The crystal structure of Ac-AChBP in complex with α-conotoxin LvIA reveals the mechanism of its selectivity towards different nAChR subtypes

The α3* nAChRs, which are considered to be promising drug targets for problems such as pain, addiction, cardiovascular function, cognitive disorders etc., are found throughout the central and peripheral nervous system. The α-conotoxin (α-CTx) LvIA has been identified as the most selective inhibitor of α3β2 nAChRs known to date, and it can distinguish the α3β2 nAChR subtype from the α6/α3β2β3 and α3β4 nAChR subtypes. However, the mechanism of its selectivity towards α3β2, α6/α3β2β3, and α3β4 nAChRs remains elusive. Here we report the co-crystal structure of LvIA in complex with Aplysia californica acetylcholine binding protein (Ac-AChBP) at a resolution of 3.4 Å. Based on the structure of this complex, together with homology modeling based on other nAChR subtypes and binding affinity assays, we conclude that Asp-11 of LvIA plays an important role in the selectivity of LvIA towards α3β2 and α3/α6β2β3 nAChRs by making a salt bridge with Lys-155 of the rat α3 subunit. Asn-9 lies within a hydrophobic pocket that is formed by Met-36, Thr-59, and Phe-119 of the rat β2 subunit in the α3β2 nAChR model, revealing the reason for its more potent selectivity towards the α3β2 nAChR subtype. These results provide molecular insights that can be used to design ligands that selectively target α3β2 nAChRs, with significant implications for the design of new therapeutic α-CTxs.

Acute urticaria with angioedema following sea hare ingestion

Seafood is one of the common causes of food allergies to adults. The sea hare Aplysia kurodai is a marine mollusk which belongs to invertebrate gastropod that has been consumed as a food in Korea. Cases of acute toxic hepatitis after ingestion of sea hares have been reported, but few cases of allergic reactions to sea hare have been reported in the literature. A 33-year-old man was referred to our Emergency Department due to urticaria and periorbital/perioral swelling after eating sea hares. Approximately 10 years ago, he experienced similar allergic reactions to it. Skin prick and intradermal tests showed strong positive responses to crude sea hare allergen extract. He was diagnosed with food allergy to sea hares. We herein report the first case of sea hare allergy after ingestion.

Specific Expression of Aplysia Phosphodiesterase 4 in Bag Cells Revealed by in situ Hybridization Analysis

Phosphodiesterases (PDEs) play a key role in the regulation of cyclic adenosine monophosphate (cAMP), which in turn mediates various cellular functions including learning and memory. We previously cloned and characterized three PDE4 isoforms (ApPDE4) from Aplysia kurodai. Using reverse transcription polymerase chain reaction (RT-PCR), we found that ApPDE4 isoforms are primarily expressed in the central nervous system. However, the detailed distribution of ApPDE4 mRNA in Aplysia individual ganglions was not evident. In this study, to determine the distribution of ApPDE4 mRNAs in Aplysia ganglions, we performed in situ hybridization (ISH) using a probe targeting ApPDE4, including the PDE catalytic domain. Interestingly, we found the strongest ISH-positive signals in the symmetrical bag cell clusters of the abdominal ganglion. The R2, R14, L7, L2 and L11 neurons in the abdominal ganglion, LP1 neuron in pleural ganglion, and metacerebral (MCC) neurons were ISH-positive. Mechanosensory neurons of the sensory cluster were also stained on the ventral aspect of the right and left pleural ganglia. Taken together, we found the detailed distribution of ApPDE4 mRNA in Aplysia ganglion and support their roles in serotonin (5-HT)-induced synaptic facilitation of Aplysia mechanosensory neurons.

Four Cases of Toxic Hepatitis after Ingestion of Sea Hare / 대한내과학회지

The sea hare is a marine mollusk in the family Aplysiidae that has long been consumed as food. Rarely, toxic hepatitis can occur after eating sea hare. We herein discuss four cases of toxic hepatitis due to sea hare ingestion and review the relevant literature.

Serotonin and Synaptic Transmission at Invertebrate Neuromuscular Junctions

The serotonergic system in vertebrates and invertebrates has been a focus for over 50 years and will likely continue in the future. Recently, genomic analysis and discovery of alternative splicing and differential expression in tissues have increased the knowledge of serotonin (5-HT) receptor types. Comparative studies can provide useful insights to the wide variety of mechanistic actions of 5-HT responsible for behaviors regulated or modified by 5-HT. To determine cellular responses and influences on neural systems as well as the efferent control of behaviors by the motor units, preparations amenable to detailed studies of synapses are beneficial as working models. The invertebrate neuromuscular junctions (NMJs) offer some unique advantages for such investigations; action of 5-HT at crustacean NMJs has been widely studied, and leech and Aplysia continue to be key organisms. However, there are few studies in insects likely due to the focus in modulation within the CNS and lack of evidence of substantial action of 5-HT at the Drosophila NMJs. There are only a few reports in gastropods and annelids as well as other invertebrates. In this review we highlight some of the key findings of 5-HT actions and receptor types associated at NMJs in a variety of invertebrate preparations in hopes that future studies will build on this knowledge base.

A Computational Model of the Temperature-dependent Changes in Firing Patterns in Aplysia Neurons

We performed experiments using Aplysia neurons to identify the mechanism underlying the changes in the firing patterns in response to temperature changes. When the temperature was gradually increased from 11degrees C to 31degrees C the firing patterns changed sequentially from the silent state to beating, doublets, beating-chaos, bursting-chaos, square-wave bursting, and bursting-oscillation patterns. When the temperature was decreased over the same temperature range, these sequential changes in the firing patterns reappeared in reverse order. To simulate this entire range of spiking patterns we modified nonlinear differential equations that Chay and Lee made using temperature-dependent scaling factors. To refine the equations, we also analyzed the spike pattern changes in the presence of potassium channel blockers. Based on the solutions of these equations and potassium channel blocker experiments, we found that, as temperature increases, the maximum value of the potassium channel relaxation time constant, taun(t) increases, but the maximum value of the probabilities of openings for activation of the potassium channels, n(t) decreases. Accordingly, the voltage-dependent potassium current is likely to play a leading role in the temperature-dependent changes in the firing patterns in Aplysia neurons.

Neuronal RNA granule contains ApCPEB1, a novel cytoplasmic polyadenylation element binding protein, in Aplysia sensory neuron

The cytoplasmic polyadenylation element (CPE)-binding protein (CPEB) binds to CPE containing mRNAs on their 3' untranslated regions (3'UTRs). This RNA binding protein comes out many important tasks, especially in learning and memory, by modifying the translational efficiency of target mRNAs via poly (A) tailing. Overexpressed CPEB has been reported to induce the formation of stress granules (SGs), a sort of RNA granule in mammalian cell lines. RNA granule is considered to be a potentially important factor in learning and memory. However, there is no study about RNA granule in Aplysia. To examine whether an Aplysia CPEB, ApCPEB1, forms RNA granules, we overexpressed ApCPEB1-EGFP in Aplysia sensory neurons. Consistent with the localization of mammalian CPEB, overexpressed ApCPEB1 formed granular structures, and was colocalized with RNAs and another RNA binding protein, ApCPEB, showing that ApCPEB1 positive granules are RNA-protein complexes. In addition, ApCPEB1 has a high turnover rate in RNA granules which were mobile structures. Thus, our results indicate that overexpressed ApCPEB1 is incorporated into RNA granule which is a dynamic structure in Aplysia sensory neuron. We propose that ApCPEB1 granule might modulate translation, as other RNA granules do, and furthermore, influence memory.

The Effect of Heat on the Spiking Patterns of the Cells in Aplysia / 의학물리

Fruitful findings have been produced from five out of sixty cells which were obtained from each 63 individual Aplisia caught at the Jeju coast. Spiking patterns of three out of five cells, such as relaxation oscillator, bursting within a short time of the inter-burst interval, chaotic bursting, period doubling sequences, bursting with long trains of action potentials separated by short silent periods, regular repeated beating or elliptic bursting, and silent states had been changed in order as the temperature was lowered to 10 degrees C from 32 degrees C. In the intervals of every about 40 minutes repeated ups and downs of temperature produced similar firing patterns at the allowable temperature ranges. The other two cells showed difference from these. The amplitudes of the action potentials of the two cells will not be highly decreased in 24 hours. Average spike frequencies, the inter-burst interval, peak to peak spike amplitude of action potentials, minimum potential values are compared and analyzed by using the computer programme. The spike frequencies according to temperature show the distribution of bell type, with maximal spike frequencies at intermediate temperatures and minimal ones at either end. The most common pattern consist of high spike frequency during falling and low one during rising temperatures.

Evaluation of the cholinomimetic actions of trimethylsulfonium, a compound present in the midgut gland of the sea hare Aplysia brasiliana (Gastropoda, Opisthobranchia)

Trimethylsulfonium, a compound present in the midgut gland of the sea hare Aplysia brasiliana, negatively modulates vagal response, indicating a probable ability to inhibit cholinergic responses. In the present study, the pharmacological profile of trimethylsulfonium was characterized on muscarinic and nicotinic acetylcholine receptors. In rat jejunum the contractile response induced by trimethylsulfonium (pD2 = 2.46 + or - 0.12 and maximal response = 2.14 + or - 0.32 g) was not antagonized competitively by atropine. The maximal response (Emax) to trimethylsulfonium was diminished in the presence of increasing doses of atropine (P<0.05), suggesting that trimethylsulfonium-induced contraction was not related to muscarinic stimulation, but might be caused by acetylcholine release due to presynaptic stimulation. Trimethylsulfonium displaced [³H]-quinuclidinyl benzilate from rat cortex membranes with a low affinity (Ki = 0.5 mM). Furthermore, it caused contraction of frog rectus abdominis muscles (pD2 = 2.70 + or - 0.06 and Emax = 4.16 + or - 0.9 g), which was competitively antagonized by d-tubocurarine (1, 3 or 10 æM) with a pA2 of 5.79, suggesting a positive interaction with nicotinic receptors. In fact, trimethylsulfonium displaced [³H]-nicotine from rat diaphragm muscle membranes with a Ki of 27.1 æM. These results suggest that trimethylsulfonium acts as an agonist on nicotinic receptors, and thus contracts frog skeletal rectus abdominis muscle and rat jejunum smooth muscle via stimulation of postjunctional and neuronal prejunctional nicotinic cholinoreceptors, respectively

Screening of antimitotic portions from sea hare using conidia of Pyricularia Oryzae / 中国中药杂志

<p><b>OBJECTIVE</b>To apply conidia of Pyricularia Oryzae to the screening of antimitotic constituents from marine animal sea hare.</p><p><b>METHOD</b>To extract and fractionate active portions from sea hare through detecting deformation of mycelia germinated from conidia of P. Oryzae P-2b, in comparison with the cytotoxic test results in vitro.</p><p><b>RESULT</b>Two active portions, of which IC50 against P388 and HL-60 was 23.4, 18.6 and 19.4, 12.5 micrograms.ml-1, respectively, were screened from this animal.</p><p><b>CONCLUSION</b>This bioassay method was efficiently applied to the primary screening of antimitotic portions from marine animals for the first time. Being convenient, speedy and cheap, the screening model is suitable for the bioassay of active constituents from marine life.</p>

Toxic, antimicrobial and hemagglutinating activities of the purple fluid of the sea hare Aplysia dactylomela Rang, 1828

The antimicrobial, hemagglutinating and toxic activities of the purple fluid of the sea hare Aplysia dactylomela are described. Intact or dialyzed purple fluid inhibited the growth of species of Gram-positive and Gram-negative bacteria and the action was not bactericidal but bacteriostatic. The active factor or factors were heat labile and sensitive to extreme pH values. The fluid preferentially agglutinated rabbit erythrocytes and, to a lesser extent, human blood cells, and this activity was inhibited by the glycoprotein fetuin, a fact suggesting the presence of a lectin. The fluid was also toxic to brine shrimp nauplii (LD 50 141.25 µg protein/ml) and to mice injected intraperitoneally (LD 50 201.8 ñ 8.6 mg protein/kg), in a dose-dependent fashion. These toxic activities were abolished when the fluid was heated. Taken together, the data suggest that the activities of the purple fluid are due primarily to substance(s) of a protein nature which may be involved in the chemical defense mechanism of this sea hare

Progressos no estudo das bases neurais da aprendizagem / Advances in the study of the neural basis of learning

O estudo dos mecanismos neurais da aprendizagem revelou considerável progresso nos últimos vinte anos. A possibilidade de experimentaçäo com invertebrados tem levado a descobertas importantes sobre as mudanças funcionais e estruturais, a nível celular, que acompanham processos simples de aprendizagem como a habituaçäo e a sensitizaçäo