Wiring patterns from auditory sensory neurons to the escape and song-relay pathways in fruit flies

Many animals rely on acoustic cues to decide what action to take next. Unraveling the wiring patterns of the auditory neural pathways is prerequisite for understanding such information processing. Here, we reconstructed the first step of the auditory neural pathway in the fruit fly brain, from primary to secondary auditory neurons, at the resolution of transmission electron microscopy. By tracing axons of two major subgroups of auditory sensory neurons in fruit flies, low-frequency tuned Johnston's organ (JO)-B neurons and high-frequency tuned JO-A neurons, we observed extensive connections from JO-B neurons to the main second-order neurons in both the song-relay and escape pathways. In contrast, JO-A neurons connected strongly to a neuron in the escape pathway. Our findings suggest that heterogeneous JO neuronal populations could be recruited to modify escape behavior whereas only specific JO neurons contribute to courtship behavior. We also found that all JO neurons have postsynaptic sites at their axons. Presynaptic modulation at the output sites of JO neurons could affect information processing of the auditory neural pathway in flies.     

A bushy cell network in the rat ventral cochlear nucleus

Geometry of the dendritic tree and synaptic organization of afferent inputs are essential factors in determining how synaptic input is integrated by neurons. This information remains elusive for one of the first brainstem neurons involved in processing of the primary auditory signal from the ear, the bushy cells (BCs) of the ventral cochlear nucleus (VCN). Here, we labeled the BC dendritic trees with retrograde tracing techniques to analyze their geometry and synaptic organization after immunofluorescence for excitatory and inhibitory synaptic markers, electron microscopy, morphometry, double tract‐tracing methods, and 3D reconstructions. Our study revealed that BC dendrites provide space for a large number of compartmentalized excitatory and inhibitory synaptic interactions. The dendritic inputs on BCs are of cochlear and noncochlear origin, and their proportion and distribution are dependent on the branching pattern and orientation of the dendritic tree in the VCN. Three‐dimensional reconstructions showed that BC dendrites branch and cluster with those of other BCs in the core of the VCN. Within the cluster, incoming synaptic inputs establish divergent multiple‐contact synapses (dyads and triads) between BCs. Furthermore, neuron–neuron connections including puncta adherentia, sarcoplasmic junctions, and gap junctions are common between BCs, which suggests that these neurons are electrically coupled. Overall, our study demonstrates the existence of a BC network in the rat VCN. This network may establish the neuroanatomical basis for acoustic information processing by individual BCs as well as for enhanced synchronization of the output signal of the VCN. J. Comp. Neurol. 516:241–263, 2009. © 2009 Wiley‐Liss, Inc.     

Neurotransmitters and gap junctions in developing neural circuits

A growing body of evidence suggests that highly correlated, spontaneous neural activity plays an important role in shaping connections in the developing nervous system prior to the maturation of sensory afferents. In this article we discuss the mechanisms involved in the generation and the regulation of spontaneous activity patterns in the developing retina and the developing neocortex. Spontaneous activity in the developing retina propagates across the ganglion cell layer as waves of action potentials and drives rhythmic increases in intracellular calcium in retinal neurons. Retinal waves are mediated by a combination of chemical synaptic transmission and gap junctions, and the circuitry responsible for generating retinal waves changes with age and between species. In the developing cortex, spontaneous calcium elevations propagate across clusters of cortical neurons called domains. Cortical domains are generated by a regenerative mechanism involving second messenger diffusion through gap junctions and subsequent calcium release from internal stores. The neocortical gap junction system is regulated by glutamate-triggered second messenger systems as well as neuromodulatory transmitters, suggesting extensive interactions between synaptic transmission and information flow through gap junctions. The interaction between gap junctions and chemical synaptic transmission observed in these developing networks represent a powerful mechanism by which activity across large groups of neurons can be correlated.     

Lack of biocytin transfer at gap junctions in the chicken vestibular nuclei

In vivo experiments were designed to test for functional gap junctions at ‘mixed' synapses that were morphologically characterized between the large-diameter, primary vestibular fibers and second-order vestibular neurons in the chicken, Gallus gallus. In previous intracellular recordings and dye injections into these neurons from brain slice preparations of chick embryos (E15/16) and also newborn hatchlings (H1-2), no evidence was obtained for functional gap junctions. Therefore, biocytin, a low molecular weight tracer that permeates gap junction channels, was extracellularly applied to either the ampullary nerves or to the vestibular ganglion of 3–6 day old hatchlings and adult chickens (9 months). This procedure resulted in the uptake of the dye and heavy staining of both the thick and thin fibers composing the vestibular nerve and in loading of vestibular efferent neurons. However, no dye transfer was observed between the large-diameter, primary vestibular fibers and second-order vestibular neurons. This observation, which was performed using a relatively non-invasive approach on intact animals, suggests that the gap junctions at these mixed synapses are probably not functional under the conditions of these experiments.     

Organization of efferent peripheral synapses at mechanosensory neurons in spiders

The mechanosensory neurons of arachnids receive diverse synaptic inputs in the periphery. The function of most of these synapses, however, is unknown. We have carried out detailed electron microscopic investigations of the peripheral synapses at sensory neurons in the compound slit sense organ VS-3 of the spider Cupiennius salei. Based on the localization of discrete presynaptic vesicle populations, it is possible to discriminate at least four different synapse types, containing either: (1) small round, electron-lucent vesicles 32 nm in diameter; (2) large round, clear 42-nm vesicles; (3) a mixture of small and large clear, round vesicles, similar in size to those in Type 1 and Type 2 synapses, respectively, and granular and dense-core vesicles; or (4) clear, round 37- to 65-nm vesicles. Combined immunocytochemical labeling at the light and the electron microscopic level suggests that gamma-aminobutyric acid (GABA) is the transmitter in many of the 32-nm vesicle synapses, and glutamate in many of the 42-nm ones. Based on vesicle type and particular synaptic configuration, various forms of presumed efferent synaptic contacts are distinguishable with the sensory neurons, the surrounding glia, and between the putative efferent fibers themselves. These include simple unidirectional synapses, reciprocal synapses, serial synapses, and convergent as well as divergent dyads. These various synaptic microcircuits are suited to serve a variety of functions. Among these are direct postsynaptic inhibition or excitation of the mechanosensory neurons, and disinhibition or sensitization via presynaptic inhibition or excitation. The observed synaptic configurations are compared with those at the crustacean muscle receptor organ. They reveal a remarkable complexity of synaptic microcircuits at spider sensilla and suggest manifold possibilities for subtle, efferent control of sensory activity.     

Electrical synapse formation disrupts calcium-dependent exocytosis, but not vesicle mobilization

Electrical coupling exists prior to the onset of chemical connectivity at many developing and regenerating synapses. At cholinergic synapses in vitro, trophic factors facilitated the formation of electrical synapses and interfered with functional neurotransmitter release in response to photolytic elevations of intracellular calcium. In contrast, neurons lacking trophic factor induction and electrical coupling possessed flash-evoked transmitter release. Changes in cytosolic calcium and postsynaptic responsiveness to acetylcholine were not affected by electrical coupling. These data indicate that transient electrical synapse formation delayed chemical synaptic transmission by imposing a functional block between the accumulation of presynaptic calcium and synchronized, vesicular release. Despite the inability to release neurotransmitter, neurons that had possessed strong electrical coupling recruited secretory vesicles to sites of synaptic contact. These results suggest that the mechanism by which neurotransmission is disrupted during electrical synapse formation is downstream of both calcium influx and synaptic vesicle mobilization. Therefore, electrical synaptogenesis may inhibit synaptic vesicles from acquiring a readily releasable state. We hypothesize that gap junctions might negatively interact with exocytotic processes, thereby diminishing chemical neurotransmission.     

Cell types and synaptic organization of the medullary electromotor nucleus in a constant frequency weakly electric fish,Sternarchus albifrons

The medullary electromotor nucleus (EMN) of Sternarchus albifrons was studied at the light and electron microscopic levels. The EMN consists of a dense meshwork of myelinated axons and glial elements with interposed large neurons; it is provided with an abundant supply of capillaries. Two types of essentially adendritic nerve cells were distinguished on the basis of size: giant neurons (approx. 70 μm in diameter) and large neurons (approx. 30 μm in diameter). Their population ratio is 1:4. Only giant cells are labelled following the injection of retrograde tracer into the spinal cord; they are therefore identified with the so-called “relay cells” of other gymnotids. Tracer experiments further suggest that the descending axons of these relay cells give off collateral branches throughout the elongated spinal electromotor nucleus. In contrast, the large cells remain unlabelled and therefore lack spinal projections; they most likely correspond to “pacemaker cells”. The perikaryal surface, including axon hillock and proximal part of initial segment of both types of EMN cells, is contacted by clusters of synaptic terminals and astrocytic processes. Two main varieties of synaptic terminals occur: (1) large endings and (2) ordinary end feet with standard size (S-type) and variable size (Sv-type) clear, spherical vesicles. The junction between large endings and EMN cells is characterized by the combination of gap junctions and surrounding intermediate junctions whose freeze-fracture characteristics were morphometrically analyzed. The large endings were formed by nodes of Ranvier as well as by fiber terminations, and synchronization within the EMN may be achieved by presynaptic fibers. Some of the contacts occur directly on the initial segment, which could allow activity to bypass the soma. It is concluded that the electromotor system of Sternarchus is comprised of a rapid conduction pathway where medullary pacemaker and relay cells as well as spinal electromotor neurons are coupled by synapses with gap junctions. In contrast to the spinal electromotor neurons, the medullary EMN cells receive synapses with morphological characteristics of chemical transmission, and the S-type and Sv-type terminals may possibly correspond to Gray's Type I and Type II synapses, respectively. These synapses may be involved in modulation of the electric organ discharge frequency.     

Activity-dependent editing of neuromuscular synaptic connections

Work over the past four decades has suggested that neural activity edits synaptic connections throughout the developing nervous system. Synaptic editing is shaped in large part by competitive interactions among different inputs innervating the same target cell that profoundly influence synaptic strength and structure. While competition plays out among presynaptic inputs that anterogradely influence their targets, postsynaptic target cells also modulate competition, in part through retrograde interactions that modulate presynaptic neurotransmitter release. One of the most useful synapses for studying how neural activity mediates synaptic editing is the connections between spinal motor neurons and skeletal muscle fibers, called neuromuscular junctions. Here we review current ideas about the role of activity in editing neuromuscular synaptic connections. The mechanisms by which activity mediates synaptic competition at these peripheral synapses are relevant to understanding how neural circuits in the central nervous system are continually altered by experience throughout life.     

Null mutation in shaking-B eliminates electrical,but not chemical,synapses in the Drosophila giant fiber system: A structural study

Mutations in the Drosophila shaking-B gene perturb synaptic transmission and dye coupling in the giant fiber escape system. The GAL4 upstream activation sequence system was used to express a neuronal-synaptobrevin-green fluorescent protein (nsyb-GFP) construct in the giant fibers (GFs); nsyb-GFP was localized where the GFs contact the peripherally synapsing interneurons (PSIs) and the tergotrochanteral motorneurons (TTMns). Antibody to Shaking-B protein stained plaquelike structures in the same regions of the GFs, although not all plaques colocalized with nsyb-GFP. Electron microscopy showed that the GF-TTMn and GF-PSI contacts contained many chemical synaptic release sites. These sites were interposed with extensive regions of close membrane apposition (3.25 nm ± 0.12 separation), with faint cross striations and a single-layered array of 41-nm vesicles on the GF side of the apposition. These contacts appeared similar to rectifying electrical synapses in the crayfish and were eliminated in shaking-B² mutants. At mutant GF-TTMn and GF-PSI contacts, chemical synapses and small regions of close membrane apposition, more similar to vertebrate gap junctions, were not affected. Gap junctions with more vertebratelike separation of membranes (1.41 nm ± 0.08) were abundant between peripheral perineurial glial processes; these were unaffected in the mutants. J. Comp. Neurol. 404:449–458, 1999. © 1999 Wiley-Liss, Inc.     

Freeze-fracture study of the large myelinated club ending synapse on the goldfish Mauthner cell: special reference to the quantitative analysis of gap junctions

The large myelinated club endings (LMCEs) of primary eighth nerve afferents form mixed synapses on the lateral dendrite of the giant Mauthner cell. The double replica freeze-fracture technique was employed to examine the intramembrane fine structure of these LMCE synapses. Morphological correlates of both chemical and electrical transmission were found at the LMCE synapses. Electrical synaptic junctions, or gap junctions, were located over much (10-20%) of the synaptic contact. These were seen in both pre-and postsynaptic membrane as tightly packed P face particle aggregates and corresponding aggregates of E face pits. Specializations characteristic of chemical synaptic junctions were most prominent at the periphery of the synaptic contact. These specializations consisted of postsynaptic E face particle aggregates which were subjacent to presynaptic active zones. The active zones were distinguishable as regions with an increased density of large particles and vesicle attachment sites represented by P face depressions and E face protuberances. Quantitative analysis of gap junction particle (connexon) number at five LMCEs revealed 24,000-106,000 connexons per LMCE. Comparison with data from electrophysiological studies of single LMCEs indicates that only a small fraction of the connexon channels are open at any given time during electrotonic transmission at an LMCE synapse.     

Spatiotemporal distribution of Connexin45 in the olivocerebellar system

The olivocerebellar system is involved in the transmission of information to maintain sensory motor coordination. Gap junctions have been described in various types of neurons in this system, including the neurons in the inferior olive that provide the climbing fibers to Purkinje cells. While it is well established that Connexin36 is necessary for the formation of these neuronal gap junctions, it is not clear whether these electrical synapses can develop without Connexin45. Here we describe the development and spatiotemporal distribution of Connexin45 in relation to that of Connexin36 in the olivocerebellar system. During development Connexin45 is expressed in virtually all neurons of the inferior olive and cerebellar nuclei. During later postnatal development and adulthood there is a considerable overlap of expression of both connexins in subpopulations of all main olivary nuclei and cerebellar nuclei as well as in the stellate cells in the cerebellar cortex. Despite this prominent expression of Connexin45, ultrastructural analysis of neuronal gap junctions in null-mutants of Connexin45 showed that their formation appears normal in contrast to that in knockouts of Connexin36. These morphological data suggest that Connexin45 may play a modifying role in widely distributed, coupled neurons of the olivocerebellar system, but that it is not essential for the creation of its neuronal gap junctions.     

Synaptic circuitry of identified neurons in the antennal lobe of Drosophila melanogaster

In Drosophila melanogaster olfactory sensory neurons (OSNs) establish synapses with projection neurons (PNs) and local interneurons within antennal lobe (AL) glomeruli. Substantial knowledge regarding this circuitry has been obtained by functional studies, whereas ultrastructural evidence of synaptic contacts is scarce. To fill this gap, we studied serial sections of three glomeruli using electron microscopy. Ectopic expression of a membrane‐bound peroxidase allowed us to map synaptic sites along PN dendrites. Our data prove for the first time that each of the three major types of AL neurons is both pre‐ and postsynaptic to the other two types, as previously indicated by functional studies. PN dendrites carry a large proportion of output synapses, with approximately one output per every three input synapses. Detailed reconstructions of PN dendrites showed that these synapses are distributed unevenly, with input and output sites partially segregated along a proximal–distal gradient and the thinnest branches carrying solely input synapses. Moreover, our data indicate synapse clustering, as we found evidence of dendritic tiling of PN dendrites. PN output synapses exhibited T‐shaped presynaptic densities, mostly arranged as tetrads. In contrast, output synapses from putative OSNs showed elongated presynaptic densities in which the T‐bar platform was supported by several pedestals and contacted as many as 20 postsynaptic profiles. We also discovered synaptic contacts between the putative OSNs. The average synaptic density in the glomerular neuropil was about two synapses/µm³. These results are discussed with regard to current models of olfactory glomerular microcircuits across species. J. Comp. Neurol. 524:1920–1956, 2016. © 2016 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc.     

Schwann cell guidance of nerve growth between synaptic sites explains changes in the pattern of muscle innervation and remodeling of synaptic sites following peripheral nerve injuries

Terminal Schwann cells (SCs) are nonmyelinating glia that are a prominent component of the neuromuscular junction (NMJ) where motor neurons form synapses onto muscle fibers. These cells play important roles not only in development and maintenance of the neuromuscular synapse but also restoring synaptic function after nerve damage. In response to muscle denervation, terminal SCs undergo dramatic changes in their gene expression patterns as well as in their morphology, such as extending elaborate processes into inter-junctional space. These SC processes serve as a path to guide axon terminal sprouts from nearby innervated junctions, promoting rapid reinnervation of denervated fibers. We studied the role of terminal SCs in synapse reformation by using two different fluorescent proteins to simultaneously label motor axons and SCs; we examined these junctions repeatedly in living animals using a fluorescence microscope. Here, we show that alterations in the patterns of muscle innervation following recovery from nerve injury can be explained by SC guidance of regenerating axons. In turn, this guidance leads to remodeling of the NMJ itself.     

Mormyromast electroreceptor organs and their afferent fibers in mormyrid fish: I. Morphology

Mormyromast electroreceptor organs are the most numerous type of electroreceptor organs in mormyrid electric fish and provide the sensory information necessary for active electrolocation. Mormyromast organs and their primary afferent fibers have not been studied very extensively. Both morphological and physiological questions remain to be answered before the neural basis of active electrolocation in mormyrids can be understood. This paper examines four different aspects of the morphology of mormyromast organs and afferent fibers: 1) Mormyromast organs in the skin. The innervation patterns for the two types of separately innervated sensory cells in the mormyromast organ are described on the basis of silver-stained whole mounts of skin. The number of sensory cells per mormyromast organ increases linearly with fish growth for both types of sensory cells. 2) Relation between peripheral sensory cell innervated and central zone of termination for mormyromast afferent fibers. The afferent fibers arising from the two types of sensory cell in the mormyromast organ project to separate zones of the electrosensory lateral line lobe, as shown by using retrograde labeling with horseradish peroxidase. 3) Central trajectories and terminal arbors of mormyromast afferent fibers. These aspects of mormyromast fibers are described by using intracellular staining of individual fibers as well as whole nerve staining of an electrosensory nerve. 4) Fine structure of mormyromast afferent terminals in the electrosensory lateral line lobe. Afferent fibers make various synaptic contacts, including contacts of a mixed type, gap junction-chemical, onto a restricted class of granule cells. The fine structure is described based on electron microscopy of horseradish-peroxidase-labeled fibers. The results provide an anatomical base for current physiological studies on mormyromast afferent fibers.     

Synaptic organization of the mushroom body calyx in Drosophila melanogaster

The calyx neuropil of the mushroom body in adult Drosophila melanogaster contains three major neuronal elements: extrinsic projection neurons, presumed cholinergic, immunoreactive to choline acetyltransferase (ChAT-ir) and vesicular acetylcholine transporter (VAChT-ir) antisera; presumed gamma-aminobutyric acid (GABA)ergic extrinsic neurons with GABA-like immunoreactivity; and local intrinsic Kenyon cells. The projection neurons connecting the calyx with the antennal lobe via the antennocerebral tract are the only source of cholinergic elements in the calyces. Their terminals establish an array of large boutons 2-7 microm in diameter throughout all calycal subdivisions. The GABA-ir extrinsic neurons, different in origin, form a network of fine fibers and boutons codistributed in all calycal regions with the cholinergic terminals and with tiny profiles, mainly Kenyon cell dendrites. We have investigated the synaptic circuits of these three neuron types using preembedding immuno-electron microscopy. All ChAT/VAChT-ir boutons form divergent synapses upon multitudinous surrounding Kenyon cell dendrites. GABA-ir elements also regularly contribute divergent synaptic input onto these dendrites, as well as occasional inputs to boutons of projection neurons. The same synaptic microcircuits involving these three neuron types are repeatedly established in glomeruli in all calycal regions. Each glomerulus comprises a large cholinergic bouton at its core, encircled by tiny vesicle-free Kenyon cell dendrites as well as by a number of GABAergic terminals. A single dendritic profile may thereby receive synaptic input from both cholinergic and GABAergic elements in close vicinity at presynaptic sites with T-bars typical of fly synapses. ChAT-ir boutons regularly have large extensions of the active zones. Thus, Kenyon cells may receive major excitatory input from cholinergic boutons and considerable postsynaptic inhibition from GABAergic terminals, as well as, more rarely, presynaptic inhibitory signaling. The calycal glomeruli of Drosophila are compared with the cerebellar glomeruli of vertebrates. The cholinergic boutons are the largest identified cholinergic synapses in the Drosophila brain and an eligible prospect for studying the genetic regulation of excitatory presynaptic function.     

Single unit recordings in the auditory nerve of congenitally deaf white cats: Morphological correlates in the cochlea and cochlear nucleus

It is well known that experimentally induced cochlear damage produces structural, physiological, and biochemical alterations in neurons of the cochlear nucleus. In contrast, much less is known with respect to the naturally occurring cochlear pathology presented by congenital deafness. The present study attempts to relate organ of Corti structure and auditory nerve activity to the morphology of primary synaptic endings in the cochlear nucleus of congenitally deaf white cats. Our observations reveal that the amount of sound-evoked spike activity in auditory nerve fibers influences terminal morphology and synaptic structure in the anteroventral cochlear nucleus. Some white cats had no hearing. They exhibited severely reduced spontaneous activity and no sound-evoked activity in auditory nerve fibers. They had no recognizable organ of Corti, presented >90% loss of spiral ganglion cells, and displayed marked structural abnormalities of endbulbs of Held and their synapses. Other white cats had partial hearing and possessed auditory nerve fibers with a wide range of spontaneous activity but elevated sound-evoked thresholds (60–70 dB SPL). They also exhibited obvious abnormalities in the tectorial membrane, supporting cells, and Reissner's membrane throughout the cochlear duct and had complete inner and outer hair cell loss in the base. The spatial distribution of spiral ganglion cell loss correlated with the pattern of hair cell loss. Primary neurons of hearing-impaired cats displayed structural abnormalities of their endbulbs and synapses in the cochlear nucleus which were intermediate in form compared to normal and totally deaf cats. Changes in endbulb structure appear to correspond to relative levels of deafness. These data suggest that endbulb structure is significantly influenced by sound-evoked auditory nerve activity. J. Comp. Neurol. 397:532–548, 1998. © 1998 Wiley-Liss, Inc.     

Ultrastructural studies of physiologically identified electrosensory afferent synapses in the gymnotiform fish, Eigenmannia

Eigenmannia is a weakly electric fish that emits a constant-frequency electric organ discharge (EOD). Probability coder (P unit) and phase coder (T unit) electroreceptive afferents differentially encode changes in EOD amplitude and phase, respectively. physiologically identified T and P units were intracellularly labelled with HRP and their terminals were examined with electron microscopy to determine their postsynaptic targets. This technique reveals that phase and amplitude are relayed to first-order electrosensory neurons by two parallel but not independent pathways. P-type afferents terminate on granular interneurons, basilar pyramidals, and polymorphic cells, electrosensory lateral line lobe targets that monitor amplitude modulations, but P-type afferents do not contact spherical cells. T-type afferents relay phase information to spherical cells and thus form a separate afferent pathway. T unit terminals do not synapse directly on basilar pyramidal cells. Collateral branches from T-type afferents, however, were also found to terminate on granule and polymorphic cells, thereby adding phase information into the amplitude channel. P- and T-type afferents exhibit cellular specificity by forming synaptic junctions with different subsets of post synaptic targets in the deep neuropil. The afferent terminals make either asymmetric chemical or gap junction synapses depending on the identity of the post synaptic target. T units contacting granule cells or polymorphic cells had not been previously described. Two possible roles of adding phase to amplitude information are discussed in terms of electrolocation.     

Chemical synapses, particle arrays, pseudo-gap junctions and gap junctions of neurons and glia in the buccal ganglion of Helisoma

The nervous system of the snail, Helisoma trivolvis, has been utilized for a wide range of studies of neuronal plasticity; however, the ultrastructural features of this tissue were previously unknown. The present study examined the nature of synaptic interactions of neurons and glia and considered several plasma membrane specializations of these cells. The symmetrical pair of buccal ganglia consisted of a ring of unipolar neurons surrounding a central neuropil. The neurons were separated by two morphologically distinct types of glia: type I were most numerous and possessed an electron-dense homogeneous cytoplasm, whereas type II glia were of lower electron density, possessed a heterogeneous cytoplasm, and appeared to be phagocytic. Gap junctions were abundant between glia and were occasionally found between neuronal processes, including those of neurons 19 injected with horseradish peroxidase (HRP). Comparison of neuron and glial gap junction widths (16.4 and 17.6 nm, respectively) in thin sections and their intramembrane particle diameters (13.1 and 13.7 nm, respectively) by freeze fracture, did not elucidate significant differences. A heterogeneous population of putative chemical synapses, similar to those reported in other molluscs, was also observed between axonal collaterals in the neuropil. Additionally, examination of freeze-fractured neuropil revealed rhombic arrays of particles localized on neuronal membranes; these arrays do not appear to form intercellular junctions but may represent postsynaptic receptor sites. Freeze fracture also revealed small, square arrays consisting of 7-9 nm diameter particles on glial membranes which may correspond to pentalaminar membrane contacts (pseudo-gap junctions) seen in thin sections between glia situated around dilated extracellular spaces (lacunae).     

Gap junction-mediated bidirectional signaling between human fetal hippocampal neurons and astrocytes

Gap junctions are clusters of intercellular channels that connect the interiors of coupled cells. In the brain, gap junctions function as electrotonic synapses between neurons and as pathways for the exchange of metabolites and second-messenger molecules between glial cells. Astrocytes, the most abundant glial cell type coupled by gap junctions, are intimately involved in the active control of neuronal activity including synaptic transmission and plasticity. Previous studies have suggested that astrocytic-neuronal signaling may involve gap junction-mediated intercellular connections; this issue remains unresolved. In this study, we demonstrate that second-trimester human fetal hippocampal neurons and astrocytes in culture are coupled by gap junctions bidirectionally; we show that human fetal neurons and astrocytes express both the same and different connexin subtypes. The formation of functional homotypic and heterotypic gap junction channels between neurons and astrocytes may add versatility to the signaling between these cell types during human hippocampal ontogeny; disruption of such signaling may contribute to CNS dysfunction during pregnancy.     

Gap junctions linking the dendritic network of GABAergic interneurons in the hippocampus.

The network of GABAergic interneurons connected by chemical synapses is a candidate for the generator of synchronized oscillations in the hippocampus. We present evidence that parvalbumin (PV)-containing GABAergic neurons in the rat hippocampal CA1 region, known to form a network by mutual synaptic contacts, also form another network connected by dendrodendritic gap junctions. Distal dendrites of PV neurons run parallel to the alveus (hippocampal white matter) and establish multiple contacts with one another at the border between the stratum oriens and the alveus. In electron microscopic serial section analysis, gap junctions could be identified clearly at 24% of these contact sites. A dendrodendritic chemical synapse and a mixed synapse also were found between PV-immunoreactive dendrites. Three-dimensional reconstruction of the dendritic arborization revealed that both PV neurons of the well known vertical type (presumptive basket cells and axoaxonic cells) and those of another horizontal type constitute the dendritic network at the light microscopic level. The extent of dendritic fields of single PV neurons in the lateral direction was 538 +/- 201 micrometer (n = 5) in the vertical type and 838 +/- 159 micrometer (n = 6) in the horizontal type. Our previous and present observations indicate that PV-containing GABAergic neurons in the hippocampus form the dual networks connected by chemical and electrical synapses located at axosomatic and dendrodendritic contact sites, respectively. Gap junctions linking the dendritic network may mediate coherent synaptic inputs to distant interneurons and thereby facilitate the synchronization of oscillatory activities generated in the interneuron network.