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P M Balaban

Publications and source records attributed to P M Balaban.

At least 37 records · Page 2Linked to original sources

Synaptic facilitation in Helix neurons depends upon postsynaptic calcium and nitric oxide.

Intracellular tetanization of premotor interneurons for withdrawal in the snail. Helix lucorum produces long-term facilitation of synaptic inputs of these neurons. Using this model of plasticity we investigated the role of calcium in postsynaptic induction of synaptic facilitation. It was shown that, from the one hand, postsynaptic injection of calcium chelator EGTA completely abolishes potentiation and, from the other hand, injection of calcium chloride into the interneuron produces facilitation-like changes in the EPSP amplitude in this neuron. Therefore, not only necessity but also sufficiency of increasing of postsynaptic calcium for induction of synaptic potentiation was demonstrated. We also showed that inhibitor of nitric oxide synthase N-omega-nitro-L-arginine prevents development of postsynaptically induced facilitation what suggests that nitric oxide may participate in investigated synaptic facilitation as a retrograde messenger. These results support the idea that both invertebrates and vertebrates have common mechanisms underlying synaptic plasticity.

Action Potentials↗

Postembryonic neurogenesis in the procerebrum of the terrestrial snail, Helix lucorum L.

Neuronogenesis during posthatching development of the procerebrum of the terrestrial snail Helix lucorum was analyzed using bromodeoxyuridine immunohistochemistry to label proliferating cells. Comparison of the distribution of labeled cells in a series of animals which differed in age at the time of incubation with bromodeoxyuridine, in survival time after incubation, and in age at sacrifice reveals a clear pattern and developmental sequence in neuron origin. First, the proliferating cells are located only at the apical portion of the procerebrum. Second, cells which are produced at any particular age remain, for the most part, confined to a single layer in the procerebrum. Third, as development proceeds, each layer of previously produced neurons is displaced toward the basal part of the procerebrum by the production of additional neurons. Our results suggest that the vast majority of the neurons (probably about 70-80%) of the snail procerebrum are produced during the first 1-2 months of posthatching development.

Animals↗

Putative neuropeptides and an EF-hand motif region are encoded by a novel gene expressed in the four giant interneurons of the terrestrial snail.

Nine giant interneurons located in the pleural and parietal ganglia of the terrestrial snail Helix lucorum L. were reported to be a key element in the network controlling withdrawal behaviour of the animal. Using a combination of complementary DNA subtraction cloning and differential screening approaches we have isolated a novel gene named HCS2 which is expressed predominantly in a subset of these interneurons. The predicted amino acid sequence of the HCS2 protein contains at the N-terminus a hydrophobic leader sequence and four putative neuropeptides, and at the C-terminus a perfect match to the consensus motif of the EF-hand family of the Ca2+-binding proteins. All four predicted neuropeptides bear a C-terminal signature sequence Tyr-Pro-Arg-X (where X is Ile, Leu, Val or Pro), and three of them are likely to be amidated. Physiological action of three synthetic peptides corresponding to the predicted mature HCS2 peptides mimics fairly well the described action of parietal interneurons on follower motoneurons controlling pneumostome closure. In situ hybridization experiments demonstrated that the HCS2 gene is selectively expressed in the four parietal giant interneurons, as well as in several small unidentified neurons. The onset of the HCS2 transcription during embryogenesis coincides temporally with the time-point when the first withdrawal responses of the embryo to tactile stimulation appear. We propose that the HCS2 gene encodes a hybrid precursor protein whose processed products act as neuromodulators or neurotransmitters mediating the withdrawal reactions of the snail, and in addition may participate in the calcium regulatory pathways or calcium homeostasis in command neurons.

Amino Acid Sequence↗

A novel neuropeptide precursor gene is expressed in the terrestrial snail central nervous system by a group of neurons that control mating behavior.

We report the isolation of a cDNA clone encoding a neuropeptide precursor named preproGFAD from the central nervous system (CNS) of the snail Helix lucorum. Analysis of the expression of this gene shows that it is neurospecific and expressed in several groups of CNS neurons. Most notable is the expression of preproGFAD gene in the right mesocerebrum, where the neurons controlling mating behavior are located. The expression in this particular region is observed in adult animals but not in juvenile ones. The preprohormone is 108 amino acids long and contains a hydrophobic leader peptide and eight Lys-Arg recognition sites for endoproteolysis. The post-translational processing of the prohormone may lead to the generation of seven tetrapeptides, Gly-Phe-Ala-Asp-COOH (GFAD). This peptide has the same sequence as two previously isolated peptides from a related snail, Achatina fulica. The first of them (achatin-I) contains D-Phe; the second (achatin-II) is its L-Phe-containing stereoisomer. Injection of synthetic D-GFAD in nanomolar concentrations into intact animals caused an increase of the heartbeat rate and opening of the genital atrium. In preparations containing CNS with intact innervation of reproductive organs, bath application of D-GFAD caused extensive movements of the penis but not of other reproductive organs. Intracellular activation of individual neurons expressing the preproGFAD gene also elicited penis movements. D-GFAD also suppressed activity of neurons modulating feeding behavior. Our data therefore indicate that the preproGFAD gene encodes the precursor of a neuropeptide that participates in the regulation of male mating behavior.

Amino Acid Sequence↗

Characterization of a cDNA clone encoding pedal peptide in the terrestrial snail.

We report the isolation of a Helix lucorum cDNA clone encoding a precursor of neuropeptides that are closely related to Aplysia and Tritonia pedal peptides (Pep). The predicted propeptide contains 20 copies of the two variants of Helix Pep interspersed with Lys-Arg endopeptidase cleavage sites. Northern blot hybridization revealed multiple Pep-hybridizing species in the Helix CNS RNA. The Pep gene was expressed by several identified serotonergic neurones in pedal and cerebral ganglia, groups of sensory neurones in procerebrum, peripheral neurones in olfactory bulb, mantle and foot, and group of neurones in pedal ganglia presumably involved in locomotion control. Pep mRNA was detected in several neurones at the early stages of nervous system development.

Amino Acid Sequence↗

The synapse between LE sensory neurons and gill motoneurons makes only a small contribution to the Aplysia gill-withdrawal reflex.

The monosynaptic connection between the mechano-sensory neurons in the LE cluster and gill motoneurons has been extensively studied and used as a model for the gill-withdrawal reflex and its behavioural plasticity. In an attempt to evaluate the contribution of this synapse to the behaviour, we used voltage-sensitive dye recording to determine the number of activated LE neurons and the number of spikes made by each neuron in response to a light touch. In five preparations, light touch activated a median of five sensory cells with a median of 1.6 spikes per cell. From a comparison of the sizes of the motoneuron synaptic potentials elicited by LE spikes and elicited by a light siphon touch, we estimate that the LE sensory neurons contribute approximately 5% of the motoneuron synaptic potential in response to this touch. This result casts doubt on the validity of using this synaptic connection as a model for gill-withdrawal behaviour. Siphon nerve recordings reveal the existence of short-latency, low-threshold neurons that may provide much of the sensory input in response to a light touch.

Animals↗

Identification of two novel genes specifically expressed in the D-group neurons of the terrestrial snail CNS.

A search for genes specifically expressed in the giant interneurons of parietal ganglia of the snail Helix lucorum yielded, among others, two genes named HDS1 and HDS2. According to data obtained by Northern hybridization and whole-mount in situ hybridization, both genes are neurospecific and expressed almost exclusively in the peptidergic D-group neurons (Sakharov, 1974) located in the right parietal ganglion. In situ hybridization of the HDS1 and HDS2 probes with CNS of several related species of the Helicoidea superfamily identified in all cases similarly located homologous groups of neurons. Sequencing of the near full-length cDNA copies of the HDS1 and HDS2 genes revealed open reading frames 107 and 102 amino acids long for HDS1 and HDS2, respectively. Both putative proteins contain a hydrophobic leader peptide and putative recognition sites for furin-like and PC-like endopeptidases. Predicted amino acid sequences of the HDS1 and HDS2 proteins were found to be moderately homologous to each other, as well as to the LYCP preprohormone expressed by the light yellow cells of the freshwater snail Lymnaea stagnalis. These results confirm an earlier hypothesis that the D-group of the Helix family and the light yellow cells of Lymnaea stagnalis represent homologous neuronal groups. Our data suggest that the HDS1 and HDS2 genes encode precursors of secreted molecules, most likely neuropeptides or neurohormones.

Amino Acid Sequence↗

Postsynaptic induction of long-term synaptic facilitation in snail central neurones.

Long-term facilitation in molluscs is believed to be induced due to purely presynaptic activations. We recorded excitatory postsynaptic potentials (EPSPs) simultaneously from two identified neurones of snail parietal ganglia. We report a non-decrementing facilitation induced by intracellular tetanization with concomitant presynaptic activation. The mean EPSP amplitude measured in 10 neurones 30-50 min after tetanization was 17% greater than in the non-tetanized control neurones. Only short-lasting (5-10 min) postsynaptic changes were found (post-tetanic hyperpolarization and resistance decrease). The facilitation was especially prominent (34%, 10 min post-tetanus) but decreased within 15 min in preparations with rapid wash-out of the external media. The data suggest that induction of long-term enhancement in molluscs depends on postsynaptic events and, like in mammals, may involve increased postsynaptic Ca2+ and subsequent release of retrograde messengers.

Animals↗

Novel gene HCS1 is specifically expressed in the giant interneurones of the terrestrial snail.

The terrestrial snail Helix lucorum is a promising model for molecular neurobiology since its central nervous system (CNS) is simple and contains many morphologically and functionally identified large neurones. Among these, the giant interneurones located in pleural and parietal ganglia are especially interesting because they trigger the withdrawal behaviour of the snail and participate in aversive conditioning. Here we describe the identification and characterization of a gene named HCS1 which is preferentially expressed in these interneurones. It encodes a putative protein 100 amino acids long containing an N-terminal hydrophobic leader peptide. No sequences with significant homology to HCS1 were found in the protein (Swiss-Prot) and nucleotide (EMBLbank) data libraries. We suppose that the product of this gene is a secreted protein, presumably a neuropeptide or a growth factor.

Amino Acid Sequence↗

Positive and negative brain zones in the snail.

Fine wire electrodes were surgically implanted in two regions of the brain of the snail Helix aspersa. To receive electrical stimulation of the brain, a tethered snail was required to displace the end of a rod. Self-stimulation delivered to the parietal ganglion resulted in non-repetition of the operant response, whereas self-stimulation delivered to the mesocerebrum resulted in an increase in response frequency. The reinforcing effect of local extracellular stimulation of two brain zones was investigated in a semi-intact preparation of a closely related species with an identical cellular map (H. lucorum). It was found that mesocerebral stimulation increased the frequency of the reinforced spontaneous movement, but decreased the frequency of the same movement if its absence was reinforced. These results allow us to attribute positive reinforcing effects to this brain area, which is involved in the control of sexual behaviour. Different results were obtained by contingent stimulation of the rostral part of the parietal ganglia, where giant cells controlling avoidance behaviour are located. Stimulation of this zone resulted in a decrease in the frequency of the ongoing spontaneous movements. These findings make possible intracellular investigations of the mechanisms of positive and negative reinforcement.

Animals↗

Influence of a stationary magnetic field on bioelectric properties of snail neurons.

Identified cells of Helix lucorum L. received 20 min exposures to 23, 120, or 200 mT stationary magnetic field (MFs). Resting potentials and input resistances were measured. Controls were instituted for temperature changes and for mechanical and other sources of artifact. Resting potentials did not change with MF exposure. Input resistances decreased significantly in normally silent cells during MF exposure, but increased significantly in spontaneously active cells. The magnitudes of changes were monotonically related to strength of the MF. Changes in excitatory postsynaptic potentials were observed during MF exposure. Elimination of perineuronal glia by proteolytic enzymes eliminated the MF effects.

Animals↗

[Localization of areas of action potential generation in neurons of the edible snail].

Site of origin of action potentials in silent and spontaneously active unipolar neurons of the snail Helix lucorum was established by stimulation of neurons isolated together with the initial part of the process from the neuropile by the sinusoidal extracellular current flowing along the soma-axon direction in the saline. The source of the spontaneous activity was in the neuronal process in all investigated neurons and one trigger zone usually dominated. Threshold of the action potential generation in response to the extracellular current in silent neurons was also connected with the axonal trigger zone activation. The low-threshold zone of action potential generation in the soma was found only in 20% of studied neurons.

Action Potentials↗

Effect of 5,7-dihydroxytryptamine on the food-aversive conditioning in the snail Helix lucorum L.

The effects of 5,7-dihydroxytryptamine (5,7-DHT), a drug which selectively ablates serotonergic terminals, were examined on acquisition of food-aversive conditioned reflex in the snail Helix lucorum. The percent of feeding reactions decreased from 80 to 15% in the conditioned group of animals after 5-8 pairings of food and electric shock. The behavioral performance of 5,7-DHT-injected animals after the same training session coincided with the data received from the unpaired control group: the percent of feeding reactions remained the same as before the training. Conditioning was carried out on the semi-intact 'lip-CNS' preparations as well. Intracellular recordings from the neurons responding to the withdrawal reaction confirmed the results of the behavioral experiments. Elaboration of associative changes was effective on preparations made from normal snails, whereas no changes were noted in 5,7-DHT-treated and pseudoconditioned animals. In 5,7-DHT-treated animals some components of the feeding behavior and withdrawal reaction changed as well. The appetitive phase duration of feeding lengthened significantly, moreover the sensitization of the withdrawal reaction evoked by rhythmic tactile stimulation disappeared in preparations made from drug-treated snails.

5,7-Dihydroxytryptamine↗

Neural mechanisms of age-dependent changes in avoidance behaviour of the snail Helix lucorum.

In order to study the emergence of different components of learning and memory, we investigated developmental changes in behavioural, neurophysiological and histochemical experiments in newborn and adult terrestrial snails (Helix lucorum L.). The absence of sensitization and inability to develop avoidance-conditioned reactions were revealed in behavioral experiments in juvenile snails under 1 month old. Investigation of neural mechanisms of this behavioral deficit showed the absence of sensitization in spike reactions to repeated nerve stimulation in command neurons for the avoidance behaviour in juvenile snails. The same dynamics of response amplitude, as in juvenile snails, was seen in adults only after selective impairment of serotonergic neurons by treatment with 5,7-dihydroxytryptamine. During 1 month after hatching, the serotonin content investigated with fluorescence techniques was very low in the nervous system. This result suggests that the absence of sensitization, as well as inability to be aversively conditioned is related to 5-hydroxytryptamine level, which changes during postnatal development in the snail.

Action Potentials↗

Functioning of identified snail neurones in electric fields.

In both silent and spontaneously active neurones of the snail Helix lucorum, depolarization and spikes were elicited by low-frequency (0.1 Hz) sinusoidal currents applied to the bath solution. Threshold voltage gradients had a range of 1-10 V m-1, which is less than gradients in the nervous tissue during synchronous activation of the neurones. It is shown that the same neurone can generate spikes in response to opposite directions of polarizing currents. Thresholds of spontaneously active neurones to extracellular currents were significantly lower than thresholds of silent cells. A simple quantitative method for evaluation of the transmembrane voltage drop evoked by an electric field is presented. The role of neuronal branches in the response was studied by electrophysiological and morphological methods.

Animals↗

[Changes in the duration of action potentials of functionally different Helix neurons in response to serotonin].

Bath application of serotonin was studied for its effect on the duration of action potentials in sensory neurons, interneurons and motoneurons participating in avoidance behaviour of snail. Serotonin application increased the action potential duration in sensory neurons, while in interneurons and motoneurons such changes were absent. An increase of the action potential duration in sensory neurons can be the mechanism of the EPSP amplitude increase recorded in interneurons during application of serotonin. Functional consequences of selective reactions in neurons with different functional role are discussed.

Action Potentials↗