Electrical synapses in mammalian CNS: Past eras,present focus and future directions

Gap junctions provide the basis for electrical synapses between neurons. Early studies in well-defined circuits in lower vertebrates laid the foundation for understanding various properties conferred by electrical synaptic transmission. Knowledge surrounding electrical synapses in mammalian systems unfolded first with evidence indicating the presence of gap junctions between neurons in various brain regions, but with little appreciation of their functional roles. Beginning at about the turn of this century, new approaches were applied to scrutinize electrical synapses, revealing the prevalence of neuronal gap junctions, the connexin protein composition of many of those junctions, and the myriad diverse neural systems in which they occur in the mammalian CNS. Subsequent progress indicated that electrical synapses constitute key elements in synaptic circuitry, govern the collective activity of ensembles of electrically coupled neurons, and in part orchestrate the synchronized neuronal network activity and rhythmic oscillations that underlie fundamental integrative processes. This article is part of a Special Issue entitled: Gap Junction Proteins edited by Jean Claude Herve.     

Electrical Transmission between Mammalian Neurons is Supported by a Small Fraction of Gap Junction Channels

Electrical synapses formed by gap junctions between neurons create networks of electrically coupled neurons in the mammalian brain, where these networks have been found to play important functional roles. In most cases, interneuronal gap junctions occur at remote dendro-dendritic contacts, making difficult accurate characterization of their physiological properties and correlation of these properties with their anatomical and morphological features of the gap junctions. In the mesencephalic trigeminal (MesV) nucleus where neurons are readily accessible for paired electrophysiological recordings in brain stem slices, our recent data indicate that electrical transmission between MesV neurons is mediated by connexin36 (Cx36)-containing gap junctions located at somato-somatic contacts. We here review evidence indicating that electrical transmission between these neurons is supported by a very small fraction of the gap junction channels present at cell-cell contacts. Acquisition of this evidence was enabled by the unprecedented experimental access of electrical synapses between MesV neurons, which allowed estimation of the average number of open channels mediating electrical coupling in relation to the average number of gap junction channels present at these contacts. Our results indicate that only a small proportion of channels (~0.1?%) appear to be conductive. On the basis of similarities with other preparations, we postulate that this phenomenon might constitute a general property of vertebrate electrical synapses, reflecting essential aspects of gap junction function and maintenance.     

Activity-dependent plasticity of electrical synapses: increasing evidence for its presence and functional roles in the mammalian brain

Gap junctions mediate electrical synaptic transmission between neurons. While the actions of neurotransmitter modulators on the conductance of gap junctions have been extensively documented, increasing evidence indicates they can also be influenced by the ongoing activity of neural networks, in most cases via local interactions with nearby glutamatergic synapses. We review here early evidence for the existence of activity-dependent regulatory mechanisms as well recent examples reported in mammalian brain. The ubiquitous distribution of both neuronal connexins and the molecules involved suggest this phenomenon is widespread and represents a property of electrical transmission in general.     

Characteristics and plasticity of electrical synaptic transmission

Electrical synapses are an omnipresent feature of nervous systems, from the simple nerve nets of cnidarians to complex brains of mammals. Formed by gap junction channels between neurons, electrical synapses allow direct transmission of voltage signals between coupled cells. The relative simplicity of this arrangement belies the sophistication of these synapses. Coupling via electrical synapses can be regulated by a variety of mechanisms on times scales ranging from milliseconds to days, and active properties of the coupled neurons can impart emergent properties such as signal amplification, phase shifts and frequency-selective transmission. This article reviews the biophysical characteristics of electrical synapses and some of the core mechanisms that control their plasticity in the vertebrate central nervous system.     

Gap junctions as electrical synapses

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

'Non-synaptic' mechanisms in seizures and epileptogenesis

The role of 'non-synaptic' mechanisms (i.e. those mechanisms that are independent of active chemical synpases) in the synchronization of neuronal activity during seizures and their possible contribution to chronic epileptogenesis are summarized. These 'non-synaptic' mechanisms include electrotonic coupling through gap junctions, electrical field effects (i.e. ephaptic transmission), and ionic interactions (e.g. increases in the extracellular concentration of K(+)). Several lines of evidence indicate that granule cells and pyramidal cells of the hippocampus, and probably other cortical neurons, can generate synchronized electrical activity after active chemical synaptic transmission has been blocked. This synchronized activity is sensitive to alterations in the size of the extracellular space, thus suggesting that electrical field effects and ionic mechanisms contribute to this synchronized activity. Recent studies also indicate that 'non-synaptic' synchronization is quite prominent early in development. Electrophysiological data from hippocampal and neocortical slices have led to a re-interpretation of the fast prepotentials (i.e. partial spikes) recorded in cortical pyramidal cells, suggesting that they may not be due to dendritic spike generation. Improvement in freeze-fracture ultrastructural techniques have led to a re-assessment of previous data on gap junctions in the nervous system and opened new approaches to the quantitative analysis and characterization of gap junctions on glia and neurons. Finally, new methods of dye/tracer coupling have the potential to provide a more rigorous basis for evaluating gap junctions and electrotonic communication between neurons in the mammalian central nervous system. Therefore, recent data continue to suggest that gap junctions and electrotonic coupling play an important role in neural integration, although additional studies using new techniques will be needed to address some of the controversial issues that have arisen over the last several decades.     

Topology of gap junction networks in C. elegans

Gap junctions are prevalent in every nervous system, but their role in information processing remains largely unknown. In C. elegans, the role of gap junctional communication in touch sensitivity has been demonstrated. In this animal, the entire complement of gap junctions in the nervous system is documented, therefore providing a good model for the computational investigation of circuit functions of gap junctions.We explored several hypotheses about the role of gap junctions in the nervous system of C. elegans by systematically analysing an anatomical database with recursive algorithms. We find that gap junctions connect different sets of neurons from those connected by chemical synapses. In addition, when analysing the topology of the gap-junction networks, we find that, surprisingly, most (92%) neurons in the worm are linked in a single gap-junction network. The worm nervous system can only be divided into smaller networks by assuming that two or more gap junctions are necessary for functional coupling or that neural activity has limited propagation. However, these groups, and others identified using algorithms with subsets or combinations of restrictive criteria, do not correspond to any known circuits identified in genetic and behavioral studies. Finally, we notice that the function of some gap junctions appears linked to their precise location on the neuronal processes. We propose that the location of the gap junctions within the neuron determines their functional role.     

Electrical coupling and neuronal synchronization in the Mammalian brain

Certain neurons in the mammalian brain have long been known to be joined by gap junctions, which are the most common type of electrical synapse. More recently, cloning of neuron-specific connexins, increased capability of visualizing cells within brain tissue, labeling of cell types by transgenic methods, and generation of connexin knockouts have spurred a rapid increase in our knowledge of the role of gap junctions in neural activity. This article reviews the many subtleties of transmission mediated by gap junctions and the mechanisms whereby these junctions contribute to synchronous firing.     

Cut-loading: A Useful Tool for Examining the Extent of Gap Junction Tracer Coupling Between Retinal Neurons

In addition to chemical synaptic transmission, neurons that are connected by gap junctions can also communicate rapidly via electrical synaptic transmission. Increasing evidence indicates that gap junctions not only permit electrical current flow and synchronous activity between interconnected or coupled cells, but that the strength or effectiveness of electrical communication between coupled cells can be modulated to a great extent^1,2. In addition, the large internal diameter (~1.2 nm) of many gap junction channels permits not only electric current flow, but also the diffusion of intracellular signaling molecules and small metabolites between interconnected cells, so that gap junctions may also mediate metabolic and chemical communication. The strength of gap junctional communication between neurons and its modulation by neurotransmitters and other factors can be studied by simultaneously electrically recording from coupled cells and by determining the extent of diffusion of tracer molecules, which are gap junction permeable, but not membrane permeable, following iontophoretic injection into single cells. However, these procedures can be extremely difficult to perform on neurons with small somata in intact neural tissue.Numerous studies on electrical synapses and the modulation of electrical communication have been conducted in the vertebrate retina, since each of the five retinal neuron types is electrically connected by gap junctions^3,4. Increasing evidence has shown that the circadian (24-hour) clock in the retina and changes in light stimulation regulate gap junction coupling^3-8. For example, recent work has demonstrated that the retinal circadian clock decreases gap junction coupling between rod and cone photoreceptor cells during the day by increasing dopamine D2 receptor activation, and dramatically increases rod-cone coupling at night by reducing D2 receptor activation^7,8. However, not only are these studies extremely difficult to perform on neurons with small somata in intact neural retinal tissue, but it can be difficult to adequately control the illumination conditions during the electrophysiological study of single retinal neurons to avoid light-induced changes in gap junction conductance.Here, we present a straightforward method of determining the extent of gap junction tracer coupling between retinal neurons under different illumination conditions and at different times of the day and night. This cut-loading technique is a modification of scrape loading^9-12, which is based on dye loading and diffusion through open gap junction channels. Scrape loading works well in cultured cells, but not in thick slices such as intact retinas. The cut-loading technique has been used to study photoreceptor coupling in intact fish and mammalian retinas^{7, 8,13}, and can be used to study coupling between other retinal neurons, as described here.     

神经组织缝隙连接

近年来神经组织中缝隙连接的分布和功能研究取得了一些显著进展。分子生物学方法的应用促进了ＧＪ结构,亚型及生物物理特性的揭示,染料偶联实验和Ｃａ＾２＋成像技术为ＧＪ的功能研究提供了直观有效的手段。ＧＪ的调控涉及ＧＪ的表达,导通性的改变等环节。ＧＪ胞间通讯的基本形式是交换第二信使和电偶联。     

Gap Junctions and Epileptic Seizures – Two Sides of the Same Coin?

Electrical synapses (gap junctions) play a pivotal role in the synchronization of neuronal ensembles which also makes them likely agonists of pathological brain activity. Although large body of experimental data and theoretical considerations indicate that coupling neurons by electrical synapses promotes synchronous activity (and thus is potentially epileptogenic), some recent evidence questions the hypothesis of gap junctions being among purely epileptogenic factors. In particular, an expression of inter-neuronal gap junctions is often found to be higher after the experimentally induced seizures than before. Here we used a computational modeling approach to address the role of neuronal gap junctions in shaping the stability of a network to perturbations that are often associated with the onset of epileptic seizures. We show that under some circumstances, the addition of gap junctions can increase the dynamical stability of a network and thus suppress the collective electrical activity associated with seizures. This implies that the experimentally observed post-seizure additions of gap junctions could serve to prevent further escalations, suggesting furthermore that they are a consequence of an adaptive response of the neuronal network to the pathological activity. However, if the seizures are strong and persistent, our model predicts the existence of a critical tipping point after which additional gap junctions no longer suppress but strongly facilitate the escalation of epileptic seizures. Our results thus reveal a complex role of electrical coupling in relation to epileptiform events. Which dynamic scenario (seizure suppression or seizure escalation) is ultimately adopted by the network depends critically on the strength and duration of seizures, in turn emphasizing the importance of temporal and causal aspects when linking gap junctions with epilepsy.     

Connexin36 mediates spike synchrony in olfactory bulb glomeruli

Neuronal synchrony is important to network behavior in many brain regions. In the olfactory bulb, principal neurons (mitral cells) project apical dendrites to a common glomerulus where they receive a common input. Synchronized activity within a glomerulus depends on chemical transmission but mitral cells are also electrically coupled. We examined the role of connexin-mediated gap junctions in mitral cell coordinated activity. Electrical coupling as well as correlated spiking between mitral cells projecting to the same glomerulus was entirely absent in connexin36 (Cx36) knockout mice. Ultrastructural analysis of glomeruli confirmed that mitral-mitral cell gap junctions on distal apical dendrites contain Cx36. Coupled AMPA responses between mitral cell pairs were absent in the knockout, demonstrating that electrical coupling, not transmitter spillover, is responsible for synchronization. Our results indicate that Cx36-mediated gap junctions between mitral cells orchestrate rapid coordinated signaling via a novel form of electrochemical transmission.     

Gap junctional communication among motor and other neurons shapes patterns of neural activity and synaptic connectivity during development

We are studying the functional roles of neuronal gap junctional coupling during development, using motor neurons and their synapses with muscle fibers as a model system. At neuromuscular synapses, several studies have shown that the relative pattern of activity among motor inputs competing for innervation of the same target muscle fiber determines how patterns of innervation are sculpted during the first weeks after birth. We asked whether gap junctional coupling among motor neurons modulates the relative timing of motor neuron activity in awake, behaving neonatal mice. We found that the activity of motor neurons innervating the same muscle is temporally correlated perinatally, during the same period that gap junction-mediated electrical and dye coupling are present. In vivo blockade of gap junctions abolished temporal correlations in motor neuron activity, without changing overall motor behavior, motor neuron activity patterns or firing frequency. Together with preliminary studies in mice lacking gap junction protein Cx40, our data suggest that developmentally regulated gap junctional coupling among motor and other neurons affects the activity in nascent neural circuits and thus in turn affects synaptic connectivity. Dynamic monitoring of dye coupling can be used to explore this possibility in normal mice and in mice lacking gap junction proteins during embryonic and neonatal development.     

Electrical synapses between GABA-releasing interneurons

Although gap junctions were first demonstrated in the mammalian brain about 30 years ago, the distribution and role of electrical synapses have remained elusive. A series of recent reports has demonstrated that inhibitory interneurons in the cerebral cortex, thalamus, striatum and cerebellum are extensively interconnected by electrical synapses. Investigators have used paired recordings to reveal directly the presence of electrical synapses among identified cell types. These studies indicate that electrical coupling is a fundamental feature of local inhibitory circuits and suggest that electrical synapses define functionally diverse networks of GABA-releasing interneurons. Here, we discuss these results, their possible functional significance and the insights into neuronal circuit organization that have emerged from them.     

Electrical synapses: a dynamic signaling system that shapes the activity of neuronal networks

Gap junctions consist of intercellular channels dedicated to providing a direct pathway for ionic and biochemical communication between contacting cells. After an initial burst of publications describing electrical coupling in the brain, gap junctions progressively became less fashionable among neurobiologists, as the consensus was that this form of synaptic transmission would play a minimal role in shaping neuronal activity in higher vertebrates. Several new findings over the last decade (e.g. the implication of connexins in genetic diseases of the nervous system, in processing sensory information and in synchronizing the activity of neuronal networks) have brought gap junctions back into the spotlight. The appearance of gap junctional coupling in the nervous system is developmentally regulated, restricted to distinct cell types and persists after the establishment of chemical synapses, thus suggesting that this form of cell-cell signaling may be functionally interrelated with, rather than alternative to chemical transmission. This review focuses on gap junctions between neurons and summarizes the available data, derived from molecular, biological, electrophysiological, and genetic approaches, that are contributing to a new appreciation of their role in brain function.     

Ultrastructure, histological distribution, and freeze-fracture immunocytochemistry of gap junctions in rat brain and spinal cord

The historical development of concepts of gap junctions as sites for electrical, ionic, and metabolic coupling is reviewed, from the initial discovery of gap junctions linking heart cells, to the current concepts that gap junctions represent 'electrotonic synapses' between neurons. The ultrastructure and immunocytochemistry of gap junctions in heart, brain, and spinal cord of adult rats is examined using conventional thin sections, negative staining, grid-mapped freeze-fracture replicas, and immunogold-labeled freeze-fracture replicas. We review evidence for neuronal gap junctions at 'mixed' (combined electrical and chemical) synapses throughout adult rat spinal cord. We also show immunogold labeling of connexin43 in astrocyte and ependymocyte gap junctions and of connexin32 in oligodendrocyte gap junctions. Ultrastructural and freeze-fracture immunocytochemical methods have provided for definitive determination of the number, size, histological distribution, and connexin composition of gap junctions between neurons in all regions of the central nervous systems of vertebrate species.     

Functional alterations in gut contractility after connexin36 ablation and evidence for gap junctions forming electrical synapses between nitrergic enteric neurons

Neurons in the enteric nervous system utilize numerous neurotransmitters to orchestrate rhythmic gut smooth muscle contractions. We examined whether electrical synapses formed by gap junctions containing connexin36 also contribute to communication between enteric neurons in mouse colon. Spontaneous contractility properties and responses to electrical field stimulation and cholinergic agonist were altered in gut from connexin36 knockout vs. wild-type mice. Immunofluorescence revealed punctate labelling of connexin36 that was localized at appositions between somata of enteric neurons immunopositive for the enzyme nitric oxide synthase. There is indication for a possible functional role of gap junctions between inhibitory nitrergic enteric neurons.     

Expression and functions of neuronal gap junctions

Gap junctions are channel-forming structures in contacting plasma membranes that allow direct metabolic and electrical communication between almost all cell types in the mammalian brain. At least 20 connexin genes and 3 pannexin genes probably code for gap junction proteins in mice and humans. Gap junctions between murine neurons (also known as electrical synapses) can be composed of connexin 36, connexin 45 or connexin 57 proteins, depending on the type of neuron. Furthermore, pannexin 1 and 2 are likely to form electrical synapses. Here, we discuss the roles of connexin and pannexin genes in the formation of neuronal gap junctions, and evaluate recent functional analyses of electrical synapses that became possible through the characterization of mouse mutants that show targeted defects in connexin genes.     

Electrotonic synapses in the mammalian spinal cord

By means of light and electron microscopy methods structural peculiarities of motor nuclei have been studied in the rat spinal cord (17 animals) on the 1st-3d and on the 10th-18th days of postnatal ontogenesis. Synaptic junctions of the gap type are revealed; they are considered as electrotonic synapses. Dendro-somatic and dendrodendritic synaptic junctions of the gap type are found. Together with the electrotonic synapses, morphologically mixed synapses of axo-somatic and axo-axonal types are disclosed; they contain, besides organells, specific for chemical synapses, close opposition areas of pre- and postsynaptic membranes of the gap junction type. Morphologically mixed synapses occur in neuropil of the motor nuclei of the spinal cord in young rats of all age groups studied. Homologous synapses are detected in the motor nuclei of the white mouse spinal cord. Synaptic junctions of the gap type in the mammalian spinal cord could be a substrate of electrical interaction between its motor neurons.     

A biophysical model of vertebrate olfactory epithelium and bulb exhibiting gap junction dependent odor-evoked spatiotemporal patterns of activity

This work describes a biophysical model of the initial stages of vertebrate olfactory system containing structures representing the olfactory epithelium and bulb. Its main novelty is the introduction of gap junctions connecting neurons both in the epithelium and bulb, and of biologically detailed dendrodendritic synapses between granule and mitral cells in the bulb. The model was used to simulate the effect of an odor presentation on the neural activity pattern in the epithelium and bulb. During the time for which an odor is presented with a constant concentration, there are spatiotemporal patterns in the epithelium and bulb generated by the couplings due to the gap junctions and/or dendrodendritic synapses. A study varying the strength of the gap junction coupling shows that the spatiotemporal patterns, both in the epithelium and bulb, are dependent of the coupling strength. It is also shown that the olfactory bulb's spatiotemporal pattern depends on the existence of the dendrodendritic connections between mitral and granule cells. If these spatiotemporal patterns really exist in the early processing stages of the olfactory system they may be used for odor coding and the gap junctions and dendrodendritic synapses might have a role on it.