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Ken Lukowiak - One of the best experts on this subject based on the ideXlab platform.
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Configural learning: a higher form of learning in Lymnaea.
The Journal of Experimental Biology, 2019Co-Authors: Cayley Swinton, Cailin M. Rothwell, Erin Swinton, Tamila Shymansky, Emily Hughes, Jack Zhang, Mili Kakadiya, Ken LukowiakAbstract:ABSTRACT Events typically occur in a specific context and the ability to assign importance to this occurrence plays a significant role in memory formation and recall. When the scent of a crayfish predator (CE) is encountered in Lymnaea stagnalis strains known to be predator experienced (e.g. the W-strain), enhancement of memory formation and depression of feeding occur, which are part of a suite of anti-predator behaviours. We hypothesized that Lymnaea possess a form of higher-order conditioning, namely configural learning. We tested this by simultaneously exposing W-strain Lymnaea to a carrot food odour (CO) and predator scent (CE). Two hours later, we operantly conditioned these snails with a single 0.5 h training session in CO to determine whether training in CO results in long-term memory (LTM) formation. A series of control experiments followed and demonstrated that only the CO+CE snails trained in CO had acquired enhanced memory-forming ability. Additionally, following CE+CO pairing, CO no longer elicited an increased feeding response. Hence, snails have the ability to undergo configural learning. Following configural learning, CO becomes a risk signal and evokes behavioural responses phenotypically similar to those elicited by exposure to CE.
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Monoamines, Insulin and the Roles They Play in Associative Learning in Pond Snails
Frontiers Media S.A., 2019Co-Authors: Yuki Totani, Hitoshi Aonuma, Ken Lukowiak, Manabu Sakakibara, Takayuki Watanabe, Dai Hatakeyama, Etsuro Ito, Akira OikeAbstract:Molluscan gastropods have long been used for studying the cellular and molecular mechanisms underlying learning and memory. One such gastropod, the pond snail Lymnaea stagnalis, exhibits long-term memory (LTM) following both classical and operant conditioning. Using Lymnaea, we have successfully elucidated cellular mechanisms of learning and memory utilizing an aversive classical conditioning procedure, conditioned taste aversion (CTA). Here, we present the behavioral changes following CTA training and show that the memory score depends on the duration of food deprivation. Then, we describe the relationship between the memory scores and the monoamine contents of the central nervous system (CNS). A comparison of learning capability in two different strains of Lymnaea, as well as the filial 1 (F1) cross from the two strains, presents how the memory scores are correlated in these populations with monoamine contents. Overall, when the memory scores are better, the monoamine contents of the CNS are lower. We also found that as the insulin content of the CNS decreases so does the monoamine contents which are correlated with higher memory scores. The present review deepens the relationship between monoamine and insulin contents with the memory score
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Strain-specific effects of crowding on long-term memory formation in Lymnaea.
Comparative biochemistry and physiology. Part A Molecular & integrative physiology, 2018Co-Authors: Shawn Dodd, Cailin M. Rothwell, Ken LukowiakAbstract:Abstract Crowding of snails is a stress that obstructs long-term memory (LTM) formation following operant conditioning of aerial respiratory behaviour in Lymnaea stagnalis. In previous experiments, snails of the same strain/population were used for both the crowding and the operant conditioning training. However, there are different strains/populations of Lymnaea stagnalis exhibiting different cognitive abilities. We asked whether Lymnaea of one strain/population are able to determine that they are of a different strain/population. We did this by asking if LTM formation would continue to be obstructed if we crowded snails with a different species of pond snail (Helisoma = Planorbella) or with different strains/populations of Lymnaea stagnalis. Using an inbred strain, the W-strain, we crowded the W-strain with seven other Lymnaea strains/populations as well as with Helisoma. The results of a 2-Way ANOVA followed by a Tukey's Post-hoc analysis showed that W-strain snails when crowded with another strain/population of Lymnaea or with Helisoma formed LTM formation. That is, the memory test session statistically met the criteria for LTM formation. Thus, one strain/population of snails determines that another strain/population is different from it. The differentness means that crowding now does not obstruct LTM formation.
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Pharmacological effects of cannabinoids on learning and memory in Lymnaea
The Journal of Experimental Biology, 2017Co-Authors: Hiroshi Sunada, Manabu Sakakibara, Takayuki Watanabe, Dai Hatakeyama, Sangmin Lee, Jeremy Forest, Etsuro Ito, Ken LukowiakAbstract:ABSTRACT Cannabinoids are hypothesized to play an important role in modulating learning and memory formation. Here, we identified mRNAs expressed in Lymnaea stagnalis central nervous system that encode two G-protein-coupled receptors ( Lymnaea CBr-like 1 and 2) that structurally resemble mammalian cannabinoid receptors (CBrs). We found that injection of a mammalian CBr agonist WIN 55,212-2 (WIN 55) into the snail before operant conditioning obstructed learning and memory formation. This effect of WIN 55 injection persisted for at least 4 days following its injection. A similar obstruction of learning and memory occurred when a severe traumatic stimulus was delivered to L. stagnalis . In contrast, injection of a mammalian CBr antagonist AM 251 enhanced long-term memory formation in snails and reduced the duration of the effects of the severe traumatic stressor on learning and memory. Neither WIN 55 nor AM 251 altered normal homeostatic aerial respiratory behaviour elicited in hypoxic conditions. Our results suggest that putative cannabinoid receptors mediate stressful stimuli that alter learning and memory formation in Lymnaea . This is also the first demonstration that putative CBrs are present in Lymnaea and play a key role in learning and memory formation.
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The role of serotonin in the enhancement of long-term memory resulting from predator detection in Lymnaea.
Journal of Experimental Biology, 2010Co-Authors: Jae Il-han, Tara Janes, Ken LukowiakAbstract:SUMMARY Serotonergic systems play important roles in modulating stress-induced arousal and vigilance behaviours. The pond snail, Lymnaea, shows multiple defensive vigilance behaviours in response to the stress associated with predator detection. Predator detection elicited by crayfish effluent (CE), increases the time to re-emerge from the shell and enhances the shadow withdrawal response. More importantly, in Lymnaea, CE enhances the ability to form long-term memory (LTM). We investigated the role of the serotonergic system in these anti-predator responses in Lymnaea. Using a serotonin-receptor antagonist, mianserin, we found that two defensive vigilance behaviours (e.g. increasing the time to re-emerge from their shell and shadow response) elicited by CE were not observed when the serotonergic system was disrupted. Also, methysergide, another serotonin antagonist, blocked the enhanced LTM formation after training in CE. Importantly, mianserin did not alter LTM formation in pond water (PW). These data suggest that a serotonergic system is activated only when Lymnaea detect a predator. When snails were trained in CE using a training procedure that in PW produces a 24-h LTM, a more persistent form of LTM (5 days) occurred. This more persistent form of LTM was abolished after mianserin treatment. Increasing 5-HT levels in the snail by the injection of 5-HT was also associated with enhanced LTM formation. Lastly, we tested whether the osphradium is implicated in CE detection and subsequent enhanced formation of LTM. Cutting the osphradial nerve to the CNS resulted in the loss of the ability to form enhanced LTM in CE. Together, these findings support the hypothesis that the serotonergic system plays a key role in modulating the predator-induced stress responses in Lymnaea.
Manabu Sakakibara - One of the best experts on this subject based on the ideXlab platform.
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Features of behavioral changes underlying conditioned taste aversion in the pond snail Lymnaea stagnalis
Invertebrate Neuroscience, 2020Co-Authors: Junko Nakai, Manabu Sakakibara, Yuki Totani, Satoshi Kojima, Etsuro ItoAbstract:Conditioned taste aversion (CTA) in the freshwater pulmonate Lymnaea stagnalis can be formed by presenting ten pairings of sucrose as the conditioned stimulus (CS) and KCl as the unconditioned stimulus (US). The CTA is consolidated to long-term memory (LTM) lasting longer than a month. In the present study, we examined the time course of protein synthesis-dependent period during the consolidation of Lymnaea CTA to LTM by pharmacological inhibition of transcription or translation. The robustness for CTA–LTM was then examined by extinction trials, i.e., repeated presentations of the CS alone. Furthermore, we evaluated the effects of the interstimulus interval (ISI) between the presentation of the CS and US. Our findings indicated that the protein synthesis-dependent period coincides with the CTA training. Repeated presentations of the CS alone after establishment of CTA did not extinguish the CTA, demonstrating the robustness of the CTA–LTM. The ISI ranged from 10 s to a few minutes, and there was no inverted U-shaped function between the ISI and the conditioned response (i.e., suppression of feeding). Thus, CTA still formed even when the presentation of the US was delayed. These features of Lymnaea CTA complement the knowledge for mammalian CTA.
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Monoamines, Insulin and the Roles They Play in Associative Learning in Pond Snails
Frontiers Media S.A., 2019Co-Authors: Yuki Totani, Hitoshi Aonuma, Ken Lukowiak, Manabu Sakakibara, Takayuki Watanabe, Dai Hatakeyama, Etsuro Ito, Akira OikeAbstract:Molluscan gastropods have long been used for studying the cellular and molecular mechanisms underlying learning and memory. One such gastropod, the pond snail Lymnaea stagnalis, exhibits long-term memory (LTM) following both classical and operant conditioning. Using Lymnaea, we have successfully elucidated cellular mechanisms of learning and memory utilizing an aversive classical conditioning procedure, conditioned taste aversion (CTA). Here, we present the behavioral changes following CTA training and show that the memory score depends on the duration of food deprivation. Then, we describe the relationship between the memory scores and the monoamine contents of the central nervous system (CNS). A comparison of learning capability in two different strains of Lymnaea, as well as the filial 1 (F1) cross from the two strains, presents how the memory scores are correlated in these populations with monoamine contents. Overall, when the memory scores are better, the monoamine contents of the CNS are lower. We also found that as the insulin content of the CNS decreases so does the monoamine contents which are correlated with higher memory scores. The present review deepens the relationship between monoamine and insulin contents with the memory score
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Pharmacological effects of cannabinoids on learning and memory in Lymnaea
The Journal of Experimental Biology, 2017Co-Authors: Hiroshi Sunada, Manabu Sakakibara, Takayuki Watanabe, Dai Hatakeyama, Sangmin Lee, Jeremy Forest, Etsuro Ito, Ken LukowiakAbstract:ABSTRACT Cannabinoids are hypothesized to play an important role in modulating learning and memory formation. Here, we identified mRNAs expressed in Lymnaea stagnalis central nervous system that encode two G-protein-coupled receptors ( Lymnaea CBr-like 1 and 2) that structurally resemble mammalian cannabinoid receptors (CBrs). We found that injection of a mammalian CBr agonist WIN 55,212-2 (WIN 55) into the snail before operant conditioning obstructed learning and memory formation. This effect of WIN 55 injection persisted for at least 4 days following its injection. A similar obstruction of learning and memory occurred when a severe traumatic stimulus was delivered to L. stagnalis . In contrast, injection of a mammalian CBr antagonist AM 251 enhanced long-term memory formation in snails and reduced the duration of the effects of the severe traumatic stressor on learning and memory. Neither WIN 55 nor AM 251 altered normal homeostatic aerial respiratory behaviour elicited in hypoxic conditions. Our results suggest that putative cannabinoid receptors mediate stressful stimuli that alter learning and memory formation in Lymnaea . This is also the first demonstration that putative CBrs are present in Lymnaea and play a key role in learning and memory formation.
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Comparative Study of Visuo-Vestibular Conditioning in Lymnaea stagnalis
The Biological bulletin, 2006Co-Authors: Manabu SakakibaraAbstract:In this review, we compare the current understanding of visuo-vestibular conditioning in Hermissenda crassicornis and Lymnaea stagnalis on the basis of behavioral, electrophysiologic, and morphologic studies. Paired presentation of a photic conditioned stimulus (CS) and an orbital rotation unconditioned stimulus (US) results in conditioned escape behavior in both species. In Hermissenda, changes in excitability of type B photoreceptors and morphologic modifications at the axon terminals follow conditioning. Caudal hair cells, which detect mechanical turbulence, have reciprocal inhibition with type B photoreceptors. In Lymnaea, the interaction between photoreceptors and hair cells is dependent on statocyst location. Furthermore, the organization of the Lymnaea eye is complex, with more than 100 photoreceptors distributed in a uniquely folded retina. Although the optimal conditions to produce long-term memory (memory persistent for >1 week) are almost identical in Hermissenda and Lymnaea, physiologic and mo...
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Electrophysiological responses to light of neurons in the eye and statocyst of Lymnaea stagnalis.
Journal of neurophysiology, 2004Co-Authors: Manabu Sakakibara, Tomoyo Aritaka, Akira Iizuka, Hiroyuki Suzuki, Tetsuro Horikoshi, Ken LukowiakAbstract:Lymnaea can be classically conditioned by pairing photic stimulation with a rotational stimulus. The electrophysiological properties of the Lymnaea photoreceptors and statocyst neurons are incomple...
Hisayo Sadamoto - One of the best experts on this subject based on the ideXlab platform.
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de novo sequencing and transcriptome analysis of the central nervous system of mollusc Lymnaea stagnalis by deep rna sequencing
PLOS ONE, 2012Co-Authors: Hisayo Sadamoto, Hironobu Takahashi, Taketo Okada, Hiromichi Kenmoku, Masao Toyota, Yoshinori AsakawaAbstract:The pond snail Lymnaea stagnalis is among several mollusc species that have been well investigated due to the simplicity of their nervous systems and large identifiable neurons. Nonetheless, despite the continued attention given to the physiological characteristics of its nervous system, the genetic information of the Lymnaea central nervous system (CNS) has not yet been fully explored. The absence of genetic information is a large disadvantage for transcriptome sequencing because it makes transcriptome assembly difficult. We here performed transcriptome sequencing for Lymnaea CNS using an Illumina Genome Analyzer IIx platform and obtained 81.9 M of 100 base pair (bp) single end reads. For de novo assembly, five programs were used: ABySS, Velvet, OASES, Trinity and Rnnotator. Based on a comparison of the assemblies, we chose the Rnnotator dataset for the following blast searches and gene ontology analyses. The present dataset, 116,355 contigs of Lymnaea transcriptome shotgun assembly (TSA), contained longer sequences and was much larger compared to the previously reported Lymnaea expression sequence tag (EST) established by classical Sanger sequencing. The TSA sequences were subjected to blast analyses against several protein databases and Aplysia EST data. The results demonstrated that about 20,000 sequences had significant similarity to the reported sequences using a cutoff value of 1e-6, and showed the lack of molluscan sequences in the public databases. The richness of the present TSA data allowed us to identify a large number of new transcripts in Lymnaea and molluscan species.
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Localization of serotonin transporter mRNA in the CNS of Lymnaea stagnalis.
Acta biologica Hungarica, 2008Co-Authors: Hisayo Sadamoto, Z Serfozo, E ItoAbstract:The serotonin transporter, SERT, is reported as a key molecule that regulates serotonergic neurotransmission. In the present study, we analyzed the localization of Lymnaea SERT (LymSERT) mRNA-containing neurons by in situ hybridization using frozen sections of the central nervous system (CNS) of Lymnaea. To precisely demonstrate the distribution of LymSERT mRNA-containing neurons, colocalization with serotonin immunoreactivity was also examined. The results showed that LymSERT mRNA was constitutively expressed and localized in the serotonin-containing neurons in the CNS.
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altered gene activity correlated with long term memory formation of conditioned taste aversion in Lymnaea
Journal of Neuroscience Research, 2006Co-Authors: Sachiyo Azami, Hisayo Sadamoto, Yutaka Fujito, Dai Hatakeyama, Akiko Wagatsuma, Takeshi Usami, Manabu Fujie, Ryo Koyanagi, Kaoru Azumi, Ken LukowiakAbstract:The pond snail Lymnaea stagnalis is capable of learning conditioned taste aversion (CTA) and then consolidating that learning into long-term memory (LTM) that persists for at least 1 month. LTM requires de novo protein synthesis and altered gene activity. Changes in gene activity in Lymnaea that are correlated with, much less causative, memory formation have not yet been identified. As a first step toward rectifying this situation, we constructed a cDNA microarray with mRNAs extracted from the central nervous system (CNS) of Lymnaea. We then, using this microarray assay, identified genes whose activity either increased or decreased following CTA memory consolidation. We also identified genes whose expression levels were altered after inhibition of the cyclic AMP response element-binding protein (CREB) that is hypothesized to be a key transcription factor for CTA memory. We found that the molluscan insulin-related peptide II (MIP II) was up-regulated during CTA-LTM, whereas the gene encoding pedal peptide preprohormone (Pep) was down-regulated by CREB2 RNA interference. We next examined mRNAs of MIP II and Pep using real-time RT-PCR with SYBR Green. The MIP II mRNA level in the CNS of snails exhibiting "good" memory for CTA was confirmed to be significantly higher than that from the CNS of snails exhibiting "poor" memory. In contrast, there was no significant difference in expression levels of the Pep mRNA between "good" and "poor" performers. These data suggest that in Lymnaea MIP II may play a role in the consolidation process that forms LTM following CTA training.
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CREB in the pond snail Lymnaea stagnalis: cloning, gene expression, and function in identifiable neurons of the central nervous system
Journal of neurobiology, 2004Co-Authors: Hisayo Sadamoto, Hanae Sato, Suguru Kobayashi, Jun Murakami, Hitoshi Aonuma, Hironori Ando, Yutaka Fujito, Kaoru Hamano, Masahiko Awaji, Ken LukowiakAbstract:The pond snail Lymnaea stagnalis is an excellent model system in which to study the neuronal and molecular substrates of associative learning and its consolidation into long-term memory. Until now, the presence of cyclic AMP (cAMP)-responsive element binding protein (CREB), which is believed to be a necessary component in the process of a learned behavior that is consolidated into long-term memory, has only been assumed in Lymnaea neurons. We therefore cloned and analyzed the cDNA sequences of homologues of CREB1 and CREB2 and determined the presence of these mRNAs in identifiable neurons of the central nervous system (CNS) of L. stagnalis. The deduced amino acid sequence of Lymnaea CREB1 is homologous to transcriptional activators, mammalian CREB1 and Aplysia CREB1a, in the C-terminal DNA binding (bZIP) and phosphorylation domains, whereas the deduced amino acid sequence of Lymnaea CREB2 is homologous to transcriptional repressors, human CREB2, mouse activating transcription factor-4, and Aplysia CREB2 in the bZIP domain. In situ hybridization revealed that only a relatively few neurons showed strongly positive signals for Lymnaea CREB1 mRNA, whereas all the neurons in the CNS contained Lymnaea CREB2 mRNA. Using one of the neurons (the cerebral giant cell) containing Lymnaea CREB1 mRNA, we showed that the injection of a CRE oligonucleotide inhibited a cAMP-induced, long-lasting synaptic plasticity. We therefore conclude that CREBs are present in Lymnaea neurons and may function as necessary players in behavioral plasticity.
Dai Hatakeyama - One of the best experts on this subject based on the ideXlab platform.
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Monoamines, Insulin and the Roles They Play in Associative Learning in Pond Snails
Frontiers Media S.A., 2019Co-Authors: Yuki Totani, Hitoshi Aonuma, Ken Lukowiak, Manabu Sakakibara, Takayuki Watanabe, Dai Hatakeyama, Etsuro Ito, Akira OikeAbstract:Molluscan gastropods have long been used for studying the cellular and molecular mechanisms underlying learning and memory. One such gastropod, the pond snail Lymnaea stagnalis, exhibits long-term memory (LTM) following both classical and operant conditioning. Using Lymnaea, we have successfully elucidated cellular mechanisms of learning and memory utilizing an aversive classical conditioning procedure, conditioned taste aversion (CTA). Here, we present the behavioral changes following CTA training and show that the memory score depends on the duration of food deprivation. Then, we describe the relationship between the memory scores and the monoamine contents of the central nervous system (CNS). A comparison of learning capability in two different strains of Lymnaea, as well as the filial 1 (F1) cross from the two strains, presents how the memory scores are correlated in these populations with monoamine contents. Overall, when the memory scores are better, the monoamine contents of the CNS are lower. We also found that as the insulin content of the CNS decreases so does the monoamine contents which are correlated with higher memory scores. The present review deepens the relationship between monoamine and insulin contents with the memory score
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Pharmacological effects of cannabinoids on learning and memory in Lymnaea
The Journal of Experimental Biology, 2017Co-Authors: Hiroshi Sunada, Manabu Sakakibara, Takayuki Watanabe, Dai Hatakeyama, Sangmin Lee, Jeremy Forest, Etsuro Ito, Ken LukowiakAbstract:ABSTRACT Cannabinoids are hypothesized to play an important role in modulating learning and memory formation. Here, we identified mRNAs expressed in Lymnaea stagnalis central nervous system that encode two G-protein-coupled receptors ( Lymnaea CBr-like 1 and 2) that structurally resemble mammalian cannabinoid receptors (CBrs). We found that injection of a mammalian CBr agonist WIN 55,212-2 (WIN 55) into the snail before operant conditioning obstructed learning and memory formation. This effect of WIN 55 injection persisted for at least 4 days following its injection. A similar obstruction of learning and memory occurred when a severe traumatic stimulus was delivered to L. stagnalis . In contrast, injection of a mammalian CBr antagonist AM 251 enhanced long-term memory formation in snails and reduced the duration of the effects of the severe traumatic stressor on learning and memory. Neither WIN 55 nor AM 251 altered normal homeostatic aerial respiratory behaviour elicited in hypoxic conditions. Our results suggest that putative cannabinoid receptors mediate stressful stimuli that alter learning and memory formation in Lymnaea . This is also the first demonstration that putative CBrs are present in Lymnaea and play a key role in learning and memory formation.
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altered gene activity correlated with long term memory formation of conditioned taste aversion in Lymnaea
Journal of Neuroscience Research, 2006Co-Authors: Sachiyo Azami, Hisayo Sadamoto, Yutaka Fujito, Dai Hatakeyama, Akiko Wagatsuma, Takeshi Usami, Manabu Fujie, Ryo Koyanagi, Kaoru Azumi, Ken LukowiakAbstract:The pond snail Lymnaea stagnalis is capable of learning conditioned taste aversion (CTA) and then consolidating that learning into long-term memory (LTM) that persists for at least 1 month. LTM requires de novo protein synthesis and altered gene activity. Changes in gene activity in Lymnaea that are correlated with, much less causative, memory formation have not yet been identified. As a first step toward rectifying this situation, we constructed a cDNA microarray with mRNAs extracted from the central nervous system (CNS) of Lymnaea. We then, using this microarray assay, identified genes whose activity either increased or decreased following CTA memory consolidation. We also identified genes whose expression levels were altered after inhibition of the cyclic AMP response element-binding protein (CREB) that is hypothesized to be a key transcription factor for CTA memory. We found that the molluscan insulin-related peptide II (MIP II) was up-regulated during CTA-LTM, whereas the gene encoding pedal peptide preprohormone (Pep) was down-regulated by CREB2 RNA interference. We next examined mRNAs of MIP II and Pep using real-time RT-PCR with SYBR Green. The MIP II mRNA level in the CNS of snails exhibiting "good" memory for CTA was confirmed to be significantly higher than that from the CNS of snails exhibiting "poor" memory. In contrast, there was no significant difference in expression levels of the Pep mRNA between "good" and "poor" performers. These data suggest that in Lymnaea MIP II may play a role in the consolidation process that forms LTM following CTA training.
Karoly Elekes - One of the best experts on this subject based on the ideXlab platform.
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effect of progesterone and its synthetic analogs on reproduction and embryonic development of a freshwater invertebrate model
Aquatic Toxicology, 2017Co-Authors: Zita Zrinyi, Gabor Maasz, Linwen Zhang, Akos Vertes, Sandor Lovas, Tibor Kiss, Karoly Elekes, Zsolt PirgerAbstract:The presence of a mixture of progestogens at ng/L concentration levels in surface waters is a worldwide problem. Only a few studies explore the effect of progestogen treatment in a mixture as opposed to individual chemicals to shed light on how non-target species respond to these contaminants. In the present study, we used an invertebrate model species, Lymnaea stagnalis, exposed to a mixture of four progestogens (progesterone, levonorgestrel, drospirenone, and gestodene) in 10ng/L concentration for 3 weeks. Data at both physiological and cellular/molecular level were analyzed using the ELISA technique, stereomicroscopy combined with time lapse software, and capillary microsampling combined with mass spectrometry. The treatment of adult Lymnaeas caused reduced egg production, and low quality egg mass on the first week, compared to the control. Starting from the second week, the egg production, and the quality of egg mass were similar in both groups. At the end of the third week, the egg production and the vitellogenin-like protein content of the hepatopancreas were significantly elevated in the treated group. At the cellular level, accelerated cell proliferation was observed during early embryogenesis in the treated group. The investigation of metabolomic changes resulted significantly elevated hexose utilization in the single-cell zygote cytoplasm, and elevated adenylate energy charge in the egg albumen. These changes suggested that treated snails provided more hexose in the eggs in order to improve offspring viability. Our study contributes to the knowledge of physiological effect of equi-concentration progestogen mixture at environmentally relevant dose on non-target aquatic species.
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Histaminergic neurons in the central and peripheral nervous system of gastropods (Helix, Lymnaea): An immunocytochemical, biochemical, and electrophysiological approach
The Journal of comparative neurology, 2004Co-Authors: Endre Roland Hegedus, Tibor Kiss, Jan Kaslin, László Hiripi, Pertti Panula, Karoly ElekesAbstract:Distribution, chemical-neuroanatomy, concentration, and uptake-release properties of histamine (HA)-containing neurons and the possible physiological effects of HA in the central and peripheral nervous system of the pulmonate snails, Helix pomatia and Lymnaea stagnalis, are described. In the CNS of both species, the distribution pattern of HA-immunoreactive (HA-IR) neurons was similar. In both species the majority were located in the buccal, cerebral, and pedal ganglia. In Helix, ∼400 HA-IR neurons were seen, whereas in Lymnaea ∼130 labeled cells were visualized. The neuropils, connectives, commissures, several peripheral nerves, and a part of the peripheral tissues (lip and foot of both species and the upper tentacles of Helix) were innervated by HA-IR elements. Numerous sensory cells were found in the tentacles, lip, and statocysts. The HA concentration values assayed by HPLC ranged from 4.8 to 47.4 pmol/mg in the different central ganglia of Helix, and from 4.3 to 18.6 pmol/mg in Lymnaea CNS, whereas the peripheral tissues contained 0.33–1 pmol/mg HA in Helix and 0.26–0.46 pmol/mg in Lymnaea. In the Lymnaea CNS, a high-affinity (37.6 μM), single component 3H-HA uptake system was demonstrated. 3H-HA release evoked by either electrical stimulation or 100 mM K+ could be prevented in Ca2+-free physiological solution. Voltage-clamp experiments indicated specific changes caused by HA in the membrane conductance of identified central neurons of Helix and Lymnaea. Exogenously applied 10-5 M HA resulted in the acceleration of locomotion (gliding by foot cilia) of Lymnaea. The findings suggest an important signaling role of HA, described here for the first time, in the nervous system of higher-order, pulmonate, gastropods, involving efferent, integrative, and sensory functions. The data can also be applied as a background for further specification of HA in the regulation of different behaviors in these species. J. Comp. Neurol. 475:391–405, 2004. © 2004 Wiley-Liss, Inc.
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Development of the nitric oxide/cGMP system in the embryonic and juvenile pond snail, Lymnaea stagnalis L. A comparative in situ hybridization, histochemical and immunohistochemical study
Journal of Neurocytology, 2002Co-Authors: Zoltán Serfőző, Zoltán Veréb, Tamás Rőszer, György Kemenes, Karoly ElekesAbstract:Recent studies have indicated that nitric oxide (NO)-induced cGMP synthesis is involved in different steps of neurogenesis in invertebrates. The development of putative NO synthetising elements was described earlier in the embryonic and juvenile pond snail, Lymnaea stagnalis , applying NADPH-diaphorase histochemistry (Serfőző et al ., 1998). In the present study, we examined the distribution of NO synthase (NOS) during Lymnaea development by in situ hybridization for Lymnaea -NOS mRNA, histochemical, and immunohistochemical techniques for the NOS and NO-stimulated cGMP. Peripheral fibers projecting to the CNS and terminating in the ganglionic neuropils showed NOS immunoreactivity from 85% of embryonic development. At the same time, a fine dot-like, immunostaining indicated the presence of cGMP in the neuropil area. In the CNS, Lymnaea -NOS mRNA positive, as well as NOS and cGMP immunoreactive perikarya were detected first during postembryonic development; their number significantly increased from P3 juvenile stage. Some of the cell groups in the CNS containing NOS immunoreactive material also displayed Lymnaea -NOS mRNA hybridization signal and were cGMP-positive. However, in the subesophageal ganglia, the distribution of Lymnaea -NOS mRNA positive cell groups did not correspond to that of the NOS immunoreactive cells. Neurons revealing transient NOS and cGMP immunoreactivity, respectively, could also be detected in this part of the CNS. In most of the ganglia the number of Lymnaea -NOS mRNA containing and cGMP immunopositive neurons, respectively, exceeded that of the NOS immunoreactive cells from P4 juvenile stage. The localization of NADPH-diaphorase reaction also correlated well with that of the NOS immunoreactivity in the developing CNS. At the periphery, colocalization of Lymnaea -NOS mRNA signal, NOS and cGMP immunoreactivities were observed in the epithelial cells of the esophagus and mantle after hatching. The findings suggest the functional maturity of the NO/cGMP signal transduction pathway at both central and peripheral levels during the development of the snail, Lymnaea stagnalis . The differences in the localization of Lymnaea -NOS mRNA labeling and NOS immunoreactivity in the CNS and PNS can be explained by the existence of different NOS isoforms, posttranslational regulation of NOS, and/or some non-specific antibody labeling.
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An octopaminergic system in the CNS of the snails, Lymnaea stagnalis and Helix pomatia.
Philosophical Transactions of the Royal Society of London. Series B: Biological Sciences, 1998Co-Authors: L. Hiripi, Ágnes Vehovszky, S. Juhos, Karoly ElekesAbstract:Octopamine (OA) levels in each ganglion of the terrestrial snail, Helix pomatia, and the pond snail, Lymnaea stagnalis, were measured by using the HPLC technique. In both species an inhomogeneous distribution of OA was found in the central nervous system. The buccal ganglia contained a concentration of OA (12.6 pmol mg-1 and 18.8 pmol mg-1) that was two to three times higher than the pedal (4.93 pmol mg-1 and 9.2 pmol mg-1) or cerebral (4.46 pmol mg-1 and 4.9 pmol mg-1) ganglia of Helix and Lymnaea, respectively, whereas no detectable amount of OA could be assayed in the visceroparietal complex. In Lymnaea ganglia, the OA uptake into the synaptosomal fraction had a high (Km1 = 4.07 ± 0.51 μM, Vmax1 = 0.56 ± 0.11 pmol mg-1 per 20 min), and a low (Km2 = 47.6 ± 5.2 μM, Vmax2 = 4.2 ± 0.27 pmol mg-1 per 20 min), affinity component. A specific and dissociable 3H-OA binding to the membrane pellet prepared from the CNS of both Helix and Lymnaea was demonstrated. The Scatchard analysis of the ligand binding data showed a one-binding site, representing a single receptor site. The Kd and Bmax values were found to be 33.7 ± 5.95 nM and 1678 ± 179 fmol g-1 tissue in Helix and 84.9 ± 17.4 nM and 3803 ± 515 fmol g-1 tissue in Lymnaea preparation. The pharmacological properties of the putative molluscan OA receptor were characterized in both species and it was demonstrated that the receptor resembled the insect OA2 rather than to the cloned Lymnaea OA receptor. Immunocytochemical labelling demonstrated the presence of OA-immunoreactive neurons and fibres in the buccal, cerebral and pedal ganglia in the central nervous system of both species investigated. Electrophysiological experiments also suggested that the Lymnaea brain possessed specific receptors for OA. Local application of OA onto the identified buccal B2 neuron evoked a hyperpolarization which could selectively be inhibited by the OAergic agents phentolamine, demethylchlordimeform and 2-chloro-4-methyl-2-(phenylimino)-imidazolidine. Among the dopamine antagonists, ergotamine reversibly inhibited the OA response, whereas sulpiride had no effect. Based on our findings, a neurotransmitter-modulator role of OA is suggested in the gastropod CNS.
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tachykinin and leucokinin related peptides in the molluscan nervous system
Acta Biologica Hungarica, 1995Co-Authors: Karoly Elekes, Tibor Kiss, L Hernadi, Y Muneoka, D R NasselAbstract:Distribution of locustatachykinin-like immunoreactive (LomTKLI) and leucokinin like immunoreactive (LKLI) neurons in the central nervous system (CNS) of the pond snail, Lymnaea stagnalis was investigated and compared to that found in Helix pomatia. Occurrence of LomTKLI neurons in different ganglia of the freshwater bivalve, Anodonta cygnea, was also studied. Similar to Helix, the Lymnaea CNS contained LomTKLI and LKLI neurons mainly in the cerebral and pedal ganglia, but the number of labelled neurons was found to be significantly lower in Lymnaea (150-184 LomTKLI and 86-104 LKLI neurons). LomTKLI elements in anodonta ganglia were mainly confined to the neuropil, whereas the immunostained perikarya were only randomly distributed. LomTKLI and LKLI neurons were also demonstrated in a similar pattern of distribution in the intestine of Lymnaea and Helix. Analyzing the membrane effects of locustatachykinin-I, leucokinin-I and anodontatachykinin, Helix neurons were found to be either depolarized or hyperpolarized. Voltage-clamp experiments revealed the role of Ca- or K-currents in peptide effects. Our results indicate that the different tachykinin- and leucokinin-systems are involved in different central and peripheral regulatory processes of the molluscan nervous system.
