Functional expression and ligand identification of homo- and heteromeric Drosophila melanogaster CO2 receptors in the Xenopus laevis oocyte system.

Carbon dioxide (CO2) is an important olfactory cue in Drosophila melanogaster and can elicit both attractive and aversive behaviors. It is detected by gustatory receptors, Gr21a and Gr63a, found in the ab1C neuron in basiconic sensilla on the third antennal segment. Volatile substances that modulate the receptors' function are of interest for pest control. While several substances block ab1C neurons or mimic the activating effect of carbon dioxide, it is not known if these substances are indeed ligands of the CO2 receptor or might act on other proteins in the receptor neuron. In this study, we used the recombinant Xenopus laevis expression system and two-electrode voltage-clamp technology to investigate the receptor function. We found that application of sodium bicarbonate evokes large inward currents in oocytes co-expressing Gr21a and Gr63a. The receptors most likely form hetromultimeric complexes. Homomultimeric receptors of Gr21a or Gr63a are sufficient for receptor functionality, although oocytes gave significantly lower current responses compared to the probable heteromultimeric receptor. We screened for putative blockers of the sodium bicarbonate response and confirmed that some of the substances identified by spike recordings of olfactory receptor neurons, such as 1-hexanol, are also blockers in the Xenopus oocyte system. We also identified a new blocking substance, citronellol, which is related to insect repellents. Many substances that activate receptor neurons were inactive in the Xenopus oocyte system, indicating that they may not be ligands for the receptor, but may act on other proteins. However, methyl pyruvate and n-hexylamine were found to be activators of the recombinant Gr21a/Gr63a receptor.

Effect of 158 herbal remedies on human TRPV1 and the two-pore domain potassium channels KCNK2, 3 and 9.

BACKGROUND AND AIM: Herbal medicines are used to treat a broad number of maladies. However, the pharmacological profile of most remedies is poorly understood. We investigated the effect of herbal remedies from kampo, traditional Chinese medicine (TCM) and other phytotherapies on human two-pore domain potassium channels (KCNK channels; TREK-1, TASK-1 and TASK-3) as well as the human TRPV1 channel. KCNK channels are responsible for the background potassium current of excitable cells, thus essential for the maintenance of the resting membrane potential. Hence, modulators of KCNK channels are of medical significance, e.g. for the treatment of sleep disorders and pain. The transient receptor potential channel TRPV1 is a pain detector for noxious heat. Agonists of this receptor are still used for the treatment of pain in ectopic applications. EXPERIMENTAL PROCEDURE: We evaluated the effect of 158 herbal remedies on these channels in a heterologous expression system (Xenopus laevis oocytes) using the two-electrode voltage-clamp technique with the aim of increasing the comprehension of their pharmacological profile. RESULTS AND CONCLUSION: Some remedies with modulating effects were identified such as Angelica pubescens (radix), which inhibit TASK-1 and TASK-3 channels. Furthermore, the modulatory effects of the most effective remedies on the two TASK family members TASK-1 and TASK-3 correlate positively, reflecting their close relation. For the TRPV1 channel Terminalia chebula and Alchemilla xanthochlora were identified as potentiators. This study identifies a variety of herbal remedies as modulators of human K2P and TRPV1 channels and gives new insights into the pharmacological profile of these herbal remedies.

Linear phase-and-frequency-modulated photonic links using optical discriminators.

We report our experimental results for linear analog optical links that use phase or frequency modulation and optical discrimination. The discriminators are based on two architectures: a cascaded MZI FIR lattice filter and a ring assisted MZI (RAMZI) IIR filter. For both types of discriminators, we demonstrate > 6 dB improvement in the link's third-order output intercept point (OIP3) over a MZM link. We show that the links have low second-order distortion when using balanced detection. Using high optical power, we demonstrate an OIP3 of 39.2 dBm. We also demonstrate 4.3dB improvement in signal compression.

The Smell of Blue Light: A New Approach toward Understanding an Olfactory Neuronal Network.

Olfaction is one of the most important senses throughout the animal kingdom. It enables animals to discriminate between a wide variety of attractive and repulsive odorants and often plays a decisive role in species specific communication. In recent years the analysis of olfactory systems both invertebrates and invertebrates has attracted much scientific interest. In this context a pivotal question is how the properties and connectivities of individual neurons contribute to a functioning neuronal network that mediates odor-guided behavior. As a novel approach to analyze the role of individual neurons within a circuitry, techniques have been established that make use of light-sensitive proteins. In this review we introduce a non-invasive, optogenetic technique which was used to manipulate the activity of individual neurons in the olfactory system of Drosophila melanogaster larvae. Both channelrhodopsin-2 and the photosensitive adenylyl cyclase PAC α in individual olfactory receptor neurons (ORNs) of the olfactory system of Drosophila larvae allows stimulating individual receptor neurons by light. Depending on which particular ORN is optogenetically activated, repulsion or attraction behavior can be induced, indicating which sensory neurons underlie which type of behavior.

Optical transmission through double-layer, laterally shifted metallic subwavelength hole arrays.

We measure the transmission of IR radiation through double-layer metal films with periodic arrays of subwavelength holes. When the two metal films are placed in sufficiently close proximity, two types of transmission resonances emerge. For the surface plasmon mode, the electromagnetic field is concentrated on the outer surface of the entire metallic layer stack. In contrast, for the guided mode, the field is confined to the gap between the two metal layers. Our measurements indicate that, as the two layers are laterally shifted from perfect alignment, the peak transmission frequency of the guided mode decreases significantly, while that of the surface plasmon mode remains largely unchanged, in agreement with numerical calculations.

Optogenetically Induced Olfactory Stimulation in Drosophila Larvae Reveals the Neuronal Basis of Odor-Aversion behavior.

Olfactory stimulation induces an odor-guided crawling behavior of Drosophila melanogaster larvae characterized by either an attractive or a repellent reaction. In order to understand the underlying processes leading to these orientations we stimulated single olfactory receptor neurons (ORNs) through photo-activation within an intact neuronal network. Using the Gal4-UAS system two light inducible proteins, the light-sensitive cation channel channelrhodopsin-2 (ChR-2) or the light-sensitive adenylyl cyclase (Pacalpha) were expressed in all or in individual ORNs of the larval olfactory system. Blue light stimulation caused an activation of these neurons, ultimately producing the illusion of an odor stimulus. Larvae were tested in a phototaxis assay for their orientation toward or away from the light source. Here we show that activation of Pacalpha expressing ORNs bearing the receptors Or33b or Or45a in blind norpA mutant larvae induces a repellent behavior away from the light. Conversely, photo-activation of the majority of ORNs induces attraction towards the light. Interestingly, in wild type larvae two ligands of Or33b and Or45a, octyl acetate and propionic ethylester, respectively, have been found to cause an escape reaction. Therefore, we combined light and odor stimulation to analyze the function of Or33b and Or45a expressing ORNs. We show that the larval olfactory system contains a designated neuronal pathway for repellent odorants and that activation of a specific class of ORNs already determines olfactory avoidance behavior.

Nanopores in solid-state membranes engineered for single molecule detection.

A nanopore is an analytical tool with single molecule sensitivity. For detection, a nanopore relies on the electrical signal that develops when a molecule translocates through it. However, the detection sensitivity can be adversely affected by noise and the frequency response. Here, we report measurements of the frequency and noise performance of nanopores

Casimir force on a surface with shallow nanoscale corrugations: geometry and finite conductivity effects.

We measure the Casimir force between a gold sphere and a silicon plate with nanoscale, rectangular corrugations with a depth comparable to the separation between the surfaces. In the proximity force approximation (PFA), both the top and bottom surfaces of the corrugations contribute to the force, leading to a distance dependence that is distinct from a flat surface. The measured Casimir force is found to deviate from the PFA by up to 10%, in good agreement with calculations based on scattering theory that includes both geometry effects and the optical properties of the material.

Measurement of the Casimir force between a gold sphere and a silicon surface with nanoscale trench arrays.

We report measurements of the Casimir force between a gold sphere and a silicon surface with an array of nanoscale, rectangular corrugations using a micromechanical torsional oscillator. At distances between 150 and 500 nm, the measured force shows significant deviations from the pairwise additive formulism, demonstrating the strong dependence of the Casimir force on the shape of the interacting bodies. The observed deviation, however, is smaller than the calculated values for perfectly conducting surfaces, possibly due to the interplay between finite conductivity and geometry effects.

Controlling the phase delay of light transmitted through double-layer metallic subwavelength slit arrays.

We demonstrate that the phase of light transmitted through double-layer subwavelength metallic slit arrays can be controlled through lateral shift of the two layers. Our samples consist of two aluminum layers, each of which contains an array of subwavelength slits. The two layers are placed in sufficient proximity to allow coupling of the evanescent fields at resonance. By changing the lateral shift between the layers from zero to half the period, the phase of the transmitted electromagnetic field is increased by pi, while the transmitted intensity remains high. Such a controllable phase delay could open new capabilities for nanophotonic devices that cannot be achieved with single-layer structures.

Small-Signal Performance and Modeling of sub-50nm nMOSFETs with f above 460-GHz.

We have fabricated and tested the performance of sub-50nm gate nMOSFETs to assess their suitability for mixed signal applications in the super high frequency (SHF) band, i.e. 3-30GHz. For a 30nm×40 µm×2 device, we found f(T) =465GHz at V(ds)=2V, V(g)=0.67V, which is the highest cut-off frequency reported for a MOSFET produced on bulk silicon substrate so far. However, our measurements of f(max) and noise figure indicate that parasitics impose limitations on SHF operation. We also present a high-frequency ac model appropriate to sub-50nm gate length nanotransistors, which incorporates the effects of the parasitics. The model accurately accounts for measurements of the S and Y parameters in the frequency range from 1 to 50GHz.

Optical transmission through double-layer metallic subwavelength slit arrays.

We present measurements of transmission of infrared radiation through double-layer metallic grating structures. Each metal layer contains an array of subwavelength slits and supports transmission resonance in the absence of the other layer. The two metal layers are fabricated in close proximity to allow coupling of the evanescent field on individual layers. The transmission of the double layer is found to be surprisingly large at particular wavelengths, even when no direct line of sight exists through the structure as a result of the lateral shifts between the two layers. We perform numerical simulations using rigorous coupled wave analysis to explain the strong dependence of the peak transmission on the lateral shift between the metal layers.

An increased receptive field of olfactory receptor Or43a in the antennal lobe of Drosophila reduces benzaldehyde-driven avoidance behavior.

Most animals orient themselves in their environment through the perception of olfactory cues. In order to gain insight into the principles of olfactory processing in Drosophila, we misexpressed olfactory receptor Or43a in additional olfactory receptor neurons of the third antennal segment using enhancer trap line GH320. The behavioral response of GH320/UAS-or43a flies was changed upon benzaldehyde application. Using the T-maze assay, misexpressing flies performed a reduced avoidance reaction to benzaldehyde as compared with wild type. This reduction of avoidance could be mimicked in wild type flies by exposing them to a mixture of benzaldehyde and ethyl acetate. We therefore conclude that the application of benzaldehyde, an identified ligand of Or43a, resulted in activation of a number of glomeruli in transformed flies in addition to glomerulus DA4, which is the regular target of Or43a expressing neurons. Our results demonstrate the relevance of specific olfactory sensory input and subsequent processing in the antennal lobe for Drosophila behavior.

Ebony protein in the Drosophila nervous system: optic neuropile expression in glial cells.

The Drosophila ebony mutation (Bridges and Morgan, [1923] Publs Carnegie Inst Wash 327:50) reveals a pleiotropic phenotype with cuticular and behavioral defects. To understand Ebony function in the nervous system, particularly in transmission of the visual signal, it is essential to know the cell type and temporal characteristics of its expression throughout development. Therefore, we raised an antiserum against an Ebony peptide to detect the protein in whole-mount and slice preparations of Drosophila. Attention was focused on ebony expression in the adult optic neuropiles of the fly. Colocalization of Ebony with neuronal or glial cell markers in frozen sections showed non-neuronal expression of ebony in the lamina and medulla neuropiles. Furthermore, colocalization with glial cell markers demonstrated glial expression of ebony in epithelial glia of the lamina and neuropile glia of the distal medulla. This finding was confirmed for the lamina epithelial glia by electron microscopic examination of immunolabeling by using the diaminobenzidine method. These glia have in common that they match the two sites of histamine release from the compound eye's photoreceptors. Possible ways in which the biochemical activity of Ebony might function with respect to histamine release are considered.

Room-temperature phosphorescence from azurin derivatives. Phosphorescence quenching in oxidized native azurin.

The tryptophan phosphorescence from a series of derivatives of Pseudomonas aeruginosa azurin has been monitored at 30 degrees C in pH 8.5 buffer solution. The phosphorescence lifetimes fall in the range of 230-270 ms for deoxygenated solutions of derivatives containing Cd(II), Cu(I), Co(II), Ni(II), Hg(II) or apoazurin. A weak signal with a lifetime of ca 130 ms is observed from solutions of oxidized native azurin, but this component is ascribed to a modified form of azurin in solution, i.e. protein heterogeneity, on the basis of the unique sensitivity to quenching by dioxygen. Aside from this minor component, the tryptophan phosphorescence in the Cu(II) protein appears to be fully quenched. The quenching is assigned an electron-transfer mechanism involving transient reduction of the metal center. The same mechanism is deemed to be responsible for fluorescence quenching in oxidized native azurin as well. These observations are of interest because aromatic groups like tryptophan may be conduits for physiological electron-transfer processes involving the copper center.

Human serum Cohn fraction IV (alpha-globulin [correction of globin] enriched) inhibits ligand binding at neurotransmitter receptors in human brain.

Human serum proteins are found in significant density in the neuropil in brains of demented individuals. The functional significance of these abnormally distributed proteins has been unknown. We now report that alpha-globulin-enriched fractions of human serum decrease the specific binding of [3H]spiroperidol at its binding sites in postmortem human frontal cortex and caudate. The substances in this serum fraction apparently exert their effect by a direct action on the binding site. Since [3H]spiroperidol labels serotoninergic and dopaminergic among other neurotransmitter receptors, these results suggest that components of human serum inhibit the binding of ligands at neurotransmitter receptors.