Gap junctions, formed by gap junction proteins (GJ), play crucial roles in cell signaling and immune responses. The structure and function of the GJ from vertebrates (called connexins) have been extensively studied. However, little is known about the proteins forming gap junctions in invertebrates (called innexins). In this study, 14 GJ genes of Chlamys nobilis were identified. GJ proteins are mainly distributed on the plasma membrane, and all proteins are hydrophilic Phylogenetic tree analysis showed that the GJ proteins in C. nobilis were distantly related to those in vertebrates but closely related to those in invertebrates. Conserved motifs analysis of these GJ proteins in C. nobilis identified to have 10 conserved motifs, similar to gap junction proteins in other bivalves. Moreover, expression profiles of CnGJ genes under chronic and acute low temperature stress were also investigated. Results showed that chronic low temperature stress had a significant effect on the expression levels of CnGJ genes, and the expression profiles of CnGJ genes showed significantly variation under acute low temperature stress. All these results indicated that CnGJ genes play important roles in environmental adaptation in scallops. The present study initially elucidated the function of gap junction genes in noble scallop C. nobilis, which provides new insights into the GJ genes in mollusks and will help us better understand their roles in environmental stress in scallops.

OBJECTIVE: Neuropathic pain is a chronic condition characterized by aberrant signaling within the somatosensory system, affecting millions of people worldwide with limited treatment options. Herein, we aim at investigating the potential of a sigma-1 receptor (σ1R) antagonist in managing neuropathic pain.
METHODS: A Chronic Constriction Injury (CCI) model was used to induce neuropathic pain. The potential of (+)-MR200 was evaluated following daily subcutaneous injections of the compound. Its mechanism of action was confirmed by administration of a well-known σ1R agonist, PRE084.
RESULTS: (+)-MR200 demonstrated efficacy in protecting neurons from damage and alleviating pain hypersensitivity in CCI model. Our results suggest that (+)-MR200 reduced the activation of astrocytes and microglia, cells known to contribute to the neuroinflammatory process, suggesting that (+)-MR200 may not only address pain symptoms but also tackle the underlying cellular mechanism involved. Furthermore, (+)-MR200 treatment normalized levels of the gap junction (GJ)-forming protein connexin 43 (Cx43), suggesting a reduction in harmful intercellular communication that could fuel the chronicity of pain.
CONCLUSIONS: This approach could offer a neuroprotective strategy for managing neuropathic pain, addressing both pain symptoms and cellular processes driving the condition. Understanding the dynamics of σ1R expression and function in neuropathic pain is crucial for clinical intervention.

This study used the brilliant cresyl blue (BCB) staining method to group buffalo oocytes (BCB+ and BCB-) and perform in vitro maturation, in vitro fertilization and embryo culture. At the same time, molecular biology techniques were used to detect gap junction protein expression and oxidative stress-related indicators to explore the molecular mechanism of BCB staining to predict oocyte developmental potential. The techniques of buffalo oocytes to analyse their developmental potential and used immunofluorescence staining to detect the expression level of CX43 protein, DCFH-DA probe staining to detect ROS levels and qPCR to detect the expression levels of the antioxidant-related genes SOD2 and GPX1. Our results showed that the in vitro maturation rate, embryo cleavage rate and blastocyst rate of buffalo oocytes in the BCB+ group were significantly higher than those in the BCB- group and the control group (p < .05). The expression level of CX43 protein in the BCB+ group was higher than that in the BCB- group both before and after maturation (p < .05). The intensity of ROS in the BCB+ group was significantly lower than that in the BCB- group (p < .05), and the expression levels of the antioxidant-related genes SOD2 and GPX1 in the BCB+ group were significantly higher than those in the BCB- group (p < .05). Brilliant cresyl blue staining could effectively predict the developmental potential of buffalo oocytes. The results of BCB staining were positively correlated with the expression of gap junction protein and antioxidant-related genes and negatively correlated with the reactive oxygen species level, suggesting that the mechanism of BCB staining in predicting the developmental potential of buffalo oocytes might be closely related to antioxidant activity.

MicroRNAs (miRNA) are important regulators of oocyte maturation, playing a key role in modulating gene expression both in a temporal- and spatial-specific manner. These small non-coding RNAs are involved in important processes during oocyte maturation, acting as messengers between the oocyte and its surrounding cumulus cells. Despite its significance, the bidirectional communication mechanism is still unknown. To test miRNA communication between oocyte and surrounding cumulus cells through the gap junctions the gap junctions were either blocked with carbenoxolone or not. MiRNA sequencing of oocytes at 1, 6, and 22 h of in vitro maturation was then performed. Among the differentially expressed miRNAs, bta-miR-21-5p, a regulator of cumulus cell viability and oocyte maturation, was the only previously known miRNA. Furthermore, by labeling a bta-miR-21-5p mimic with FAM, crossing of this miRNA through the gap junctions within the cumulus-oocyte complex could be visualized and internalization in the oocyte was confirmed by RT-qPCR. In conclusion, this study provides, for the first time, evidence that miRNA communication within the bovine cumulus-oocyte complex is enabled through the gap junctional network.

Intravascularly transplanted bone marrow cells, including bone marrow mononuclear cells (BM-MNC) and mesenchymal stem cells, transfer water-soluble molecules to cerebral endothelial cells via gap junctions. After transplantation of BM-MNC, this fosters hippocampal neurogenesis and enhancement of neuronal function. Herein, we report the impact of transplanted BM-MNC on neural stem cells (NSC) in the brain. Surprisingly, direct transfer of water-soluble molecules from transplanted BM-MNC and peripheral mononuclear cells to NSC in the hippocampus was observed already 10 min after cell transplantation, and transfer from BM-MNC to GFAP-positive cortical astrocytes was also observed. In vitro investigations revealed that BM-MNC abolish the expression of hypoxia-inducible factor-1α in astrocytes. We suggest that the transient and direct transfer of water-soluble molecules between cells in circulation and NSC in the brain may be one of the biological mechanisms underlying the repair of brain function.

The histochemical Falck-Hillarp method for the localization of dopamine (DA), noradrenaline (NA) and serotonin in the central nervous system (CNS) of rodents was introduced in the 1960s. It supported the existence of chemical neurotransmission in the CNS. The monoamine neurons in the lower brain stem formed monosynaptic ascending systems to the telencephalon and diencephalon and monoamine descending systems to the entire spinal cord. The monoamines were early on suggested to operate via synaptic chemical transmission in the CNS. This chemical transmission reduced the impact of electrical transmission. In 1969 and the 1970s indications were obtained that important modes of chemical monoamine communication in the CNS also took place through the extra-synaptic fluid, the extracellular fluid, and long-distance communication in the cerebrospinal fluid involving diffusion and flow of transmitters like DA, NA and serotonin. In 1986, this type of transmission was named volume transmission (VT) by Agnati and Fuxe and their colleagues, also characterized by transmitter varicosity and receptor mismatches. The short and long-distance VT pathways were characterized by volume fraction, tortuosity and clearance. Electrical transmission also exists in the mammalian CNS, but chemical transmission is in dominance. One electrical mode is represented by electrical synapses formed by gap junctions which represent low resistant passages between nerve cells. It allows for a more rapid passage of action potentials between nerve cells compared to chemical transmission. The second mode is based on the ability of synaptic currents to generate electrical fields to modulate chemical transmission. One aim is to understand how chemical transmission can be integrated with electrical transmission and how putative (aquaporin water channel, dopamine D2R and adenosine A2AR) complexes in astrocytes can significancy participate in the clearance of waste products from the glymphatic system. VT may also help accomplish the operation of the acupuncture meridians essential for Chinese medicine in view of the indicated existence of extracellular VT pathways.

Gap junctions transmit electrical signals in neurons and serve metabolic coupling and chemical communication. Gap junctions are made of intercellular channels with large pores, allowing ions and small molecules to permeate. In the mammalian retina, intercellular coupling fulfills many vital functions in visual signal processing but is also implicated in promoting cell death after insults, such as excitotoxicity or hypoxia. Conversely, some studies also suggested a role for retinal gap junctions in neuroprotection. Recently, gap junctions were also advocated as conduits for therapeutic drug delivery in neurodegenerative disorders. This requires the permeation of rather large molecules through retinal gap junctions. However, the permeability of retinal networks for molecules >0.6 kDa has not been tested systematically. Here, we used the cut-loading method and probed gap junctional networks in the mouse retina for their permeability to cGMP and cAMP coupled to Biotin, using the well-characterized tracer Neurobiotin as control. Biotin-cGMP and -cAMP have a molecular weight of >0.8 kDa. We show that they cannot pass the gap junctions of horizontal cells but can permeate through the gap junctions of specific amacrine cells in the inner retina. These amacrine cells do not comprise AII amacrine cells and nitric oxide-releasing amacrine cells but some unknown type. In summary, we show that some retinal gap junctions are large enough to let molecules >0.8 kDa pass, making the intercellular delivery of therapeutic agents - already successfully exploited, for example, in cancer - also feasible in neurodegenerative diseases.

UNASSIGNED: Gap junctions are dynamically modulated bridges allowing the transcellular passage of ions and small molecules with a molecular mass of up to 1 kDa, a mechanism utilized for molecular communication purposes by living cells. This same mechanism is also exploited by scientists to reveal the existence of gap junction contacts by the cell-to-cell movement of tracers. However, multiple labeling experiments require the availability of multiple gap junction-permeable tracers.
UNASSIGNED: To this end, we utilized the well-known transient OFF alpha retinal ganglion cell (RGC)-coupled array as a model system to study and compare the transjunctional movement of neurobiotin (NB), a commonly used tracer, and serotonin, a recently identified tracer.
UNASSIGNED: Although the transjunctional movement of serotonin has been established in cell cultures, here we show, for the first time, that serotonin is also a potent tracer in in vitro tissue. In addition, serotonin is lighter than the classical gap junction-permeable NB, and thus, we expected that tracer movement would be comparable to or better than that of serotonin. We found that intracellular serotonin injections result in the labeling of the coupled transient OFF alpha RGC array very similar to those of the classical NB-labeled arrays. Both serotonin and NB-injected transient OFF alpha RGCs displayed the well-known pattern with coupled RGCs and a cohort of coupled wide-field amacrine cells (ACs).
UNASSIGNED: By using morphological characteristics, we confirm that the serotonin and the NB-coupled AC arrays are identical, and thereby confirm that serotonin is a potent gap junction-permeable tracer and can be readily used as an alternative to NB in in vitro tissue. Moreover, serotonin can be utilized in parallel with other dyes or tracers, enabling the use of multiple labels in the same material.

In the monkey retina, there are two distinct types of axon-bearing horizontal cells, known as H1 and H2 horizontal cells (HCs). In this study, cell bodies were prelabled using 4\',6-diamidino-2-phenylindole (DAPI), and both H1 and H2 horizontal cells were filled with Neurobiotin™ to reveal their coupling, cellular details, and photoreceptor contacts. The confocal analysis of H1 and H2 HCs was used to assess the colocalization of terminal dendrites with glutamate receptors at cone pedicles. After filling H1 somas, a large coupled mosaic of H1 cells was labeled. The dendritic terminals of H1 cells contacted red/green cone pedicles, with the occasional sparse contact with blue cone pedicles observed. The H2 cells were also dye-coupled. They had larger dendritic fields and lower densities. The dendritic terminals of H2 cells preferentially contacted blue cone pedicles, but additional contacts with nearly all cones within the dendritic field were still observed. The red/green cones constitute 99% of the input to H1 HCs, whereas H2 HCs receive a more balanced input, which is composed of 58% red/green cones and 42% blue cones. These observations confirm those made in earlier studies on primate horizontal cells by Dacey and Goodchild in 1996. Both H1 and H2 HCs were axon-bearing. H1 axon terminals (H1 ATs) were independently coupled and contacted rod spherules exclusively. In contrast, the H2 axon terminals contacted cones, with some preference for blue cone pedicles, as reported by Chan and Grünert in 1998. The primate retina contains three independently coupled HC networks in the outer plexiform layer (OPL), identified as H1 and H2 somatic dendrites, and H1 ATs. At each cone pedicle, the colocalization of both H1 and H2 dendritic tips with GluA4 subunits close to the cone synaptic ribbons indicates that glutamate signaling from the cones to H1 and H2 horizontal cells is mediated by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors.

Electrical synapses, formed of gap junctions, are ubiquitous components of the central nervous system (CNS) that shape neuronal circuit connectivity and dynamics. In the retina, electrical synapses can create a circuit, control the signal-to-noise ratio in individual neurons, and support the coordinated neuronal firing of ganglion cells, hence, regulating signal processing at the network, single-cell, and dendritic level. We, the authors, and Steve Massey have had a long interest in gap junctions in retinal circuits, in general, and in the network of photoreceptors, in particular. Our combined efforts, based on a wide array of techniques of molecular biology, microscopy, and electrophysiology, have provided fundamental insights into the molecular structure and properties of the rod/cone gap junction. Yet, a full understanding of how rod/cone coupling controls circuit dynamics necessitates knowing its operating range. It is well established that rod/cone coupling can be greatly reduced or eliminated by bright-light adaptation or pharmacological treatment; however, the upper end of its dynamic range has long remained elusive. This held true until Steve Massey\'s recent interest for connectomics led to the development of a new strategy to assess this issue. The effort proved effective in establishing, with precision, the connectivity rules between rods and cones and estimating the theoretical upper limit of rod/cone electrical coupling. Comparing electrophysiological measurements and morphological data indicates that under pharmacological manipulation, rod/cone coupling can reach the theoretical maximum of its operating range, implying that, under these conditions, all the gap junction channels present at the junctions are open. As such, channel open probability is likely the main determinant of rod/cone coupling that can change momentarily in a time-of-day- and light-dependent manner. In this article we briefly review our current knowledge of the molecular structure of the rod/cone gap junction and of the mechanisms behind its modulation, and we highlight the recent work led by Steve Massey. Steve\'s contribution has been critical toward asserting the modulation depth of rod/cone coupling as well as elevating the rod/cone gap junction as one of the most suitable models to examine the role of electrical synapses and their plasticity in neural processing.