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PubMed2000

Seasonal plasticity of synaptic connections between identified neurones in Lymnaea.

Here we investigate the synaptic connectivity of the giant dopamine containing neurone (RPeDI) of Lymnaea stagnalis during the winter months, in wild and laboratory bred animals. RPeD1 is one of the three neurones forming the respiratory central pattern generator (CPG) in Lymnaea and initiates ventilation under normal circumstances. Many of the follower cells of RPeD1 are ventilatory motor neurones. The connections of RPeD1 to its follower cells were investigated using standard intracellular recording techniques and dopamine was applied to the follower cells using a puffer pipette. During February and early March, RPeD1 was functionally disconnected from its follower cells, but connections reappeared towards the end of March. Most functionally disconnected cells failed to respond to applied dopamine, consistent with the hypothesis that there is down regulation of dopamine receptors in the follower cells of RPeD1 in the winter months. Behaviourally, Lymnaea that survive the winter, are not active at this time and do not indulge in lung ventilation, but stay quiescent. Thus functional disconnection of neurones from the CPG may be either a cause or a consequence of this change in behaviour.

Animals↗
PubMed1989

Effects of in vitro lead exposure on voltage-sensitive calcium channels differ among cell types in central neurons of Lymnaea stagnalis.

The effects of acute in vitro lead exposure on slowly inactivating voltage-sensitive calcium channels in central neurons of the freshwater pond snail Lymnaea stagnalis were studied under voltage clamp. Three physiologically distinct cell types were used: two subsets of the B cell cluster (Bpos and Bneg) and the pedal giant neuron (RPeD1). In Bpos neurons, 5 nM free Pb2+ irreversibly inhibited current flow through calcium channels by 38 +/- 10%. In Bneg neurons, 5 nM free Pb2+ slightly inhibited inward currents (12 +/- 6%) and may have shifted their voltage dependence to more depolarized voltages. The inhibition and voltage shift were irreversible. In RPeD1 neurons, Pb2+ caused a small, statistically insignificant inhibition of inward current (5 nM free Pb2+; 18 +/- 19%; 30 nM free Pb2+: 31 +/- 23%). The effects of Pb2+ were fully reversible. These data indicate that (1) voltage-sensitive calcium channels in Lymnaea neurons are inhibited by nanomolar concentrations of free Pb2+; (2) there are multiple types of calcium channels in Lymnaea neurons; and (3) the effects of in vitro lead exposure differ qualitatively among channel types.

Animals↗
PubMed1994

The effect of Ambrosia maritima (Damsissa) on the viability of Lymnaea cailliaudi; an experimental study.

The present work aimed at studying the effect of Ambrosia maritima (Damsissa) on the viability of Lymnaea snails. Lymnaea snails used in these trials were acclimatized to laboratory conditions before use in the toxicity test. The snails were exposed to various concentrations (100-3000 mg/l) of dry powdered Damsissa, Ambrosia maritima was lethal to Lymnaea snails at concentration of 3000 mg/L after one day and at 100 mg/l after 14 days.

Animals↗
PubMed2004

Central localization of plasticity involved in appetitive conditioning in Lymnaea.

Learning to associate a conditioned (CS) and unconditioned stimulus (US) results in changes in the processing of CS information. Here, we address directly the question whether chemical appetitive conditioning of Lymnaea feeding behavior involves changes in the peripheral and/or central processing of the CS by using extracellular recording techniques to monitor neuronal activity at two stages of the sensory processing pathway. Our data show that appetitive conditioning does not affect significantly the overall CS response of afferent nerves connecting chemosensory structures in the lips and tentacles to the central nervous system (CNS). In contrast, neuronal output from the cerebral ganglia, which represent the first central processing stage for chemosensory information, is enhanced significantly in response to the CS after appetitive conditioning. This demonstrates that chemical appetitive conditioning in Lymnaea affects the central, but not the peripheral processing of chemosensory information. It also identifies the cerebral ganglia of Lymnaea as an important site for neuronal plasticity and forms the basis for detailed cellular studies of neuronal plasticity.

Animals↗
PubMed1996

Inhibition of the respiratory pattern-generating neurons by an identified whole-body withdrawal interneuron of Lymnaea stagnalis

Respiration and the whole-body withdrawal are two incompatible behaviors in the freshwater snail Lymnaea stagnalis. Whole-body withdrawal behavior is believed to be higher on the behavioral hierarchy than respiratory behavior. A central pattern generator (CPG) underlies respiratory behavior; whole-body withdrawal is mediated by a network of electrically coupled neurons. In this study, we provide evidence that the behavioral hierarchy between the whole-body withdrawal and the respiratory behaviors is established at the interneuronal level. We demonstrate that an identified whole-body withdrawal interneuron inhibits both muscular and neuronal components of the respiratory behavior in Lymnaea stagnalis. A pair of identified, electrically coupled interneurons, termed left and right pedal dorsal 11 (L/RPeD11), coordinates the whole-body withdrawal behavior in Lymnaea stagnalis. In the present study, RPeD11 inhibited spontaneously occurring respiratory CPG activity in isolated brain preparations. In addition, electrical stimulation of RPeD11 in a semi-intact preparation also inhibited respiratory CPG interneuron RPeD1. The synaptic connections between RPeD11 and the respiratory CPG neurons RPeD1 and visceral dorsal 4 (VD4) persisted in the presence of high-Ca2+/high-Mg2+ saline, suggesting the possibility that they may be monosynaptic. In a semi-intact preparation (lung­mantle, pneumostome and central nervous system), electrical stimulation of RPeD11 induced pneumostome and columellar muscle contractions while inhibiting the activity of RPeD1. Moreover, mechanical stimulation of the respiratory orifice, the pneumostome, excited RPeD11, while its effects on the respiratory CPG neuron (RPeD1) were inhibitory. To determine the monosynaptic nature of connections between RPeD11 and the respiratory CPG neurons in the intact nervous system, we constructed these synapses in culture. RPeD11 and individual respiratory interneurons were isolated from their respective ganglia and co-cultured under conditions that support neurite outgrowth. Following neuritic overlap, RPeD11 was found to establish inhibitory synapses with the respiratory interneurons, supporting the hypothesis that these synaptic connections are likely to be monosynaptic in the intact central nervous system.

Journal Article↗
PubMed1999

Development of catecholaminergic neurons in the pond snail, Lymnaea stagnalis: I. Embryonic development of dopamine-containing neurons and dopamine-dependent behaviors.

The embryonic development of the catecholaminergic system of the pond snail, Lymnaea stagnalis, was investigated by using chromatographic and histochemical methods. High performance liquid chromatography suggested that dopamine was the only catecholamine present in significant concentrations throughout the embryonic development of Lymnaea. Dopamine first became detectable at about embryonic stage (E) 15 (15% of embryonic development) and then increased in amount during early development to reach about 120-140 fmol per animal by around E40. Dopamine content remained stable during mid-embryogenesis (E40-65), increased slowing for the next couple of days, and then increased rapidly to culminate at about 400 fmol per animal by hatching. The detection of aldehyde- and glyoxylate-induced fluorescence and of tyrosine hydroxylaselike immunoreactivity indicated that the first catecholaminergic cells appeared in the late trochophore or early veliger stage of embryonic development (E32-35). The paired perikarya of these transient apical catecholaminergic (TAC) neurons were located beneath the apical plate, remained outside of the central ganglia during embryogenesis, and no longer contained detectable catecholamines close to hatching. TAC neurons bore cilia on the ends of short processes that penetrated the overlying epithelium; their long processes branched repeatedly under the ciliated apical plate. Several smaller catecholaminergic cells first appeared in the anterior margin of the foot at a stage when the embryos began to metamorphose from the veliger form (E55). Similar bipolar cells later appeared in the tentacle and lips. The axons of all of these small peripheral cells projected centrally and terminated within the neuropil of different central ganglia. Central catecholaminergic neurons, including RPeD1, differentiated only after metamorphosis was complete (E75). Development of locomotor, respiratory, and feeding behaviors correlated with maturation of catecholaminergic neurons, as indicated by histology and chromatography.

Animals↗
PubMed1996

Atrophy and degeneration of peptidergic neurons and cessation of egg laying in the aging pond snail Lymnaea stagnalis.

The morphology of the neuroendocrine caudodorsal cells (CDCs), which are involved in the regulation of female reproduction in the pond snail Lymnaea stagnalis, was studied in young (200 to 234 days of age) and old (400 to 500 days) animals. Lucifer Yellow fills of ventral CDCs showed that in young animals ventral CDCs branch ipsilaterally as well as contralaterally in the cerebral commissure. In old animals these branches were reduced at different degrees and in some cases even lacking completely, leaving only an axon crossing the commissure. Immunocytochemical stainings with antibodies against CDC peptides (CDCH-I and alpha CDCP) corroborated the finding that ventral CDCs degenerate. Among the other types of CDCs (dorsal, lateral), degeneration was found as well. The immunocytochemical findings showed that in old animals the axon terminals of the CDCs were strongly stained, indicating that they are packed with secretory vesicles containing peptides. It was also found that these darkly stained, peptide-containing axon terminals protruded into the perineurium. These findings suggest that accumulation of peptides in the terminals of the CDCs of old animals may be due to the impaired release. The relationship between atrophy and degeneration of CDCs and cessation of egg-laying activity in Lymnaea is discussed.

Aging↗
PubMed2005

Ltrk is differentially expressed in developing and adult neurons of the Lymnaea central nervous system.

The Trk receptor family plays diverse roles in both development and plasticity of the vertebrate nervous system. Ltrk is a related receptor that is expressed in the CNS of the mollusk Lymnaea, although little is known of its cellular distribution. This study provides three independent lines of evidence (based on RT-PCR, in situ hybridization, and immunohistochemistry) that Ltrk is universally expressed by neurons and dorsal body cells of both the juvenile and the adult Lymnaea CNS. The highest level of expression by neuronal somata occurs in the late juvenile stage, whereas axon collaterals express high levels throughout the animal's life span. Our data support multifunctional roles for Ltrk that parallel those of its mammalian counterparts.

Age Factors↗
PubMed2005

Epidermal growth factor-dependent enhancement of axonal regeneration in the pond snail Lymnaea stagnalis: role of phagocyte survival.

Peripheral nerve injury triggers complex responses from neuronal as well as from multiple nonneuronal cell types. These responses are coordinated by a wide spectrum of secreted and nonsecreted factors, including growth factors, cytokines, and cell adhesion molecules. These molecules originate from different sources and act both locally at the site of injury as well as centrally at the location of the neuronal cell bodies. One of the signal systems frequently implicated in this process is the epidermal growth factor (EGF) family and its receptors. Expression of members of this family as well as that of EGF-receptors is upregulated in different cell types after peripheral nerve injury. However, the functional significance of this response is unclear. Using a simple invertebrate model system (Lymnaea stagnalis), the present study implicates the EGF/EGF-receptor system in the survival of ionized calcium-binding adaptor molecule 1 (Iba1)-positive phagocytes that reside in the nervous system. We show that inhibiting the EGF-signaling pathway enhances cell death in this type of cell, an effect paralleled by a substantial reduction in axonal regeneration. Therefore, complementing our previous observation that Lymnaea EGF provides trophic support to axotomized neurons, the present results emphasize the significance of nonneuronal actions of EGF receptor ligands in axonal regeneration. Thus, we add a novel perspective to the ongoing discussion on the functional significance of the EGF signaling system in the injury responses of the nervous system.

Animals↗
PubMed1989

Postembryonic development of serotoninlike immunoreactivity in the central nervous system of the snail, Lymnaea stagnalis.

Posthatching growth in the pond snail Lymnaea stagnalis involves approximately a 20-fold increase in the linear dimensions of the ganglia composing the central nervous system. Developmental change within the population of neurons exhibiting serotoninlike immunoreactivity (SLIR) was examined in order to explain this growth in cellular terms. The study indicates that at least two factors contribute to the growth of the nervous system. First, SLIR cells approximately double in number from the 200-250 cells in hatchlings to the complement found in animals approaching sexual maturity. Much of this increase in cell number occurred within identifiable discrete clusters of neurons with different clusters adding cells at different rates and at different times. The number of SLIR cells also increased in more diffuse populations, particularly along the medial aspects of the paired pedal and the right parietal ganglion. No identified cells were added postembryonically. In addition to the increases in neuron numbers, posthatching development in Lymnaea also involves the growth of individual cells. All cells examined showed continuous somatic growth during posthatching development, but different identified cells and different cell clusters were characterized by different rates of relative growth. Together, the results highlight the complexity of postembryonic development in the snail by indicating the temporal and spatial specificity for both cell addition and cell growth within the nervous system.

Age Factors↗
PubMed1991

Dopamine-immunoreactive neurones in the central nervous system of the pond snail Lymnaea stagnalis.

The distribution of dopamine and dopamine-immunoreactive neurones was studied in the central nervous system of the snail Lymnaea stagnalis. The results from immunocytochemical labelling were compared with those from the application of the glyoxylic acid fluorescence method and 6-hydroxydopamine-induced pigment labelling. Comparisons were also made between the number of dopamine immunoreactive neurones and the dopamine content of the ganglia, measured by high-performance liquid chromatography. Dopamine immunocytochemistry proved to be superior to the other two histochemical techniques in terms of specificity and sensitivity. The 6-hydroxydopamine-induced pigment labelling failed to prove a useful tool for the in vivo identification of all dopamine-containing neurones. The distribution and number of dopamine-immunoreactive neurones and levels of biochemically measured dopamine in specific ganglia showed a close correspondence. By using the results of the dopamine immunocytochemistry and glyoxylic acid technique, a detailed map of dopamine-containing neurones was constructed. Dopamine-containing inter- and intra-ganglionic axon tracts were also demonstrated. The mapping of dopamine-containing neurones will facilitate further neurophysiological analysis of dopaminergic neural mechanisms in Lymnaea.

Animals↗
PubMed1994

Neuropeptides myomodulin, small cardioactive peptide, and buccalin in the central nervous system of Lymnaea stagnalis: purification, immunoreactivity, and artifacts.

The neuropeptides myomodulin, small cardioactive peptide (SCP), and buccalin are widely distributed in the phylum Mollusca and have important physiological functions. Here, we describe the detailed distribution of each class of peptide in the central nervous system (CNS) of the snail Lymnaea stagnalis by the use of immunocytochemical techniques combined with dye-marking of electrophysiologically identified neurons. We report the isolation and structural characterization of a Lymnaea myomodulin, PMSMLRLamide, identical to myomodulin A of Aplysia californica. Myomodulin immunoreactivity was localized in all 11 ganglia, in their connectives, and in peripheral nerves. In many cases, myomodulin immunoreactivity appeared localized in neuronal clusters expressing FMRFamide-like peptides, but also in a large number of additional neurons. Double-labelling experiments demonstrated myomodulin immunoreactivity in the visceral white interneuron, involved in regulation of cardiorespiration. SCP-like immunoreactivity also appeared in all ganglia, and double-labelling experiments revealed that in many locations it was specifically associated with clusters expressing distinct exons of the FMRFamide gene that are differentially expressed in the CNS. Characterization of the two types of SCP-antisera used in this study, however, suggested that they cross-reacted with both FMRFamide and N-terminally extended FMRFamide-like peptides. Selective preadsorption with these cross-reacting peptides resulted in elimination of the widespread staining and retention of bona fide SCP immunoreactivity in the buccal and pedal ganglia only. Buccalin immunoreactivity was limited to the buccal and pedal ganglia. It did not coincide with the distribution of either myomodulin or SCP. Most immunoreactive clusters were found in the pedal ganglia.

Amino Acid Sequence↗
PubMed1994

Neural network controlling feeding in Lymnaea stagnalis: immunocytochemical localization of myomodulin, small cardioactive peptide, buccalin, and FMRFamide-related peptides.

This paper investigates the distribution of four classes of neuropeptides, myomodulin, small cardioactive peptide (SCP), buccalin, and FMRFamide, in central neurons forming the network that underlies feeding behavior in the snail Lymnaea stagnalis. Intracellular dye-marking and immunocytochemical analysis, using antisera to the different classes of peptides, were applied to identified neurons of all three levels of the hierarchy of the circuitry: modulatory interneurons (cerebral giant cells, CGC; slow oscillator, SO), central pattern generator (CPG) interneurons (N1, N2, N3), motoneurons (B1-B10), and their peripheral target organs. Myomodulin immunoreactivity was detected in the CGC interneurons, in the SO, and in ventral N2-type CPG interneurons. Several large buccal motoneurons, the paired B1, B2, B3, B7, and neurons located in the dorsal posterior area (putative B4 cluster types) were also myomodulin immunoreactive. Target organs of buccal motoneurons, the buccal mass, salivary glands, and oesophagus contained myomodulin-immunopositive fibers. SCP appeared in N2-type interneurons and was found colocalized with myomodulin in the B1 and B2 motoneurons. SCP-containing neurons in the B4 cluster area were also detected. The buccal mass and salivary glands exhibited SCP-immunoreactive fibers. Buccalin immunoreactivity was scarce in the buccal ganglia and was identified only in N1-type interneurons and three pairs of dorsal posterior neurons. In the periphery, immunoreactive fibers were localized in the oesophagus only. None of the buccal neuronal types examined revealed immunoreactivity to SEQPDVDDYLRDVVLQSEEPLY ("SEEPLY"), a peptide encoded in the FMRFamide precursor protein of Lymnaea. SEEPLY immunoreactivity was confined to a pair of novel ventral neurons with projections to the laterobuccal nerve innervating the buccal mass. Immunoreactive fibers were also traced in this organ.

Amino Acid Sequence↗
PubMed1996

A new Ig-superfamily member, molluscan defence molecule (MDM) from Lymnaea stagnalis, is down-regulated during parasitosis.

To survive the attacks of the internal defence system (IDS) of their host, parasites have developed various strategies to manipulate the IDS. We present evidence that the avian schistosome parasite Trichobilharzia ocellata affects gene expression in the granular cells, a cell type of the IDS of the intermediate host, the mollusc Lymnaea stagnalis. From a differential screening, a clone was isolated encoding a protein named molluscan defence molecule (MDM), which encompasses five C2-like immunoglobulin (Ig) domains. The protein shares a domain organization and high amino acid sequence identity with hemolin, an Ig-family member of the insect IDS. Interestingly, both MDM and hemolin have highest sequence identity with neural cell adhesion molecules, but lack the typical fibronectin repeats and motifs for membrane anchors. We find that the expression of the MDM gene is gradually down-regulated during the course of parasitosis to approximately 21% compared to the non-parasitized level, 8 weeks post-infection. Based on our findings, we suggest that MDM is involved in the proper function of the Lymnaea IDS, and that down-regulation of MDM is part of the parasite-induced disabling on non-self recognition.

Amino Acid Sequence↗
PubMedDate not supplied

Histopathological effects in tissues of snail Lymnaea stagnalis (Gastropoda, Pulmonata) exposed to sublethal concentration of Thiodan and recovery after exposure.

Histopathological alterations induced by Thiodan in three tissues, namely, digestive gland, foot and mantle, of the freshwater snail Lymnaea stagnalis were investigated. Specimens of Lymnaea stagnalis were exposed to 0.36% and 0.72% Thiodan 35 EC, a commercial grade of endosulfan, for 96 h followed by a recovery period of 30 days. Thiodan caused significant dose dependent histopathological changes in all the tissues of the snail. Irreversible necrotic changes occurred in the digestive gland of the snail following Thiodan exposure. Degenerative changes in the muscle fiber of the foot, protein and pigment cells of the foot and the connective tissue element of the mantle were recovered after 30 days of recovery of the snail in pesticide-free freshwater.

Animals↗
PubMed1991

Neuropeptide schistosomin inhibits hormonally-induced ovulation in the freshwater snail Lymnaea stagnalis.

This study examines the interaction between the caudodorsal cell hormone (CDCH) and schistosomin, a peptide secreted by the central nervous system of the snail (Lymnaea stagnalis) infected with the avian schistosome Trichobilharzia ocellata. Non-infected snails were injected with synthetic as well as native CDCH in the absence or presence of purified schistosomin. The response to 2 pmol of synthetic CDCH was blocked for 90% by coinjection with 3.5 pmol of schistosomin. The ovulation-inducing activity of extracts of cerebral commissures (the storage area of native CDCH) was also blocked by schistosomin. The degree of inhibition (65%), however, was less than that observed with synthetic CDCH. These results show that schistosomin inhibits ovulation and egg laying in Lymnaea. This explains the decrease or absence of egg laying in schistosome-infected freshwater snails.

Animals↗
PubMed1991

Nerve growth factor (NGF) induces sprouting of specific neurons of the snail, Lymnaea stagnalis.

Nerve growth factor (NGF) was examined for its ability to elicit sprouting by adult molluscan neurons. Motoneurons and interneurons (but not neurosecretory cells) from Lymnaea exhibited a sprouting response to murine 2.5S NGF in defined medium with a half-maximal response at about 150 ng/mL. Furthermore, an NGF antiserum blocked sprouting by all normally responsive neurons. We tested whether an NGF-like molecule is a component of conditioned medium (CM) by attempting to preabsorb its sprout-inducing activity with NGF antiserum. Treatment of CM with immune (but not nonimmune) serum largely blocked the response of motoneurons, but not that of neurosecretory cells, to CM. We conclude that NGF exerts neurotrophic activity on specific adult Lymnaea neurons, and suggest the possibility that an NGF-like molecule may exist in the molluscan nervous system.

Absorption↗
PubMed1998

Associative learning of visual and vestibular stimuli in Lymnaea.

A conditioned withdrawal response was characterized in the pond snail Lymnaea stagnalis. Using light as the conditioned stimulus and high-speed orbital rotation as the unconditioned stimulus, experimental animals were trained with 30 paired presentations of light and orbital rotation per day for 3 days. After training, all experimental animals responded to light with a withdrawal response, the conditioned response. Control animals exposed to the same number of explicitly unpaired presentations of light and orbital rotation, light alone, or no stimulation did not respond to light. Thirty paired presentations per day for 2 days produced less than optimal acquisition of the conditioned withdrawal response. Neither 45 paired presentations per day for 2 days nor 90 paired presentations for 1 day resulted in complete acquisition of the conditioned withdrawal response. The conditioned withdrawal response observed following 30 paired presentations per day for 3 to 5 days persisted to Day 10, regardless of the number of training days. As a measure of savings, reacquisition of the conditioned response after extinction was investigated. After the conditioned withdrawal response was extinguished, only 2 to 5 paired presentations of light and orbital rotation were required for reacquisition of the conditioned response for most animals. This study further establishes Lymnaea as an animal model of basic associative learning.

Animals↗