sharp wave-burst discharges, also occurred during the on-going γ-frequency oscillation. These spontaneous burst discharges were not seen in WT mice. Burst discharges in the Cx36KO mice occurred with a mean frequency of 0.23 ± 0.11 Hz and were accompanied by a series of fast (approximately 60–115 Hz) population spikes or “ripple” oscillations in many recordings. Intracellular recordings from CA3 pyramidal cells showed that the burst discharges consisted of a depolarizing response and presumed coupling potentials (spikelets) could occasionally be se...
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selective impairment of hippocampal gamma oscillations in Connexin 36 knock out mouse in vivo
The Journal of Neuroscience, 2003Co-Authors: D L Buhl, Sheriar G Hormuzdi, Hannah Monyer, Kenneth D Harris, Gyorgy BuzsakiAbstract:The physiological roles of neuronal gap junctions in the intact brain are not known. The recent generation of the Connexin-36 knock-out (Cx36 KO) mouse has offered a unique opportunity to examine this problem. Recent in vitro recordings in Cx36 KO mice suggested that Cx36 gap junction contributes to various oscillatory patterns in the theta (∼5–10 Hz) and gamma (∼30–80 Hz) frequency ranges and affects certain aspects of high-frequency (>100 Hz) patterns. However, the relevance of these pharmacologically induced patterns to the intact brain is not known. We recorded field potentials and unit activity in the CA1 stratum pyramidale of the hippocampus in the behaving wild-type (WT) and Cx36 KO mice. Fast-field “ripple” oscillations (140–200 Hz) were present in both WT and KO mice and did not differ significantly in power, intraepisode frequency, or probability of occurrence. Thus, fast-field oscillations either may not require electrical synapses or may be mediated by a hitherto unknown class of gap junctions. Theta oscillations, recorded during either wheel running or rapid eye movement sleep, were not different either. However, the power in the gamma frequency band and the magnitude of theta-phase modulation of gamma power were significantly decreased in KO mice compared with WT controls during wheel running. This suggests that Cx36 interneuronal gap junctions selectively contribute to gamma oscillations.
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impaired electrical signaling disrupts gamma frequency oscillations in Connexin 36 deficient mice
Neuron, 2001Co-Authors: Sheriar G Hormuzdi, Isabel Pais, Fiona E N Lebeau, Stephen K Towers, Andrei Rozov, Eberhard H Buhl, Miles A Whittington, Hannah MonyerAbstract:Abstract Neural processing occurs in parallel in distant cortical areas even for simple perceptual tasks. Associated cognitive binding is believed to occur through the interareal synchronization of rhythmic activity in the γ (30–80 Hz) range. Such oscillations arise as an emergent property of the neuronal network and require conventional chemical neurotransmission. To test the potential role of gap junction-mediated electrical signaling in this network property, we generated mice lacking Connexin 36, the major neuronal Connexin. Here we show that the loss of this protein disrupts γ frequency network oscillations in vitro but leaves high frequency (150 Hz) rhythms, which may involve gap junctions between principal cells (Schmitz et al., 2001), unaffected. Thus, specific Connexins differentially deployed throughout cortical networks are likely to regulate different functional aspects of neuronal information processing in the mature brain.
Feliksas F Bukauskas - One of the best experts on this subject based on the ideXlab platform.
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modulation of Connexin 36 gap junction channels by intracellular ph and magnesium ions
Frontiers in Physiology, 2018Co-Authors: Lina Rimkute, Tadas Kraujalis, Mindaugas Snipas, Nicolas Palaciosprado, Vaidas Jotautis, Vytenis Arvydas Skeberdis, Feliksas F BukauskasAbstract:Connexin-36 (Cx36) protein forms gap junction (GJ) channels in pancreatic beta cells and is also the main Cx isoform forming electrical synapses in the adult mammalian brain. Cx36 GJs can be regulated by intracellular pH (pHi) and cytosolic magnesium ion concentration ([Mg2+]i), which can vary significantly under various physiological and pathological conditions. However, the combined effect and relationship of these two factors over Cx36-dependent coupling have not been previously studied in detail. Our experimental results in HeLa cells expressing Cx36 show that changes in both pHi and [Mg2+]i affect junctional conductance (gj) in an interdependent manner; in other words, intracellular acidification cause increase or decay in gj depending on whether [Mg2+]i is high or low, respectively, and intracellular alkalization cause reduction in gj independently of [Mg2+]i. Our experimental and modelling data support the hypothesis that Cx36 GJ channels contain two separate gating mechanisms, and both are differentially sensitive to changes in pHi and [Mg2+]i. Using recombinant Cx36 we found that two glutamate residues in the N-terminus could be partly responsible for the observed interrelated effect of pHi and [Mg2+]i. Single mutations of glutamate at positions 8 and 12 attenuated the stimulatory effect of intracellular acidification at high [Mg2+]i and double mutation reversed stimulatory effect to inhibition. Moreover, Cx36*E8Q lost the initial increase of gj at low [Mg2+]i and double mutation lost the sensitivity to high [Mg2+]i. These results suggest that E8 and E12 are involved in regulation of Cx36 GJ channels by Mg2+ and H+ ions.
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the effect of arachidonic acid on junctional conductance and gating of Connexin 36 gap junction channels and their modulation by n alkanols
Biophysical Journal, 2013Co-Authors: Alina Marandykina, Nicolas Palaciosprado, Lina Rimkutė, Arvydas Skeberdis, Feliksas F BukauskasAbstract:Arachidonic acid (AA) is one of major components of membrane phospholipids and is actively involved in the regulation of junctional conductance (gj). N-alkanols are well known inhibitors of gap junction (GJ) channels and have been shown to reduce neurological tremors. In this study, we examined function of Cx36 GJ channels, which are expressed in neurons and β-cells of pancreas, under control conditions and application of factors modulating concentration of AA in the plasma membrane. We found that in HeLa cells expressing Cx36-EGFP, short carbon chain alkanols (SCCAs), such as pentanol, hexanol and heptanol, increased gj by ∼3-fold. Conversely, long carbon chain alkanols (LCCAs), such as octanol, nonanol and decanol, uncoupled cells fully. We demonstrate that under control conditions only ∼0.003 of Cx36 GJ channels assembled in junctional plaques are functional, and this fraction increases by SCCAs and fatty acid free bovine serum albumin (BSA). BSA increased gj by 1.6-fold with EC50 of 2.3 M, while its modified form 1,2-cyclohexanedione (BSA-CHD), which does not bind AA, was ineffective. Voltage sensitive gating of Cx36 GJs was reduced by SCCA and BSA that explain in part their gj-enhancing effect. The inhibition of Cx36 GJ channels by AA can be rescued by BSA but not by BSA-CHD. MAFP and thapsigargin, inhibitor and activator of AA synthesis via phospholipase A2, increased and reduced gj, respectively. We assume that endogenous AA is one of key factors leading to low functional efficacy of Cx36 GJ channels under control conditions. Furthermore, we suggest that gj-enhancing effect of BSA and MAFP may be related with reduction of AA levels, while SCCAs limit AA's accessibility to its binding site on Cx36.
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functional expression of the murine Connexin 36 gene coding for a neuron specific gap junctional protein
The Journal of Membrane Biology, 2000Co-Authors: Barbara Teubner, Feliksas F Bukauskas, Joachim Degen, Goran Sohl, Martin Guldenagel, E B Trexler, Vytautas Verselis, C I De Zeeuw, Christine A Kozak, Elisabeth PetraschparwezAbstract:The mouse Connexin 36 (Cx36) gene was mapped on chromosome 2 and an identical transcriptional start site was determined in brain and retina on exon I. Rabbit polyclonal antibodies to the presumptive cytoplasmic loop of the Cx36 protein recognized in immunohistochemical analyses Cx36 expression in the retina, olfactory bulb, hippocampus, inferior olive and cerebellum. In olivary neurons strong punctate labeling at dendritic cell contacts and weaker labeling in the cytoplasm of dendrites were shown by immuno electron microscopy. After expression of mouse Cx36 cDNA in human HeLa cells, neurobiotin transfer was increased 1.8-fold and electrical conductance at least 15-fold compared to untransfected HeLa cells. No Lucifer Yellow transfer was detected in either untransfected or Cx36 transfected HeLa cells. Single Cx36 channels in transfected HeLa cells showed a unitary conductance of 14.3 ± 0.8 pS. The sensitivity of Cx36 channels to transjunctional voltage was low in both HeLa-Cx36 cells and Xenopus oocytes expressing mouse Cx36. No increased transfer of neurobiotin was detected in heterotypic gap junctions formed by Cx36 and 9 other Connexins expressed in HeLa cells. Our results suggest that Cx36 channels function as electrical synapses for transmission of electrical and metabolic signals between neurons in the central nervous system.