Selective Effects of an NMDA Glutamate Receptor Antagonist on the Sensory Input from Chemoreceptors in the Snail's Head during Acquisition of Nociceptive Sensitization

Experiments on semi-intact preparations from common snails were used to study the characteristics of the actions of MK-801, an antagonist of NMDA glutamate receptors, on the plasticity of various sensory inputs to defensive behavior command neurons LPl1 and RPl1 during acquisition of nociceptive sensitization. Application of sensitizing stimuli to the head or foot of control snails led to depression of neuron responses to tactile and chemical sensory stimulation during the short-term stage and marked facilitation of these responses during the long-term stage of sensitization. Application of sensitizing stimuli to the snail's head during administration of MK-801 led to marked depression of responses to chemical stimulation of the head in both the short-term and long-term stages of sensitization. In addition, blockade of NMDA receptors during application of sensitizing stimuli to the foot or head had no effect on changes in neuron responses elicited by chemical stimulation of the snail's foot and by tactile stimulation of the foot or head. It is suggested that NMDA-like glutamate receptors are selectively involved in the mechanism of induction of plasticity of synaptic inputs to command neurons LPl1 and RPl1, excited by chemical sensory stimulation of the head – a skin receptor zone specific for these neurons.     

The Critical Role of Intracellular Calcium in the Mechanisms of Plasticity of Common Snail Defensive Behavior Command Neurons LPl1 and RPl1 in Nociceptive Sensitization

Studies on semi-intact common snail preparations addressed the involvement of intracellular calcium in changes in the excitability and responses to sensory stimuli of defensive behavior command neurons LPl1 and RPl1 during the acquisition of nociceptive sensitization. Application of sensitizing stimuli to the heads of control snails led to depolarization of neuron membranes, increases in neuron excitability, and depression of the responses of neurons to sensory stimuli during the short-term stage, and marked facilitation of responses in the long-term stage of sensitization. Acquisition of sensitization during profound hyperpolarization of neurons led to suppression of the increase in excitability, along with depression of responses to chemical stimulation of the head in the short- and long-term stages of sensitization. Neuron responses to tactile stimulation of the head and foot showed synaptic facilitation, similar to that seen in neurons of control animals. Acquisition of sensitization during intracellular injection of the calcium chelators EGTA and BAPTA led to suppression of synaptic facilitation in the responses of neurons to both chemical and tactile stimulation. In these conditions, membrane excitability increased to a greater extent than in neurons of control animals. The results of these experiments suggest that changes in responses to sensory stimulation in sensitized snails are associated with postsynaptic calcium-dependent mechanisms of plasticity in neurons LPl1 and RPl1.     

Effects of antisense oligonucleotides to mRNA for the early gene zif268 on the mechanisms of synapse-specific plasticity

Acquisition of nociceptive sensitization in common snails was accompanied by long-term facilitation of the responses of defensive behavior command neuron LPl1 to sensory stimulation of chemoreceptors on the head and mechanoreceptors on the head and foot. Acquisition of sensitization during intracellular administration of antisense oligonucleotides to mRNA encoding the early gene zif268 showed suppression of synaptic facilitation in the responses of neuron LPl1 to tactile and chemical stimulation of the snail’s head. Synaptic facilitation in the responses to tactile stimulation of the foot developed as in neurons of control sensitized animals. These results suggest that the early gene zif268 is selectively involved in the mechanisms of the specific regulation of the synaptic inputs of neuron LPl1 from sensory receptors on the snail’s head. __________ Translated from Zhurnal Vysshei Nervnoi Deyatel’nosti imeni I. P. Pavlova, Vol. 56, No. 4, pp. 499–505, July–August, 2006.     

Neuronal mechanisms of site-specific nociceptive sensitization in the common snail

Comparative studies of the electrophysiological effects of site-specific nociceptive sensitization were performed on LPl1 and RPl1 command neurons in the common snail, after placing concentrated quinine solution (10%), on the animal's head or mantle ridge. Greater synaptic facilitation was observed in command neuron responses to tactile test stimuli applied to the same part of the body as the sensitizing stimulus, as compared with the level of synaptic facilitation in responses to tactile stimulation of other parts of the body. Synaptic facilitation of responses of neurons LPl1 and RPl1 to tactile stimulation of the head appeared 1 h after sensitizing stimulation and consisted of two phases: the first phase was characterized by a peak-like increase in the area of slow EPSP (sEPSP) and lasted 1 h; the second phase was characterized by maintenance of a relatively stable level of facilitation of sEPSP, which lasted to the end of the observation period (2–3 h). Synaptic facilitation of neuron responses to tactile stimulation of the mantle ridge appeared 40–60 min, after facilitation of responses to test stimulation of the snail's head, and was characterized by maintenance of a relatively stable level of sEPSP facilitation. It is suggested that the specificity of synaptic facilitation occurring in snail defensive behavior command neurons during the period of long-term nociceptive sensitization is associated with the genetic regulation of the various synaptic “inputs” to neurons. Translated from Zhurnal Vysshei Nervnoi Deyatel'nosti, Vol. 47, No. 6, pp. 994–1003, November–December, 1997.     

The Effects of Antibodies Against Proteins of the s100 Group on Neuron Plasticity in Sensitized and Non-Sensitized Snails

The effects of antibodies to a total fraction of s100 proteins and protein s100b on the activity of defensive behavior command neurons LPl1 and RPl1 were studied in common snails, using non-sensitized animals and animals which had acquired nociceptive sensitization. In non-sensitized snails, application of antibodies against s100 or s100b (0.1 mg/ml) induced membrane depolarization, increased membrane permeability, and suppressed slow excitatory postsynaptic potentials in the responses of neurons to sensory stimulation. Acquisition of sensitization in snails in the presence of antibodies to s100 or s100b (0.1 mg/ml) led to significantly less marked facilitation of synaptic transmission and smaller increases in neuron membrane excitability than in cells of control sensitized animals. The difference in synaptic facilitation in the neurons of control sensitized snails and neurons in sensitized snails given antibody was comparable with the magnitude of synaptic depression due to antibody in non-sensitized animals. At a dose of 0.01 mg/ml, antibody had no effect on these measures of neuron activity. It is suggested that s100 proteins, particularly s100b, are involved in the mechanisms regulating excitability, the membrane potential, and synaptic transmission in command neurons in untrained snails, as well as in the mechanism of plasticity of the electrogenic membranes of nerve cells during the acquisition of nociceptive sensitization.     

Selective involvement of opioids in the mechanisms of synapse-specific plasticity in the common snail during the acquisition of sensitization

Studies on defensive behavior command neurons LPl1 and RPl1 in semi-intact common snail preparations were performed to investigate the effects of the opioid peptide metenkephalin and the opioid antagonist naloxone on the effects of nociceptive sensitization. Application of nociceptive stimuli to the snails head elicited marked reversible membrane depolarization along with depression of neuron responses to sensory stimulation during the short-term stage of sensitization and facilitation of responses in the long-term stage. Metenkephalin at a dose of 10 µM but not at a dose of 0.1 µM partially suppressed responses to nociceptive stimuli. Acquisition of sensitization during exposure to metenkephalin at doses of 10 and 0.1 µM led to complete suppression of the facilitation of responses to tactile stimulation of the head. Facilitation of responses to chemical stimulation of the head and tactile stimulation of the foot in these conditions was similar to that of neurons in control sensitized animals. Acquisition of sensitization during exposure to metenkephalin and/or naloxone elicited selective suppression of facilitation of responses to chemical stimulation of the head but had no effect on facilitation of responses to tactile stimulation of the head and foot. Met-enkephalin and naloxone had no effect on the depression of neuron responses evoked by sensory stimulation in the short-term stage of sensitization. It is suggested that during the acquisition of sensitization in the common snail, opioids are involved in controlling the mechanism of nociception and in the mechanisms of selective induction of long-term plasticity of the synaptic inputs to command neurons activated by tactile and chemical stimulation of the animals head.Translated from Zhurnal Vysshei Nervnoi Deyatelnosti, Vol. 53, No. 6, pp. 766–774, November–December, 2003.     

Synaptic dynamics on different time scales in a parallel fiber feedback pathway of the weakly electric fish

Synaptic dynamics comprise a variety of interacting processes acting on a wide range of time scales. This enables a synapse to perform a large array of computations, from temporal and spatial filtering to associative learning. In this study, we describe how changing synaptic gain via long-term plasticity can act to shape the temporal filtering of a synapse through modulation of short-term plasticity. In the weakly electric fish, parallel fibers from cerebellar granule cells provide massive feedback inputs to the pyramidal neurons of the electrosensory lateral line lobe. We demonstrate a long-term synaptic enhancement (LTE) of these synapses that is biochemically similar to the presynaptic long-term potentiation expressed by parallel fibers in the mammalian cerebellum. Using a novel stimulation protocol and a simple modeling paradigm, we then quantify the changes in short-term plasticity during the induction of LTE and show that these changes can be explained by gradual changes in only one model parameter, that which is associated with the baseline probability of transmitter release. These changes lead to a shift in the spike frequency preference of the synapse, suggesting that long-term plasticity is not only involved in controlling the gain of the parallel fiber synapse, but also provides a means of controlling synaptic filtering over multiple time scales.     

NMDA Glutamate Receptor Antagonists Selectively Affect the Synaptic Mechanisms of Nociceptive Sensitization in Snails

Defensive behavior command neurons LPl1 and RPl1 were studied in semi-intact snail preparations to investigate the effects of N-methyl-D-aspartate (NMDA) receptor antagonists on the mechanisms of nociceptive sensitization. Application of sensitizing stimuli to the heads of control snails led to membrane depolarization and increased excitability, and also depressed the responses of neurons to tactile and chemical sensory stimuli in the short-term stage and facilitated responses in the long-term stage of sensitization. Development of sensitization in conditions of exposure to the NMDA receptor antagonists AP5 or MK-801 produced changes in the membrane potential and membrane excitability of command neurons similar to those seen in neurons of control sensitized snails. In addition, changes in the responses of command neurons to tactile stimulation of the head and foot and chemical stimulation of the foot in these conditions were also similar to those seen in neurons of control animals. Acquisition of sensitization during administration of NMDA receptor antagonists led to pronounced depression of responses to chemical test sensory stimulation of the snails' heads in both the short-term and long-term stages of sensitization. Thus, in sensitized snails, NMDA glutamate receptor antagonists selectively acted on the mechanisms of induction of plasticity the synaptic inputs of command neurons mediating excitation from chemical sensory stimuli from the animal's head.     

Studies of the Rf = 0.58 protein in common snail command neurons during learning

Levels of the acidic brain-specific Rf = 0.58 protein in neurons of the subglottal complex of ganglia were studied in the common snail during the process of acquisition of defensive food aversion. Levels were significantly increased in neurons LPa3 and RPa3 at both the early and late stages of learning. There was a tendency to increased protein levels in neurons LPl1 and RPl2, while there were no changes in levels in neuron RPa5 and pool D. The level of involvement of defensive behavior command neurons appears to be determined by the specific involvement of their receptor and effector fields.     

A cholinergic model synapse to elucidate protein function at presynaptic terminals

A large number of proteins have been identified at nerve terminals and a cascade of protein-protein interactions has been suggested to be involved in cycling of synaptic vesicle states. To explore protein function in presynaptic terminals, only a few unique synapses such as the squid giant synapse, the calyx of Held synapse and the hippocampal neuron autapse have been used. The squid giant synapse and the calyx of Held are useful to introduce reagents into their large presynaptic terminals and the hippocampal neuron autapse is a good system to modify a protein level by exogenous DNA or RNA. The cholinergic synapse formed between superior cervical ganglion (SCG) neurons in long-term culture is a useful model for a fast synapse. The axon of the large cell body contacts with soma of neighboring neurons. The architecture of synaptic connections makes it possible to introduce reagents into the presynaptic terminals by diffusion from a cell body within a short time. Introduction of exogenous cDNA or siRNA performed by microinjection into a SCG neuron allows us to modulate the level of the protein of interest or to express mutant proteins in the neuron. Here, we describe use of the model SCG neuronal synapse to elucidate function of presynaptic proteins in mediating synaptic transmission.     

Selective Effects of Antibodies to Protein SMP-69 on the Activity of Defensive Behavior Command Neurons in the Common Snail

The effects of antibodies to the serotonin-modulated protein SMP-69 on the activity of defensive behavior command neurons LP11 and RP11 in semi-intact preparations from common snails were studied. Antibody to SMP-69 increased membrane excitability and facilitated neuron responses to chemical sensory stimulation by application of dilute quinine solution to the animal's head, these effects being seen at 1-1.5 h. The synaptic effects of the antibodies were specific, as they had no influence on responses induced by tactile stimulation of the head. The neuronal effects of antibody SMP-69 were similar to changes in the activity of cells LP11 and RP11 induced by serotonin and cAMP, and to changes seen when snails acquired nociceptive sensitization. It seems likely that a protein homologous to mammalian SMP-69 is involved in the mechanisms controlling excitability and long-term specific plasticity of the synaptic inputs to neurons LP11 and RP11 from chemoreceptors on the snail's head.     

The Selective Effect of a Protein Kinase C Inhibitor on Synaptic Plasticity in Defensive Behavior Command Neurons During Development of Sensitization in the Snail

Studies of defensive behavior command neurons LP11 and RP11 in semi-intact common snail preparations addressed the effects of the protein kinase C antagonist polymyxin B on the effect of nociceptive sensitization. Neurons in control snails responded to application of nociceptive stimuli to the head with membrane depolarization, increases in excitability, and depression of neuron responses to sensory stimulation during the short-term stage, with marked facilitation of responses during the long-term stage of sensitization. Acquisition of sensitization in the presence of polymyxin B resulted in partial suppression of responses to nociceptive stimuli. Changes in command neuron membrane excitability in these conditions, as well as changes in responses to tactile stimulation of the foot and chemical stimulation of the head, were similar to those seen in neurons of sensitized control animals. The inhibitor also had no effect on short-term depression of neuron responses induced by tactile stimulation of the head. In addition, acquisition of sensitization during administration of polymyxin B led to complete suppression of the facilitation of responses to tactile stimulation of the snail's head during the long-term stage of sensitization. It is suggested that in sensitized common snails, protein kinase C is involved in controlling the mechanisms of nociception and is also involved in the mechanisms of selective induction of plasticity in the synaptic inputs of command neurons, which are activated by tactile stimulation of the animal's head.     

Estradiol affects axo-somatic contacts of neuroendocrine cells in the arcuate nucleus of adult rats

It has been shown that gonadal steroids have the capacity to induce synaptic plasticity in certain areas of the nervous system. Previously we have demonstrated that due to the effect of estradiol there is a transient decrease in the number of GABAergic axo-somatic synapses in the arcuate nucleus. By using systemic application of the tracer Fluorogold we retrogradely labeled a subpopulation of arcuate neurons that project to the median eminence. We than applied the disector method for synapse quantification and found that these "hypophysiotropic neurons" receive less axo-somatic inputs. We found that 17beta-estradiol induced a decrease in the numerical density of axo-somatic contacts of these retrogradely-labeled neoroendocrine cells.Our data support the hypothesis that the hormonally driven morphological synaptic plasticity is neuron specific within the arcuate nucleus and plays a decisive role in the regulation of anterior pituitary.     

Presynaptic efficacy directs normalization of synaptic strength in layer 2/3 rat neocortex after paired activity

Paired neuronal activity is known to induce changes in synaptic strength that result in the synapse in question having different properties to unmodified synapses. Here we show that in layer 2/3 excitatory connections in young adult rat cortex paired activity acts to normalize the strength and quantal parameters of connections. Paired action potential firing produces long-term potentiation in only a third of connections, whereas a third remain with their amplitude unchanged and a third exhibit long-term depression. Furthermore, the direction of plasticity can be predicted by the initial strength of the connection: weak connections potentiate and strong connections depress. A quantal analysis reveals that changes in synaptic efficacy were predominantly presynaptic in locus and that the key determinant of the direction and magnitude of synaptic modification was the initial release probability (P(r)) of the synapse, which correlated inversely with change in P(r) after pairing. Furthermore, distal synapses also exhibited larger potentiations including postsynaptic increases in efficacy, whereas more proximal inputs did not. This may represent a means by which distal synapses preferentially increase their efficacy to achieve equal weighting at the soma. Paired activity thus acts to normalize synaptic strength, by both pre- and postsynaptic mechanisms.     

In vitro formation and activity-dependent plasticity of synapses between Helix neurons involved in the neural control of feeding and withdrawal behaviors

Short-term activity-dependent synaptic plasticity has a fundamental role in short-term memory and information processing in the nervous system. Although the neuronal circuitry controlling different behaviors of land snails of the genus Helix has been characterized in some detail, little is known about the activity-dependent plasticity of synapses between identified neurons regulating specific behavioral acts. In order to study homosynaptic activity-dependent plasticity of behaviorally relevant Helix synapses independently of heterosynaptic influences, we sought to reconstruct them in cell culture. To this aim, we first investigated in culture the factors regulating synapse formation between Helix neurons, and then we studied the short-term plasticity of in vitro-reconstructed monosynaptic connections involved in the neural control of salivary secretion and whole-body withdrawal. We found that independently of extrinsic factors, cell-cell interactions are seemingly sufficient to trigger the formation of electrical and chemical synapses, although mostly inappropriate--in their type or association--with respect to the in vivo synaptic connectivity. The presence of ganglia-derived factors in the culture medium was required for the in vitro reestablishment of the appropriate in vivo-like connectivity, by reducing the occurrence of electrical connections and promoting the formation of chemical excitatory synapses, while apparently not influencing the formation of inhibitory connections. These heat-labile factors modulated electrical and chemical synaptogenesis through distinct protein tyrosine kinase signal transduction pathways. Taking advantage of in vitro-reconstructed synapses, we have found that feeding interneuron-efferent neuron synapses and mechanosensory neuron-withdrawal interneuron synapses display multiple forms of short-term enhancement-like facilitation, augmentation and posttetanic potentiation as well as homosynaptic depression. These forms of plasticity are thought to be relevant in the regulation of Helix feeding and withdrawal behaviors by inducing dramatic activity-dependent changes in the strength of input and output synapses of high-order interneurons with a crucial role in the control of Helix behavioral hierarchy.