Snail Lymnaea Research Articles

Possible roles of Wnt in the shell growth of the pond snail Lymnaea stagnalis

Although the mechanisms of molluscan shell growth have been studied using mathematical models, little is known about the molecular basis underpinning shell morphogenesis. Here, we performed Wnt activation experiments to elucidate the potential roles of Wnt signaling in the shell growth of Lymnaea stagnalis. In general, we observed following three types of shell malformations in both dose- and developmental stage-dependent manners: (i) cap-shaped shell, (ii) cap-shaped shell with hydropic soft tissues, and (iii) compressed shell with a smaller number of coiling. We analyzed the morphologies of these malformed shells using the growing tube model, revealing that the compressed malformations show significantly larger values for T (torsion), with no significant changes in the values for the remaining parameters E (expansion) and C (curvature). We also found that cap-shaped malformations have significantly larger values for E, suggesting that the effects of BIO on shell formation may change during growth. Since the changes in T and/or E parameter values can greatly alter the shell morphologies from a planispiral or a cap-shaped one to various three-dimensional helices, changes in shell developmental processes possibly controlled by Wnt signaling may account for at least a part of the evolution of diverse shell forms in molluscs.

Quercetin, the new stress buster: Investigating the transcriptional and behavioral effects of this flavonoid on multiple stressors using Lymnaea stagnalis

Growing evidence suggests that a flavonoid-rich diet can prevent or reverse the effects of stressors, although the underlying mechanisms remain poorly understood. One common and abundant flavonoid found in numerous foods is quercetin. This study utilizes the pond snail Lymnaea stagnalis, a valid model organism for learning and memory, and a simple but robust learning paradigm—operant conditioning of aerial respiration—to explore the behavioral and transcriptional effects of different stressors on snails' cognitive functions and to investigate whether quercetin exposure can prevent stress effects on learning and memory formation. Our findings demonstrate that three different stressors—severe food deprivation, lipopolysaccharide injection (an inflammatory challenge), and fluoride exposure (a neurotoxic agent)—block memory formation for operant conditioning and affect the expression levels of key targets related to stress response, energy balance, and immune response in the snails' central ring ganglia. Remarkably, exposing snails to quercetin for 1 h before stress presentation prevents these effects at both the behavioral and transcriptional levels, demonstrating the potent stress-preventive properties of quercetin. Despite the evolutionary distance from humans, L. stagnalis has proven to be a valuable model for studying conserved mechanisms by which bioactive compounds like quercetin mitigate the adverse effects of various stressors on cognitive functions across species. Moreover, these findings offer insights into quercetin's potential for mitigating stress-induced physiological and cognitive impairments.

Hot and cold exposure triggers distinct transcriptional and behavioral responses in laboratory-inbred pond snails

Animals exhibit remarkable behavioral and molecular adaptations to cope with thermal stressors, which are crucial for survival in variable environments that are exacerbated by climate change. Aquatic poikilotherms like our model organism—the pond snail Lymnaea stagnalis—face significant challenges due to their dependence on external temperatures. Our study provides valuable insights into the different behavioral and molecular responses of lab-inbred snails to cold and heat shock stressors (i.e., 4 °C and 30 °C), particularly in the context of learning and memory formation. We found that while short-term (1 h) cold exposure transiently upregulated the expression levels of HSP70 and HSP40 in the snail’s central ring ganglia, prolonged cold exposure (24 h) resulted in a significant downregulation of LymMIPII and an upregulation of LymMIPR. These data suggest, albeit at the transcriptional level, the existence of a negative feedback loop necessary for sustaining cellular functions when metabolic demands might shift towards conserving energy during prolonged cold exposure. At the behavioral level, we found that, compared to heat shock, cold exposure did not result in a Garcia effect (i.e., a “special form” of conditioned taste aversion). The difference in memory outcomes was associated with changes in the expression levels of selected targets involved in neuronal plasticity and the stress response. While both cold and heat shock upregulated the HSP levels in the snail's central ring ganglia, cold exposure did not affect the expression levels of the neuroplasticity genes LymGRIN1 and LymCREB1, contrasting with heat shock’s neurogenic effects. Overall, this study provides insights into L. stagnalis's adaptive responses to thermal stressors, emphasizing different molecular strategies for coping with heat versus cold challenges in aquatic environments. These findings contribute to our understanding of thermal biology and stress physiology in aquatic organisms, underscoring the importance of molecular mechanisms in shaping species' resilience in dynamic environments.

Characterization of two carbonic anhydrase isoforms in the pulmonate snail (Lymnaea Stagnalis) and their involvement in Molluskan calcification

Calcifying organisms are suffering from negative impacts induced by climate change, such as CO2-induced acidification, which may impair external calcified structures. Freshwater mollusks have the potential to suffer more from CO2-induced acidification than marine calcifiers due to the lower buffering capacity of many freshwater systems. One of the most important enzymes contributing to the biomineralization reaction is carbonic anhydrase (CA), which catalyzes the reversible conversion of CO2 to bicarbonate, the major carbon source of the calcareous structure in calcifiers. In this study we characterized two α-CA isoforms (LsCA1 and LsCA4) from the freshwater snail Lymnaea stagnalis using a combination of gene sequencing, gene expression, phylogenetic analysis and biochemical assays. Both CA isoforms demonstrated high expression levels in the mantle tissue, the major site for biomineralization. Furthermore, expression of LsCA4 during development parallels shell formation. The primary protein structure analysis, active site configuration and the catalytic activity of LsCA4 together suggest that the LsCA4 is embedded in the apical and basolateral membranes of mantle cells; while LsCA1 is proposed to be cytosolic and might play an important role in acid-base regulation. These findings of LsCA isoforms form a strong basis for a more detailed physiological understanding of the effects of elevated CO2 on calcification in freshwater mollusks.

Population genetic structure in a self-compatible hermaphroditic snail is driven by drift independently of its contemporary mating system.

Genetic drift, gene flow, and natural selection commonly influence population genetic diversity. In populations of self-compatible hermaphrodites, the mating system (e.g., self-fertilization) further reduces individual heterozygosity. Furthermore, selfing, as a form of inbreeding, significantly impacts genetic drift by reducing effective population size (N e). This can potentially accelerate genetic drift, particularly in small populations where self-fertilization is likely during founder events. To investigate the roles of genetic drift and contemporary mating system in populations of the freshwater snail Lymnaea stagnalis, we examined their effective population sizes (N e) and Tajima's D values, which reflect genetic drift over extended time periods, as well as estimates of within-population selfing rates and pairwise relatedness reflecting contemporary mating system. We used 4054 SNP markers obtained using restriction site associated DNA (RAD) sequencing from individuals in five snail populations originating from geographically closely located ponds. We found strong population genetic structure and differences in genetic diversity among populations. Covariation between genetic diversity and N e estimates and Tajima's D values suggested drift being an important determinant of genetic diversity and structure in these populations. However, this effect was independent of the contemporary mating system, as indicated by the similarity of selfing rates and relatedness estimates among populations. Thus, founder events (possibly including historical inbreeding) and/or drift due to small sizes of L. stagnalis populations are likely to explain their genetic structure and limit within-population genetic diversity.

How to reduce fear in a snail: Take an aspirin, call me in the morning

Aspirin (Acetylsalicylic acid, ASA), one of the widely used non-steroid anti-inflammatory drugs can easily end up in sewage effluents and thus it becomes necessary to investigate the effects of aspirin on behaviour of aquatic organisms. Previous studies in mammals have shown ASA to alter fear and anxiety-like behaviours. In the great pond snail Lymnaea stagnalis, ASA has been shown to block a ‘sickness state’ induced by lipopolysaccharide injection which upregulates immune and stress-related genes thus altering behavioural responses. In Lymnaea, eliciting physiological stress may enhance memory formation or block its retrieval depending on the stimulus type and intensity. Here we examine whether ASA will alter two forms of associative-learning memory in crayfish predator-experienced Lymnaea when ASA exposure accompanies predator-cue-induced stress during the learning procedure. The two trainings procedures are: 1) operant conditioning of aerial respiration; and 2) a higher form of learning, called configural learning, which here is dependent on evoking a fear response. We show here that ASA alone does not alter homeostatic aerial respiration, feeding behaviour or long-term memory (LTM) formation of operantly conditioned aerial respiration. However, ASA blocked the enhancement of LTM formation normally elicited by training snails in predator cue. ASA also blocked configural learning, which makes use of the fear response elicited by the predator cue. Thus, ASA alters how Lymnaea responds cognitively to predator detection.

Behavioral and transcriptional effects of carnosine in the central ring ganglia of the pond snail Lymnaea stagnalis.

Carnosine is a naturally occurring endogenous dipeptide with well-recognized anti-inflammatory, antioxidant, and neuroprotective effects at the central nervous system level. To date, very few studies have been focused on the ability of carnosine to rescue and/or enhance memory. Here, we used a well-known invertebrate model system, the pond snail Lymnaea stagnalis, and a well-studied associative learning procedure, operant conditioning of aerial respiration, to investigate the ability of carnosine to enhance long-term memory (LTM) formation and reverse memory obstruction caused by an immune challenge (i.e., lipopolysaccharide [LPS] injection). Exposing snails to 1 mM carnosine for 1 h before training in addition to enhancing memory formation resulted in a significant upregulation of the expression levels of key neuroplasticity genes (i.e., glutamate ionotropic receptor N-methyl-d-aspartate [NMDA]-type subunit 1-LymGRIN1, and the transcription factor cAMP-response element-binding protein 1-LymCREB1) in snails' central ring ganglia. Moreover, pre-exposure to 1 mM carnosine before an LPS injection reversed the memory deficit brought about by inflammation, by preventing the upregulation of key targets for immune and stress response (i.e., Toll-like receptor 4-LymTLR4, molluscan defense molecule-LymMDM, heat shock protein 70-LymHSP70). Our data are thus consistent with the hypothesis that carnosine can have positive benefits on cognitive ability and be able to reverse memory aversive states induced by neuroinflammation.

Different stressors uniquely affect the expression of endocannabinoid-metabolizing enzymes in the central ring ganglia of Lymnaea stagnalis.

The endocannabinoid system (ECS) plays an important role in neuroprotection, neuroplasticity, energy balance, modulation of stress, and inflammatory responses, acting as a critical link between the brain and the body's peripheral regions, while also offering promising potential for novel therapeutic strategies. Unfortunately, in humans, pharmacological inhibitors of different ECS enzymes have led to mixed results in both preclinical and clinical studies. As the ECS has been highly conserved throughout the eukaryotic lineage, the use of invertebrate model organisms like the pond snail Lymnaea stagnalis may provide a flexible tool to unravel unexplored functions of the ECS at the cellular, synaptic, and behavioral levels. In this study, starting from the available genome and transcriptome of L. stagnalis, we first identified putative transcripts of all ECS enzymes containing an open reading frame. Each predicted protein possessed a high degree of sequence conservation to known orthologues of other invertebrate and vertebrate organisms. Sequences were confirmed by qualitative PCR and sequencing. Then, we investigated the transcriptional effects induced by different stress conditions (i.e., bacterial LPS injection, predator scent, food deprivation, and acute heat shock) on the expression levels of the enzymes of the ECS in Lymnaea's central ring ganglia. Our results suggest that in Lymnaea as in rodents, the ECS is involved in mediating inflammatory and anxiety-like responses, promoting energy balance, and responding to acute stressors. To our knowledge, this study offers the most comprehensive analysis so far of the ECS in an invertebrate model organism.

Potentilla fulgens: organic column purified extract and its effect on enzyme inhibition in the vector snail Lymnaea acuminata

Fasciola hepatica and F. gigantica are parasitic trematode that causes fascioliasis in cattle and humans. The vector snail Lymnaea acuminata is an intermediate host of the Fasciola species. The major preventive step against fascioliasis is the control of the host snail L. acuminata. Effect of sub-lethal concentration of (20% and 60% of 96h LC50) column purified extract of Potentilla fulgens on acid phosphatase (ACPase), alkaline phosphatase (ALPase), and acetylcholinesterase (AChE) inhibition in nervous tissue of host L. acuminata were studied. Alteration in the enzymes of ACPase, ALPase, and AChE in nervous tissue was time and concentration-dependent. Withdrawal experiment demonstrates that there was a significant recovery in ACPase, ALPase, and AChE inhibition in nervous tissues of L. acuminata were withdrawn from 7 days and treated with 20 and 60% of 96h LC50 for 96h. A significant recovery in the levels of ACPase, ALPase, and AChE was observed in the nervous tissue of L. acuminata. . KEYWORDS :Lymnaea acuminata; Potentilla fulgens; ACPase; ALPase; AChE; Fasciola

Histopathological Evaluation of the Curative Effects of EDTA on Lymnaea stagnalis Exposed to Subacute Cadmium.

We investigated the therapeutic effects of EDTA application for 14 and 28 days on cadmium (Cd) induced pond snail Lymnaea stagnalis (Linnaeus, 1758). The sublethal concentration of cadmium (63.4mg/l Cd) caused tissue damages to the snail after an exposure for 28 days.In the groups treated with EDTA, the concentration of Cd in the foot, mantle and hepatopancreas tissues showed significantly decreased during the recovery period. The curative effects of EDTA on Cd-induced damage were assessed using a scoring system. Cadmium exposure led to histopathological changes including increased mucositis, pigment and protein cells, foot epithelium desquamation, muscle fibril damage, connective tissue cell atrophy, and increased lipid vacuoles in the mantle and hepatopancreas. However, these changes were less severe in snails treated with EDTA (2.00 mL/L for 28day), indicating that EDTA reduces their susceptibility to heavy metal toxicity.

Nutrient enrichment, propagule pressure, and herbivory interactively influence the competitive ability of an invasive alien macrophyte Myriophyllum aquaticum.

Freshwater ecosystems are susceptible to invasion by alien macrophytes due to their connectivity and various plant dispersal vectors. These ecosystems often experience anthropogenic nutrient enrichment, favouring invasive species that efficiently exploit these resources. Propagule pressure (reflecting the quantity of introduced individuals) and habitat invasibility are key determinants of invasion success. Moreover, the enemy release hypothesis predicts that escape from natural enemies, such as herbivores, allows alien species to invest more resources to growth and reproduction rather than defense, enhancing their invasive potential. Yet, the combined impact of propagule pressure, herbivory, and nutrient enrichment on the competitive dynamics between invasive alien macrophytes and native macrophyte communities is not well understood due to a paucity of studies. We conducted a full factorial mesocosm experiment to explore the individual and combined effects of herbivory, nutrient levels, propagule pressure, and competition on the invasion success of the alien macrophyte Myriophyllum aquaticum into a native macrophyte community comprising Vallisneria natans, Hydrilla verticillata, and Myriophyllum spicatum. This setup included varying M. aquaticum densities (low vs. high, simulating low and high propagule pressures), two levels of herbivory by the native snail Lymnaea stagnalis (herbivory vs no-herbivory), and two nutrient conditions (low vs. high). Myriophyllum aquaticum was also grown separately at both densities without competition from native macrophytes. The invasive alien macrophyte M. aquaticum produced the highest shoot and total biomass when simultaneously subjected to conditions of high-density intraspecific competition, no herbivory, and low-nutrient availability treatments. Moreover, a high propagule pressure of M. aquaticum significantly reduced the growth of the native macrophyte community in nutrient-rich conditions, but this effect was not observed in nutrient-poor conditions. These findings indicate that M. aquaticum has adaptive traits enabling it to flourish in the absence of herbivory (supporting the enemy release hypothesis) and in challenging environments such as intense intraspecific competition and low nutrient availability. Additionally, the findings demonstrate that when present in large numbers, M. aquaticum can significantly inhibit the growth of native macrophyte communities, particularly in nutrient-rich environments. Consequently, reducing the propagule pressure of M. aquaticum could help control its spread and mitigate its ecological impact. Overall, these findings emphasize that the growth and impacts of invasive alien plants can vary across different habitat conditions and is shaped by the interplay of biotic and abiotic factors.

Sleep-like State in Pond Snails Leads to Enhanced Memory Formation.

To test the hypothesis that a sleep-like quiescent state enhances memory consolidation in the pond snail Lymnaea stagnalis, we interposed a period in which snails experienced either a quiescent, sleeping state or an active, non-sleeping state following escape behavior suppression learning (EBSL). During EBSL training, the number of escapes made by a snail from a container was significantly suppressed using an external aversive stimulus (punishment). After training, the snails were divided into two groups. One group of snails was allowed to move freely and to experience a sleep-like quiescent state for 3 h in distilled water. The other group was stimulated with a sucrose solution every 10 min to keep them active (i.e., non-sleeping). In the memory test, escape behavior was suppressed in the group that experienced the quiescent state, whereas the suppression was not observed in snails that were kept active. Additionally, the latency of the first escape in the memory test was shorter in the snails kept active than in those that experienced the quiescent state. Together, these data are consistent with the hypothesis that a sleep-like quiescent state enhances EBSL memory consolidation in L. stagnalis.

A translational and multidisciplinary approach to studying the Garcia effect, a higher form of learning with deep evolutionary roots.

Animals, including humans, learn and remember to avoid a novel food when its ingestion is followed, hours later, by sickness - a phenomenon initially identified during World War II as a potential means of pest control. In the 1960s, John Garcia (for whom the effect is now named) demonstrated that this form of conditioned taste aversion had broader implications, showing that it is a rapid but long-lasting taste-specific food aversion with a fundamental role in the evolution of behaviour. From the mid-1970s onward, the principles of the Garcia effect were translated to humans, showing its role in different clinical conditions (e.g. side-effects linked to chemotherapy). However, in the last two decades, the number of studies on the Garcia effect has undergone a considerable decline. Since its discovery in rodents, this form of learning was thought to be exclusive to mammals; however, we recently provided the first demonstration that a Garcia effect can be formed in an invertebrate model organism, the pond snail Lymnaea stagnalis. Thus, in this Commentary, after reviewing the experiments that led to the first characterization of the Garcia effect in rodents, we describe the recent evidence for the Garcia effect in L. stagnalis, which may pave the way for future studies in other invertebrates and mammals. This article aims to inspire future translational and ecological studies that characterize the conserved mechanisms underlying this form of learning with deep evolutionary roots, which can be used to address a range of different biological questions.

Lipid Profile of the Great Pond Snail Lymnaea stagnalis (Mollusca: Gastropoda) in Lakes with Different Degrees of Anthropogenic Pollution

Lipid Profile of the Great Pond Snail Lymnaea stagnalis (Mollusca: Gastropoda) in Lakes with Different Degrees of Anthropogenic Pollution

In Vivo: Anthelmintic Activity of Phyllanthus Emblica Against Larvae of Fasciola Gigantica

The research article illustrates the in vivo anthelmintic larvicidal activities of the Phyllanthus emblica against sporocyst, redia, and cercaria larvae of Fasciola gigantica. Fascioliases are distributed among cattle and human populations worldwide. The spreading of Fasciola can be controlled by the break of the life cycle in the host snail. The fresh-water snail Lymnaea acuminata is an intermediate host of the F. gigantica. The exposure of bark powder and, various preparations, of P. emblica, was observed in vivo for up to 8h at various concentrations against F. gigantica larvae. The toxic effect of various preparations of P. emblica was observed at 2, 4, 6, and 8h of the exposure period. The in vivo exposure of bark powder of P. emblica was more effective against redia larvae after 2h, 4h, 6h, and 8h the LC50 values were 74.63, 65.42, 72.16 and 71.03 mg/l, respectively. The column extract of bark powder of P. emblica against sporocyst, redia, and cercaria in 2h exposure the LC50 value was noted 65.36, 70.32, and 69.36 mg/l and 8h exposure the LC50 value was 56.23, 66.10, and 67.13 mg/l, respectively. The various formulations of P. emblica bark powder can be used for the control of fascioliasis.

Dominant gingers - Discovery and inheritance of a new shell polymorphism in the great pond snail Lymnaea stagnalis.

Color polymorphism is a classic study system for evolutionary genetics. One of the most color-polymorphic animal taxa is mollusks, but the investigation of the genetic basis of color determination is often hindered by their life history and the limited availability of genetic resources. Here, we report on the discovery of shell color polymorphism in a much-used model species, the great pond snail Lymnaea stagnalis. While their shell is usually beige, some individuals from a Greek population show a distinct red shell color, which we nicknamed Ginger. Moreover, we found that the inheritance fits simple, single-locus Mendelian inheritance with dominance of the Ginger allele. We also compared crucial life-history traits between Ginger and wild-type individuals, and found no differences between morphs. We conclude that the relative simplicity of this polymorphism will provide new opportunities for a deeper understanding of the genetic basis of shell color polymorphism and its evolutionary origin.

Cadmium impact on the growth and survival rate of great pond snail (Lymnaea stagnalis) in the chronic experiment

Cadmium impact on the growth and survival rate of great pond snail (Lymnaea stagnalis) in the chronic experiment

The Morphometric Analysis of the Effects of Hydrogen Peroxide on the Off Springs of Helix Lymnaea Model

Background: Hydrogen peroxide free radical is a potent mutagen and teratogen that can obviously affects the color of the offspring's of the land snail Helix Lymnaea. Aim: To assess the morphometric changes of the offspring's of the land snail Helix Lymnaea after parents exposure for H2O2 for three years. Materials and Methods: A twenty four Helix Lymnaea snails were randomly divided into two groups (12 in each).Group A is the control whereas group B is the H2O2 test group. All were adults (4-6 weeks age) of the same size 5-7 mm length, all were hermaphrodites. The two groups were followed for three years for ten generations for different morphometric parameters Results: There was a significant reduction in the movement score in the group treated by H2O2 in comparison with the control group 25% vs 8.3 % respectively p = 3.8, chi square = 0.8. The eating activity of the offspring's had a minute change noticed in between the two groups 95% in H2O2 exposed vs 92% in the control group. The morphometric outcomes mainly the albino offsprings were none in the control group that showed no teratogenicity in comparison with 60% in H2O2 group, p < 0.05. The rate of successful eggs hatching was significantly differed between the control and the H2O2 groups, p = 0.017. There was also an obvious difference in the mortality rate between the control and H2O2 group, however this difference was not significant p = 0.2. Conclusion: There was a significant teratogenic effect of H2O2 on the offsprings of the snail Helix Lymnaea. This effects may extend to include different free radical producing substances.

Proactive and retroactive interference with associative memory consolidation in the snail Lymnaea is time and circuit dependent

Interference-based forgetting occurs when new information acquired either before or after a learning event attenuates memory expression (proactive and retroactive interference, respectively). Multiple learning events often occur in rapid succession, leading to competition between consolidating memories. However, it is unknown what factors determine which memory is remembered or forgotten. Here, we challenge the snail, Lymnaea, to acquire two consecutive similar or different memories and identify learning-induced changes in neurons of its well-characterized motor circuits. We show that when new learning takes place during a stable period of the original memory, proactive interference only occurs if the two consolidating memories engage the same circuit mechanisms. If different circuits are used, both memories survive. However, any new learning during a labile period of consolidation promotes retroactive interference and the acquisition of the new memory. Therefore, the effect of interference depends both on the timing of new learning and the underlying neuronal mechanisms.

Bellamya chinensis and Lymnaea stagnalis mortality in the presence of Macrobdella decora

Food webs are complex and each trophic level guides the dynamic stability of the whole food web. As humans cause the loss of piscivores and zooplanktivores from lake food webs, parasitic leeches, such as Macrobdella decora, lose potential host organisms. In the absence of a host, it has been suggested M. decora will parasitize freshwater snails, but there is a lack of research supporting this. At the University of Notre Dame Environmental Research Center – East, the snail species Bellamya chinensis and Lymnaea stagnalis have similar niches and are prevalent in the lakes, so they were used as research species. Trials were run to observe the mortality of the two snail species in the presence of leeches. The snail species were tested while in the same tank and different tanks. Leeches did parasitize B. chinensis. However, L. stagnalis did have a significantly higher average mortality than B. chinensis in the same tank (treatment: p-value: 0.022; control: p-value: 0.044) and in different tanks (p-value: 0.001), but leech presence did not significantly (p-value: 0.25) change snail mortality. An exact reason for snail species being the factor that controlled snail mortality could not be determined. However, a possible explanation was due to unfiltered fecal material and copper contamination in the trial tanks. Copper is a lethal metal to most gastropods.