Gap Junction Coupling Research Articles

Network Architecture of Gap Junctional Coupling among Parallel Processing Channels in the Mammalian Retina.

Gap junctions are ubiquitous throughout the nervous system, mediating critical signal transmission and integration, as well as emergent network properties. In mammalian retina, gap junctions within the Aii amacrine cell-ON cone bipolar cell (CBC) network are essential for night vision, modulation of day vision, and contribute to visual impairment in retinal degenerations, yet neither the extended network topology nor its conservation is well established. Here, we map the network contribution of gap junctions using a high-resolution connectomics dataset of an adult female rabbit retina. Gap junctions are prominent synaptic components of ON CBC classes, constituting 5%-25% of all axonal synaptic contacts. Many of these mediate canonical transfer of rod signals from Aii cells to ON CBCs for night vision, and we find that the uneven distribution of Aii signals to ON CBCs is conserved in rabbit, including one class entirely lacking direct Aii coupling. However, the majority of gap junctions formed by ON CBCs unexpectedly occur between ON CBCs, rather than with Aii cells. Such coupling is extensive, creating an interconnected network with numerous lateral paths both within, and particularly across, these parallel processing streams. Coupling patterns are precise with ON CBCs accepting and rejecting unique combinations of partnerships according to robust rulesets. Coupling specificity extends to both size and spatial topologies, thereby rivaling the synaptic specificity of chemical synapses. These ON CBC coupling motifs dramatically extend the coupled Aii-ON CBC network, with implications for signal flow in both scotopic and photopic retinal networks during visual processing and disease.SIGNIFICANCE STATEMENT Electrical synapses mediated by gap junctions are fundamental components of neural networks. In retina, coupling within the Aii-ON CBC network shapes visual processing in both the scotopic and photopic networks. In retinal degenerations, these same gap junctions mediate oscillatory activity that contributes to visual impairment. Here, we use high-resolution connectomics strategies to identify gap junctions and cellular partnerships. We describe novel, pervasive motifs both within and across classes of ON CBCs that dramatically extend the Aii-ON CBC network. These motifs are highly specific with implications for both signal processing within the retina and therapeutic interventions for blinding conditions. These findings highlight the underappreciated contribution of coupling motifs in retinal circuitry and the necessity of their detection in connectomics studies.

Stress gates an astrocytic energy reservoir to impair synaptic plasticity

Astrocytes support the energy demands of synaptic transmission and plasticity. Enduring changes in synaptic efficacy are highly sensitive to stress, yet whether changes to astrocyte bioenergetic control of synapses contributes to stress-impaired plasticity is unclear. Here we show in mice that stress constrains the shuttling of glucose and lactate through astrocyte networks, creating a barrier for neuronal access to an astrocytic energy reservoir in the hippocampus and neocortex, compromising long-term potentiation. Impairing astrocytic delivery of energy substrates by reducing astrocyte gap junction coupling with dominant negative connexin 43 or by disrupting lactate efflux was sufficient to mimic the effects of stress on long-term potentiation. Furthermore, direct restoration of the astrocyte lactate supply alone rescued stress-impaired synaptic plasticity, which was blocked by inhibiting neural lactate uptake. This gating of synaptic plasticity in stress by astrocytic metabolic networks indicates a broader role of astrocyte bioenergetics in determining how experience-dependent information is controlled.

Differential Contribution of Gap Junctions to the Membrane Properties of ON- and OFF-Bipolar Cells of the Rat Retina

Gap junctions are ubiquitous within the retina, but in general, it remains to be determined whether gap junction coupling between specific cell types is sufficiently strong to mediate functionally relevant coupling via electrical synapses. From ultrastructural, tracer coupling and immunolabeling studies, there is clear evidence for gap junctions between cone bipolar cells, but it is not known if these gap junctions function as electrical synapses. Here, using whole-cell voltage-clamp recording in rat (male and female) retinal slices, we investigated whether the gap junctions of bipolar cells make a measurable contribution to the membrane properties of these cells. We measured the input resistance (RN) of bipolar cells before and after applying meclofenamic acid (MFA) to block gap junctions. In the presence of MFA, RN of ON-cone bipolar cells displayed a clear increase, paralleled by block of the electrical coupling between these cells and AII amacrine cells in recordings of coupled cell pairs. For OFF-cone and rod bipolar cells, RN did not increase in the presence of MFA. The results for rod bipolar cells are consistent with the lack of gap junctions in these cells. However, for OFF-cone bipolar cells, our results suggest that the morphologically identified gap junctions between these cells do not support a junctional conductance that is sufficient to mediate effective electrical coupling. Instead, these junctions might play a role in chemical and/or metabolic coupling between subcellular compartments.

The amino terminal domain plays an important role in transjunctional voltage-dependent gating kinetics of Cx45 gap junctions

Gap junction (GJ) channels formed by Cx45 exist in nodal cells in the heart where the action potential propagation is the slowest. The cellular mechanisms of slow propagation speed (or longer junctional delay) in nodal cells could be a combination of several factors, including lack of voltage-gated sodium channels, smaller cell size, and a lower GJ coupling conductance of Cx45. Compared to other cardiac GJs, Cx45 GJs possess not only the lowest unitary channel conductance, but also the highest extent and the fastest kinetics of the transjunctional voltage-dependent gating (Vj-gating) together with a slow recovery. These unique gating properties could make Cx45 GJs more vulnerable for dynamic uncoupling to a much lower coupling level, especially when junctional delay is lengthened and/or the heart rate is elevated. The molecular mechanisms determining the Vj-gating properties of Cx45 (a connexin belongs to γ group) GJs have not been studied. Previous functional studies on the amino terminal (NT) domain chimeras or point variants of other connexins belong to α or β group showed that their NT domains played an important role in determining their Vj-gating properties. The crystal and cryo-electron microscope structures of homologous connexin GJs showed that the NT domain lines the GJ pore, a position that could serve a role in Vj-sensing and gating. We hypothesize that the residues in the NT domain of Cx45 are important for its Vj-gating properties. Protein sequence alignment of human Cx45 NT domain with the connexins in the α and β groups revealed that the second and the eighth residues in Cx45 are different from most of these connexins. We generated a total of 14 variants on these two residues and studied their ability to form functional GJs and their Vj-gating properties in model cells. Our results revealed an important role of these two residues on fast Vj-gating kinetics and formation of morphological and functional GJ channels. In contrast, no Vj-gating change was observed on a GFP tagged Cx45 at its carboxyl terminus.

Constitutive deletion of astrocytic connexins aggravates kainate-induced epilepsy.

The astroglial gap junctional network formed by connexin (Cx) channels plays a central role in regulating neuronal activity and network synchronization. However, its involvement in the development and progression of epilepsy is not yet understood. Loss of interastrocytic gap junction (GJ) coupling has been observed in the sclerotic hippocampus of patients with mesial temporal lobe epilepsy (MTLE) and in mouse models of MTLE, leading to the suggestion that it plays a causative role in the pathogenesis. To further elucidate this clinically relevant question, we investigated consequences of astrocyte disconnection on the time course and severity of kainate-induced MTLE with hippocampal sclerosis (HS) by comparing mice deficient for astrocytic Cx proteins with wild-type mice (WT). Continuous telemetric EEG recordings and video monitoring performed over a period of 4 weeks after epilepsy induction revealed substantially higher seizure and interictal spike activity during the chronic phase in Cx deficient versus WT mice, while the severity of status epilepticus was not different. Immunohistochemical analysis showed that, despite the elevated chronic seizure activity, astrocyte disconnection did not aggravate the severity of HS. Indeed, the extent of CA1 pyramidal cell loss was similar between the experimental groups, while astrogliosis, granule cell dispersion, angiogenesis, and microglia activation were even reduced in Cx deficient as compared to WT mice. Interestingly, seizure-induced neurogenesis in the adult dentate gyrus was also independent of astrocytic Cxs. Together, our data indicate that constitutive loss of GJ coupling between astrocytes promotes neuronal hyperexcitability and attenuates seizure-induced histopathological outcomes.

Epileptiform Neuronal Discharges Impair Astrocyte Syncytial Isopotentiality in Acute Hippocampal Slices

Astrocyte syncytial isopotentiality is a physiological mechanism resulting from a strong electrical coupling among astrocytes. We have previously shown that syncytial isopotentiality exists as a system-wide feature that coordinates astrocytes into a system for high efficient regulation of brain homeostasis. Neuronal activity is known to regulate gap junction coupling through alteration of extracellular ions and neurotransmitters. However, the extent to which epileptic neuronal activity impairs the syncytial isopotentiality is unknown. Here, the neuronal epileptiform bursts were induced in acute hippocampal slices by removal of Mg2+ (Mg2+ free) from bath solution and inhibition of γ-aminobutyric acid A (GABAA) receptors by 100 µM picrotoxin (PTX). The change in syncytial coupling was monitored by using a K+ free-Na+-containing electrode solution ([Na+]p) in the electrophysiological recording where the substitution of intracellular K+ by Na+ ions dissipates the physiological membrane potential (VM) to ~0 mV in the recorded astrocyte. However, in a syncytial coupled astrocyte, the [Na+]p induced VM loss can be compensated by the coupled astrocytes to a quasi-physiological membrane potential of ~73 mV. After short-term exposure to this experimental epileptic condition, a significant closure of syncytial coupling was indicated by a shift of the quasi-physiological membrane potential to −60 mV, corresponding to a 90% reduction of syncytial coupling strength. Consequently, the closure of syncytial coupling significantly decreased the ability of the syncytium for spatial redistribution of K+ ions. Altogether, our results show that epileptiform neuronal discharges weaken the strength of syncytial coupling and that in turn impairs the capacity of a syncytium for spatial redistribution of K+ ions.

Danegaptide Enhances Astrocyte Gap Junctional Coupling and Reduces Ischemic Reperfusion Brain Injury in Mice.

Ischemic stroke is a complex and devastating event characterized by cell death resulting from a transient or permanent arterial occlusion. Astrocytic connexin43 (Cx43) gap junction (GJ) proteins have been reported to impact neuronal survival in ischemic conditions. Consequently, Cx43 could be a potential target for therapeutic approaches to stroke. We examined the effect of danegaptide (ZP1609), an antiarrhythmic dipeptide that specifically enhances GJ conductance, in two different rodent stroke models. In this study, danegaptide increased astrocytic Cx43 coupling with no significant effects on Cx43 hemichannel activity, in vitro. Using matrix-assisted laser desorption ionization imaging mass spectrometry (MALDI IMS) the presence of danegaptide within brain tissue sections were detected one hour after reperfusion indicating successful transport of the dipeptide across the blood brain barrier. Furthermore, administration of danegaptide in a novel mouse brain ischemia/reperfusion model showed significant decrease in infarct volume. Taken together, this study provides evidence for the therapeutic potential of danegaptide in ischemia/reperfusion stroke.

Purinergic modulation of neuronal gap junction circuits in the CNS of the leech.

Gap junctions (GJs) are widely distributed in brains across the animal kingdom. To visualize the GJ- coupled networks of two major mechanosensory neurons in the ganglia of medicinal leeches, we injected these cells with the GJ-permeable tracer Neurobiotin. When diffusion time was limited to only 30min, tracer coupling was highly variable for both cells, suggesting a possible modulation of GJ permeability. In invertebrates the innexins (homologs of vertebrate pannexins) form the GJs. Because extracellular adenosine triphosphate (ATP) modulates pannexin and leech innexin hemichannel permeability and is released by leech glial cells following injury, we tested the effects of bath application of ATP after the injection of Neurobiotin and observed a significant increase in the number of neurons tracer coupled to the sensory neurons. This effect required the elevation of intracellular Ca2+ and could be produced by bath application of caffeine. Conversely, scavenging endogenous extracellular ATP with the ATPase apyrase decreased the number of coupled cells. ATP also increased electrical conductance and tracer permeability between the bilateral Retzius neurons. This modulatory effect of ATP on GJ coupling was blocked by siRNA knockdown of a P1-like adenosine receptor. Finally, exposure of leech ganglia to extracellular ATP induced a characteristic low frequency (<0.3Hz) rhythmic bursting activity that was roughly synchronous among multiple neurons, a behavior that was significantly attenuated by the GJ blocker octanol. These findings highlight the mediation by ATP of a robust physiological mechanism for modifying neuronal circuits by rapidly recruiting neurons into active networks and entraining synchronized bursting activity.

Uncoupling of the Astrocyte Syncytium Differentially Affects AQP4 Isoforms.

The water channel protein aquaporin-4 (AQP4) and the gap junction forming proteins connexin-43 (Cx43) and connexin-30 (Cx30) are astrocytic proteins critically involved in brain water and ion homeostasis. While AQP4 is mainly involved in water flux across the astrocytic endfeet membranes, astrocytic gap junctions provide syncytial coupling allowing intercellular exchange of water, ions, and other molecules. We have previously shown that mice with targeted deletion of Aqp4 display enhanced gap junctional coupling between astrocytes. Here, we investigate whether uncoupling of the astrocytic syncytium by deletion of the astrocytic connexins Cx43 and Cx30 affects AQP4 membrane localization and expression. By using quantitative immunogold cytochemistry, we show that deletion of astrocytic connexins leads to a substantial reduction of perivascular AQP4, concomitant with a down-regulation of total AQP4 protein and mRNA. Isoform expression analysis shows that while the level of the predominant AQP4 M23 isoform is reduced in Cx43/Cx30 double deficient hippocampal astrocytes, the levels of M1, and the alternative translation AQP4ex isoform protein levels are increased. These findings reveal a complex interdependence between AQP4 and connexins, which are both significantly involved in homeostatic functions and astrogliopathologies.

Adherens Junction Engagement Regulates Functional Patterning of the Cardiac Pacemaker Cell Lineage

Cardiac Pacemaker Cells (CPCs) rhythmically initiate the electrical impulses that drive heart contraction. CPCs display the highest rate of spontaneous depolarization in the heart despite being subjected to inhibitory electrochemical conditions that should theoretically suppress their activity. While several models have been proposed to explain this apparent paradox, the actual molecular mechanisms that allow CPCs to overcome electronic barriers to their function remain largely unexplored. Here we have traced CPC development at single cell resolution and uncovered a series of cytoarchitectural patterning events that are critical for proper pacemaking. Specifically, our data reveal that CPCs dynamically modulate adherens junction (AJ) engagement to control characteristics including surface area, volume, and gap junctional coupling. This allows CPCs to adopt a structural configuration that supports their overall excitability. Thus, our data have identified a previously unrecognized role for local cellular mechanics in patterning critical morphological features that are necessary for CPC electrical activity.

Development of a sexual dimorphism in a central pattern generator driving a rhythmic behavior: The role of glia-mediated potassium buffering in the pacemaker nucleus of the weakly electric fish Apteronotus leptorhynchus.

Central pattern generators play a critical role in the neural control of rhythmic behaviors. One of their characteristic features is the ability to modulate the oscillatory output. An important yet little-studied type of modulation involves the generation of oscillations that are sexually dimorphic in frequency. In the weakly electric fish Apteronotus leptorhynchus, the pacemaker nucleus serves as a central pattern generator that drives the electric organ discharge of the fish in a one-to-one fashion. Males discharge at higher frequencies than females-a sexual dimorphism that develops under the influence of steroid hormones. The two principal neurons that constitute the oscillatory network of the pacemaker nucleus are the pacemaker and relay cells. Whereas the number and size of the pacemaker and relay cells are sexually monomorphic, pronounced sex-dependent differences exist in the morphology, and subcellular properties of astrocytes, which form a syncytium closely associated with these neurons. In females, compared to males, the astrocytic syncytium covers a larger area surrounding the pacemaker and relay cells and exhibits higher levels of expression of connexin-43 expression. The latter indicates a strong gap-junction coupling of the individual cells within the syncytium. It is hypothesized that these sex-specific differences result in an increased capacity for buffering of extracellular potassium ions, thereby lowering the potassium equilibrium potential, which, in turn, leads to a decrease in the oscillation frequency. This hypothesis has received strong support from simulations based on computational models of individual neurons and the whole neural network of the pacemaker nucleus.

Limited contribution of astroglial gap junction coupling to buffering of extracellular K+ in CA1 stratum radiatum.

Astrocytes form large networks, in which individual cells are connected via gap junctions. It is thought that this astroglial gap junction coupling contributes to the buffering of extracellular K+ increases. However, it is largely unknown how the control of extracellular K+ by astroglial gap junction coupling depends on the underlying activity patterns and on the magnitude of extracellular K+ increases. We explored this dependency in acute hippocampal slices (CA1, stratum radiatum) by direct K+ -sensitive microelectrode recordings and acute pharmacological inhibition of gap junctions. K+ transients evoked by synaptic and axonal activity were largely unaffected by acute astroglial uncoupling in slices obtained from young and adult rats. Iontophoretic K+ -application enabled us to generate K+ gradients with defined spatial properties and magnitude. By varying the K+ -iontophoresis position and protocol, we found that acute pharmacological uncoupling increases the amplitude of K+ transients once their initial amplitude exceeded ~10 mM. Our experiments demonstrate that the contribution of gap junction coupling to buffering of extracellular K+ gradients is limited to large and localized K+ increases.

Gap Junction Coupling Shapes the Encoding of Light in the Developing Retina.

Detection of ambient illumination in the developing retina prior to maturation of conventional photoreceptors is mediated by intrinsically photosensitive retinal ganglion cells (ipRGCs) and is critical for driving several physiological processes, including light aversion, pupillary light reflexes, and photoentrainment of circadian rhythms. The strategies by which ipRGCs encode variations in ambient light intensity at these early ages are not known. Using unsupervised clustering of two-photon calcium responses followed by inspection of anatomical features, we found that the population activity of the neonatal retina could be modeled as six functional groups that were composed of mixtures of ipRGC subtypes and non-ipRGC cell types. By combining imaging, whole-cell recording, pharmacology, and anatomical techniques, we found that functional mixing of cell types is mediated in part by gap junction coupling. Together, these data show that both cell-autonomous intrinsic light responses and gap junction coupling among ipRGCs contribute to the proper encoding of light intensity in the developing retina.

How Heterogeneity in Glucokinase and Gap-Junction Coupling Determines the Islet [Ca2+] Response

Understanding how cell subpopulations in a tissue impact overall system function is challenging. There is extensive heterogeneity among insulin-secreting β-cells within islets of Langerhans, including their insulin secretory response and gene expression profile, and this heterogeneity can be altered in diabetes. Several studies have identified variations in nutrient sensing between β-cells, including glucokinase (GK) levels, mitochondrial function, or expression of genes important for glucose metabolism. Subpopulations of β-cells with defined electrical properties can disproportionately influence islet-wide free-calcium activity ([Ca2+]) and insulin secretion via gap-junction electrical coupling. However, it is poorly understood how subpopulations of β-cells with altered glucose metabolism may impact islet function. To address this, we utilized a multicellular computational model of the islet in which a population of cells deficient in GK activity and glucose metabolism was imposed on the islet or in which β-cells were heterogeneous in glucose metabolism and GK kinetics were altered. This included simulating GK gene (GCK) mutations that cause monogenic diabetes. We combined these approaches with experimental models in which gck was genetically deleted in a population of cells or GK was pharmacologically inhibited. In each case, we modulated gap-junction electrical coupling. Both the simulated islet and the experimental system required 30–50% of the cells to have near-normal glucose metabolism, fewer than cells with normal KATP conductance. Below this number, the islet lacked any glucose-stimulated [Ca2+] elevations. In the absence of electrical coupling, the change in [Ca2+] was more gradual. As such, electrical coupling allows a large minority of cells with normal glucose metabolism to promote glucose-stimulated [Ca2+]. If insufficient numbers of cells are present, which we predict can be caused by a subset of GCK mutations that cause monogenic diabetes, electrical coupling exacerbates [Ca2+] suppression. This demonstrates precisely how metabolically heterogeneous β-cell populations interact to impact islet function.

4969Enhanced gap junction coupling organised and terminated acute ventricular fibrillation in ex-vivo perfused hearts

Abstract Background The underlying mechanism of ventricular fibrillation (VF) remains unclear. There are both experimental and clinical data to support the existence of rotational drivers (RDs), though other opposing studies suggest that VF is the result of disorganized myocardial activation. Abnormal electrical coupling between cardiomyocytes through gap junctions (GJ) has been considered an important factor in the genesis and maintenance of VF and pre-treatment with GJ couplers, rotigaptide (RTG), has been shown to reduce VF inducibility. Purpose We hypothesized that the degree of GJ coupling determines the underlying mechanism of VF, and that changes in GJ coupling can shift or modify the predominant mechanism of fibrillation along the spectrum between disorganised activity and organised drivers. We proposed that increased organisation of VF is critical to its termination. Methods Thirty Sprague-Dawley rat hearts were explanted, perfused ex-vivo and acute VF was induced with burst pacing and 30μM pinacidil. Optical mapping of transmembrane potential was performed at baseline and the effects of GJ coupling on VF dynamics were studied in an acute VF model by perfusing with increasing concentrations of a GJ uncoupler; carbenoxolone (0–50μM, CBX, n=10) or a GJ coupling-enhancer; RTG (0–80nM, n=10). A chronic diffuse fibrosis model (n=10) was generated with 4 weeks of in-vivo angiotensin infusion (500nm/kg/min). Fibrillation dynamics were quantified using phase analysis, phase singularity (PS) tracking and our novel method of global fibrillation organisation quantification, frequency dominance index (FDI), which is a power ratio of highest amplitude dominant frequency in the frequency spectrum. Results RTG increased average rotations per RD (Baseline: 2.86±0.10 vs 80nM: 5.66±0.43, p&lt;0.001) whilst CBX caused a reduction (Baseline: 3.77±0.39 vs 50μM: 0.26±0.26, p&lt;0.001). Maximum rotations for a RD increased with RTG (5.4±0.45 vs 48.20±12.32, p&lt;0.001) and decreased with CBX (8.0±1.3 vs 0.3±0.3, p&lt;0.001). Proportion of time PSs were detected in VF increased with RTG (0.44±0.06 vs 0.93±0.02, p&lt;0.001) and decreased with CBX (0.61±0.9 vs 0.03±0.02, p&lt;0.001). RTG reduced meander of longest duration RD (20.6±1.68 vs 11.51±0.77 pixels, p&lt;0.001) for PS &gt;5 rotations. FDI increased with RTG (0.53±0.04 vs 0.78±0.3, p&lt;0.001) and decreased with CBX (0.60±0.05 vs 0.17±0.03, p&lt;0.001). In the diffuse fibrosis group, in comparison to baseline RTG 80nM increased FDI (0.35 vs 0.65, p&lt;0.001) and terminated VF in 40% of hearts. Conclusion The degree of GJ coupling is a key determinant of the underlying mechanism of VF. RTG organised fibrillation and stabilised RDs in a concentration-dependent manner whilst CBX disorganised VF. Enhancing GJ coupling with RTG in diseased hearts with fibrosis can terminate VF and may be a potential therapeutic target in acute VF. Acknowledgement/Funding BHF Programme Grant PG/16/17/32069

All-trans retinoic acid exposure increases connexin 43 expression in cumulus cells and improves embryo development in bovine oocytes.

In developing follicles, cellular coupling within cumulus-oocyte complexes (COCs) creates a functional syncytium allowing for the passage of small molecules. In many species, intercellular coupling between granulosa cells results from the expression of connexin 43 (CX43 or Gja1) and the formation of gap junctional plaques. Previously, our lab has shown that oocytes with a higher developmental potential had higher CX43 expression in their cumulus cells compared with developmentally incompetent oocytes. All-trans retinoic acid (ATRA) has been shown to increase CX43 expression in several different cell types. In this study we investigated the effect of ATRA treatment, during maturation, on CX43 expression and localization in cumulus cells and the developmental competence of bovine oocytes. COCs and granulosa cells exposed to ATRA during maturation had significantly higher CX43 expression and increased gap junctional coupling, respectively. In addition, there was a significant increase in the maturation, cleavage, and blastocyst rates in ATRA treated COCs. Data from these studies suggest that not only can CX43 be used as a biomarker for oocyte health, it can also potentially be manipulated using ATRA to increase the number of oocytes achieving developmental competence.

Targeting Gap Junctions: New Insights into the Treatment of Major Depressive Disorder.

Major depressive disorder (MDD) is a multifactorial chronic and debilitating mood disease with high lifetime prevalence and associated with excess mortality. Treatments for this disease are not effective in all patients showing the need to find new therapeutic targets. This review aims to update our knowledge on the involvement of astroglial gap junctions and hemichannels in MDD and to show how they have become potential targets for the treatment of this pathology. The method applied in this review includes a systematic compilation of the relevant literature. The use of rodent models of depression, gene analysis of hippocampal tissues of MDD patients and post-mortem studies on the brains from MDD patients suggest that astrocytic gap junction dysfunction may be a part of MDD etiologies. Chronic antidepressant treatments of rats, rat cultured cortical astrocytes and human astrocytoma cell lines support the hypothesis that the up-regulation of gap junctional coupling between astrocytes could be an underlying mechanism for the therapeutic effect of antidepressants. However, two recent functional studies suggest that connexin43 hemichannel activity is a part of several antidepressants' mode of action and that astrocyte gap junctional intercellular communication and hemichannels exert different effects on antidepressant drug response. Even if they emerge as new therapeutic targets for new and more active treatments, further studies are needed to decipher the sophisticated and respective role of astrocytic gap junctions and hemichannels in MDD.

Evidence of decreased gap junction coupling between astrocytes and oligodendrocytes in the anterior cingulate cortex of depressed suicides.

Glial dysfunction is a major pathophysiological feature of mood disorders. While altered astrocyte (AS) and oligodendrocyte-lineage (OL) functions have been associated with depression, the crosstalk between these glial cell types has never been assessed in that context. AS are potent regulators of myelination, in part through gap junction (GJ) channels formed by the heterotypic coupling of AS-specific (Cx30 and Cx43) and OL-specific (Cx32 and Cx47) connexins. This study therefore aimed at addressing the integrity of AS/OL coupling in the anterior cingulate cortex (ACC) of depressed suicides. Using immunofluorescence and confocal imaging, we characterized the distribution of Cx30 and mapped its expression onto OL somas, myelinated axons, and brain vasculature in postmortem brain samples from depressed suicides (N = 48) and matched controls (N = 23). Differential gene expression of key components of the GJ nexus was also screened through RNA-sequencing previously generated by our group, and validated by quantitative real-time PCR. We show that Cx30 expression localized onto OL cells and myelinated fibers is decreased in deep cortical layers of the ACC in male-depressed suicides. This effect was associated with decreased expression of OL-specific connexins, as well as the downregulation of major connexin-interacting proteins essential for the scaffolding, trafficking, and function of GJs. These results provide a first evidence of impaired AS/OL GJ-mediated communication in the ACC of individuals with mood disorders. These changes in glial coupling are likely to have significant impact on brain function, and may contribute to the altered OL function previously reported in this brain region.

Abstract 5150: Adipose-derived stem cell disruption of gap junction intercellular communication in obesity-associated endometrial cancer

Abstract Endometrial cancer is the most common gynecologic cancer. It is divided into two major types, a less aggressive endometrioid type and a more aggressive serous type. Recent clinical observations underscore obesity as a major risk factor for endometrial cancer. In this study, we hypothesized that obesity can result in long term epigenetic modifications in the endometrial compartment, leading to the promotion of endometrial cancer. We examined the role of adipose-derived stem cells (ASCs), which are adipocyte progenitors, in the endometrial tumor microenvironment. By staining for ASCs in a tumor tissue microarray, we observed ASC infiltration that was specific to samples from obese patients, correlating with body mass index (R2=0.77, p&amp;lt;0.001). Using a co-culture system to simulate ASC microenvironmental effects and transcriptomic analysis, we showed that exposure of immortalized endometrial epithelial cells (EECs) to ASC secreted factors led to the repression of cell-cell communication pathways, most notably gap junctions and related factors tight junction proteins TJP and PKC genes. This gene repression was associated with induction of DNA Methylation in the promoter of the major GJ gene GJA1 (encoding connexin 43, Cx43) in the ASC-exposed EECs. Importantly, we further demonstrated this DNA hypermethylation in the promoters of the GJA1, TJP2 and PKC genes in primary endometrial tumors from obese patients compared to non-obese patients. We assessed the effects of epigenetic regulation on gap junction intercellular communication (GJIC) and cell-cell interactions using cellular calcein dye transfer assays and atomic force microscopy (for nano-scale assessment of cell-cell adhesion) by treating endometrial cancer cells with a demethylating agent (DAC). DAC treatment resulted in increased level of cell-cell adhesiveness and communication via gap junction coupling. Specific reactivation of GJA1 (Cx43) by expression vector in endometrial cancer cells led to decreased cellular motility. Because we found that PAI-1 is a major adipokine in the ASC secretome, inhibition of PAI-1 in ASC-exposed EECs led to a cell population expression profile with lower GJ expression, based on single-cell PCR studies. Collectively, the data demonstrate multi-scale regulation of cellular communication via paracrine actions and direct cell-cell coupling via gap junctions by epigenetic silencing influenced by ASCs. This leads to disruption of cellular homeostasis and enhanced motility, promoting endometrial cancer in obese patients. Citation Format: Li-Ling Lin, Srikanth Polusani, Guangcun Huang, Chun-Lin Lin, Chiou-Miin Wang, Nicholas Lucio, Mikhail Kolonin, Alexes Daquinag, Pawel Osmulski, Bruce Nicholson, Edward Kost, Tim Huang, Nameer B. Kirma. Adipose-derived stem cell disruption of gap junction intercellular communication in obesity-associated endometrial cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5150.

Synchronization between neural circuits connected by hybrid synapse

Reliable neural circuits provide feasible ways to estimate the information encoding and signal exchange between neurons so that more physical effects can be considered and explained. Electrical synapse benefits signal propagation and neurons can give rapid response selection under an external stimulus. Chemical synapse plays a standing biological function by the continuous release of neurotransmitter. Gap junction coupling can estimate the effect of electrical synapse coupling as resistor-based voltage coupling while chemical synapse coupling shows a complex form in synapse currents. In this paper, two FitzHugh–Nagumo neuron circuits are connected by resistor and induction coil, and thus field coupling is activated to realize synchronization. It gives possible guidance to explain the synapse function from physical view. These circuit equations are obtained and scale transformation is applied to get the dimensionless dynamical systems. Furthermore, the same scheme is used to discuss synchronization realization between Hindmarsh–Rose neuron models. It is found that synchronization can be suppressed when the coupling resistor is in series with the induction coil in which only one coupling channel is open. However, complete synchronization can be enhanced when the coupling resistor is paralleled with the induction coil because two coupling channels are activated simultaneously. As a result, appropriate adjustment and selection for the coupling electric devices can induce transition from synchronization to desynchronization between neural circuits, which indicates the self-adaption of synapse when two types of synapse are activated together.