Evaluation of the DREAM Technique for a High-Throughput Deorphanization of Chemosensory Receptors in Drosophila

In the vinegar fly Drosophila melanogaster, the majority of olfactory receptors mediating the detection of volatile chemicals found in their natural habitat have been functionally characterized (deorphanized) in vivo. In this process, receptors have been assigned ligands leading to either excitation or inhibition in the olfactory sensory neuron where they are expressed. In other, non-drosophilid insect species, scientists have not yet been able to compile datasets about ligand–receptor interactions anywhere near as extensive as in the model organism D. melanogaster, as genetic tools necessary for receptor deorphanization are still missing. Recently, it was discovered that exposure to artificially high concentrations of odorants leads to reliable alterations in mRNA levels of interacting odorant receptors in mammals. Analyzing receptor expression after odorant exposure can, therefore, help to identify ligand–receptor interactions in vivo without the need for other genetic tools. Transfer of the same methodology from mice to a small number of receptors in D. melanogaster resulted in a similar trend, indicating that odorant exposure induced alterations in mRNA levels are generally applicable for deorphanization of interacting chemosensory receptors. Here, we evaluated the potential of the DREAM (Deorphanization of receptors based on expression alterations in mRNA levels) technique for high-throughput deorphanization of chemosensory receptors in insect species using D. melanogaster as a model. We confirmed that in some cases the exposure of a chemosensory receptor to high concentration of its best ligand leads to measureable alterations in mRNA levels. However, unlike in mammals, we found several cases where either confirmed ligands did not induce alterations in mRNA levels of the corresponding chemosensory receptors, or where gene transcript-levels were altered even though there is no evidence for a ligand–receptor interaction. Hence, there are severe limitations to the suitability of the DREAM technique for deorphanization as a general tool to characterize olfactory receptors in insects

Linking Symptom Inventories using Semantic Textual Similarity

An extensive library of symptom inventories has been developed over time to measure clinical symptoms, but this variety has led to several long standing issues. Most notably, results drawn from different settings and studies are not comparable, which limits reproducibility. Here, we present an artificial intelligence (AI) approach using semantic textual similarity (STS) to link symptoms and scores across previously incongruous symptom inventories. We tested the ability of four pre-trained STS models to screen thousands of symptom description pairs for related content - a challenging task typically requiring expert panels. Models were tasked to predict symptom severity across four different inventories for 6,607 participants drawn from 16 international data sources. The STS approach achieved 74.8% accuracy across five tasks, outperforming other models tested. This work suggests that incorporating contextual, semantic information can assist expert decision-making processes, yielding gains for both general and disease-specific clinical assessment

ENIGMA and global neuroscience: A decade of large-scale studies of the brain in health and disease across more than 40 countries

This review summarizes the last decade of work by the ENIGMA (Enhancing NeuroImaging Genetics through Meta Analysis) Consortium, a global alliance of over 1400 scientists across 43 countries, studying the human brain in health and disease. Building on large-scale genetic studies that discovered the first robustly replicated genetic loci associated with brain metrics, ENIGMA has diversified into over 50 working groups (WGs), pooling worldwide data and expertise to answer fundamental questions in neuroscience, psychiatry, neurology, and genetics. Most ENIGMA WGs focus on specific psychiatric and neurological conditions, other WGs study normal variation due to sex and gender differences, or development and aging; still other WGs develop methodological pipelines and tools to facilitate harmonized analyses of "big data" (i.e., genetic and epigenetic data, multimodal MRI, and electroencephalography data). These international efforts have yielded the largest neuroimaging studies to date in schizophrenia, bipolar disorder, major depressive disorder, post-traumatic stress disorder, substance use disorders, obsessive-compulsive disorder, attention-deficit/hyperactivity disorder, autism spectrum disorders, epilepsy, and 22q11.2 deletion syndrome. More recent ENIGMA WGs have formed to study anxiety disorders, suicidal thoughts and behavior, sleep and insomnia, eating disorders, irritability, brain injury, antisocial personality and conduct disorder, and dissociative identity disorder. Here, we summarize the first decade of ENIGMA's activities and ongoing projects, and describe the successes and challenges encountered along the way. We highlight the advantages of collaborative large-scale coordinated data analyses for testing reproducibility and robustness of findings, offering the opportunity to identify brain systems involved in clinical syndromes across diverse samples and associated genetic, environmental, demographic, cognitive, and psychosocial factors

No Evidence for Ionotropic Pheromone Transduction in the Hawkmoth Manduca sexta.

Insect odorant receptors (ORs) are 7-transmembrane receptors with inverse membrane topology. They associate with the conserved ion channel Orco. As chaperon, Orco maintains ORs in cilia and, as pacemaker channel, Orco controls spontaneous activity in olfactory receptor neurons. Odorant binding to ORs opens OR-Orco receptor ion channel complexes in heterologous expression systems. It is unknown, whether this also occurs in vivo. As an alternative to this ionotropic transduction, experimental evidence is accumulating for metabotropic odor transduction, implicating that insect ORs couple to G-proteins. Resulting second messengers gate various ion channels. They generate the sensillum potential that elicits phasic-tonic action potentials (APs) followed by late, long-lasting pheromone responses. Because it is still unclear how and when Orco opens after odor-OR-binding, we used tip recordings to examine in vivo the effects of the Orco antagonist OLC15 and the amilorides MIA and HMA on bombykal transduction in the hawkmoth Manduca sexta. In contrast to OLC15 both amilorides decreased the pheromone-dependent sensillum potential amplitude and the frequency of the phasic AP response. Instead, OLC15 decreased spontaneous activity, increased latencies of phasic-, and decreased frequencies of late, long-lasting pheromone responses Zeitgebertime-dependently. Our results suggest no involvement for Orco in the primary transduction events, in contrast to amiloride-sensitive channels. Instead of an odor-gated ionotropic receptor, Orco rather acts as a voltage- and apparently second messenger-gated pacemaker channel controlling the membrane potential and hence threshold and kinetics of the pheromone response

Scheme of Orco functions in hawkmoth olfactory sensilla (modified after review: [2]).

<p>MsexOrco is suggested to play no role for the primary events of odor/pheromone transduction. In the dendritic cilia Orco acts as “chaperon”, locating and maintaining odor-binding olfactory receptors (OR_x) in membranes of cilia, possibly Ca²⁺/calmoduline-dependently, as shown recently [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166060#pone.0166060.ref049" target="_blank">49</a>]. Odor/pheromone binding to ORs activates a phospholipase (PLCβ) that hydrolyzes phospholipids (PIP₂). The metabotropic odor/pheromone transduction generates rises in IP₃ which open first a Ca²⁺ channel. The rise in intracellular Ca²⁺ starts a cascade of ion channel openings (not shown) that generate the depolarizing receptor potential. The resulting odor-dependent voltage-changes gate Orco in the soma of the olfactory receptor neurons. There, MsexOrco acts as pacemaker channel that controls membrane potential oscillations and, thus, spike threshold and response kinetics. It remains to be studied whether MsexOrco is also gated via cAMP and cGMP, or via protein kinase C-dependent phosphorylation. BL basal lamina, CI cilium, CU cuticle, D dendrite, GL glia, HL hemolymph, ORN olfactory receptor neuron, P pore, RE receptor lymph, TE thecogen cell, TO tormogen cell, TR trichogen cell.</p

OLC15, MIA, and HMA affect different targets of the pheromone transduction cascade.

<p>(A) The bombykal (50 ms; 1 μg on filter paper)—dependent AP response consists of at least 3 phases which occur at characteristic time windows after the pheromone stimulus (red arrow), also at other pheromone doses employed. In the absence of pheromone, olfactory receptor neurons (ORNs) show low rates of spontaneous activity (yellow). ORNs respond to pheromone with a fast, phasic (red), and a delayed, tonic (blue) series of APs (enlargement). After a period of inhibition the late, long-lasting pheromone response (LLPR, green) begins and can last for several seconds up to minutes. (B, C) In contrast to both amilorides, Orco antagonist OLC15 does not affect the primary, phasic, or the tonic pheromone response in tip-recordings of intact hawkmoth antennae, during activity- (B) and rest-phase (C). The bombykal-dependent sensillum potential (DC) and phasic action potential response (AC) were not affected by OLC15 (10 μM), as compared to controls (DMSO) (B, C). In contrast, both parameters of the primary pheromone response were decreased Zeitgebertimer (ZT)-dependently by MIA (10 μM; shown only at activity (B)) and HMA (10 μM; shown only at rest (C)).</p

Orco agonist VUAA1 activates MsexOrco in olfactory receptor neurons (ORNs) of hawkmoths.

<p>(A) Two weeks-old primary cell cultures of pupal antennae contain two antennal cells strongly filled with Fura-2 (red arrows), attached to less-stained non-neuronal cells (yellow arrows) with long processes. According to its soma size of about 10 μm and its fine processes (not stained) the smaller of the two strongly stained cells is an ORN. (B) In Ca²⁺ imaging studies VUAA1 dose-dependently (1 μM, 10 μM, 100 μM) increased intracellular Ca²⁺ levels in the small ORN shown in A, identifying it as an Orco-expressing ORN. Primary cell cultures of ORNs from 2 day-old pupae were grown for two weeks <i>in vitro</i> before stimulation. At this stage the ORNs matured and were shown previously to become pheromone-responsive [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166060#pone.0166060.ref035" target="_blank">35</a>].</p

The conserved ion channel Orco is present in developing and adult hawkmoth antennae.

<p>(A) Antisera against moth Orco stain both somata of olfactory receptor neurons (ORNs) under long pheromone-sensitive trichoid sensilla in adult male antennae of the hawkmoth <i>Manduca sexta</i>. Dendrites and cilia, which extend into the cuticular hairs of the sensilla, are Orco-immunoreactive (green), while axons of the ORNs are not stained. A 27 μm maximum projection of a confocal image is shown. Grey: autofluorescence of the cuticle. Scalebar: 20 μm. C, cuticle of the antenna; le, leading edge of the antenna; te, trailing edge of the antenna. (B) Antisera against moth Orco recognize a protein of the approximate predicted molecular mass of Orco (~54 kDa). Western blots of pupal male <i>M</i>. <i>sexta</i> antennae at different days after pupation (animals eclose at pupal stage 18) show that Orco is strongly expressed at early pupal stages 1–8 and in the adult antenna. However, Orco-immunoreactive protein is hardly detectable at pupal stages 10–17 while two lower molecular weight bands are recognized by the anti-Orco antiserum (stars). We do not know, whether these lower molecular weight proteins are splice variants, degradation products of Orco, or unrelated proteins. As a control, tissues from optic lobes and muscles were used, which show no signal near the expected molecular weight of MsexOrco. A adult male antennae, kDa kilodalton, M muscle tissue of adult males; OL optic lobes.</p

Statistics for current injection experiments.

<p>Statistics for current injection experiments.</p