Biomedical subjects

P M Balaban

Publications and source records attributed to P M Balaban.

At least 19 recordsLinked to original sources

PubMed2006

Intracellular localization of the HCS2 gene products in identified snail neurons in vivo and in vitro.

1. The HCS2 (Helix command specific 2) gene expressed in giant command neurons for withdrawal behavior of the terrestrial snail Helix lucorum encodes a unique hybrid precursor protein that contains a Ca-binding (EF-hand motif) protein and four small peptides (CNP1-CNP4) with similar Tyr-Pro-Arg-X aminoacid sequence at the C terminus. Previous studies suggest that under conditions of increased intracellular Ca(2+) concentration the HCS2 peptide precursor may be cleaved, and small physiologically active peptides transported to the release sites. In the present paper, intracellular localization of putative peptide products of the HCS2-encoded precursor was studied immunocytochemically by means of light and electron microscopy. 2. Polyclonal antibodies against the CNP3 neuropeptide and a Ca-binding domain of the precursor protein were used for gold labeling of ultrathin sections of identified isolated neurons maintained in culture for several days, and in same identified neurons freshly isolated from the central nervous system. 3. In freshly isolated neurons, the gold particles were mainly localized over the cytoplasmic secretory granules, with the density of labeling for the CNP3 neuropeptide being two-fold higher than for the calcium-binding domain. In cultured neurons, both antibodies mostly labeled clusters of secretory granules in growth cones and neurites of the neuron. The density of labeling for cultured neurons was the same for both antibodies, and was two-fold higher than for the freshly isolated from the central nervous system neurons. 4. The immunogold particles were practically absent in the bodies of cultured neurons. 5. The data obtained conform to the suggestion that the HCS2 gene products are transported from the cell body to the regions of growth or release sites.

Animals↗

PubMed2006

Regulation of tentacle length in snails by odor concentration.

The upper tentacle of the snail, bearing the olfactory organ, produces complex movements when the snail explores a new environment. Tentacle trajectories were reconstructed in the presence and absence of odors using two simultaneous video recordings. Reconstructions showed that in the absence of odor, snails constantly scanned the surrounding space with the extended tentacles. Presentation of an odor elicited rapid flexion, independent of the odor concentration, accompanied by concentration-dependent tentacle contractions. Activation of identified motoneuron MtC3 is known to elicit tentacle contraction. Recordings made in semi-intact preparations showed that the dynamics and duration of the spike activity of MtC3 produced in response to odors correlated with the degree of tentacle contraction in response to odors. These data suggest that the central motoneuron MtC3, which triggers tentacle contraction, is involved in controlling the margins of the scanning field. Slow contraction or extension of the tentacle, associated with the level of MtC3 activity, may operate to tune the snail's investigative behavior to the conditions of the sensory environment.

Action Potentials↗

PubMed2006

Protein synthesis-dependent reactivation of a contextual conditioned reflex in the common snail.

We report here a study of the effects of blockade of protein synthesis with anisomycin during reactivation of a contextual conditioned reflex in the common snail. The amplitudes of the defensive reactions of snails to standard tactile stimulation before training were identical in two conditions: 1) testing of responses of snails fixed by the shell to a plastic ball floating in water and 2) on the surface of the terrarium glass. After applying electric shocks to the snails' skin for 5 days, a significant difference in responses reflecting the formation of a contextual conditioned reflex was seen in only one of the contexts. Placing trained snails in the same context (reminding) two days after training with simultaneous injection of anisomycin led to significant weakening of training, while control injections of physiological saline produced no such changes. These data suggest that the mechanisms of memory consolidation after training and reminding are not identical.

Animals↗

PubMed2006

Functions of peptide CNP4, encoded by the HCS2 gene, in the nervous system of Helix lucorum.

The aims of the present work were to study the role of neuropeptide CNP4, encoded by the HCS2 gene (which is expressed mainly in parietal command interneurons), in controlling the activity of the respiratory system, and also to study the effects of this neuropeptide on isolated defensive behavior neurons in prolonged culture. The influence of the command interneuron on the pneumostoma included a direct effect consisting of closure and a delayed effect consisting of intensification of respiratory movements. Application of neuropeptide CNP4 produced a pattern similar to the delayed effects seen on stimulation of the command interneuron, i.e., significant increases in the frequency and intensity of pneumostoma movements and strengthening of the rhythmic activity of the pneumostoma motoneuron. Studies of the effects of neuropeptide CNP4 on isolated neurons after prolonged culture showed that neuron process growth correlated with the presence of the neuropeptide in the medium. Identification of the location of the HCS2 precursor protein and neuropeptide CNP4 in isolated command interneurons after prolonged culture showed that that only those parts of the cell showing active process growth were immunopositive. Thus, neuropeptide CNP4 appears to be a secreted neuropeptide controlling respiratory system activity, which may also be involved in rearrangements of the network controlling defensive behavior in Helix snails.

Animals↗

PubMed2006

Caspase-like activity is essential for long-term synaptic plasticity in the terrestrial snail Helix.

Although caspase activity in the nervous system of mollusks has not been described before, we suggested that these cysteine proteases might be involved in the phenomena of neuroplasticity in mollusks. We directly measured caspase-3 (DEVDase) activity in the Helix lucorum central nervous system (CNS) using a fluorometrical approach and showed that the caspase-3-like immunoreactivity is present in the central neurons of Helix. Western blots revealed the presence of caspase-3-immunoreactive proteins with a molecular mass of 29 kDa. Staurosporin application, routinely used to induce apoptosis in mammalian neurons through the activating cleavage of caspase-3, did not result in the appearance of a smaller subunit corresponding to the active caspase in the snail. However, it did increase the enzyme activity in the snail CNS. This suggests differences in the regulation of caspase-3 activity in mammals and snails. In the snail CNS, the caspase homolog seems to possess an active center without activating cleavage typical for mammals. In electrophysiological experiments with identified snail neurons, selective blockade of the caspase-3 with the irreversible and cell-permeable inhibitor of caspase-3 N-benzyloxycarbonyl-Asp(OMe)-Glu(OMe)-Val-Asp-(OMe)-fluoro-methylketone prevented development of the long-term stage of synaptic input sensitization, suggesting that caspase is necessary for normal synaptic plasticity in snails. The results of our study give the first direct evidence that the caspase-3-like activity is essential for long-term plasticity in the invertebrate neurons. This activity is presumably involved in removing inhibitory constraints on the storage of long-term memory.

Animals↗

PubMed2006

[Commonality of molecular mechanisms of neuroplasticity and neuropathology: integrative approach].

Analysis of our own data and of the results from other groups brings about a concept concerning community of the basic molecular mechanisms involved in neuroplasticity and neuropathology at different levels. Along with the fundamental significance of this idea for understanding of processes taking place both in normal brain and in neuropathological conditions, the concept is of principal importance for practical application. Failure in development of the "pathogenetically directed" approaches to treatment of neural diseases (e. g. stroke) are, in particular, related to the neglecting of identity of the basic molecular mechanism underlying both normal brain functioning and neuropathological conditions resulting in intervention into these common mechanisms (NMDA receptors blockade, inhibitor of the so called "apoptotic" proteases).

Animals↗

PubMed2005

Ephaptic feedback in identified synapses in mollusk neurons.

The possible existence of intrasynaptic ephaptic feedback in the invertebrate CNS was studied. Intracellular recordings were made of excitatory postsynaptic potentials and currents arising on activation of the recently described monosynaptic connection between identified neurons in the snail CNS. In the presence of ephaptic feedback, tetanization of the postsynaptic neuron with hyperpolarizing impulses should activate presynaptic calcium channels, thus increasing the amplitude of excitatory postsynaptic potential, while sufficiently strong postsynaptic hyperpolarization applied during generation of the excitatory postsynaptic current should induce "supralinear" increases in its amplitude, as has been observed previously in rat hippocampal neurons. The first series of experiments involved delivery of 10 trains of hyperpolarizing postsynaptic impulses (40-50 mV, duration 0.5 sec, frequency 1 Hz, train duration 45 sec); significant changes in the amplitude of excitatory postsynaptic were not seen. In the second series of experiments, changes in the amplitude of the excitatory postsynaptic current were studied during hyperpolarization of the postsynaptic neuron. At a potential of -100 mV, the amplitude of the excitatory postsynaptic current increased significantly more than predicted by its "classical" linear relationship with membrane potential. This "supralinear" increase in the amplitude of the excitatory postsynaptic potential can be explained by the operation of ephaptic feedback and is the first evidence for this phenomenon in CNS synapses of invertebrates.

Animals↗

PubMed2004

Postsynaptic calcium contributes to reinforcement in a three-neuron network exhibiting associative plasticity.

We show that activation of a single serotonergic cell is sufficient to trigger long-term associative enhancement of synaptic input to the withdrawal interneuron in a simple network consisting of three interconnected identified cells in the nervous system of terrestrial snail Helix. 1,2-bis (2-aminophenoxy) Ethane-N,N,N',N'-tetraacetic acid (BAPTA) injection in the postsynaptic neuron abolishes the pairing-specific enhancement of synaptic input. Activation of a single modulatory cell that we used to reinforce the synaptic input induced an increase of the intracellular [Ca2+] in the ipsilateral withdrawal interneuron without any changes of its membrane potential or input resistance. Similar changes in intracellular [Ca2+] were observed in the same withdrawal interneuron under bath application of 10(-5) m serotonin. Responses to repeated glutamate applications to the soma of synaptically isolated withdrawal interneurons increased after 10 min of serotonin or thapsigargin bath application, but were absent in conditions of preliminary BAPTA intracellular injection, significantly decreased under heparin injection. Thus, activity of a single modulatory cell may mediate reinforcement via an increase of [Ca2+] in the postsynaptic cell in a simple network consisting of neurons with defined behavioural roles.

Animals↗

PubMed2003

Helix peptide immunoreactivity pattern in the nervous system of juvenile aplysia.

Distribution of neurons immunopositive to antibody against the small peptides encoded by the Helix Command-Specific 2 (HCS2) gene in the central nervous system of juvenile Aplysia californica was investigated. The HCS2 gene is specifically expressed in the withdrawal behavior neurons of the terrestrial snail Helix lucorum. In Aplysia, 20-25 immunopositive neuronal somata were observed on dorsal surface of each pleural ganglion (including a giant pleural neuron). The HCS2-encoded peptide immunopositive fibers were observed in neuropiles of all ganglia and in many nerves. Functional significance of Aplysia immunopositive cells is discussed.

Animals↗

PubMed2003

Synaptic contact between mechanosensory neuron and withdrawal interneuron in terrestrial snail is mediated by L-glutamate-like transmitter.

The properties of the monosynaptic input from mechanosensory neurons to withdrawal interneurons were examined in Helix lucorum. The instantaneous I-V relation of the excitatory postsynaptic current in withdrawal interneurons was nonlinear, having a plateau region between -40 and -60 mV. On application of the blocker of vertebrate N-methyl-D-aspartate (NMDA) receptors AP5, or reduction of the Mg(2+) concentration, the current-voltage relation became more linear, suggesting that Mg(2+) may partially block the ion channel underlying the EPSC at voltages ranging from -40 to around -60 mV and the involvement of NMDA-like receptors. DNQX and 6-cyano-7-nitroquinoxaline-2,3-dione, which are known to block the glutamate non-NMDA receptors in mammals, significantly depress in a dose-dependent manner the actions of the natural transmitter. Exogenous L-glutamate applications mimicked the action of the mechanosensory neuron transmitter.

Animals↗

PubMed2003

Phase-dependent coordination of two motor programs in the buccal ganglion of a pteropod mollusk.

Rhythmic activity in two independent structures of the digestive apparatus of Clione limacina--the radula and the hooks--is coordinated by neural networks in the buccal ganglion during feeding behavior. Optical recording of neuron activity in the buccal ganglion, which allows simultaneous recording of large numbers of neurons, showed that the activity of all neurons producing volley discharges can be assigned to only two phases of a single rhythm. Instead of the four theoretically possible phases of rhythmic neural activity, all experiments yielded recordings of biphasic activity, even in conditions of electrical stimululation of the cerebrobuccal connectives, which triggers rhythmic movements of this apparatus in preparations. These data demonstrate the phase-dependent coordination of two independent rhythmic food-procuring movements in Clione.

Animals↗

PubMed2002

Cellular mechanisms of behavioral plasticity in terrestrial snail.

Functional organization of networks underlying withdrawal, feeding, and respiration in terrestrial gastropod snail Helix are described. Tracking the changes during non-associative and associative modifications of behavior, analysis of plasticity mechanisms in identified neurons involved in these networks allowed to formulate several conceptual principles which are not widely accepted. The review will present data underlying the following principles: 1. Command neuron concept can be applied only to all-or-none behavior. 2. Habituation is an active down-regulation process opposite to up-regulating sensitization. All long-term behavioral changes at least in part are associative. 3. Reinforcement is a motivational state mediated by neuromodulatory neurons and can be produced by activity of a single modulatory neuron. 4. Non-addressed ('soft-wired') neuromodulatory influences are necessary for acquisition of memory, while retention of memory depends mostly on 'hard-wired' local changes in synaptic connectivity. 5. Retrieval of declarative (sensory) and procedural (motor) memory involves different functional classes of neurons.

Animals↗

PubMed2001

Comparison of reactive processes in the rat brain elicited by xenotransplantation of nervous tissues of chicken or pulmonate snail.

It is known that a histocompatibility system is not developed to the same extent in lower invertebrates as in vertebrate animals. We assumed that the xenografts from the newborn invertebrate nervous system would not exert destructive effects on the brain of the vertebrate recipient even without immunosuppressive therapy. In search of brain xenografts (XG) capable to survive in the brain of a recipient without intensive immunosuppression, we transplanted ganglia of terrestrial snails into the rat brain. We compared effects of transplantation of the XG taken from anterior brain of the 18-day embryo chicken (XGC) and from ganglia of a newborn terrestrial pulmonate snail (Helix aspersa L., XGSn). Part of the XGSn were stained by vital fluorescent dyes Bisbenzimid or Fast Blue before grafting. The XGSn were implanted into the neocortex parenchyma in each hemisphere. Rat brains with the XGC were examined 5 days after, and brains with the XGSn - 5 and 28 days after the transplantation. Nonstained sections with the XGSn labeled with fluorescent dyes prior to transplantation were investigated in fluorescent microscope and stained later with tionin and cresyl-violet. Quantitative videoimage analysis of lymphocyte aggregations, reactive gliosis, morphology of the XG areas, and implantation trace was performed. It was found that the XGSn transplantation did not elicit in the rat brain an intensive immunological conflict 5 and 28 days after transplantation. In contrast, the XGC rapidly elicited a strong immune response resulting in massive obliterations in the rat brain and were rejected in 5 days. Labeled snail glia and vessels were observed in the stained XGSn 28 days after transplantation by fluorescence imaging. Putative snail vessels grew into the rat brain from the place of snail tissue transplantation serving the humoral integration of the XG and the host brain. Migration of molluscan glial cells was observed in the brain of recipients.

Animals↗

PubMed2001

A single serotonergic modulatory cell can mediate reinforcement in the withdrawal network of the terrestrial snail.

A cluster of 40 serotonergic cells in the rostral part of pedal ganglia of the terrestrial snail Helix lucorum was shown previously to participate in the modulation of withdrawal behavior and to be necessary during the acquisition of aversive withdrawal conditioning in intact snails. Local extracellular stimulation of the serotonergic cells paired with a test stimulus elicited a pairing-specific increase (the difference between paired and explicitly unpaired sessions was significant, p <.01) of synaptic responses to test stimulation in the premotor interneurons involved in withdrawal. This result suggested participation of serotonergic cells in mediating the reinforcement in the withdrawal network. Intracellular stimulation of only one identified Pd4 cell from the pedal group of serotonergic neurons paired with a test stimulus also significantly increased (the difference between paired and explicitly unpaired sessions was significant, p <.05) synaptic responses to paired nerve stimulation in same premotor interneurons involved in withdrawal. Morphological investigation of a cluster of pedal serotonergic neurons showed that only the Pd4 cell had branches in the parietal ganglia neuropile where the synapses of premotor withdrawal interneurons and of presynaptic neurons are located. The data suggest that a single serotonergic cell can mediate the reinforcement in the withdrawal network of the terrestrial snail. Patterns of responses of the Pd4 cells to tactile and chemical stimuli conform to the suggestion.

Animals↗

PubMed2001

Participation of GABA in establishing behavioral hierarchies in the terrestrial snail.

GABA-immunoreactive fibers were observed in the neuropile of each ganglion of Helix lucorum, while GABA-immunoreactive neural somata were found only in the buccal, cerebral, and pedal ganglia. Bath application of 10(-5) M GABA to the preparation "buccal mass-buccal ganglia" elicited a sequence of radula movements characteristic of feeding behavior. Corresponding bursts of activity were recorded in the buccal nerves under GABA application and in the buccal neurons recorded optically. In preparations of isolated central nervous system, the bath applications of GABA (10(-5) to 10(-4) M) elicited no changes in synaptic input of the premotor interneurons involved in the withdrawal behavior. However, a significant decrease in amplitude of the synaptic input and in the number of spikes in responses elicited by the test nerve stimulation was observed in metacerebral serotonergic neurons involved in modulating the feeding behavior. GABA application inhibited the spontaneous spike activity in some pedal serotonergic neurons involved in the network underlying withdrawal responses and evoked bursting activity in the other neurons of this functional group. The effects of GABA application on mechanically isolated serotonergic neurons suggest that the primary effect of GABA is inhibition. Thus, our results give evidence of the putative role of GABA in activating the feeding behavior and in the synergistic suppression of serotonergic modulation of the withdrawal behavior and serotonergic modulation of feeding, which has corresponded to the observed behavioral suppression of withdrawal reactions during feeding.

Animals↗

PubMed2001

Up- and down-regulation of Helix command-specific 2 (HCS2) gene expression in the nervous system of terrestrial snail Helix lucorum.

A novel gene named Helix command-specific 2 (HCS2) was shown to be expressed predominantly in four giant parietal interneurons involved in withdrawal behavior of the terrestrial snail Helix lucorum L. and several single neurons in other ganglia. Decrease in spontaneous electrophysiological activity of neurons in the isolated CNS by 24h incubation in saline with elevated Mg(2+) concentration significantly decreased the number of HCS2-expressing neurons. Five short-term serotonin applications (each of 10microM), during a 24h incubation of the nervous system in saline induced expression of the HCS2 gene in many cells in cerebral, parietal, pleural and pedal ganglia. Dopamine applications under similar conditions were not effective. Application of anisomycin or cycloheximide, known to block protein synthesis, did not prevent the induction of HCS2 expression under serotonin influence. Skin injury elicited a significant increase in the number of HCS2-expressing cells 24h later in pleural and cerebral ganglia. Incubation of the isolated nervous system preparations for three days in culture medium elicited close to a maximum increase in number of HCS2-expressing cells. Elevation of the normal Mg(2+) concentration in the culture medium significantly decreased the number of cells demonstrating HCS2 expression. Application of the cAMP activator forskolin (10microM) increased the expression under Mg(2+), indicating that cAMP was involved in the up-regulation of HCS2. Application of thapsigargin (10microM), known to release Ca(2+) from intracellular stores, was also effective in increasing expression, suggesting participation of Ca(2+) in regulation of HCS2 expression. Cellular groups expressing the HCS2 gene under different conditions seem to be functionally related since it was demonstrated earlier that some neurons constituting these clusters are involved in the withdrawal behavior and the response of the organism to stress stimuli. From these results we suggest that the HCS2 pattern of expression can be down-regulated by a decrease in synaptic activity in the nervous system, and up-regulated by external noxious inputs, as well as the application of neurotransmitters and second messengers known to be involved in the withdrawal behavior and maintenance of isolated ganglia in culture medium. When up-regulated, the HCS2 expression appears, at least in part in neurons, to be involved in the withdrawal behavior.

Animals↗

PubMed2001

Ontogenesis of the snail, Helix aspersa: embryogenesis timetable and ontogenesis of GABA-like immunoreactive neurons in the central nervous system.

Late stages of embryogenesis in the terrestrial snail Helix aspersa L. were studied and a developmental timetable was produced. The distribution of gamma-aminobutyric acid-like immunoreactive (GABA-ir) elements in the CNS of the snail was studied from embryos to adulthood in wholemounts. In adults, approximately 226 GABA-ir neurons were located in the buccal, cerebral and pedal ganglia. The population of GABA-ir cells included four pairs of buccal neurons, three neuronal clusters in the pedal ganglia, two clusters and six single neurons in the cerebral ganglia. GABA-ir fibers were observed in all ganglia and in some nerves. The first detected pair of GABA-ir cells in the embryos appeared in the buccal ganglia at about 63-64% of embryonic development. Five pairs of GABA-ir cell bodies were observed in the cerebral ganglia at about 64-65% of development. During the following 30% of development three more pairs of GABA-ir neurons were detected in the buccal ganglia and over fifteen cells were detected in each cerebral ganglion. At the stage of 70% of development, the first pair of GABA-ir neurons was found in the pedal ganglia. In the suboesophageal ganglion complex, GABA-ir fibers were first detected at about 90% of embryonic development. In the posthatching period, the quantity of GABA-ir neurons reached the adult status in four days in the cerebral ganglia, and in three weeks in the pedal ganglia. In juveniles, transient expression of GABA was found in the pedal ganglia (fourth cluster).

Age Factors↗

PubMed2001

Development of neuronal ganglion xenografts from gastropoda in rat brain.

Survival of neuronal ganglia from newborn snail (Helix aspera L.) in the brain of adult rats was studied. Snail ganglion survived in the brain of warm-blooded animals for 6 months without inducing immune conflict. At early stages (5 days) after transplantation, xenografts increased in size and were several times larger than native ganglia from 10-day-old snails, thereafter (on days 28 and 180) they became smaller still surpassing the sizes of ganglia from snail of the corresponding age. Rapid enlargement of the xenograft was due to cell reactive processes in the ganglion. Deep penetration of large vessels from xenografts to rat brain was observed.

Animals↗