Appetitive Associative Olfactory Learning in Drosophila Larvae

In the following we describe the methodological details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with genetic interference, provides a handle to analyze the neuronal and molecular fundamentals of specifically associative learning in a simple larval brain.Organisms can use past experience to adjust present behavior. Such acquisition of behavioral potential can be defined as learning, and the physical bases of these potentials as memory traces^1-4. Neuroscientists try to understand how these processes are organized in terms of molecular and neuronal changes in the brain by using a variety of methods in model organisms ranging from insects to vertebrates^5,6. For such endeavors it is helpful to use model systems that are simple and experimentally accessible. The Drosophila larva has turned out to satisfy these demands based on the availability of robust behavioral assays, the existence of a variety of transgenic techniques and the elementary organization of the nervous system comprising only about 10,000 neurons (albeit with some concessions: cognitive limitations, few behavioral options, and richness of experience questionable)^7-10.Drosophila larvae can form associations between odors and appetitive gustatory reinforcement like sugar^11-14. In a standard assay, established in the lab of B. Gerber, animals receive a two-odor reciprocal training: A first group of larvae is exposed to an odor A together with a gustatory reinforcer (sugar reward) and is subsequently exposed to an odor B without reinforcement ⁹. Meanwhile a second group of larvae receives reciprocal training while experiencing odor A without reinforcement and subsequently being exposed to odor B with reinforcement (sugar reward). In the following both groups are tested for their preference between the two odors. Relatively higher preferences for the rewarded odor reflect associative learning - presented as a performance index (PI). The conclusion regarding the associative nature of the performance index is compelling, because apart from the contingency between odors and tastants, other parameters, such as odor and reward exposure, passage of time and handling do not differ between the two groups⁹.     

On the inactivity of Candida antartica lipase B towards strong acids

Candida antarctica lipase B (CalB, Novozyme 435) was evaluated as catalyst for the conversion of so-called edible acids (e.g. malic and tartaric acid). While transesterification using these acyl donors proceeds smoothly, albeit with low regioselectivity, esterification is hardly catalyzed.As major reason for CalB inactivation the high acidity of edible acids was identified leading to irreversible inactivation of the biocatalyst. Furthermore, indication exist that all acids exhibiting a pK_a value below 4.8 cause irreversible inactivation of CalB.     

Comparative ecological niche models predict the invasive spread of variable-leaf milfoil (<Emphasis Type="Italic">Myriophyllum</Emphasis><Emphasis Type="Italic">heterophyllum</Emphasis>) and its potential impact on closely related native species

Invasive species can alter patterns of biodiversity by displacing closely related native species that occupy similar habitats. We used multivariate ecological niche modeling to determine the potential spread and displacement of native congeners by the invasive aquatic plant, variable-leaf milfoil (VLM, Myriophyllum heterophyllum) in New Hampshire, USA. We show that VLM occurs almost exclusively in “higher order” lakes characterized as large, low elevation systems with relatively high pH, alkalinity and conductivity. In contrast, native milfoils occur across a broad range of lake orders. The strong association between lake order and VLM invasions suggests that VLM is most likely to displace native milfoils in higher order lakes. However, the mechanism by which VLM occurs in higher order lakes—higher propagule pressure versus higher growth and survivorship—is unclear. We therefore caution that native species may ultimately be susceptible to displacement from lower order lakes if the current distribution of VLM reflects higher propagule pressures. Our model provides a valuable tool for the prioritization of monitoring efforts.     

Using 18S rDNA to resolve diaptomid copepod (Copepoda: Calanoida: Diaptomidae) phylogeny: an example with the North American genera

The phylogenetic relationships among the numerous genera of diaptomid copepods remain elusive due to difficulties in obtaining sufficient numbers of phylogenetically informative morphological characters for cladistic analysis. Molecular phylogenetic techniques offer high potential to resolve phylogenetic relationships in the absence of sufficient morphological characters because of the ease in which many characters can be unambiguously coded. I present the first molecular phylogeny for diaptomid copepod genera using 18S rDNA. Specifically, I test Light’s (1939) hypothesis regarding the interrelationships among the North American diaptomid genera. The 18S phylogeny is remarkably consistent with Light’s hypothesis. The endemic North American genera represent a monophyletic group exclusive of the non-endemic genera. Moreover, his hypothesized basal genus for the North America genera, Hesperodiaptomus, is the basal genus in this analysis. However, his Leptodiaptomus group is not reciprocally monophyletic with his Hesperodiaptomus group, but is rather a derived member of the latter group. Finally, the genus Mastigodiaptomus is found to be more closely allied with the non-endemic genera, as Light suggested. This phylogeny contributes heavily to the understanding of phylogenetic relationships among North American diaptomids and has large implications for the systematics of diaptomids in general. The use of 18S rDNA sequences in phylogenetic analyses of diaptomid copepods can be used to confirm the monophyly of recognized genera, the interrelationships among genera, and subsequent biogeographic interpretation of the family’s diversification. The use of molecular data, such as 18S rDNA sequences, to test phylogenetic hypotheses based on a very limited number of morphological characters will be a particularly useful approach to phylogenetic analysis in this system.     

The neural substrate of spectral preference in Drosophila

Drosophila vision is mediated by inputs from three types of photoreceptor neurons; R1-R6 mediate achromatic motion detection, while R7 and R8 constitute two chromatic channels. Neural circuits for processing chromatic information are not known. Here, we identified the first-order interneurons downstream of the chromatic channels. Serial EM revealed that small-field projection neurons Tm5 and Tm9 receive direct synaptic input from R7 and R8, respectively, and indirect input from R1-R6, qualifying them to function as color-opponent neurons. Wide-field Dm8 amacrine neurons receive input from 13-16 UV-sensing R7s and provide output to projection neurons. Using a combinatorial expression system to manipulate activity in different neuron subtypes, we determined that Dm8 neurons are necessary and sufficient for flies to exhibit phototaxis toward ultraviolet instead of green light. We propose that Dm8 sacrifices spatial resolution for sensitivity by relaying signals from multiple R7s to projection neurons, which then provide output to higher visual centers.     

Genome-wide investigation of light and carbon signaling interactions in Arabidopsis

Background  

Light and carbon are two essential signals influencing plant growth and development. Little is known about how carbon and light signaling pathways intersect or influence one another to affect gene expression.     

Clostridium difficile toxin A induces expression of the stress-induced early gene product RhoB

Clostridium difficile toxin A monoglucosylates the Rho family GTPases Rho, Rac, and Cdc42. Glucosylation leads to the functional inactivation of Rho GTPases and causes disruption of the actin cytoskeleton. A cDNA microarray revealed the immediate early gene rhoB as the gene that was predominantly up-regulated in colonic CaCo-2 cells after treatment with toxin A. This toxin A effect was also detectable in epithelial cells such as HT29 and Madin-Darby canine kidney cells, as well as NIH 3T3 fibroblasts. The expression of RhoB was time-dependent and correlated with the morphological changes of cells. The up-regulation of RhoB was approximately 15-fold and was based on the de novo synthesis of the GTPase because cycloheximide completely inhibited the toxin A effect. After 8 h, a steady state was reached, with no further increase in RhoB. The p38 MAPK inhibitor SB202190 reduced the expression of RhoB, indicating a participation of the p38 MAPK in this stress response. Surprisingly, newly formed RhoB protein was only partially glucosylated by toxin A, sparing a pool of potentially active RhoB, as checked by sequential C3bot-catalyzed ADP-ribosylation. A pull-down assay in fact revealed a significant amount of active RhoB in toxin A-treated cells that was not present in control cells. We demonstrate for the first time that toxin A has not only the property to inactivate the GTPases RhoA, Rac1, and Cdc42 by glucosylation, but it also has the property to generate active RhoB that likely contributes to the overall picture of toxin treatment.     

Microarray-based gene expression profiling to elucidate cellular responses to nitric oxide--a review from an analytical and biomedical point of view

Nitric oxide (NO) produced by NO synthases (NOS) regulates a wide range of cellular functions. Analysis by gene arrays provides valuable information for identifying important elements of the cellular responses to NO. Such screening tools might be useful to elucidate NO-responsive regulators, which play a central role in mediating NO effects. Although the final importance of a particular gene is determined by the encoded protein and further protein modifications, measurements of RNA levels have proven to be partly valuable in identifying the molecular changes that occur in cells. Microarray technology permits large-scale and genome-wide analysis of gene expression from multiple samples. We review the current knowledge of the use of microarray gene expression screening in elucidating the effects of NO on various cells and tissues. We also point out the limitations of general microarray-based gene expression analyses and especially when investigating the effects of NO.     

Complex postglacial recolonization inferred from population genetic structure of mottled sculpin Cottus bairdii in tributaries of eastern Lake Michigan,U.S.A.

This study used analyses of the genetic structure of a non‐game fish species, the mottled sculpin Cottus bairdii to hypothesize probable recolonization routes used by cottids and possibly other Laurentian Great Lakes fishes following glacial recession. Based on samples from 16 small streams in five major Lake Michigan, U.S.A., tributary basins, significant interpopulation differentiation was documented (overall F_ST = 0·235). Differentiation was complex, however, with unexpectedly high genetic similarity among basins as well as occasionally strong differentiation within basins, despite relatively close geographic proximity of populations. Genetic dissimilarities were identified between eastern and western populations within river basins, with similarities existing between eastern and western populations across basins. Given such patterns, recolonization is hypothesized to have occurred on three occasions from more than one glacial refugium, with a secondary vicariant event resulting from reduction in the water level of ancestral Lake Michigan. By studying the phylogeography of a small, non‐game fish species, this study provides insight into recolonization dynamics of the region that could be difficult to infer from game species that are often broadly dispersed by humans.