Target innervation is necessary for neuronal polyploidization in the terrestrial slug Limax

The brain of gastropod mollusks contains many giant neurons with polyploid genomic DNAs. Such DNAs are generated through repeated DNA endoreplication during body growth. However, it is not known what triggers DNA endoreplication in neurons. There are two possibilities: (1) DNAs are replicated in response to some unknown molecules in the hemolymph that reflect the nutritive status of the animal; or (2) DNAs are replicated in response to some unknown factors that are retrogradely transported through axons from the innervated target organs. We first tested whether hemolymph with rich nutrition could induce DNA endoreplication. We tested whether the transplanted brain exhibits enhanced DNA endoreplication like an endogenous brain does when transplanted into the homocoel of the body of a slug whose body growth is promoted by an increased food supply. However, no enhancement was observed in the frequency of DNA endoreplication when we compared the transplanted brains in the growth‐promoted and growth‐suppressed host slugs, suggesting that the humoral environment is irrelevant to triggering the body growth‐dependent DNA endoreplication. Next, we tested the requirement of target innervation by surgically dissecting a unilateral posterior pedal nerve of an endogenous brain. Substantially lower number of neurons exhibited DNA endoreplication in the pedal ganglion ipsilateral to the dissected nerve. These results support the view that enhanced DNA endoreplication is mediated by target innervation and is not brought about through the direct effect of humoral factors in the hemolymph during body growth. © 2013 Wiley Periodicals, Inc. Develop Neurobiol 73: 609–620, 2013     

Taxonomic and regional patterns in benthic macroinvertebrate elemental composition in streams

1. Ecological stoichiometry has been used to better understand dynamics in consumer growth and the role of consumer‐recycled nutrients because it focuses on more than one element. Most research has focused on pelagic rather than benthic consumers. Variation in elemental composition among benthic consumer taxa would suggest that taxa differ in their susceptibility to nutrient limitation or in their role in recycling nutrients. 2. We collected benthic macroinvertebrates from streams in two regions (Indiana–Michigan and Wisconsin, U.S.A.) to examine taxonomic and regional variation in benthic macroinvertebrate body carbon (C), nitrogen (N), and phosphorus (P) concentrations and ratios. 3. Elemental composition varied little within taxa common to both regions. In contrast, elemental composition differed greatly among taxa and appeared to be related to phylogeny. The elemental composition of macroinvertebrates clustered into three distinct groups: insects, mollusks, and crustaceans. To a lesser extent, insects and mollusks also differed in elemental composition among genera. 4. Functional feeding groups (FFGs) differed in elemental composition, with predators having a higher N content than other groups. Substantial elemental imbalances between C and N were found between most primary consumers and their likely food sources, and the magnitude of the imbalance depended in part on the FFG. 5. Our results support an assumption of most ecological stoichiometry models that, within a species, the elemental composition of aquatic invertebrates is relatively constant. Variation in elemental composition among taxa at various higher taxonomic levels suggests that susceptibility of stream invertebrates to nutrient limitation and their role in nutrient cycling will strongly depend on phylogeny.     

Modulation of two oscillatory networks in the peripheral olfactory system by γ‐aminobutyric acid,glutamate, and acetylcholine in the terrestrial slug Limax marginatus

The digit‐like extensions (the digits) of the tentacular ganglion of the terrestrial slug Limax marginatus are the cell body rich region in the primary olfactory system, and they contain primary olfactory neurons and projection neurons that send their axons to the olfactory center via the tentacular nerves. Two cell clusters (the cell masses) at the bases of the digits form the other cell body rich regions. Although the spontaneous slow oscillations and odor responses in the tentacular nerve have been studied, the origin of the oscillatory activity is unknown. In the present study, we examined the contribution of the neurons in the digits and cell masses to generation of the tentacular nerve oscillations by surgical removal from the whole tentacle preparations. Both structures contributed to the tentacular oscillations, and surgical isolation of the digits from the whole tentacle preparations still showed spontaneous oscillations. To analyze the dynamics of odor‐processing circuits in the digits and tentacular ganglia, we studied the effects of γ‐aminobutyric acid, glutamate, and acetylcholine on the circuit dynamics of the oscillatory network(s) in the peripheral olfactory system. Bath or local puff application of γ‐aminobutyric acid to the cell masses decreased the tentacular nerve oscillations, whereas the bath or local puff application of glutamate and acetylcholine to the digits increased the digits' oscillations. Our results suggest the existence of two intrinsic oscillatory circuits that respond differentially to endogenous neurotransmitters in the primary olfactory system of slugs. © 2004 Wiley Periodicals, Inc. J Neurobiol 59: 304–318, 2004     

云南澄江早寒武世黑林铺组软体动物太阳女神螺

云南澄江及梅树村早寒武世黑林铺组（玉案山段及石岩头段）的帽形化石,被命名为云南太阳女神螺（新种）,这是澄江生物群内软体动物化石的初次发现。根据梅树村及澄江的地层层位,该种由梅树村阶上部可延至筇竹寺阶,玉案山段软体动物稀少的原因与环境的波动,如潮汐影响近海沉积环境有关。     

红树林区软体动物生态位的三种分析方法

利用广东湛江红树林保护区定量取样获取的软体动物密度数据,根据Shannon-Wiener和Pj．anka公式,分别选用潮带和季节（方法1）、红树植物群落和季节（方法2）、红树植物群落和潮带（方法3）3种不同环境因子组合确定环境资源位点数,从而计算研究区软体动物生态位宽度和物种间重叠值。结果表明,3种分析方法计算结果有差别,采用方法1、方法2和方法3计算的物种生态位宽度分别为0～1．96、0～2．16、0～1．64;生态位重叠指数为0～0．1的种对,方法1、方法2和方法3分别占25．1％、32．8％和21．6％,生态位重叠指数为0．1～0．5的种对,分别占31．6％、36．2％和22．3％,生态位重叠指数〉0．5的种对分别占43．396、3196和56．1％。可见,方法3计算出的生态位重叠指数比其它2种方法的大。物种聚类和排序也验证了3种分析方法的计算结果有差别。将计算结果与实际定量取样比较,表明以红树植物群落和季节为准确定环境资源位点数的分析方法与实际相符,适合红树林区软体动物的生态位研究。     

Catalytic properties and mode of action of endo-(1-->3)-beta-D-glucanase and beta-D-glucosidase from the marine mollusk Littorina kurila

A complex of the enzymes from the liver of the marine mollusk Littorina kurila that hydrolyzes laminaran was investigated. Two (1-->3)-beta-d-glucanases (G-I and G-II) were isolated. The molecular mass of G-I as estimated by gel-permeation chromatography and SDS-PAGE analysis was 32 and 40kDa, respectively. The G-II molecular mass according to SDS-PAGE analysis was about 200kDa. The pH optimum for both G-I and G-II was pH 5.4. The G-I had narrow substrate specificity and hydrolyzed only the (1-->3)-beta-d-glucosidic bonds in the mixed (1-->3),(1-->6)- and (1-->3),(1-->4)-beta-d-glucans down to glucose and glucooligosaccharides. This enzyme acted with retention of the anomeric configuration and catalyzed a transglycosylation reaction. G-I was classified as the glucan endo-(1-->3)-beta-d-glucosidase (EC 3.2.1.39). G-II exhibited both exo-glucanase and beta-d-glucoside activities. This enzyme released from the laminaran glucose as a single product, but retained the anomeric center configuration and possessed transglycosylation activity. The hydrolysis rate of glucooligosaccharides by G-I decreased with an increase of the substrate's degree of polymerization. In addition to (1-->3)-beta-d-glucanase activity, the enzyme had the ability to hydrolyze p-nitrophenyl beta-d-glucoside and beta-d-glucobioses: laminaribiose, gentiobiose, and cellobiose, with the rate ratio of 50:12:1. G-II may correspond to beta-d-glucoside glucohydrolase (EC 3.2.1.21).     

Nitric oxide release from a single cell affects filopodial motility on growth cones of neighboring neurons

Nitric oxide (NO), a gaseous messenger, has been reported to be involved in a variety of functions in the nervous system, ranging from neuronal pathfinding to learning and memory. We have shown previously that the application of NO via NO donors to growth cones of identified Helisoma buccal neurons B5 in vitro induces an increase in filopodial length, a decrease in filopodial number, and a slowing in neurite advance. It is unclear, however, whether NO released from a physiological source would affect growth cone dynamics. Here we used cell bodies of identified neurons known to express the NO synthesizing enzyme nitric oxide synthase (NOS) as a source of constitutive NO production and tested their effect on growth cones of other cells in a sender-receiver paradigm. We showed that B5 cell bodies induced a rapid increase in filopodial length in NO-responsive growth cones, and that this effect was blocked by the NOS inhibitor 7-NI, suggesting that the effect was mediated by NO. Inhibition of soluble guanylyl cyclase (sGC) with ODQ blocked filopodial elongation induced by B5 somata, confirming that NO acted via sGC. We also demonstrate that the effect of NO was reversible and that a cell releasing NO can affect growth cones over a distance of at least 100 microm. Our results suggest that NO released from a physiological source can affect the motility of nearby growth cones and thus should be considered a signaling molecule with the potential to affect the outcome of neuronal pathfinding in vivo.     

Isolation and partial characterization of neuropeptides that mimic prolonged inhibition produced by bag cell neurons in Aplysia

The bag cell neurons of the marine mollusk Aplysia are part of a neural system that utilizes four neuropeptides as neurotansmitters. The peptides, derived from the egglaying hormone/bag cell peptide (ELH/BCP) precursor protein, are released during a 20-min burst discharge of the bag cells and produce several types of responses in various abdominal ganglion neurons. In the identified neurons L3 and L6, bag cell activity produces prolonged inhibition that lasts for more than 2 h. One of the bag cell peptides, alpha-BCP, mediates an early component of the inhibition in these neurons. To identify the co-transmitter mediating the prolonged component of inhibition, we purified material from an acid extract of abdominal ganglia using molecular sizing high-pressure liquid chromatography (HPLC) on TSK 250-125 followed by two steps of reverse-phase HPLC on C4 or C18. We isolated three inhibitory factors that mimic the prolonged component of inhibition. Mass spectroscopy and partial amino acid sequence analysis indicate one factor is ELH [2-36], that is, ELH that lacks the first, N-terminal amino acid. This inhibitory activity was similar in potency to that of ELH and is the first to be described for an ELH related peptide. The two other factors were approximately 3,300 and 4,700 Da and were effective at 10- and 50-fold lower concentration, respectively, than ELH or its fragment. Amino acid composition analysis suggests that they are not derived from the ELH/BCP precursor protein. The 4,700 Da factor is effective at the lowest concentration and produces an effect that lasts as long as 100 min. Therefore, it is the best candidate for the true inhibitory transmitter. Because the inhibited neurons in nervate the kidney, the function of prolonged inhibition may be to regulate kidney function during egg laying.     

MicroRNA Regulation in Extreme Environments:Differential Expression of MicroRNAs in the Intertidal Snail Littorina littorea During Extended Periods of Freezing and Anoxia

Several recent studies of vertebrate adaptation to environmental stress have suggested roles for microRNAs (miRNAs) in regulating glo- bal suppression of protein synthesis and/or restructuring protein expression patterns. The present study is the first to characterize stress-responsive alterations in the expression of miRNAs during natural freezing or anoxia exposures in an invertebrate species, the intertidal gastropod Littorina littorea. These snails are exposed to anoxia and freezing conditions as their environment constantly fluctuates on both a tidal and seasonal basis. The expression of selected miRNAs that are known to influence the cell cycle, cellular signaling pathways, carbohydrate metabolism and apoptosis was evaluated using RT-PCR. Compared to controls, significant changes in expression were observed for miR-1a-1, miR-34a and miR-29b in hepatopancreas and for miR-1a-1, miR-34a, miR-133a, miR-125b, miR-29b and miR-2a in foot muscle after freezing exposure at 6 °C for 24 h (P < 0.05). In addition, in response to anoxia stress for 24 h, significant changes in expression were also observed for miR-1a-1, miR-210 and miR-29b in hepatopancreas and for miR-1a-1, miR-34a, miR-133a, miR-29b and miR-2a in foot muscle (P < 0.05). Moreover, protein expression of Dicer, an enzyme responsible for mature microRNA processing, was increased in foot muscle during freezing and anoxia and in hepatopancreas during freezing. Alterations in expression of these miRNAs in L. littorea tissues may contribute to organismal survival under freezing and anoxia.