Aminergic modulation of graded synaptic transmission in the lobster stomatogastric ganglion

Graded chemical synaptic transmission is important for establishing the motor patterns produced by the pyloric central pattern generator (CPG) circuit of the lobster stomatogastric ganglion (Raper, 1979; Anderson and Barker, 1981; Graubard et al., 1983). We examined the modulatory effects of the amines dopamine (DA), serotonin (5-HT), and octopamine (Oct) on graded synaptic transmission at all the central chemical synapses made by the pyloric dilator (PD) neuron onto its follower cells, using synaptic input-output curves measured from cell somata. DA strongly reduced the graded synaptic strength at all the PD synapses. DA reduction of chemical synaptic strength from PD onto the inferior cardiac (IC) neuron could change the sign of synaptic interaction between these 2 cells from inhibitory to excitatory by uncovering a weak electrical connection. 5-HT had weaker and more variable effects, reducing graded synaptic strength from the PD onto the lateral pyloric and pyloric neurons and enhancing the weak synapse from the PD to the IC cell. Oct strongly enhanced the graded synaptic strength at all the PD central synapses. Oct enhancement of graded synaptic strength between the PD and IC cells could also change the sign of the interaction: weak, excitatory electrical coupling, which was sometimes dominant before Oct, was masked by the enhanced chemical inhibitory interaction during Oct application. Measurements of electrical coupling between 2 PD cells and between 2 postsynaptic cells suggest that Oct does not change the input resistance of these cells and may act directly at the PD synapses. The effects of DA and 5-HT are most easily explained by their general reductions in pre- and postsynaptic input resistance. DA, 5-HT, and Oct each produce a distinct pyloric motor pattern (Flamm and Harris-Warrick, 1986a). These amine-induced motor patterns may be explained by the unique actions of each amine on the intrinsic membrane properties of different pyloric CPG neurons (Flamm and Harris-Warrick, 1986b) and by modulation of graded synaptic transmission between the pyloric neurons.     

Ultrastructure of the electrotonic and chemical components of the lateral-to-motor and medial-to-motor synapses in crayfish nerve cord

Lateral-to-motor and medial-to-motor synapses in crayfish nerve cords are composed of an electrical and a chemical component. The presynaptic terminals showed localized clusters of synaptic vesicles, electron-dense areas, coated pits, and coated vesicles. In thin sections, active zones were defined by electron-dense regions where synaptic vesicles attached and, in freeze-fracture replicas, by clusters of intramembrane particles localized in bands with vesicle openings on the sides of these bands. The cytoplasmic surface of the postsynaptic membrane opposite the active zones was coated with electron-dense material that in freeze-fracture replicas was seen as an increase in intramembrane particles located in the external leaflet (EF-face). This specialization of the postsynaptic membrane may correspond to the neurochemical receptor. Also, pre- and postsynaptic membranes were separated by a wider extracellular gap than those of adjacent nonsynaptic regions and electrical synapses or gap junctions. Synaptic vesicles were located exclusively at the synaptic regions by means of a cytoskeleton that was different for the electrical and the chemical components. The vesicles associated with the electrical component were anchored to a cytoskeleton composed of a beaded layer of densities located parallel to the membrane. This cytoskeleton maintained the synaptic vesicles separated from the presynaptic membrane by a distance of 13 +/- 2 nm. The synaptic vesicles associated with the chemical component were anchored to electron-dense regions formed by filaments arranged in bundles, anchored to the presynaptic membrane. Vesicles lined both sides positioned to discharge their contents into the extracellular space and to replace the discharged vesicles.     

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.     

Special forms of synaptic junctions in cultures of dissociated mammalian spinal cord and spinal ganglia

The ultrastructural study of cells in the monolayer culture of dissociated spinal cord and spinal ganglia of mouse embryos has been performed. The results obtained show that in the course of differentiation of completely isolated cells of the spinal cord special forms of synaptic contacts may appear. They are typical of the spinal cord of phylogenetically inferior animal species. These "mixed" synaptic contacts having properties both of electrical and of chemical synapses seem to represent phylogenetically determined processes of synaptogenesis.     

Transient electrical coupling regulates formation of neuronal networks

Electrical synapses are abundant before and during developmental windows of intense chemical synapse formation, and might therefore contribute to the establishment of neuronal networks. Transient electrical coupling develops and is then eliminated between regenerating Helisoma motoneurons 110 and 19 during a period of 48-72 h in vivo and in vitro following nerve injury. An inverse relationship exists between electrical coupling and chemical synaptic transmission at these synapses, such that the decline in electrical coupling is coincident with the emergence of cholinergic synaptic transmission. In this study, we have generated two- and three-cell neuronal networks to test whether predicted synaptogenic capabilities were affected by previous synaptic interactions. Electrophysiological analyses demonstrated that synapses formed in three-cell neuronal networks were not those predicted based on synaptogenic outcomes in two-cell networks. Thus, new electrical and chemical synapse formation within a neuronal network is dependent on existing connectivity of that network. In addition, new contacts formed with established networks have little impact on these existing connections. These results suggest that network-dependent mechanisms, particularly those mediated by gap junctional coupling, regulate synapse formation within simple neural networks.     

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.     

Ultrastructural correlates of electrical-chemical synaptic transmission in goldfish cranial motor nuclei

The output connections of the cranial relay neurons, part of the Mauthner cell network, were examined in goldfish with light and electron microscopic techniques. Either lucifer yellow or horseradish peroxidase (HRP) was injected into cranial relay neuron axons to demonstrate that they diverge to several motor nuclei and to many motoneurons within one nucleus. Retrograde transport of the enzyme from injections of mandibular muscles was used to label the trigeminal motoneurons. In the electron microscope, cranial relay neuron processes were distinguished by the granular appearance of the electron-opaque polymer formed enzymatically by HRP, while the retrogradely labeled motoneurons had the polymer enclosed in lysosomes. The cranial relay neuron terminals contained many presynaptic vesicles which concentrated the HRP reaction product. Active zones and synaptic clefts were evident. At some synapses, both gap junctions and presynaptic vesicles were found. The mechanism of synaptic transmission was investigated by simultaneous recording with two intracellular microelectrodes from cranial relay neuron-motoneuron pairs. Composite postsynaptic potentials in a trigeminal motoneuron were evoked by intracellular stimulation of a cranial relay neuron axon. The earliest excitatory postsynaptic potential (EPSP) component had a latency of 0.25 msec and had a peak amplitude that was not depressed by repetitive stimulation. A second component had larger peak amplitudes which were reduced easily by repetitive stimulation. Antidromic action potentials were not transmitted from motoneurons to the cranial relay neuron axons. Thus, both electrical and chemical transmission probably occur at the cranial relay neuron-motoneuron synapses. Since the cranial relay neurons fire synchronously and receive excitatory chemical synapses, the function of the gap junctions and electrical transmission is unclear. Perhaps the importance of these gap junctions is more for transport of small molecules than for impulse transmission.     

Electrical synapses, a personal perspective (or history)

Gap junctions are the morphological substrate of one class of electrical synapse. This memoir records the author's involvement in the development of our knowledge of the physiology and ultrastructure of electrical synapses. The answer to whether neurotransmission is electrical or chemical is either. One lesson is that Occam's razor sometimes cut too deep; the nervous system does its operations in a number of different ways and a unitarian approach can lead one astray [M.V.L. Bennett, Nicked by Occam's razor: unitarianism in the investigation of synaptic transmission, Biol. Bull. 168 (1985) 159–167]. Electrical synapses can do many things that chemical synapses can do, and do them just as slowly. The new molecular, cellular and physiological techniques will clarify where gap junctions and electrical coupling do and do not occur and permit experimental manipulation with high specificity.     

Morphologic changes in the axo-dendritic synapses of the leech following excitatory conduction block in a medium with decreased ionic concentration

Reactive changes in the synaptic apparatus of the leech neuropile were studied in low ionic strength media using supravital methylene blue staining. Special preparation developed in the laboratory permitted obtaining clearly identified axodendritic synapses in the leech. It was shown that the reactive changes were pronounced during a decrease in the electrical conductance and block in the abdominal chain. Certain correlations between the structural alterations in axodendritic synapses and phasic reactive changes in axosomatic synapses were observed.     

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.     

Ultrastructure of the circuit providing,input to the crayfish lateral giant neurons

Labeled or otherwise identified neurons of the crayfish lateral giant escape reaction circuit were examined electron microscopically and the findings compared to expectations from physiology. Terminals of primary afferents contained clear, approximately 45 nm, irregularly round synaptic vesicles, while sensory interneuron terminals had slightly larger, 50 nm, more strictly round vesicles, permitting tentative classification based on anatomical criteria. Excitatory synapses on the lateral giants, believed from physiology to be electrical, generally had some gap junctions, but these were almost invariably paralleled by more prominent chemical junctional regions of unknown function. There may also be a class of interneurons making purely chemical synapses on the lateral giants. Synapses from primary afferents to sensory interneurons, believed from physiology to be cholinergic, had purely chemical morphology. Synapses with narrow elongated vesicles, similar to GABAergic vesicles seen in other neurons, frequently occurred on terminals of primary afferents. These synapses provide a basis for known presynaptic inhibition of afferent input. Consistent with physiology, such inhibitors sometimes also contacted the postsynaptic targets of the primary afferents and sometimes received input from other primary afferents. Afferent terminals also received some input from profiles richin large dense cored vesicles. Presumptive inhibitory input found on proximal dendrites of lateral giants provides a basis for known recurrent inhibition. However, similar inhibitory synapses that sometimes received local input from excitors of the lateral giants were also found distally mixed with excitatory inputs. These provide a basis for recently discovered distal inhibitory input following excitation and for tonic inhibition. © 1993 Wiley-Liss, Inc.     

Non-synaptic release from dense-cored vesicles occurs at all terminal types in crayfish neuropile

In the crayfish neuropile, dense-cored vesicles (DCV) have been found in chemical terminals, mixed in with round or pleomorphic agranular synaptic vesicles, as well as in electrical terminals and neurohemal endings. DCV release their content at unspecialized non-synaptic sites. The simultaneous exocytosis of DCV and synaptic vesicles seems to be the rule in chemical terminals. DCV in specific terminals suggest non-synaptic communication. In chemical and electrical terminals, the content of DCV could have a neuromodulatory function.     

Origin and function of spiral fibers projecting to the goldfish mauthner cell

Two neuron types contact the Mauthner cell (M cell) in the axon cap, a specialized region of high electrical resistance surrounding the initial segment of the M cell axon. One type produces a mixed electrical and chemical inhibition of the M cell. The second sends axons into the central core of the axon cap, where they spiral around the initial segment making both conventional synapses and gap junction contacts. The origin and synaptic effects of these spiral fibers have not been studied previously. When goldfish M cells were filled with Lucifer yellow, presynaptic spiral fibers were seen in the axon cap. These fibers could be traced back through the medial longitudinal fasciculus to their somata, near the contralateral fifth nerve motor nucleus. The same somata were labeled by horseradish peroxidase injected extracellularly into the axon cap. Recordings were made in the axon cap and the M cell after stimulation of hindbrain areas near the spiral fiber somata and axons. Extracellularly, a negative potential was observed close to the termination of the spiral fibers and termed the spiral fiber potential (SFP). Intracellularly, a graded, short latency depolarization of the M cell corresponded to the SFP and could cause the M cell to spike. This depolarization did not shunt the membrane, indicating that it may be produced through gap junctions. Intracellular responses to hindbrain stimulation also had a chloride-dependent, second component that shunted the membrane during paired-pulse testing. This inhibitory second component was probably evoked by cells other than the spiral fiber cells themselves. © 1994 Wiley-Liss, Inc.     

Horizontal cell electrical coupling in the giant danio: synaptic modulation by dopamine and synaptic maintenance by calcium

Electrical synapses, and their structural manifestation, gap junctions, are critical elements of retinal circuitry. These synapses are subject to both rapid modulation and slower structural changes by physiological signals which mediate changes in the adaptational state of the retina. The electrical synapses of fish retinal horizontal cells are an excellent preparation for in vitro studies of electrical synapses. We have examined the rapid modulation of electrical coupling by dopamine and effects on the expression and maintenance of electrical synapses by cell calcium in pairs of horizontal cells isolated from retinas of the giant danio (Danio aquipinnatus). We report that rapid modulation by dopamine reduces junctional conductance by modifying gap junction channel gating, while maintaining cells in reduced calcium medium, and lowering; intracellular calcium concentration, results in the loss of electrical coupling. The effects of calcium on synaptic maintenance may be related to structural changes observed in horizontal cell electrical synapses during light adaptation.     

Dendritic spine dynamics regulate the long-term stability of synaptic plasticity

Long-term synaptic plasticity requires postsynaptic influx of Ca2? and is accompanied by changes in dendritic spine size. Unless Ca2? influx mechanisms and spine volume scale proportionally, changes in spine size will modify spine Ca2? concentrations during subsequent synaptic activation. We show that the relationship between Ca2? influx and spine volume is a fundamental determinant of synaptic stability. If Ca2? influx is undercompensated for increases in spine size, then strong synapses are stabilized and synaptic strength distributions have a single peak. In contrast, overcompensation of Ca2? influx leads to binary, persistent synaptic strengths with double-peaked distributions. Biophysical simulations predict that CA1 pyramidal neuron spines are undercompensating. This unifies experimental findings that weak synapses are more plastic than strong synapses, that synaptic strengths are unimodally distributed, and that potentiation saturates for a given stimulus strength. We conclude that structural plasticity provides a simple, local, and general mechanism that allows dendritic spines to foster both rapid memory formation and persistent memory storage.     

Distribution of synaptic vesicle proteins in the mammalian retina identifies obligatory and facultative components of ribbon synapses

The mammalian retina contains two synaptic layers. The outer plexiform layer (OPL) is primarily composed of ribbon synapses while the inner plexiform layer (IPL) comprises largely conventional synapses. In presynaptic terminals of ribbon synapses, electron-dense projections called ribbons are present at the synaptic plasma membranes. Ribbons bind synaptic vesicles and guide them to the synaptic membrane for fusion. In this manner, ribbons are thought to accelerate the delivery of vesicles for continuous exocytosis. In recent years, a large number of synaptic proteins has been described but it is not known if these protein colocalize in the same types of synapses. In previous studies, several proteins essential for synaptic function were not detected in ribbon synapses, suggesting that the mechanism of synaptic vesicle exocytosis may be very different in ribbon and conventional synapses. Using confocal laser scanning microscopy, we have now systematically investigated the protein composition of ribbon synapses. Our results show that, of the 19 synaptic proteins investigated, all except synapsin and rabphilin are obligatorily present in ribbon synapses. For example, rab3 which was reported to be absent from ribbon synapses, was found in bovine, rat and mouse ribbon synapses using multiple independent antibodies. In addition, we found staining in these synapses for PSD-95 and NMDA receptors, which suggested a similar design for the postsynaptic component in ribbon and conventional synapses. Our data show that ribbon synapses are more conventional in composition than reported, that most synaptic proteins are colocalized to the same type of synapse, and that synapsin and rabphilin are likely to be dispensible for basic synaptic functions.     

The Uniform and Nonuniform Nature of Slow and Rapid Scaling in Embryonic Motoneurons

Neurons regulate the strength of their synapses in response to a perturbation to stabilize neuronal signaling through a form of homeostatic plasticity known as synaptic scaling. The process of scaling has the potential to alter all of a cell''s miniature postsynaptic current (mPSC) amplitudes by a single multiplicative factor (uniform scaling), and in doing so could change action potential-dependent or evoked synaptic strength by that factor. However, recent studies suggest that individual synapses scale with different scaling factors (nonuniform). This could complicate the simple multiplicative transform from mPSC scaling to the evoked response. We have previously identified a slow AMPAergic and GABAergic synaptic scaling in chick embryo motoneurons following 2 d in vivo perturbations inhibiting neuronal activity or GABA_AR function, and now show a rapid form of scaling following NMDAR blockade in vitro. Slow GABAergic scaling appeared to be of a classical uniform pattern. Alternatively, other forms of rapid and slow scaling demonstrated a uniform and nonuniform component in their mPSC amplitude distributions. Slow and rapid AMPAergic scaling was mediated by insertion of GluA2-lacking AMPA receptors. The nonuniform pattern of scaling may contribute to the observed complexity of the changes in evoked responses. Scaling-induced changes in mPSC amplitudes were not associated with changes in probability of release (Pr). Together, our results demonstrate a new rapid form of scaling in embryonic motoneurons, that slow and rapid scaling is not purely uniform, and that upscaling does not translate to an increase in evoked responses in a simple manner.SIGNIFICANCE STATEMENT Different forms of homeostatic plasticity are thought to play a critical role in maintaining neural function. For example, altering the amplitudes of spontaneous currents through a form of homeostatic plasticity known as synaptic scaling could affect evoked transmission; however, this is rarely tested. Here we demonstrate two forms of scaling and show that in many cases synaptic strength scales differently for distinct synapses within an embryonic motoneuron. These results have functional consequences for evoked synaptic strength and suggest that, like Hebbian plasticity, scaling can change relative synaptic strengths within a cell. Furthermore, our results demonstrate how different forms of homeostatic plasticity influence neuronal communication as the nascent spinal network is first established in the embryonic period.     

Synaptic development of the mouse dorsal lateral geniculate nucleus

The dorsal lateral geniculate nucleus (dLGN) of the mouse has emerged as a model system in the study of thalamic circuit development. However, there is still a lack of information regarding how and when various types of retinal and nonretinal synapses develop. We examined the synaptic organization of the developing mouse dLGN in the common pigmented C57/BL6 strain, by recording the synaptic responses evoked by electrical stimulation of optic tract axons, and by investigating the ultrastructure of identified synapses. At early postnatal ages (<P12), optic tract evoked responses were primarily excitatory. The full complement of inhibitory responses did not emerge until after eye opening (>P14), when optic tract stimulation routinely evoked an excitatory postsynaptic potential/inhibitory postsynaptic potential (EPSP/IPSP) sequence, with the latter having both a GABA_A and GABA_B component. Electrophysiological and ultrastructural observations were consistent. At P7, many synapses were present, but synaptic profiles lacked the ultrastructural features characteristic of the adult dLGN, and little γ‐aminobutyric acid (GABA) could be detected by using immunocytochemical techniques. In contrast, by P14, GABA staining was robust, mature synaptic profiles of retinal and nonretinal origin were easily distinguished, and the size and proportion of synaptic contacts were similar to those of the adult. The emergence of nonretinal synapses coincides with pruning of retinogeniculate connections, and the transition of retinal activity from spontaneous to visually driven. These results indicate that the synaptic architecture of the mouse dLGN is similar to that of other higher mammals, and thus provides further support for its use as a model system for visual system development. J. Comp. Neurol. 518:622–635, 2010. © 2009 Wiley‐Liss, Inc.     

Synaptic organization in the pacemaker nucleus of a medium-frequency weakly electric fish,Eigenmannia sp

The medullary command nucleus (MCN) of the medium-frequency weakly electric fish,Eigenmannia sp., contains two types of neurones, namely large and small cells, which are embedded in a neuropile of large and small myelinated fibers. Using serial semi-thin and ultra-thin sectioning, combined with HRP labelling established that both cell types possess rich dendritic arborization and large myelinated axons. Only the axons of the large cells leave the nucleus and these contribute the unique output of the MCN. Axon branching has been observed only in the axons of small cells and their collaterals show an exclusively intranuclear course. Two types of synaptic terminals have been found on large as well as on small cells: (1) large club endings forming both gap (electronic) junctions and polarized chemical synapses, which often appear at the same junction constituting morphologically mixed synapses; and (2) small bouton-like terminals forming exclusively chemical synaptic contacts. No differences between the two neuron types could be detected with respect to the arrangement of the synaptic contacts: club endings and small bouton-like terminals synapse on dendritic processes as well as on perikarya, while the unmyelinated initial segments were always found to be free of synaptic contacts. Large and small cells were found to be simultaneously connected by the same club ending or small bouton-like terminal: in the case of club endings by means of gap junctions and chemical synapses, whereas in the case of boutons by chemical synapses only. Club endings sometimes form gap junctions with each other. The possible role of these unusual synaptic connections in local synchronization is suggested. Club endings originate from the large axons of small cells, while small bouton-like terminals originate from the fine myelinated fibers of extranuclear origin. InEigenmannia, small cells, being connected to large cells as well as to each other by axo-somatic and axodendritic synapses, can be considered as the pacemaker cells of the MCN whereas large cells are relay cells. Small bouton-like terminals may convey exogeneous impulses towards the MCN exerting modulatory effects at both pacemaker and relay cell levels. The greater variety of ultrastructural correlates established in the MCN ofEigenmannia, in comparison withSternarchus⁵ (see also ref. 16), suggests increased modulation possibilities in the former fish's EOD behaviour.