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NMDA receptor (NMDAR) and GABA neuronal dysfunctions are observed in animal models of autism spectrum disorders, but how these dysfunctions impair social cognition and behavior remains unclear. We report here that NMDARs in cortical parvalbumin (Pv)-positive interneurons cooperate with gap junctions to promote high-frequency (>80 Hz) Pv neuronal burst firing and social cognition. Shank2–/– mice, displaying improved sociability upon NMDAR activation, show impaired cortical social representation and inhibitory neuronal burst firing. Cortical Shank2–/– Pv neurons show decreased NMDAR activity, which suppresses the cooperation between NMDARs and gap junctions (GJs) for normal burst firing. Shank2–/– Pv neurons show compensatory increases in GJ activity that are not sufficient for social rescue. However, optogenetic boosting of Pv neuronal bursts, requiring GJs, rescues cortical social cognition in Shank2–/– mice, similar to the NMDAR-dependent social rescue. Therefore, NMDARs and gap junctions cooperate to promote cortical Pv neuronal bursts and social cognition., How NMDAR and GABA neuronal dysfunctions result in impaired social behaviour is unclear. Here, the authors show that NMDARs and gap junctions in cortical PV interneurons modulate burst firing, affecting social behaviour.

Oligodendrocyte precursor cells (OPCs) are not merely a transitory progenitor cell type, but rather a distinct and heterogeneous population of glia with various functions in the developing and adult central nervous system. In this review, we discuss the fate and function of OPCs in the brain beyond their contribution to myelination. OPCs are electrically sensitive, form synapses with neurons, support blood–brain barrier integrity, and mediate neuroinflammation. We explore how sex and age may influence OPC activity, and we review how OPC dysfunction may play a primary role in numerous neurological and neuropsychiatric diseases. Finally, we highlight areas of future research.

Modulation of gap junction-mediated electrical synapses is a common form of neural plasticity. However, the behavioral consequence of the modulation and the underlying molecular cellular mechanisms are not understood. Here, using a C. elegans circuit of interneurons that are connected by gap junctions, we show that modulation of the gap junctions facilitates olfactory learning. Learning experience weakens the gap junctions and induces a repulsive sensory response to the training odorants, which together decouple the responses of the interneurons to the training odorants to generate learned olfactory behavior. The weakening of the gap junctions results from downregulation of the abundance of a gap junction molecule, which is regulated by cell-autonomous function of the worm homologs of a NMDAR subunit and CaMKII. Thus, our findings identify the function of a gap junction modulation in an in vivo model of learning and a conserved regulatory pathway underlying the modulation., Neural plasticity underlies learning and is regulated by modulation of gap junctions. Here, the authors show that NMDAR- and CaMKII-mediated cell autonomous downregulation of gap junction molecules in a circuit of interneurons facilitates olfactory learning affecting behavior in C. elegans.

Plasticity within the neuronal networks of the brain underlies the ability to learn and retain new information. The initial discovery of synaptic plasticity occurred by measuring synaptic strength in vivo, applying external stimulation and observing an increase in synaptic strength termed long-term potentiation (LTP). Many of the molecular pathways involved in LTP and other forms of synaptic plasticity were subsequently uncovered in vitro. Over the last few decades, technological advances in recording and imaging in live animals have seen many of these molecular mechanisms confirmed in vivo, including structural changes both pre- and postsynaptically, changes in synaptic strength, and changes in neuronal excitability. A well-studied aspect of neuronal plasticity is the capacity of the brain to adapt to its environment, gained by comparing the brains of deprived and experienced animals in vivo, and in direct response to sensory stimuli. Multiple in vivo studies have also strongly linked plastic changes to memory by interfering with the expression of plasticity and by manipulating memory engrams. Plasticity in vivo also occurs in the absence of any form of external stimulation, i.e., during spontaneous network activity occurring with brain development. However, there is still much to learn about how plasticity is induced during natural learning and how this is altered in neurological disorders.

Summary Electrical synapses are established between specific neurons and within distinct subcellular compartments, but the mechanisms that direct gap junction assembly in the nervous system are largely unknown. Here, we show that a developmental program tunes cAMP signaling to direct the neuron-specific assembly and placement of electrical synapses in the C. elegans motor circuit. We use live-cell imaging to visualize electrical synapses in vivo and an optogenetic assay to confirm that they are functional. In ventral A class (VA) motor neurons, the UNC-4 transcription factor blocks expression of cAMP antagonists that promote gap junction miswiring. In unc-4 mutants, VA electrical synapses are established with an alternative synaptic partner and are repositioned from the VA axon to soma. cAMP counters these effects by driving gap junction trafficking into the VA axon for electrical synapse assembly. Thus, our experiments establish that cAMP regulates gap junction trafficking for the biogenesis of functional electrical synapses.

The retinas of many species show regional specialisations that are evident in the differences in the processing of visual input from different parts of the visual field. Regional specialisation is thought to reflect an adaptation to the natural visual environment, optical constraints, and lifestyle of the species. Yet, little is known about regional differences in synaptic circuitry. Here, we were interested in the topographical distribution of connexin-36 (Cx36), the major constituent of electrical synapses in the retina. We compared the retinas of mice, rats, and cats to include species with different patterns of regional specialisations in the analysis. First, we used the density of Prox1-immunoreactive amacrine cells as a marker of any regional specialisation, with higher cell density signifying more central regions. Double-labelling experiments showed that Prox1 is expressed in AII amacrine cells in all three species. Interestingly, large Cx36 plaques were attached to about 8–10% of Prox1-positive amacrine cell somata, suggesting the strong electrical coupling of pairs or small clusters of cell bodies. When analysing the regional changes in the volumetric density of Cx36-immunoreactive plaques, we found a tight correlation with the density of Prox1-expressing amacrine cells in the ON, but not in the OFF sublamina in all three species. The results suggest that the relative contribution of electrical synapses to the ON- and OFF-pathways of the retina changes with retinal location, which may contribute to functional ON/OFF asymmetries across the visual field.

A significant component of the repetitive dynamics during locomotion in vertebrates is generated within the spinal cord. The legged locomotion of mammals is most likely controled by a hierarchical, multi-layer spinal network structure, while the axial circuitry generating the undulatory swimming motion of animals like lamprey is thought to have only a single layer in each segment. Recent experiments have suggested a hybrid network structure in zebrafish larvae in which two types of excitatory interneurons (V2a-I and V2a-II) both make first-order connections to the brain and last-order connections to the motor pool. These neurons are connected by electrical and chemical synapses across segments. Through computational modeling and an asymptotic perturbation approach we show that this interleaved interaction between the two neuron populations allows the spinal network to quickly establish the correct activation sequence of the segments when starting from random initial conditions and to reduce the dependence of the intersegmental phase difference (ISPD) of the oscillations on the swimming frequency. The latter reduces the frequency dependence of the waveform of the swimming motion. In the model the reduced frequency dependence is largely due to the different impact of chemical and electrical synapses on the ISPD and to the significant spike-frequency adaptation that has been observed experimentally in V2a-II neurons, but not in V2a-I neurons. Our model makes experimentally testable predictions and points to a benefit of the hybrid structure for undulatory locomotion that may not be relevant for legged locomotion.

SummaryThe coordination of activity between brain cells is a key determinant of neural circuit function; nevertheless, approaches that selectively regulate communication between two distinct cellular components of a mammalian circuit remain sparse. To address this gap, we developed a novel class of gap junctions by selectively engineering two connexin proteins found inMorone americana(white perch fish): connexin34.7 (Cx34.7) and connexin35 (Cx35). By iteratively exploiting protein mutagenesis, a novelin vitroassay of connexin docking, and computational modeling of connexin hemichannel interactions, we uncovered a pattern of structural motifs that contribute to hemichannel docking compatibility. Targeting these motifs, we designed Cx34.7 and Cx35 hemichannels that dock with each other, but not with themselves, nor with other major connexins expressed in the mammalian central nervous system. We validated these hemichannelsin vivousingC. elegansand mice, demonstrating that they can facilitate communication across neural circuits composed of pairs of distinct cell types and modify behavior accordingly. Thus, we establish a potentially translational approach, ‘Long-termintegration ofCircuits using connexins’ (LinCx), for context-precise circuit-editing with unprecedented spatiotemporal specificity in mammals.

Experimental data published earlier confirming the representations of the reticular theory of the organization of the nervous system are analyzed. It is noted that Sherrington’s evidence has refuted the main concept of the neuronal theory of Ramon y Cajal, the law of dynamic polarization of neurons. After the detection of gap junctions (GJs), the adhesion of neurites, their high electrical and metabolic permeability, and their cumulative interaction were detected. Data demonstrating that GJs are able to transform into syncytial perforations are given, and somatic GJs, under favorable conditions, can convert neurons into dikaryons. It is assumed that all the phenomena described are consecutive stages of a single process of reconstruction of the outer cell membrane of the neuron.

The dorsal portion of the descending octaval nucleus (dDO), the main first-order auditory nucleus in jawed fish, includes four lateral and three medial neuronal populations that project to the auditory midbrain. One medial population and one lateral population contain neurons that receive a remarkably large axon terminal from the utricular branch of the octaval nerve. Immunocytochemistry for connexin 35 (Cx35) was used to determine whether this connection includes electrical synapses. Although Cx35 was not localized to these large contacts, it was observed in the three other lateral dDO populations. Another first-order nucleus, the dorsal portion of the anterior octaval nucleus (dAO), primitively projects to the auditory midbrain in jawed fishes and contains neurons positive for Cx35. Utricular branch terminals were coincident with some Cx35 puncta in dDO and dAO. The results are discussed in light of what is known about the occurrence of electrical synapses in first-order auditory and vestibular nuclei in fish and tetrapods.

Most research on neurodegenerative diseases has focused on neurons, yet glia help form and maintain the synapses whose loss is so prominent in these conditions. To investigate the contributions of glia to Huntington's disease (HD), we profiled the gene expression alterations of Drosophila expressing human mutant Huntingtin (mHTT) in either glia or neurons and compared these changes to what is observed in HD human and HD mice striata. A large portion of conserved genes are concordantly dysregulated across the three species; we tested these genes in a high-throughput behavioral assay and found that downregulation of genes involved in synapse assembly mitigated pathogenesis and behavioral deficits. To our surprise, reducing dNRXN3 function in glia was sufficient to improve the phenotype of flies expressing mHTT in neurons, suggesting that mHTT's toxic effects in glia ramify throughout the brain. This supports a model in which dampening synaptic function is protective because it attenuates the excitotoxicity that characterizes HD., eLife digest When a neuron dies, through injury or disease, the body loses all communication that passes through it. The brain compensates by rerouting the flow of information through other neurons in the network. Eventually, if the loss of neurons becomes too great, compensation becomes impossible. This process happens in Alzheimer's, Parkinson's, and Huntington's disease. In the case of Huntington's disease, the cause is mutation to a single gene known as huntingtin. The mutation is present in every cell in the body but causes particular damage to parts of the brain involved in mood, thinking and movement. Neurons and other cells respond to mutations in the huntingtin gene by turning the activities of other genes up or down, but it is not clear whether all of these changes contribute to the damage seen in Huntington's disease. In fact, it is possible that some of the changes are a result of the brain trying to protect itself. So far, most research on this subject has focused on neurons because the huntingtin gene plays a role in maintaining healthy neuronal connections. But, given that all cells carry the mutated gene, it is likely that other cells are also involved. The glia are a diverse group of cells that support the brain, providing care and sustenance to neurons. These cells have a known role in maintaining the connections between neurons and may also have play a role in either causing or correcting the damage seen in Huntington's disease. The aim of Onur et al. was to find out which genes are affected by having a mutant huntingtin gene in neurons or glia, and whether severity of Huntington’s disease improved or worsened when the activity of these genes changed. First, Onur et al. identified genes affected by mutant huntingtin by comparing healthy human brains to the brains of people with Huntington's disease. Repeating the same comparison in mice and fruit flies identified genes affected in the same way across all three species, revealing that, in Huntington's disease, the brain dials down glial cell genes involved in maintaining neuronal connections. To find out how these changes in gene activity affect disease severity and progression, Onur et al. manipulated the activity of each of the genes they had identified in fruit flies that carried mutant versions of huntingtin either in neurons, in glial cells or in both cell types. They then filmed the flies to see the effects of the manipulation on movement behaviors, which are affected by Huntington’s disease. This revealed that purposely lowering the activity of the glial genes involved in maintaining connections between neurons improved the symptoms of the disease, but only in flies who had mutant huntingtin in their glial cells. This indicates that the drop in activity of these genes observed in Huntington’s disease is the brain trying to protect itself. This work suggests that it is important to include glial cells in studies of neurological disorders. It also highlights the fact that changes in gene expression as a result of a disease are not always bad. Many alterations are compensatory, and try to either make up for or protect cells affected by the disease. Therefore, it may be important to consider whether drugs designed to treat a condition by changing levels of gene activity might undo some of the body's natural protection. Working out which changes drive disease and which changes are protective will be essential for designing effective treatments.

SUMMARYShank3 mutations contribute to intellectual disability. Because SHANK3 is a protein scaffold that helps organize the multiprotein network of the glutamatergic postsynaptic density (PSD), alterations in chemical synaptic transmission are implicated. Electrical synaptic transmission is a second form of synaptic transmission, enabled by intercellular channels comprised of connexin36 that support direct electrical communication among neurons, electrical brain rhythms, and neurocognitive states. Using multiplex proteomics, we report that two autism-related mutations of mouse Shank3 disrupt the glutamatergic PSD differently, but have in common the disruption of an association between NMDA-type glutamate-receptors (NMDARs) and connexin36. Mutation of Shank3 exons 13-16 most robustly dissociated connexin36 from NMDARs while impairing electrical synaptic transmission and the synchrony of an electrical rhythm in mouse inferior olive. We suggest that electrical synapses are a component of an “extended PSD” sensitive to Shank3 mutations that produce intellectual disability, at least in part, by impairing electrical synaptic transmission.

Shank3 mutations contribute to intellectual disability. Because SHANK3 is a protein scaffold that helps organize the multiprotein network of the glutamatergic postsynaptic density (PSD), alterations in chemical synaptic transmission are implicated. Electrical synaptic transmission is a second form of synaptic transmission, enabled by intercellular channels comprised of connexin36 that support direct electrical communication among neurons, electrical brain rhythms, and neurocognitive states. Using multiplex proteomics, we report that two autism-related mutations of mouse Shank3 disrupt the glutamatergic PSD differently, but have in common the disruption of an association between NMDA-type glutamate-receptors (NMDARs) and connexin36. Mutation of Shank3 exons 13-16 most robustly dissociated connexin36 from NMDARs while impairing electrical synaptic transmission and the synchrony of an electrical rhythm in mouse inferior olive. We suggest that electrical synapses are a component of an “extended PSD” sensitive to Shank3 mutations that produce intellectual disability, at least in part, by impairing electrical synaptic transmission.

Embryogenesis, as well as regeneration, is increasingly recognized to be orchestrated by an interplay of transcriptional and bioelectric networks. Spatiotemporal patterns of resting potentials direct the size, shape, and locations of numerous organ primordia during patterning. These bioelectrical properties are established by the function of ion channels and pumps that set voltage potentials of individual cells, and gap junctions (electrical synapses) that enable physiological states to propagate across tissue networks. Functional experiments to probe the roles of bioelectrical states can be carried out by targeting endogenous ion channels during development. Here, we describe protocols, optimized for the highly tractable Xenopus laevis embryo, for molecular genetic targeting of ion channels and connexins based on CRISPR, and monitoring of resting potential states using voltage-sensing fluorescent dye. Similar strategies can be adapted to other model species.

Electrical synaptic transmission relies on neuronal gap junctions containing channels constructed by Connexins. While at chemical synapses neurotransmitter-gated ion channels are critically supported by scaffolding proteins, it is unknown if channels at electrical synapses require similar scaffold support. Here, we investigated the functional relationship between neuronal Connexins and Zonula Occludens 1 (ZO1), an intracellular scaffolding protein localized to electrical synapses. Using model electrical synapses in zebrafish Mauthner cells, we demonstrated that ZO1 is required for robust synaptic Connexin localization, but Connexins are dispensable for ZO1 localization. Disrupting this hierarchical ZO1/Connexin relationship abolishes electrical transmission and disrupts Mauthner cell-initiated escape responses. We found that ZO1 is asymmetrically localized exclusively postsynaptically at neuronal contacts where it functions to assemble intercellular channels. Thus, forming functional neuronal gap junctions requires a postsynaptic scaffolding protein. The critical function of a scaffolding molecule reveals an unanticipated complexity of molecular and functional organization at electrical synapses., eLife digest Neurons ‘talk’ with each another at junctions called synapses, which can either be chemical or electrical. Communication across a chemical synapse involves a ‘sending’ neuron releasing chemicals that diffuse between the cells and subsequently bind to specialized receptors on the receiving neuron. These complex junctions involve a large number of well-studied molecular actors. Electrical synapses, on the other hand, are believed to be simpler. There, neurons are physically connected via channels formed of ‘connexin’ proteins, which allow electrically charged ions to flow between the cells. However, it is likely that other proteins help to create these structures. In particular, recent evidence shows that without a structurally supporting ‘scaffolding’ protein called ZO1, electrical synapses cannot form in the brain of a tiny freshwater fish known as zebrafish. As their name implies, scaffolding proteins help cells organize their internal structure, for example by anchoring other molecules to the cell membrane. By studying electrical synapses in zebrafish, Lasseigne, Echeverry, Ijaz, Michel et al. now show that these structures are more complex than previously assumed. In particular, the experiments reveal that ZO1 proteins are only present on one side of electrical synapses; despite their deceptively symmetrical anatomical organization, these junctions can be asymmetric, like their chemical cousins. The results also show that ZO1 must be present for connexins to gather at electrical synapses, whereas the converse is not true. This suggests that when a new electrical synapse forms, ZO1 moves into position first: it then recruits or stabilizes connexins to form the channels connecting the two cells. In many animals with a spine, electrical synapses account for about 20% of all neural junctions. Understanding how these structures form and work could help to find new treatments for disorders linked to impaired electrical synapses, such as epilepsy.

The term “synapse” represents the junction between nerve cells. There are two major types of synapses namely chemical...

In nervous systems, there are two main modes of transmission for the propagation of activity between cells. Synaptic transmission relies on close contact at chemical or electrical synapses while volume transmission is mediated by diffusible chemical signals and does not require direct contact. It is possible to wire complex neuronal networks by both chemical and synaptic transmission. Both types of networks are ubiquitous in nervous systems, leading to the question which of the two appeared first in evolution. This paper explores a scenario where chemically organized cellular networks appeared before synapses in evolution, a possibility supported by the presence of complex peptidergic signalling in all animals except sponges. Small peptides are ideally suited to link up cells into chemical networks. They have unlimited diversity, high diffusivity and high copy numbers derived from repetitive precursors. But chemical signalling is diffusion limited and becomes inefficient in larger bodies. To overcome this, peptidergic cells may have developed projections and formed synaptically connected networks tiling body surfaces and displaying synchronized activity with pulsatile peptide release. The advent of circulatory systems and neurohemal organs further reduced the constraint imposed on chemical signalling by diffusion. This could have contributed to the explosive radiation of peptidergic signalling systems in stem bilaterians. Neurosecretory centres in extant nervous systems are still predominantly chemically wired and coexist with the synaptic brain.This article is part of the theme issue ‘Basal cognition: multicellularity, neurons and the cognitive lens’.

Calmodulin binding is a nearly universal property of gap junction proteins, imparting a calcium-dependent uncoupling behavior that can serve in an emergency to decouple a stressed cell from its neighbors. However, gap junctions that function as electrical synapses within networks of neurons routinely encounter large fluctuations in local cytoplasmic calcium concentration, frequent uncoupling would be impractical and counterproductive. We have studied the properties and functional consequences of calmodulin binding to the electrical synapse protein Connexin 35 (Cx35 or gjd2b), homologous to mammalian Connexin 36 (Cx36 or gjd2). We find that specializations in Cx35 calmodulin binding sites make it relatively impervious to moderately high levels of cytoplasmic calcium. Calmodulin binding to a site in the C-terminus causes uncoupling when calcium reaches low micromolar concentrations, a behavior prevented by mutations that eliminate calmodulin binding. However, milder stimuli promote calcium/calmodulin-dependent protein kinase II activity that potentiates coupling without interference from calmodulin binding. A second calmodulin binding site in the end of the Cx35 cytoplasmic loop, homologous to a calmodulin binding site present in many connexins, binds calmodulin with very low affinity and stoichiometry. Together, the calmodulin binding sites cause Cx35 to uncouple only at extreme levels of intracellular calcium.

An innexin N terminus confers strong rectification property to electrical synapses through a ball-and-receptor mechanism., Rectifying electrical synapses (RESs) exist across animal species, but their rectification mechanism is largely unknown. We investigated why RESs between AVA premotor interneurons and A-type cholinergic motoneurons (A-MNs) in Caenorhabditis elegans escape circuit conduct junctional currents (Ij) only in the antidromic direction. These RESs consist of UNC-7 innexin in AVA and UNC-9 innexin in A-MNs. UNC-7 has multiple isoforms differing in the length and sequence of the amino terminus. In a heterologous expression system, only one UNC-7 isoform, UNC-7b, can form heterotypic gap junctions (GJs) with UNC-9 that strongly favor Ij in the UNC-9 to UNC-7 direction. Knockout of unc-7b alone almost eliminated the Ij, whereas AVA-specific expression of UNC-7b substantially rescued the coupling defect of unc-7 mutant. Neutralizing charged residues in UNC-7b amino terminus abolished the rectification property of UNC-7b/UNC-9 GJs. Our results suggest that the rectification property results from electrostatic interactions between charged residues in UNC-7b amino terminus.

Purpose To characterize newly discovered electrical synapses, formed by connexin (Cx) 36 and 45, between neighbouring axons within the optic nerve head. Methods Twenty‐five Wistar rats were killed by CO 2 inhalation. Proximal and distal optic nerve (ON) stumps were collected and processed for immunostainings, electron microscopy (EM) with immunogold labelling, PCR and Western blots (WB). Additional 15 animals were deeply anaesthetized, and flash visual evoked potentials (fVEP) after retrobulbar injection of saline (negative control) or 100 μ m meclofenamic acid solution (gap junctions’ blocker) were recorded. Human paraffin cross‐sections of eyeballs for immunostainings were obtained from the Human Eye Biobank for Research. Results Immunostainings of both rat and human ON revealed the presence of Cx45 and 36 colocalizing with β3‐tubulin, but not with glial fibrillary acidic protein (GFAP). In WB, Cx36 content in optic nerve was approximately halved when compared with retina (0.58 ± 0.005 in proximal stump and 0.44 ± 0.02 in distal stump), Cx45 showed higher levels (0.68 ± 0.01 in proximal stump and 0.9 ± 0.07 in distal stump). In immunogold‐EM of optic nerve sections, we found electric synapses (formed mostly by Cx45) directly coupling neighbouring axons. In fVEP, blocking of gap junctions with meclofenamic acid resulted in significant prolongation of the latency of P1 wave up to 160% after 30 min (p

SUMMARY Our understanding of nervous system function is limited by our ability to identify and manipulate neuronal subtypes within intact circuits. We show that the Gbx2CreERT2-IRES-EGFP mouse line labels two amacrine cell (AC) subtypes in the mouse retina that have distinct morphological, physiological, and molecular properties. Using a combination of RNA-seq, genetic labeling, and patch clamp recordings, we show that one subtype is GABAergic that receives excitatory input from On bipolar cells. The other population is a non-GABAergic, non-glycinergic (nGnG) AC subtype that lacks the expression of standard neurotransmitter markers. Gbx2+ nGnG ACs have smaller, asymmetric dendritic arbors that receive excitatory input from both On and Off bipolar cells. Gbx2+ nGnG ACs also exhibit spatially restricted tracer coupling to bipolar cells (BCs) through gap junctions. This study identifies a genetic tool for investigating the two distinct AC subtypes, and it provides a model for studying synaptic communication and visual circuit function., Graphical Abstract, In Brief Investigations into neural circuit development and function are limited by the lack of genetic tools to label and perturb individual neuronal subtypes. Using the Gbx2CreERT2 mouse line, Kerstein et al. identify two amacrine cell subtypes in the mouse retina and explore their distinct molecular, morphological, and physiological characteristics.

Neuronal loss can considerably diminish neural circuit function, impairing normal behavior by disrupting information flow in the circuit. We reasoned that by rerouting the flow of information in the damaged circuit it may be possible to offset these negative outcomes. We examined this possibility using the well-characterized chemosensory circuit of the nematode worm C. elegans. In this circuit, a main sensory neuron class sends parallel outputs to several interneuron classes. We found that the removal of one of these interneuron classes impairs chemotaxis to attractive odors, revealing a prominent path for information flow in the circuit. To alleviate these deficiencies, we sought to reinforce a remaining neural pathway. We used genetically engineered electrical synapses for this purpose, and observed the successful recovery of chemotaxis performance. However, we were surprised to find that the recovery was largely mediated by inadvertently formed left-right lateral electrical connections within individual neuron classes. Our analysis suggests that these additional electrical synapses help restore circuit function by amplifying weakened neuronal signals in the damaged circuit. These results demonstrate the power of genetically engineered synapses to regulate information flow and signal intensity in damaged neural circuits.

In this review article, we will describe the recent advances made towards understanding the molecular and cell biological mechanisms of electrical synapse formation. New evidence indicates that electrical synapses, which are gap junctions between neurons, can have complex molecular compositions including protein asymmetries across joined cells, diverse morphological arrangements, and overlooked similarities with other junctions, all of which indicate new potential roles in neurodevelopmental disease. Aquatic organisms, and in particular the vertebrate zebrafish, have proven to be excellent models for elucidating the molecular mechanisms of electrical synapse formation. Zebrafish will serve as our main exemplar throughout this review and will be compared with other model organisms. We highlight the known cell biological processes that build neuronal gap junctions and compare these with the assemblies of adherens junctions, tight junctions, non-neuronal gap junctions, and chemical synapses to explore the unknown frontiers remaining in our understanding of the critical and ubiquitous electrical synapse.

Gap junctions are physical channels that connect adjacent cells, permitting the flow of small molecules/ions between the cytoplasms of the coupled units. Innexin/innexin-like proteins are responsible for the formation of invertebrate gap junctions. Within the nervous system, gap junctions often function as electrical synapses, providing a means for coordinating activity among electrically coupled neurons. While some gap junctions allow the bidirectional flow of small molecules/ions between coupled cells, others permit flow in one direction only or preferentially. The complement of innexins present in a gap junction determines its specific properties. Thus, understanding innexin diversity is key for understanding the full potential of electrical coupling in a species/system. The decapod crustacean cardiac ganglion (CG), which controls cardiac muscle contractions, is a simple pattern-generating neural network with extensive electrical coupling among its circuit elements. In the lobster, Homarus americanus, prior work suggested that the adult neuronal innexin complement consists of six innexins (Homam-Inx1-4 and Homam-Inx6-7). Here, using a H. americanus CG-specific transcriptome, we explored innexin complement in this portion of the lobster nervous system. With the exception of Homam-Inx4, all of the previously described innexins appear to be expressed in the H. americanus CG. In addition, transcripts encoding seven novel putative innexins (Homam-Inx8-14) were identified, four (Homam-Inx8-11) having multiple splice variants, e.g., six for Homam-Inx8. Collectively, these data indicate that the innexin complement of the lobster nervous system in general, and the CG specifically, is likely significantly greater than previously reported, suggesting the possibility of expanded gap junction diversity and function in H. americanus.

Gap junction connections between neurons play critical roles in the development of the nervous system. However, studies on the sensory experience–driven plasticity during the critical period rarely examine the involvement of gap junction connections. ON-OFF direction selective ganglion cells (ooDSGCs) in the mouse retina that prefer upward motion are connected by gap junctions throughout development. Here, we show that after exposing the mice to a visual environment dominated by upward motion from eye-opening to puberty, ooDSGCs that respond preferentially to upward motion show enhanced spike synchronization, while downward motion training has the opposite effect. The effect is long-term, persisting at least three months after the training. Correlated activity during training is tightly linked to this effect: Cells trained by stimuli that promote higher levels of activity correlation show stronger gap junction connection after the training, while stimuli that produce very low activity correlation leave the cells with much weaker gap junction connections afterwards. Direct investigation of the gap junction connections among upward motion–preferring ooDSGCs show that both the percentage of electrically coupled ooDSGCs and the strength of the coupling are affected by visual motion training. Our results demonstrate that in the retina, one of the peripheral sensory systems, gap junction connections can be shaped by experience during development., Gap junction connections between upward motion–preferring direction selective ganglion cells can be shaped by early visual experience; upward motion training leads to enhanced connectivity, while downward motion greatly suppresses the connection, suggesting a form of activity-dependent plasticity.

Connections between neurons called synapses are the key components underlying all nervous system functions of animals and humans. However, important genetic information on the formation and plasticity of one type, the electrical (gap junction-mediated) synapse, is severely understudied, especially in invertebrates. In the present study, we set forth to identify and characterize the gap junction-encoding gene innexin in the central nervous system (CNS) of the mollusc pond snailLymnaea stagnalis(L. stagnalis). With PCR, 3’ and 5’ RACE, and BLAST searches, we identified eight innexin genes in theL. stagnalisnervous system namedLst Inx1-8. Phylogenetic analysis revealed that theL. stagnalisinnexin genes originated from a single copy in the common ancestor of molluscan species by multiple gene duplication events and have been maintained inL. stagnalissince they were generated. The paralogous innexin genes demonstrate distinct expression patterns among tissues. In addition, one paralog,Lst Inx1, exhibits heterogeneity in cells and ganglia, suggesting the occurrence of functional diversification after gene duplication. These results introduce possibilities to study an intriguing potential relationship between innexin paralog expression and cell-specific functional outputs such as heterogenic ability to form channels and exhibit synapse plasticity. TheL. stagnalisCNS contains large neurons and a functionally defined network for behaviors; with the introduction ofL. stagnalisin the gap junction field, we are providing novel opportunities to combine genetic research with direct investigation of functional outcomes at the cellular, synaptic, and behavioral levels.Summary StatementBy characterizing the gap junction gene innexin inLymnaea stagnalis, we open opportunities for novel studies on the regulation, plasticity, and evolutionary function of electrical synapses throughout the animal kingdom.

Changes to the structure of nodes of Ranvier in the normal-appearing white matter (NAWM) of multiple sclerosis (MS) brains are associated with chronic inflammation. We show that the paranodal domains in MS NAWM are longer on average than control, with Kv1.2 channels dislocated into the paranode. These pathological features are reproduced in a model of chronic meningeal inflammation generated by the injection of lentiviral vectors for the lymphotoxin-α (LTα) and interferon-γ (IFNγ) genes. We show that tumour necrosis factor (TNF), IFNγ, and glutamate can provoke paranodal elongation in cerebellar slice cultures, which could be reversed by an N-methyl-D-aspartate (NMDA) receptor blocker. When these changes were inserted into a computational model to simulate axonal conduction, a rapid decrease in velocity was observed, reaching conduction failure in small diameter axons. We suggest that glial cells activated by pro-inflammatory cytokines can produce high levels of glutamate, which triggers paranodal pathology, contributing to axonal damage and conduction deficits., Current thinking on the mechanisms by which multiple sclerosis gives rise to cumulative neurological disability revolves largely around focal lesions of inflammation and demyelination. However, some of the debilitating symptoms, such as severe fatigue, might be better explained by a more diffuse pathology. This study shows that paranodes in the white matter become abnormal as a result of neuroinflammation, which may be the result of the action of cytokines that cause glia to release glutamate.

The circadian neuronal circuit of Drosophila melanogaster is made up of about 150 neurons, distributed bilaterally and distinguished into 7 clusters. Multiple lines of evidence suggest that coherent rhythms in behaviour are brought about when these clusters function as a network. Although chemical modes of communication amongst circadian neurons have been well-studied, there has been no report of communication via electrical synapses made up of gap junctions. Here, we report for the first time that gap junction proteins - Innexins play crucial roles in determining the period of free-running activity rhythms in flies. Our experiments reveal the presence of gap junction protein INNEXIN2 in the ventral lateral neurons. RNA-interference based knockdown of its expression in circadian pacemakers slows down the speed of locomotor activity rhythm. Concomitantly, we find alterations in the oscillation of a core-clock protein PERIOD and in the output molecule Pigment Dispersing Factor in the circadian pacemaker neuron network.

Electrical synaptic transmission via gap junctions underlies direct and rapid neuronal communication in the central nervous system. The diversity of functional roles played by electrical synapses is perhaps best exemplified in the vertebrate retina, in which gap junctions are expressed by each of the five major neuronal types. These junctions are highly plastic; they are dynamically regulated by ambient illumination and circadian rhythms acting through light-activated neuromodulators. The networks formed by electrically coupled neurons provide plastic, reconfigurable circuits positioned to play key and diverse roles in the transmission and processing of visual information at every retinal level. Recent work indicates gap junctions also play a role in the progressive cell death and aberrant activity seen in various pathological conditions of the retina. Gap junctions thus form potential targets for novel neuroprotective therapies in the treatment of neurodegenerative retinal diseases such as glaucoma and ischemic retinopathies.

International audience; The developing barrel cortex reveals a rich repertoire of neuronal activity patterns, which have been also found in other sensory neocortical areas and in other species including the somatosensory cortex of preterm human infants. The earliest stage is characterized by asyn-chronous, sparse single-cell firing at low frequencies. During the second stage neurons show correlated firing, which is initially mediated by electrical synapses and subsequently transforms into network bursts depending on chemical synapses. Activity patterns during this second stage are synchronous plateau assemblies, delta waves, spindle bursts and early gamma oscillations (EGOs). In newborn rodents spindle bursts and EGOs occur spontaneously or can be elicited by sensory stimulation and synchronize the activity in a barrel-related columnar network with topo-graphic organization at the day of birth. Interfering with this early activity causes a disturbance in the development of the cortical architecture, indicating that spindle bursts and EGOs influence the formation of cortical columns. Early neuronal activity also controls the rate of programed cell death in the developing barrel cortex, suggesting that spindle bursts and EGOs are physiological activity patterns particularly suited to suppress apoptosis. It remains to be studied in more detail how these different neocortical activity patterns control early developmental processes such as formation of synapses, microcircuits, topographic maps and large-scale networks. This article is part of a Special Issue entitled: Barrel Cortex. Ó

Simple Summary Electrical synapses are ubiquitous in nervous systems, where they coordinate the activity of neural networks in time. However, their role in the execution and learning of complex behaviors remains unknown. Electrical synapses have remained unexplored in the songbird brain, which provides a model to link the production and learning of a complex behavior to the synaptic structure of defined neural circuits. Here, we show that GJD2 mRNA, coding for the major channel-forming electrical synapse protein connexin 36 (Cx36), is extensively expressed in the two nuclei that control song production, HVC and RA and their embedding regions. Our in situ hybridizations, together with the analysis of published transcriptomics data, demonstrate that electrical synapses are a general and widespread feature of song premotor nuclei in songbirds, where they show brain region-specific, cell type-specific expression patterns, dynamic during neuronal differentiation. We propose songbirds as a suitable model to investigate the contribution of the major vertebrate electrical synapse protein, Cx36, to the production and learning of motor skills in vertebrates. Abstract Birdsong is a precisely timed animal behavior. The connectivity of song premotor neural networks has been proposed to underlie the temporal patterns of neuronal activity that control vocal muscle movements during singing. Although the connectivity of premotor nuclei via chemical synapses has been characterized, electrical synapses and their molecular identity remain unexplored. We show with in situ hybridizations that GJD2 mRNA, coding for the major channel-forming electrical synapse protein in mammals, connexin 36, is expressed in the two nuclei that control song production, HVC and RA from canaries and zebra finches. In canaries’ HVC, GJD2 mRNA is extensively expressed in GABAergic and only a fraction of glutamatergic cells. By contrast, in RA, GJD2 mRNA expression is widespread in glutamatergic and GABAergic neurons. Remarkably, GJD2 expression is similar in song nuclei and their respective embedding brain regions, revealing the widespread expression of GJD2 in the avian brain. Inspection of a single-cell sequencing database from zebra and Bengalese finches generalizes the distributions of electrical synapses across cell types and song nuclei that we found in HVC and RA from canaries, reveals a differential GJD2 mRNA expression in HVC glutamatergic subtypes and its transient increase along the neurogenic lineage. We propose that songbirds are a suitable model to investigate the contribution of electrical synapses to motor skill learning and production.

The circadian neuronal circuit ofDrosophila melanogasteris made up of about 150 neurons, distributed bilaterally and distinguished into 7 clusters. Multiple lines of evidence suggest that coherent rhythms in behaviour are brought about when these clusters function as a network. Although chemical modes of communication amongst circadian neurons have been well-studied, there has been no report of communication via electrical synapses made up of gap junctions. Here, we report for the first time that gap junction proteins – Innexins play crucial roles in determining the period of free-running activity rhythms in flies. Our experiments reveal the presence of gap junction protein INNEXIN2 in the ventral lateral neurons. RNA-interference based knockdown of its expression in circadian pacemakers slows down the speed of locomotor activity rhythm. Concomitantly, we find alterations in the oscillation of a core-clock protein PERIOD and in the output molecule Pigment Dispersing Factor in the circadian pacemaker neuron network.

In the central nervous system (CNS), connexin and pannexin gap junctions and hemichannels are an integral component of homeostatic neuronal excitability and synaptic plasticity. Neuronal connexin (Cx) gap junctions form electrical synapses across biochemically similar GABAergic networks, allowing rapid and extensive inhibition in response to principle neuron excitation. Glial Cx gap junctions link astrocytes and oligodendrocytes in the pan-glial network that is responsible for removing excitotoxic ions and metabolites. In addition, Cx gap junctions between glia help constrain excessive excitatory activity in neurons and facilitate astrocyte Ca2+ slow wave propagation. Pannexins (Panxs) do not form gap junctions in vivo, but Panx hemichannels participate in autocrine and paracrine gliotransmission alongside connexin hemichannels. ATP and other gliotransmitters released by Cx and Panx hemichannels maintain physiologic glutamatergic tone by strengthening synapses and mitigating aberrant high frequency bursting. Under pathological depolarizing and inflammatory conditions, gap junctions and hemichannels become dysregulated, resulting in aberrant neuronal firing and seizure. In this review, we present known contributions of Cxs and Panxs to physiologic neuronal excitation and explore how disruption of gap junctions and hemichannels lead to aberrant glutamatergic transmission, purinergic signaling, and seizures.

Key points Electrical synapses are modified by various forms of activity, including paired activity in coupled neurons and tetanization of the input to coupled neurons. We show that plasticity of electrical synapses that results from paired spiking activity in coupled neurons depends on calcium influx and calcium-initiated signalling pathways. Plasticity that results from tetanization of input fibres does not depend on calcium influx or dynamics. These results imply that electrically coupled neurons have distinct sets of mechanisms for adjusting coupling according to the specific type of activity they experience. Abstract Recent results have demonstrated modification of electrical synapse strength by varied forms of neuronal activity. However, the mechanisms underlying plasticity induction in central mammalian neurons are unclear. Here we show that the two established inductors of plasticity at electrical synapses in the thalamic reticular nucleus - paired burst spiking in coupled neurons, and mGluR-dependent tetanization of synaptic input - are separate pathways that converge at a common downstream endpoint. Using occlusion experiments and pharmacology in patched pairs of coupled neurons in vitro, we show that burst-induced depression depends on calcium entry via voltage-gated channels, is blocked by BAPTA chelation, and recruits intracellular calcium release on its way to activation of phosphatase activity. In contrast, mGluR-dependent plasticity is independent of calcium entry or calcium dynamics. Together, these results show that the spiking-initiated mechanisms underlying electrical synapse plasticity are similar to those that induce plasticity at chemical synapses, and offer the possibility that calcium-regulated mechanisms may also lead to alternate outcomes, such as potentiation. Because these mechanistic elements are widely found in mature neurons, we expect them to apply broadly to electrical synapses across the brain, acting as the crucial link between neuronal activity and electrical synapse strength.

Electrical synapses are finding increasing representation and importance in our understanding of signaling in the nervous system. In contrast to chemical synapses, at which molecules are evolutionary conserved, vertebrate and invertebrate electrical synapses represent molecularly different structures that share a common communicating strategy that allows them to serve very similar functions. A better understanding of differences and commonalities regarding the structure, function and regulation of vertebrate and invertebrate electrical synapses will lead to a better understanding of the properties and functional diversity of this modality of synaptic communication. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 517-521, 2017.

Electrical synapses are neuronal gap junctions that are ubiquitous across brain regions and species. The biophysical properties of most electrical synapses are relatively simple-transcellular channels allow nearly ohmic, bidirectional flow of ionic current. Yet these connections can play remarkably diverse roles in different neural circuit contexts. Recent findings illustrate how electrical synapses may excite or inhibit, synchronize or desynchronize, augment or diminish rhythms, phase-shift, detect coincidences, enhance signals relative to noise, adapt, and interact with nonlinear membrane and transmitter-release mechanisms. Most of these functions are likely to be widespread in central nervous systems. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 610-624, 2017.

Rhythmic neuronal activity of multiple frequency bands has been described in many brain areas and attributed to numerous brain functions. Among these, little is known about the mechanism and role of infra-slow oscillations, which have been demonstrated recently in the mouse accessory olfactory bulb (AOB). Along with prolonged responses to stimuli and distinct network connectivity, they inexplicably affect the AOB processing of social relevant stimuli. Here, we show that assemblies of AOB mitral cells are synchronized by lateral interactions through chemical and electrical synapses. Using a network model, we demonstrate that the synchronous oscillations in these assemblies emerge from interplay between intrinsic membrane properties and network connectivity. As a consequence, the AOB network topology, in which each mitral cell receives input from multiple glomeruli, enables integration of chemosensory stimuli over extended time scales by interglomerular synchrony of infra-slow bursting. These results provide a possible functional significance for the distinct AOB physiology and topology. Beyond the AOB, this study presents a general model for synchronous infra-slow bursting in neuronal networks.SIGNIFICANCE STATEMENTInfra-slow rhythmic neuronal activity with a very long (>10 s) duration has been described in many brain areas, but little is known about the role of this activity and the mechanisms that produce it. Here, we combine experimental and computational methods to show that synchronous infra-slow bursting activity in mitral cells of the mouse accessory olfactory bulb (AOB) emerges from interplay between intracellular dynamics and network connectivity. In this novel mechanism, slow intracellular Na+dynamics endow AOB mitral cells with a weak tendency to burst, which is further enhanced and stabilized by chemical and electrical synapses between them. Combined with the unique topology of the AOB network, infra-slow bursting enables integration and binding of multiple chemosensory stimuli over a prolonged time scale.

In this study, we used the peristaltic crawling ofDrosophilalarvae as a model to study how motor patterns are regulated by central circuits. We built an experimental system that allows simultaneous application of optogenetics and calcium imaging to the isolated ventral nerve cord (VNC). We then investigated the effects of manipulating local activity of motor neurons (MNs) on fictive locomotion observed as waves of MN activity propagating along neuromeres. Optical inhibition of MNs with halorhodopsin3 in a middle segment (A4, A5, or A6), but not other segments, dramatically decreased the frequency of the motor waves. Conversely, local activation of MNs with channelrhodopsin2 in a posterior segment (A6 or A7) increased the frequency of the motor waves. Since peripheral nerves mediating sensory feedback were severed in the VNC preparation, these results indicate that MNs send signals to the central circuits to regulate motor pattern generation. Our results also indicate segmental specificity in the roles of MNs in motor control. The effects of the local MN activity manipulation were lost inshaking-B2(shakB2) orogre2, gap-junction mutations inDrosophila, or upon acute application of the gap junction blocker carbenoxolone, implicating electrical synapses in the signaling from MNs. Cell-type-specific RNAi suggestedshakBandogrefunction in MNs and interneurons, respectively, during the signaling. Our results not only reveal an unexpected role for MNs in motor pattern regulation, but also introduce a powerful experimental system that enables examination of the input–output relationship among the component neurons in this system.SIGNIFICANCE STATEMENTMotor neurons are generally considered passive players in motor pattern generation, simply relaying information from upstream interneuronal circuits to the target muscles. This study shows instead that MNs play active roles in the control of motor generation by conveying information via gap junctions to the central pattern-generating circuits in larvalDrosophila, providing novel insights into motor circuit control. The experimental system introduced in this study also presents a new approach for studying intersegmentally coordinated locomotion. Unlike traditional electrophysiology methods, this system enables the simultaneous recording and manipulation of populations of neurons that are genetically specified and span multiple segments.

Connexin36 (Cx36) constituent gap junctions (GJ) throughout the brain connect neurons into functional syncytia. In the retina they underlie the transmission, averaging and correlation of signals prior conveying visual information to the brain. This is the first study that describes retinal bipolar cell (BC) GJs in the human inner retina, whose function is enigmatic even in the examined animal models. Furthermore, a number of unique features (e.g. fovea, trichromacy, midget system) necessitate a reexamination of the animal model results in the human retina. Well-preserved postmortem human samples of this study are allowed to identify Cx36 expressing BCs neurochemically. Results reveal that both rod and cone pathway interneurons display strong Cx36 expression. Rod BC inputs to AII amacrine cells (AC) appear in juxtaposition to AII GJs, thus suggesting a strategic AII cell targeting by rod BCs. Cone BCs serving midget, parasol or koniocellular signaling pathways display a wealth of Cx36 expression to form homologously coupled arrays. In addition, they also establish heterologous GJ contacts to serve an exchange of information between parallel signaling streams. Interestingly, a prominent Cx36 expression was exhibited by midget system BCs that appear to maintain intimate contacts with bistratified BCs serving other pathways. These findings suggest that BC GJs in parallel signaling streams serve both an intra- and inter-pathway exchange of signals in the human 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 comprised 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.

Connexin-36 (Cx36) electrical synapses strengthen transmission in a calcium/calmodulin (CaM)/calmodulin-dependent kinase II (CaMKII)-dependent manner similar to a mechanism whereby the N-methyl-D-aspartate (NMDA) receptor subunit NR2B facilitates chemical transmission. Since NR2B&ndash, microtubule interactions recruit receptors to the cell membrane during plasticity, we hypothesized an analogous modality for Cx36. We determined that Cx36 binding to tubulin at the carboxy-terminal domain was distinct from Cx43 and NR2B by binding a motif overlapping with the CaM and CaMKII binding motifs. Dual patch-clamp recordings demonstrated that pharmacological interference of the cytoskeleton and deleting the binding motif at the Cx36 carboxyl-terminal (CT) reversibly abolished Cx36 plasticity. Mechanistic details of trafficking to the gap-junction plaque (GJP) were probed pharmacologically and through mutational analysis, all of which affected GJP size and formation between cell pairs. Lys279, Ile280, and Lys281 positions were particularly critical. This study demonstrates that tubulin-dependent transport of Cx36 potentiates synaptic strength by delivering channels to GJPs, reinforcing the role of protein transport at chemical and electrical synapses to fine-tune communication between neurons.

In order to respond to changing environments and fluctuations in internal states, animals adjust their behavior through diverse neuromodulatory mechanisms. In this study we show that electrical synapses between the ASH primary quinine-detecting sensory neurons and the neighboring ASK neurons are required for modulating the aversive response to the bitter tastant quinine in C. elegans. Mutant worms that lack the electrical synapse proteins INX-18 and INX-19 become hypersensitive to dilute quinine. Cell-specific rescue experiments indicate that inx-18 operates in ASK while inx-19 is required in both ASK and ASH for proper quinine sensitivity. Imaging analyses find that INX-19 in ASK and ASH localizes to the same regions in the nerve ring, suggesting that both sides of ASK-ASH electrical synapses contain INX-19. While inx-18 and inx-19 mutant animals have a similar behavioral phenotype, several lines of evidence suggest the proteins encoded by these genes play different roles in modulating the aversive quinine response. First, INX-18 and INX-19 localize to different regions of the nerve ring, indicating that they are not present in the same synapses. Second, removing inx-18 disrupts the distribution of INX-19, while removing inx-19 does not alter INX-18 localization. Finally, by using a fluorescent cGMP reporter, we find that INX-18 and INX-19 have distinct roles in establishing cGMP levels in ASK and ASH. Together, these results demonstrate that electrical synapses containing INX-18 and INX-19 facilitate modulation of ASH nociceptive signaling. Our findings support the idea that a network of electrical synapses mediates cGMP exchange between neurons, enabling modulation of sensory responses and behavior., Author summary Animals are constantly adjusting their behavior to respond to changes in the environment or to their internal state. This behavior modulation is achieved by altering the activity of neurons and circuits through a variety of neuroplasticity mechanisms. Chemical synapses are known to impact neuroplasticity in several different ways, but the diversity of mechanisms by which electrical synapses contribute is still being investigated. Electrical synapses are specialized sites of connection between neurons where ions and small signaling molecules can pass directly from one cell to the next. By passing small molecules through electrical synapses, neurons may be able to modify the activity of their neighbors. In this study we identify two genes that contribute to electrical synapses between two sensory neurons in C. elegans. We show that these electrical synapses are crucial for proper modulation of sensory responses, as without them animals are overly responsive to an aversive stimulus. In addition to pinpointing their sites of action, we present evidence that they may be contributing to neuromodulation by facilitating passage of the small molecule cGMP between neurons. Our work provides evidence for a role of electrical synapses in regulating animal behavior.

Summary Human pluripotent stem cells can be rapidly converted into functional neurons by ectopic expression of proneural transcription factors. Here we show that directly reprogrammed neurons, despite their rapid maturation kinetics, can model teratogenic mechanisms that specifically affect early neurodevelopment. We delineated distinct phases of in vitro maturation during reprogramming of human neurons and assessed the cellular phenotypes of valproic acid (VPA), a teratogenic drug. VPA exposure caused chronic impairment of dendritic morphology and functional properties of developing neurons, but not those of mature neurons. These pathogenic effects were associated with VPA-mediated inhibition of the histone deacetylase (HDAC) and glycogen synthase kinase-3 (GSK-3) pathways, which caused transcriptional downregulation of many genes, including MARCKSL1, an actin-stabilizing protein essential for dendritic morphogenesis and synapse maturation during early neurodevelopment. Our findings identify a developmentally restricted pathogenic mechanism of VPA and establish the use of reprogrammed neurons as an effective platform for modeling teratogenic pathways.

The complete description of the expression of gap junction proteins in the nervous system of the worm reveals a great complexity of their distribution amongst different neuronal classes, opening an unprecedented opportunity to expose the functional diversity of electrical synapses.

Common and rare variants of the CACNA1C voltage-gated calcium channel gene have been associated with autism and other neurodevelopmental disorders including schizophrenia, bipolar disorder and ADHD. However, little is known about how CACNA1C variants affect cellular processes to alter neurodevelopment. The Timothy syndrome mutation is a rare de novo gain-of-function variant in CACNA1C that causes autism with high penetrance, providing a powerful avenue into investigating the role of CACNA1C variants in neurodevelopmental disorders. Here, we use egl-19, the C. elegans homolog of CACNA1C, to investigate the role of voltage-gated calcium channels in autism. We show that an egl-19(gof) mutation that is equivalent to the Timothy syndrome mutation can alter axon targeting and affect behavior in C. elegans. We find that wildtype egl-19 negatively regulates axon termination. The egl-19(gof) mutation represses axon termination to cause axon targeting defects that lead to the misplacement of electrical synapses and alterations in habituation to light touch. Moreover, genetic interactions indicate that the egl-19(gof) mutation functions with genes that promote selective autophagy to cause defects in axon termination and behavior. These results reveal a novel genetic mechanism whereby a de novo mutation in CACNA1C can drive alterations in circuit formation and behavior., Author summary Autism is a disorder that affects neuronal development, leading to alterations in cognition and behavior. Imaging studies have revealed alterations in axonal connectivity as a key feature of autism. However, the underlying perturbations in cell biology that drive these alterations remain largely unknown. To address this issue, we have taken advantage of the Timothy syndrome mutation, a variant in a voltage-gated calcium channel that has the unusual property of causing autism with high penetrance. We identify a role for wild-type voltage-gated calcium channels in regulating axon targeting in C. elegans. Moreover, we find that two different versions of the Timothy syndrome mutation disrupt axon targeting. Our results suggest that the Timothy syndrome mutations disrupt axon targeting and behavior by interacting with genes that promote selective autophagy, the process through which cellular components are selected for degradation. These results reveal a mechanism through which variants in voltage-gated calcium channels can cause the disruptions in axonal connectivity that underlie autism.

Electrical synapses between neurons, also known as gap junctions, are direct cell membrane channels between adjacent neurons. Gap junctions play a role in the synchronization of neuronal network activity; however, their involvement in cognition has not been well characterized. Three-hour olfactory associative memory in Drosophila has two components: consolidated anesthesia-resistant memory (ARM) and labile anesthesia-sensitive memory (ASM). Here, we show that knockdown of the gap junction gene innexin5 (inx5) in mushroom body (MB) neurons disrupted ARM, while leaving ASM intact. Whole-mount brain immunohistochemistry indicated that INX5 protein was preferentially expressed in the somas, calyxes, and lobes regions of the MB neurons. Adult-stage-specific knockdown of inx5 in αβ neurons disrupted ARM, suggesting a specific requirement of INX5 in αβ neurons for ARM formation. Hyperpolarization of αβ neurons during memory retrieval by expressing an engineered halorhodopsin (eNpHR) also disrupted ARM. Administration of the gap junction blocker carbenoxolone (CBX) reduced the proportion of odor responsive αβ neurons to the training odor 3 hours after training. Finally, the α-branch-specific 3-hour ARM-specific memory trace was also diminished with CBX treatment and in inx5 knockdown flies. Altogether, our results suggest INX5 gap junction channels in αβ neurons for ARM retrieval and also provide a more detailed neuronal mechanism for consolidated memory in Drosophila., Author summary One of the most important questions in the neuroscience is how the brain process memory. Memory formation requires neuronal communication in the brain via synaptic transmissions, which include chemical and electrical synapses. Unlike the chemical synapses, the biological functions of electrical synapses in memory formation remain poorly understood. Here, we revealed that electrical synapses between mushroom body (MB) αβ neurons in Drosophila are critical for consolidated memory retrieval. We also showed that the electrical synapses are important for the branch-specific modification of calcium influx into the αβ neurons during memory retrieval. Our results provide novel insights into the molecular mechanisms and synaptic networks underlying memory retrieval.

Neuronal networks generating hindlimb locomotion are located in the spinal cord. The mechanisms underlying spinal rhythmogenesis are unknown but network activity and interconnectivity of excitatory interneurons likely play prominent roles. Here, we investigate interconnectivity within the Shox2 interneuron population, a subset of which has been suggested to be involved in locomotor rhythm generation, using paired recordings in isolated spinal cords or slices from transgenic mice. Sparse unidirectional connections consistent with chemical synaptic transmission and prominent bidirectional connections mediated by electrical synapses were present within distinct subsets of Shox2 interneurons. Moreover, bidirectional electrical connections were preferentially found between functionally-related Shox2 interneurons. Though prevalent in neonatal mice, electrical coupling began to decline in incidence and strength in mice ~ 3 weeks of age. Overall, our data suggest that gap junctional coupling promotes synchronization of Shox2 interneurons, and may be implicated in locomotor rhythmicity in developing mice.

Intercellular gap junction channels and single-membrane channels have been reported to regulate electrical synapse and the brain function. Innexin is known as a gap junction-related protein in invertebrates and is involved in the formation of intercellular gap junction channels and single-cell membrane channels. Multiple isoforms of innexin protein in each species enable the precise regulation of channel function. In molluscan species, sequence information of innexins is still limited and the sequences of multiple innexin isoforms have not been classified. This study examined the innexin transcripts expressed in the central nervous system of the terrestrial slugLimax valentianusand identified 16 transcripts of 12 innexin isoforms, including the splicing variants. We performed phylogenetic analysis and classified the isoforms with other molluscan innexin sequences. Next, the phosphorylation, N-glycosylation, and S-nitrosylation sites were predicted to characterize the innexin isoforms. Further, we identified 16 circular RNA sequences of nine innexin isoforms in the central nervous system ofLimax. The identification and classification of molluscan innexin isoforms provided novel insights for understanding the regulatory mechanism of innexin in this phylum.

SUMMARY Mutations in genes essential for synaptic function, such as the presynaptic adhesion molecule Neurexin1α (Nrxn1α), are strongly implicated in neuropsychiatric pathophysiology. As the input nucleus of the basal ganglia, the striatum integrates diverse excitatory projections governing cognitive and motor control, and its impairment may represent a recurrent pathway to disease. Here, we test the functional relevance of Nrxn1α in striatal circuits by employing optogenetic-mediated afferent recruitment of dorsal prefrontal cortical (dPFC) and parafascicular thalamic connections onto dorsomedial striatal (DMS) spiny projection neurons (SPNs). For dPFC-DMS circuits, we find decreased synaptic strength specifically onto indirect pathway SPNs in both Nrxn1α+/− and Nrxn1α−/− mice, driven by reductions in neurotransmitter release. In contrast, thalamic excitatory inputs to DMS exhibit relatively normal excitatory synaptic strength despite changes in synaptic N-methyl-D-aspartate receptor (NMDAR) content. These findings suggest that dysregulation of Nrxn1α modulates striatal function in an input- and target-specific manner., Graphical Abstract, In brief Davatolhagh and Fuccillo demonstrate that loss of Nrxn1α, a synaptic cell adhesion molecule, leads to divergent synaptic phenotypes within dorsal medial striatum for inputs from dorsal prefrontal cortex and parafascicular thalamic nucleus. These findings suggest input-specific imbalances in striatal circuit activity as a key perturbation in models of neurodevelopmental disorders.