Rhythmic Changes in Synapse Numbers in Drosophila melanogaster Motor Terminals

Previous studies have shown that the morphology of the neuromuscular junction of the flight motor neuron MN5 in Drosophila melanogaster undergoes daily rhythmical changes, with smaller synaptic boutons during the night, when the fly is resting, than during the day, when the fly is active. With electron microscopy and laser confocal microscopy, we searched for a rhythmic change in synapse numbers in this neuron, both under light:darkness (LD) cycles and constant darkness (DD). We expected the number of synapses to increase during the morning, when the fly has an intense phase of locomotion activity under LD and DD. Surprisingly, only our DD data were consistent with this hypothesis. In LD, we found more synapses at midnight than at midday. We propose that under LD conditions, there is a daily rhythm of formation of new synapses in the dark phase, when the fly is resting, and disassembly over the light phase, when the fly is active. Several parameters appeared to be light dependent, since they were affected differently under LD or DD. The great majority of boutons containing synapses had only one and very few had either two or more, with a 70∶25∶5 ratio (one, two and three or more synapses) in LD and 75∶20∶5 in DD. Given the maintenance of this proportion even when both bouton and synapse numbers changed with time, we suggest that there is a homeostatic mechanism regulating synapse distribution among MN5 boutons.     

Circadian rhythms in screening pigment and invaginating organelles in photoreceptor terminals of the housefly's first optic neuropile

Screening pigment granules occur in the synaptic terminals of photoreceptors in the fly's (Musca domestica, L.) compound eye. The granules resemble ommochrome granules in the overlying photoreceptor cell body. There are also two types of invagination into receptor terminals: capitate projections (from glial cells) and invaginations from neighboring receptor terminals. The number of profiles of these organelles in the first optic neuropile, the lamina, have been counted using single-section quantitative electron microscopic methods. Pigment granules are concentrated proximally in the terminal, toward the brain. The numbers change, increasing during the night (1 h after lights off) up to values more than twice the number 1 h after lights on, apparently by longitudinal migration of granules from the cell body into the terminal. Flies entrained to day/night conditions and then held under constant darkness continue to exhibit changes in the numbers of profiles. Even though overall there were 80–90% fewer granule profiles than under day/night conditions, the numbers attained a peak many times higher at the end of the subjective day. Thus, the changes are endogenous, showing circadian rhythmicity. Although their significance is unknown, these changes parallel previously described circadian rhythms in the receptor terminals and their lamina monopolar-cell targets. The invaginations from receptor terminals were more numerous under day/night conditions than under constant darkness, and cycled in constant darkness, peaking at the end of subjective night. Capitate projections, by contrast, failed to change significantly under the experimental conditions analyzed, a lack of responsiveness they share with photoreceptor tetrad synapses. © 1997 John Wiley & Sons, Inc. J Neurobiol 32: 517–529, 1997     

Stability and variability of synapses in the adult molluskan CNS

Synaptic transmission was examined between identified neurons in the central nervous system (CNS) of the freshwater mollusk, Lymnaea stagnalis. Four identified neurons were used: Right Pedal Dorsal one (RPeD1; a dopaminergic respiratory interneuron), Visceral Dorsal two and three (VD2/3), and Visceral Dorsal four (VD4; a cardiorespiratory interneuron). Neuron RPeD1 synapses onto both VD2/3 and VD4, while VD4 makes a reciprocal synapse onto RPeD1. When compared from animal to animal, the connections were variable in sign. Previously, we demonstrated that, in a given animal, the RPeD1 --> VD4 synapse could be either inhibitory, biphasic, or undetectable. The present study now expands this concept of variability by showing that the RPeD1 --> VD2/3 synapse was either excitatory or undetectable from animal to animal, while the synapse from VD4 to RPeD1 was observed as inhibitory, biphasic, depolarizing, excitatory, or undetectable. Next, we used 1-day organ culture to determine if the variability observed between animals is a product of ongoing change to the sign of these identified synapses and whether or not the extent of change could be influenced by the culture conditions. Changes to the sign of transmission occurred within minutes and, more commonly, after 24-h organ culture. All three synapses were investigated before and after 1-day organ culture, in either defined medium (DM) or brain-conditioned medium (CM). Regardless of culture conditions, the RPeD1 --> VD2/3 synapse showed no change of sign, i.e., it was relatively stable. However, the synapses between RPeD1 and VD4 did change sign, and when cultured in CM, the VD4 --> RPeD1 synapse changed significantly more than in DM. These data indicate that variability of some synapses reflects changes at these synapses. This is the first report that specific synapses in an adult CNS can change sign, and that the sign of transmission can be modulated by environmental conditions.     

The interaction between the synapses of a single motor fiber

It has been shown that stimulation of synapses of the giant motor fibers of the third roots of Cambarus clarkii can block transmission at other synapses located on the same fiber. Peripherally located synapses block most synapses which are more centrally located. The reverse is true in a small number of cases. Possible reasons for this difference are discussed. It was further found that the two medial giant fibers in fresh, carefully dissected, preparations show a functional connection in the brain. It is probable that, under natural conditions, both medial giant fibers are always active at the same time.     

A Machine Learning Method for the Prediction of Receptor Activation in the Simulation of Synapses

Chemical synaptic transmission involves the release of a neurotransmitter that diffuses in the extracellular space and interacts with specific receptors located on the postsynaptic membrane. Computer simulation approaches provide fundamental tools for exploring various aspects of the synaptic transmission under different conditions. In particular, Monte Carlo methods can track the stochastic movements of neurotransmitter molecules and their interactions with other discrete molecules, the receptors. However, these methods are computationally expensive, even when used with simplified models, preventing their use in large-scale and multi-scale simulations of complex neuronal systems that may involve large numbers of synaptic connections. We have developed a machine-learning based method that can accurately predict relevant aspects of the behavior of synapses, such as the percentage of open synaptic receptors as a function of time since the release of the neurotransmitter, with considerably lower computational cost compared with the conventional Monte Carlo alternative. The method is designed to learn patterns and general principles from a corpus of previously generated Monte Carlo simulations of synapses covering a wide range of structural and functional characteristics. These patterns are later used as a predictive model of the behavior of synapses under different conditions without the need for additional computationally expensive Monte Carlo simulations. This is performed in five stages: data sampling, fold creation, machine learning, validation and curve fitting. The resulting procedure is accurate, automatic, and it is general enough to predict synapse behavior under experimental conditions that are different to the ones it has been trained on. Since our method efficiently reproduces the results that can be obtained with Monte Carlo simulations at a considerably lower computational cost, it is suitable for the simulation of high numbers of synapses and it is therefore an excellent tool for multi-scale simulations.     

NMDA receptor subunits: diversity, development and disease

N-methyl-D-aspartate receptors (NMDARs) are present at many excitatory glutamate synapses in the central nervous system and display unique properties that depend on their subunit composition. Biophysical, pharmacological and molecular methods have been used to determine the key features conferred by the various NMDAR subunits, and have helped to establish which NMDAR subtypes are present at particular synapses. Recent studies are beginning to address the functional significance of NMDAR diversity under normal and pathological conditions.     

Mechanisms of synaptic plasticity

We have shown that the synapse maturation phase of synaptogenesis is a model for synaptic plasticity that can be particularly well-studied in chicken forebrain because for most forebrain synapses, the maturation changes occur slowly and are temporally well-separated from the synapse formation phase. We have used the synapse maturation phase of neuronal development in chicken forebrain to investigate the possible link between changes in the morphology and biochemical composition of the postsynaptic density (PSD) and the functional properties of glutamate receptors overlying the PSD. Morphometric studies of PSDs in forebrains and superior cervical ganglia of chickens and rats have shown that the morphological features of synapse maturation are characteristic of a synaptic type, but that the rate at which these changes occur can vary between types of synapses within one animal and between synapses of the same type in different species. We have investigated, during maturation in the chicken forebrain, the properties of the N-methyl-D-aspartate (NMDA) subtype of the glutamate receptors, which are concentrated in the junctional membranes overlying thick PSDs in the adult. There was no change in the number of NMDA receptors during maturation, but there was an increase in the rate of NMDA-stimulated uptake of 45Ca2+ into brain prisms. This functional change was not seen with the other ionotropic subtypes of the glutamate receptor and was NMDA receptor-mediated. The functional change also correlated with the increase in thickness of the PSD during maturation that has previously been shown to be due to an increase in the amount of PSD associated Ca(2+)-calmodulin stimulated protein kinase II (CaM-PK II). Our results provide strong circumstantial evidence for the regulation of NMDA receptors by the PSD and implicate changing local concentrations of CaM-PK II in this process. The results also indicate some of the ways in which properties of existing synapses can be modified by changes at the molecular level.     

Reconstitution in bilayer lipid membranes of the crab Potamon transcaspicum spider venom sensitive glutamate receptors

Membrane proteins have been isolated from neuromuscular synapses of the crab Potamon transcaspicum using a specific blocker of glutamatergic synapses, the neurotoxin of the spider Argiope lobata. These membrane components have been shown to induce glutamate-sensitive conductance in bilayer lipid membranes (BLM) in the presence of sodium ions. As an agent blocking desensitization of glutamatergic synapses, concanavalin A was shown to enhance the conductance and to abolish desensitization. Diethylester of glutamic acid as a blocker of glutamatergic synapses inhibited glutamate-induced conductance. No similar change in conductance was seen when BLM was modified by a membrane proteins -neurotoxin complex. Conductance current fluctuations induced by these receptor protein components were monitored by the single-channel registration method.     

Effect of hypoxia on the structure of the presynaptic grid of the interneuronal contacts of the rat neocortex

Hypoxic effect (a 6 minute asphyxia) on the presynaptic grid structure and on the amount of synapses in the neocortex has been studied in white rats using the method of selective staining of synapses with phosphoric tungsten acid. Neurofilamentous formations of the presynaptic grid appeared to be most labile structures. Dense projections of the presynaptic grid are most sensitive to hypoxia: their height, distinctness of the frame, and the intensity degree of phosphoric tungsten acid staining decrease. The content of intermediate form contacts with low indistict dense projections increases with the increase in the number of light type changed synapses. The principle organization of the presynaptic grid does not change in the posthypoxic period: hexagonal division of dense projections and places of vesicular attachment are kept. The hypertrophy of the presynaptic grid is also kept during that process.     

The Effect of Acute Unilateral Denervation Injury on Purinergic Signaling in the Cholinergic Synapse

Biophysics - Previously, we found a change in the efficiency of the modulatory action of ATP under the influence of some non-physiological factors at neuromuscular synapses in rodents. This study...     

Neural control of the corpus allatum in the Colorado potato beetle, Leptinotarsa decemlineata: An electron microscope study utilizing the in vitro tannic acid Ringer incubation method

The release of material from neurosecretory synapses in the corpora allata of the Colorado potato beetle, Leptinotarsa decemlineata was visualised by an electron microscope procedure which involved tissue incubation in tannic acid. Using morphometry, the frequency of exocytosis phenomena was quantified in beetles kept under two different photoregimes. The number of exocytosis phenomena in the neurosecretory synapses in the corpora allata of beetles kept under short days was significantly higher than that of beetles reared under long-day conditions. In addition, the corpus allatum gland cells appeared to be more richly innervated by neurosecretory synapses under short-day than under long-day conditions. Previous studies using the in vitro radiochemical assay showed that the corpus allatum activity of short-day beetles is at least partly restrained by neurally mediated factors. The present morphological data strongly imply that this corpus allatum inhibitory substance is released from the neurosecretory synapses.     

Neuronal networks and memory: role of the hippocampus

Learning and memory are related both to cognitive processes and to neurobiological mechanisms. The human pathology focused on the role of the hippocampus and animal experiments have analyzed its implications. The most usually admitted hypothesis is that memories are underlied by distributed specific neural networks defined through the strengthening of certain synapses, under the action of the flow of information during learning. The best candidate for this strengthening of the synapses is a change in synaptic plasticity similar to the artificial phenomenon of long-term potentiation. During memory processes, the hippocampus would play a particular role in information processing (analyzing novelty and significance of the information) and would allow the specification of the neural network, mainly in the cortical territories. We report data in olfactory learning in rats comforting these hypotheses. Considering neurochemistry of memory processes, specific synaptic changes and neuromodulatory processes must be distinguished. We report data about vasopressin illustrating both kinds of mechanisms in the hippocampus.     

Interaction between immunoglobulin G and peroxidase-like iron oxide nanoparticles: Physicochemical and structural features of the protein

The study is devoted to the oxidative modification of immunoglobulin G (IgG) on the surface of peroxidase-like iron oxide magnetic nanoparticles (MNPs) under conditions of induced reactive oxygen species (ROS) generation and without them. A pronounced change of thermodynamic parameters of denaturation has been detected for IgG in solutions containing MNPs under hydrogen peroxide action during 24 h of incubation. Dynamic light scattering measurements and UV–Visible spectrophotometry have been used to show aggregation in these solutions. Ferromagnetic resonance (FMR) was used to compare IgG coating thickness on individual MNPs under conditions of induced ROS generation and without them. The similarity between IgG adsorption on MNPs under these conditions after 24 h of incubation has been confirmed by the fluorescence measurements. The sites of IgG oxidative modifications that take place on MNPs surface and some evidences of the influence of oxidative modification and adsorption on the chemical structure of IgG were revealed by HPLC MS/MS analysis.     

Modulation of glutamate mobility reveals the mechanism underlying slow-rising AMPAR EPSCs and the diffusion coefficient in the synaptic cleft

Fast- and slow-rising AMPA receptor-mediated EPSCs occur at central synapses. Fast-rising EPSCs are thought to be mediated by rapid local release of glutamate. However, two controversial mechanisms have been proposed to underlie slow-rising EPSCs: prolonged local release of transmitter via a fusion pore, and spillover of transmitter released rapidly from distant sites. We have investigated the mechanism underlying slow-rising EPSCs and the diffusion coefficient of glutamate in the synaptic cleft (Dglut) at cerebellar mossy fiber-granule cell synapses using a combination of diffusion modeling and patch-clamp recording. Simulations show that modulating Dglut has different effects on the peak amplitudes and time courses of EPSCs mediated by these two mechanisms. Slowing diffusion with the macromolecule dextran slowed slow-rising EPSCs and had little effect on their amplitude, indicating that glutamate spillover underlies these currents. Our results also suggest that under control conditions Dglut is approximately 3-fold lower than in free solution.     

Deviation of the living system from the stationary state during oogenesis

Summary The changes in respiration and glycolysis of whole oocytes and homogenates of oocytes during oogenesis have been studied.The respiration rate of whole oocytes increases during oocyte growth and decreases during oocyte maturation. The respiration rate of homogenates also increases during oocyte growth and does not change during egg maturation. At all oogenesis stages the respiration rate of homogenates is higher than the respiration rate of whole oocytes.Respiration intensity increases during the small growth stage and decreases during the following stages of oogenesis. Respiration intensity of homogenates under optimal conditions changes in a similar way. Respiration intensity under physiological conditions diminishes during oogenesis from 70% at the small growth stage to 42% in unfertilised eggs.The rate of glycolysis in whole oocytes and homogenates of oocytes increases during the growth period of oocytes but does not change during egg maturation.Glycolysis intensity of the whole oocytes increases at the large growth stage—stage of cytoplasmic vacuolisation—and becomes less during the following stages. Glycolysis intensity in homogenates under optimal conditions is much higher than the glycolysis intensity of whole oocytes and it decreases slightly during oogenesis. The efficiency of glycolysis in oocytes under physiological conditions is very low. It increases from the stage of cytoplasmic vacuolisation (3.6%) to the stage at which vitellogenesis starts (20%) and diminishes at the following stages.The data obtained are considered in the light of the Prigogine and Wiame interpretation of a thermodynamic theory of development.     

Action of strychine on evoked potentials and postsynaptic responses of cat sensomotor cortical neurons

Acute experiments on immobilized cats lightly anesthetized with pentobarbital showed that application of strychine to the cortical surface inhibits slow negative potentials arising during direct and primary responses of the sensomotor cortex and corresponding IPSPs in pyramidal neurons. Iontophoretic applications of strychine blocks predominantly the early component of the IPSP, during which the input resistance under normal conditions is significantly less than during the late component of the IPSP, indicating that these components differ in their genesis. It is concluded that individual components of cortical evoked potentials have a common genesis, and that the slow negative potential is the dipole reflection of the IPSP in pyramidal neurons; the early component of the IPSP, moreover, is generated as a result of activation of axo-somatic inhibitory synapses, whereas the late component is generated as a result of activation of axo-dendritic synapses. The mediators in these inhibitory synapses may be different.I. S. Beritashvili Institute of Physiology, Academy of Sciences of the Georgian SSR, Tbilisi. Translated from Neirofiziologiya, Vol. 16, No. 4, pp. 480–487, July–August, 1984.     

Organization of the stomatogastric ganglion of the spiny lobster

Summary Three direct synaptic connections occur between neurons in the gastric and pyloric systems of the stomatogastric ganglion ofPanulirus interruptus. Two synapses are inhibitory, and one is electrical. This electrical synapse is both excitatory and inhibitory at different times. These synapses, and others within each system, let the two systems interact under some conditions. The synapses also form multisynaptic pathways which modulate the firing of many neurons in both systems. The consequences of these multisynaptic pathways are described and discussed.I thank Allen I. Selverston, Karen Sigvardt, Eve Marder, David Russell and Mary Chamberlin for criticizing a draft of this paper, Forrest Gompf and Doug Tissdale for technical support, and Nina Pollack and Betty Jorgensen for laboratory assistance. The research was supported by USPHS grant NS-12295 to BM and USPHS grant NS-09322 to Alien I. Selverston. BM was a USPHS NIH Postdoctoral Fellow in A.I. Selverston's laboratory during part of this research and is now a Research Fellow of the Alfred P. Sloan Foundation.     

Cloning of the first sn1-DAG lipases points to the spatial and temporal regulation of endocannabinoid signaling in the brain

Diacylglycerol (DAG) lipase activity is required for axonal growth during development and for retrograde synaptic signaling at mature synapses. This enzyme synthesizes the endocannabinoid 2-arachidonoyl-glycerol (2-AG), and the CB1 cannabinoid receptor is also required for the above responses. We now report on the cloning and enzymatic characterization of the first specific sn-1 DAG lipases. Two closely related genes have been identified and their expression in cells correlated with 2-AG biosynthesis and release. The expression of both enzymes changes from axonal tracts in the embryo to dendritic fields in the adult, and this correlates with the developmental change in requirement for 2-AG synthesis from the pre- to the postsynaptic compartment. This switch provides a possible explanation for a fundamental change in endocannabinoid function during brain development. Identification of these enzymes may offer new therapeutic opportunities for a wide range of disorders.     

Effect of malathion on ultrastructure of synapses in forebrain of the goldfish and rainbow trout

An electron microscope study of the effect of sublethal concentrations of malathion on the length of active zones has been performed in synapses of forebrain in the goldfish and rainbow trout. There was established a statistically significant reversible decrease of the main values of the length of sections of active zones in asymmetric and symmetric synapses under action of the toxicant. Effect of malathion increases proportionally to an increase of its concentration. In the both fish species the response to malathion was more pronounced in the asymmetric synapses compared to the symmetric ones. The obtained results allow considering that malathion causes changes in configuration and dimensions of the active zones of the cholinergic and probably GABAergic synapses. An adaptive character of these changes is discussed.     

The synaptic changes in the structures of the hyperstriatal section of the chicken brain during imprinting, visual deprivation and intermittent photic stimulation]

Morphometric analysis of synapses was conducted in medial areas of the ventral and accessory hyperstriatum in the right and left hemispheres of chickens brain after: 1) rhythmic optical stimulation beginning from the 18-th day of incubation, 2) presentation of flashing stimuli on the 10-12-th hours of life, 3) keeping under constant illumination in the first day or 4) in darkness during 24 hours after hatching. On electronograms of ultrathin slices parameters were recorded of synapses numerical density, mean length and width of postsynaptic condensations, number of synaptic vesicles per active zone in the presynapse and mean length of the whole axodendrite contact. The results of the studies have shown that early visual experience leads to significant changes of synaptic complex in the studied brain regions. Plasticity of synapses is discussed during stimulation and morphofunctional interrelations between hyperstriatal areas which are connected with different aspects of processing and storage of the visual information.