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Background: In view of demographic change, rising retirement age, and a growing shortage of skilled workers, it is increasingly important to prevent widespread diseases such as type 2 diabetes or its risk factor metabolic syndrome. Since the workplace is an important setting for preventive measures and little is known about incident cases in the working population, the aim of this study was to identify vulnerable occupational groups for whom these interventions are particularly appropriate. Therefore, we investigated the 10-year incidence of type 2 diabetes and metabolic syndrome across occupational groups in Germany., Methods: Employees of the population-based Gutenberg-Health-Study (GHS) were examined at baseline (2007-2012) and 10 years later. We calculated age- and sex-standardised incidence rates and standardised incidence ratios (SIR) with a 95% confidence interval (CI) for occupations, job complexity levels, and supervisory and managerial positions. 5954 persons at risk for type 2 diabetes and 5103 at risk for metabolic syndrome were observed., Results: Between baseline and follow-up, 388 cases of type 2 diabetes and 1104 cases of metabolic syndrome occurred, and standardised incidences were 6.9% and 22.6%, respectively. The highest incidence of type 2 diabetes was observed in the occupational group "food production and processing" (20.7%) with a threefold increased incidence (SIR = 3.0, 95% CI 1.8-4.7) compared to the total working population of the GHS. Employees in "metal production, processing and construction" had the highest incidence of metabolic syndrome and a two times higher SIR (48.5%; SIR = 2.1, 95% CI 1.4-2.9). There was also a high incidence of both type 2 diabetes and metabolic syndrome in "cleaners" (16.5% and 34.8%) and "drivers and mobile plant operators" (14.8% and 41.2%). An increased incidence of type 2 diabetes and metabolic syndrome was observed with decreasing job complexity levels., Conclusions: This study shows wide differences in the incidence of type 2 diabetes and metabolic syndrome between occupational groups and highlights the vulnerability of certain occupations. As the workplace is an important platform for interventions, the findings of this study could guide the development of more nuanced and effective workplace health initiatives to promote a healthier workforce for the future., Competing Interests: Declarations. Ethics approval and consent to participate: The local ethics committee (Ethics Commission of the State Chamber of Physicians of Rhineland-Palatine) (#837.020.07(5555)) and the local federal data safety commissioners approved the study in full compliance with the Declaration of Helsinki. Written informed consent was obtained from all participants. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Channelrhodopsins (ChRs) are light-gated ion channels widely used to optically activate or silence selected electrogenic cells, such as individual brain neurons. Here, we describe identifying and characterizing a set of anion-conducting ChRs (ACRs) from diverse taxa and representing various branches of the ChR phylogenetic tree. The Mantoniella squamata ACR (MsACR1) showed high sensitivity to yellow-green light ( λ max at 555 nm) and was further engineered for optogenetic applications. A single amino-acid substitution that mimicked red-light-sensitive rhodopsins like Chrimson shifted the photosensitivity 20 nm toward red light and accelerated photocurrent kinetics. Hence, it was named red and accelerated ACR, raACR. Both wild-type and mutant are capable optical silencers at low light intensities in mouse neurons in vitro and in vivo , while raACR offers a higher temporal resolution., Competing Interests: JO, AR, TF, CG, RP, OB, MP, PH No competing interests declared, (© 2023, Oppermann et al.)

The model plant Physcomitrium patens has played a pivotal role in enhancing our comprehension of plant evolution and development. However, the current genome harbours numerous regions that remain unfinished and erroneous. To address these issues, we generated an assembly using Oxford Nanopore reads and Hi-C mapping. The assembly incorporates telomeric and centromeric regions, thereby establishing it as a near telomere-to-telomere genome except a region in chromosome 1 that is not fully assembled due to its highly repetitive nature. This near telomere-to-telomere genome resolves the chromosome number at 26 and provides a gap-free genome assembly as well as updated gene models to aid future studies using this model organism., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Imaging of living synapses has relied for over two decades on the overexpression of synaptic proteins fused to fluorescent reporters. This strategy alters the stoichiometry of synaptic components and ultimately affects synapse physiology. To overcome these limitations, here a nanobody is presented that binds the calcium sensor synaptotagmin-1 (NbSyt1). This nanobody functions as an intrabody (iNbSyt1) in living neurons and is minimally invasive, leaving synaptic transmission almost unaffected, as suggested by the crystal structure of the NbSyt1 bound to Synaptotagmin-1 and by the physiological data. Its single-domain nature enables the generation of protein-based fluorescent reporters, as showcased here by measuring spatially localized presynaptic Ca 2+ with a NbSyt1- jGCaMP8 chimera. Moreover, the small size of NbSyt1 makes it ideal for various super-resolution imaging methods. Overall, NbSyt1 is a versatile binder that will enable imaging in cellular and molecular neuroscience with unprecedented precision across multiple spatiotemporal scales., (© 2023 The Authors. Small Methods published by Wiley-VCH GmbH.)

Focal cortical epilepsies are frequently refractory to available anticonvulsant drug therapies. One key factor contributing to this state is the limited availability of animal models that allow to reliably study focal cortical seizures and how they recruit surrounding brain areas in vivo. In this study, we selectively expressed the inhibitory chemogenetic receptor, hM4D, in GABAergic neurons in focal cortical areas using viral gene transfer. GABAergic silencing using Clozapine-N-Oxide (CNO) demonstrated reliable induction of local epileptiform events in the electroencephalogram signal of awake freely moving mice. Anesthetized mice experiments showed consistent induction of focal epileptiform-events in both the barrel cortex (BC) and the medial prefrontal cortex (mPFC), accompanied by high-frequency oscillations, a known characteristic of human seizures. Epileptiform-events showed propagation indication with favored propagation pathways: from the BC on 1 hemisphere to its counterpart and from the BC to the mPFC, but not vice-versa. Lastly, sensory whisker-pad stimulation evoked BC epileptiform events post-CNO, highlighting the potential use of this model in studying sensory-evoked seizures. Combined, our results show that targeted chemogenetic inhibition of GABAergic neurons using hM4D can serve as a novel, versatile, and reliable model of focal cortical epileptic activity suitable for systematically studying cortical ictogenesis in different cortical areas., (© The Author(s) 2022. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The Locus coeruleus (LC) modulates various neuronal circuits throughout the brain. Its unique architectural organization encompasses a net of axonal innervation that spans the entire brain, while its somatic core is highly compact. Recent research revealed an unexpected cellular input specificity within the nucleus that can give rise to various network states that either broadcast norepinephrine signals throughout the brain or pointedly modulate specific brain areas. Such adaptive input-output functions likely surpass our existing network models that build upon a given synaptic wiring configuration between neurons. As the distances between noradrenergic neurons in the core of the LC are unusually small, neighboring neurons could theoretically impact each other via volume transmission of NE. We therefore set out to investigate if such interaction could be mediated through noradrenergic alpha2-receptors in a spiking neuron model of the LC. We validated our model of LC neurons through comparison with experimental patch-clamp data and identified key variables that impact alpha2-mediated inhibition of neighboring LC neurons. Our simulation confirmed a reliable autoinhibition of LC neurons after episodes of high neuronal activity that continue even after neuronal activity subsided. Additionally, dendro-somatic synapses inhibited spontaneous spiking in the somatic compartment of connected neurons in our model. We determined the exact position of hundreds of LC neurons in the mouse brain stem via a tissue clearing approach and, based on this, further determined that 25 percent of noradrenergic neurons have a neighboring LC neuron within less than a 25-micrometer radius. By modeling NE diffusion, we estimated that more than 15 percent of the alpha2-adrenergic receptors fraction can bind NE within such a diffusion radius. Our spiking neuron model of LC neurons predicts that repeated or long-lasting episodes of high neuronal activity induce partitioning of the gross LC network and reduce the spike rate in neighboring neurons at distances smaller than 25 μm. As these volume-mediating neighboring effects are challenging to test with the current methodology, our findings can guide future experimental approaches to test this phenomenon and its physiological consequences.

Objectives: Sedentary behaviour is a modifiable risk factor for cardiovascular health. Although long periods of sedentary behaviour take place at work, evidence of the relationship between such occupational sitting and cardiometabolic health risks remains limited. This systematic review aimed to update the evidence on the associations of occupational sitting with cardiovascular outcomes and cardiometabolic risk factors based on longitudinal studies., Design: Systematic review., Setting: Workplace., Population: Employees aged 18-65 years., Primary and Secondary Outcomes: Primary outcomes were cardiovascular diseases and cardiometabolic risk markers. The secondary outcome was all-cause mortality., Data Sources: Ten databases, including PubMed, Web of Science and CINAHL (search January 2018, updated February 2019)., Data Extraction and Synthesis: Data were screened, extracted and appraised by three independent reviewers following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines., Results: Studies were markedly heterogeneous in terms of measurement of occupational sitting, cardiometabolic risk factors and cardiovascular morbidity and mortality, so that standards were hardly identifiable and limiting the value of the evidence. The review included 27 high or acceptable quality publications. Of the eight high-quality publications from seven cohorts, three cohort studies found significant associations of occupational sitting with primary outcomes. Additionally, one study described an association with the secondary outcome. Another high-quality publication found an association between occupational sitting and ischaemic heart disease in a subgroup already at risk due to hypertension. For sex/gender analysis, 11 of the 27 high and acceptable quality publications reported sex-stratified results. Five of these found sex differences., Conclusions: Evidence regarding the association of occupational sitting with cardiometabolic health risks was limited because of the lack of standardised measurements for occupational sitting. Occupational sitting combined with an overall sedentary lifestyle was associated with an elevated relative risk for several cardiometabolic outcomes. There is an urgent need for standardised measurements of occupational sitting to facilitate meta-analysis. Sex/gender aspects of this relationship require further investigation., Competing Interests: Competing interests: TK reports having contributed to an advisory board of CoLucid and a research project funded by Amgen, for which the Charité Universitätsmedizin Berlin received unrestricted compensation. He further reports having received honoraria from Lilly, Newsenselab and Total for providing methodological advice, from Novartis and from Daiichi Sankyo for providing a lecture on neuroepidemiology and research methods, and from the BMJ for editorial services. All other authors declare no competing interests., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Given its non-invasive nature, there is increasing interest in the use of transcutaneous vagus nerve stimulation (tVNS) across basic, translational and clinical research. Contemporaneously, tVNS can be achieved by stimulating either the auricular branch or the cervical bundle of the vagus nerve, referred to as transcutaneous auricular vagus nerve stimulation(VNS) and transcutaneous cervical VNS, respectively. In order to advance the field in a systematic manner, studies using these technologies need to adequately report sufficient methodological detail to enable comparison of results between studies, replication of studies, as well as enhancing study participant safety. We systematically reviewed the existing tVNS literature to evaluate current reporting practices. Based on this review, and consensus among participating authors, we propose a set of minimal reporting items to guide future tVNS studies. The suggested items address specific technical aspects of the device and stimulation parameters. We also cover general recommendations including inclusion and exclusion criteria for participants, outcome parameters and the detailed reporting of side effects. Furthermore, we review strategies used to identify the optimal stimulation parameters for a given research setting and summarize ongoing developments in animal research with potential implications for the application of tVNS in humans. Finally, we discuss the potential of tVNS in future research as well as the associated challenges across several disciplines in research and clinical practice., Competing Interests: EK and SK are employed by company SzeleSTIM GmbH. JS received honoraria from SzeleSTIM GmbH and owns patents in the field of the auricular vagus nerve stimulation. EK, SK, and JS are shareholders of SzeleSTIM GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer TS declared a shared affiliation, with no collaboration, with one of the authors HJ, to the handling editor at the time of review., (Copyright © 2021 Farmer, Strzelczyk, Finisguerra, Gourine, Gharabaghi, Hasan, Burger, Jaramillo, Mertens, Majid, Verkuil, Badran, Ventura-Bort, Gaul, Beste, Warren, Quintana, Hämmerer, Freri, Frangos, Tobaldini, Kaniusas, Rosenow, Capone, Panetsos, Ackland, Kaithwas, O'Leary, Genheimer, Jacobs, Van Diest, Schoenen, Redgrave, Fang, Deuchars, Széles, Thayer, More, Vonck, Steenbergen, Vianna, McTeague, Ludwig, Veldhuizen, De Couck, Casazza, Keute, Bikson, Andreatta, D'Agostini, Weymar, Betts, Prigge, Kaess, Roden, Thai, Schuster, Montano, Hansen, Kroemer, Rong, Fischer, Howland, Sclocco, Sellaro, Garcia, Bauer, Gancheva, Stavrakis, Kampusch, Deuchars, Wehner, Laborde, Usichenko, Polak, Zaehle, Borges, Teckentrup, Jandackova, Napadow and Koenig.)

Correct brain wiring depends on reliable synapse formation. Nevertheless, signaling codes promoting synaptogenesis are not fully understood. Here, we report a spinogenic mechanism that operates during neuronal development and is based on the interaction of tumor necrosis factor receptor-associated factor 6 (TRAF6) with the synaptic cell adhesion molecule neuroplastin. The interaction between these proteins was predicted in silico and verified by co-immunoprecipitation in extracts from rat brain and co-transfected HEK cells. Binding assays show physical interaction between neuroplastin's C-terminus and the TRAF-C domain of TRAF6 with a K d value of 88 μM. As the two proteins co-localize in primordial dendritic protrusions, we used young cultures of rat and mouse as well as neuroplastin-deficient mouse neurons and showed with mutagenesis, knock-down, and pharmacological blockade that TRAF6 is required by neuroplastin to promote early spinogenesis during in vitro days 6-9, but not later. Time-framed TRAF6 blockade during days 6-9 reduced mEPSC amplitude, number of postsynaptic sites, synapse density and neuronal activity as neurons mature. Our data unravel a new molecular liaison that may emerge during a specific window of the neuronal development to determine excitatory synapse density in the rodent brain., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Vemula, Malci, Junge, Lehmann, Rama, Hradsky, Matute, Weber, Prigge, Naumann, Kreutz, Seidenbecher, Gundelfinger and Herrera-Molina.)

Background: Overweight, obesity and the conditions resulting from them have become one of the major challenges for health systems all over the world. High dropout rates are particularly common among the participants in outpatient obesity intervention programmes (OIP). Limited research has examined retention in OIP., Methods: Based on the data of a 24-month Bundeswehr outpatient OIP with a total of 630 participants, predisposing factors (n = 30) for continued participation beyond an early stage (0-3 months) and beyond a later stage (0-6 months) were analysed by means of a logistic regression analysis. In order to correct for multiple-comparison, the p-value was adjusted (p* < 0.0017)., Results: Three hundred out of 630 participants continued to participate beyond an early stage and 205 beyond a later stage. Besides an age between 40 and 50 at the beginning of the outpatient OIP, it was possible to show that knowledge of one's blood pressure and a positive lifestyle prior to participation in the programme (higher level of sporting activity) were predisposing factors for early dropout (>3 months) in the intervention programme., Discussion: The possible predisposing factors examined accounted for about 35% of the variance in a rough estimate. In order to improve long-term participation in workplace outpatient OIP, the study focused on how the motivation of older people and employees with an unfavourable cardiovascular risk profile could be improved., Competing Interests: Sammito is an active Bundeswehr Medical Service officer and works for the Federal Ministry of Defence. The other authors are not affected by conflicts of interest., (© 2020 Published by Elsevier Inc.)

Holistic approaches to workplace health promotion (WHP) within the military setting are challenging. In 2015, the German Ministry of Defense initiated a 6-month pilot study of WHP in the Federal Armed Forces. The pilot study was to identify organizational challenges that should be addressed before the Ministry implemented a comprehensive occupational health management policy in all departments. Eleven diverse departments were selected to participate in a WHP program that addressed physical activity, diet, stress management and addiction prevention. As part of the evaluation concept, we interviewed coordinators, and department heads focusing on transfer factors from the perspective of the implementers. All coordinators and their department heads or deputies participated in semi-structured face-to-face on-site interviews. The data were analyzed based on qualitative content analysis. The coordinators (officers with sports science degree) seemed fully prepared and capable to master the new task. They experienced difficulties in adapting WHP activities to local structures and needs, and complications in administering modular activities. Department heads described conflict regarding human resources between the military mission and the implementation of WHP. Commitment to WHP was a strong facilitator. The interviews identified various barriers related to support by middle management (supervisors) and specific work conditions (e.g. shift work). If occupational health management is to be successfully implemented on a large scale, conceptional and practically collaboration is necessary between WHP and occupational safety and health, and organization and leadership, respectively. Supervisors will benefit from open communication about compensation for the release time of their subordinates to attend WHP.

Actively transcribed genes adopt a unique chromatin environment with characteristic patterns of enrichment. Within gene bodies, H3K36me3 and cytosine DNA methylation are elevated at exons of spliced genes and have been implicated in the regulation of pre-mRNA splicing. H3K36me3 is further responsive to splicing, wherein splicing inhibition led to a redistribution and general reduction over gene bodies. In contrast, little is known of the mechanisms supporting elevated DNA methylation at actively spliced genic locations. Recent evidence associating the de novo DNA methyltransferase Dnmt3b with H3K36me3-rich chromatin raises the possibility that genic DNA methylation is influenced by splicing-associated H3K36me3. Here, we report the generation of an isogenic resource to test the direct impact of splicing on chromatin. A panel of minigenes of varying splicing potential were integrated into a single FRT site for inducible expression. Profiling of H3K36me3 confirmed the established relationship to splicing, wherein levels were directly correlated with splicing efficiency. In contrast, DNA methylation was equivalently detected across the minigene panel, irrespective of splicing and H3K36me3 status. In addition to revealing a degree of independence between genic H3K36me3 and DNA methylation, these findings highlight the generated minigene panel as a flexible platform for the query of splicing-dependent chromatin modifications., (Published by Oxford University Press on behalf of Nucleic Acids Research 2017.)

Strigolactones (SLs) are well known for their role in repressing shoot branching. In pea, increased transcript levels of SL biosynthesis genes are observed in stems of highly branched SL deficient (ramosus1 (rms1) and rms5) and SL response (rms3 and rms4) mutants indicative of negative feedback control. In contrast, the highly branched rms2 mutant has reduced transcript levels of SL biosynthesis genes. Grafting studies and hormone quantification led to a model where RMS2 mediates a shoot-to-root feedback signal that regulates both SL biosynthesis gene transcript levels and xylem sap levels of cytokinin exported from roots. Here we cloned RMS2 using synteny with Medicago truncatula and demonstrated that it encodes a putative auxin receptor of the AFB4/5 clade. Phenotypes similar to rms2 were found in Arabidopsis afb4/5 mutants, including increased shoot branching, low expression of SL biosynthesis genes and high auxin levels in stems. Moreover, afb4/5 and rms2 display a specific resistance to the herbicide picloram. Yeast-two-hybrid experiments supported the hypothesis that the RMS2 protein functions as an auxin receptor. SL root feeding using hydroponics repressed auxin levels in stems and down-regulated transcript levels of auxin biosynthesis genes within one hour. This auxin down-regulation was also observed in plants treated with the polar auxin transport inhibitor NPA. Together these data suggest a homeostatic feedback loop in which auxin up-regulates SL synthesis in an RMS2-dependent manner and SL down-regulates auxin synthesis in an RMS3 and RMS4-dependent manner.

Reversible silencing of neuronal activity is a powerful approach for isolating the roles of specific neuronal populations in circuit dynamics and behavior. In contrast with neuronal excitation, for which the majority of studies have used a limited number of optogenetic and chemogenetic tools, the number of genetically encoded tools used for inhibition of neuronal activity has vastly expanded. Silencing strategies vary widely in their mechanism of action and in their spatial and temporal scales. Although such manipulations are commonly applied, the design and interpretation of neuronal silencing experiments present unique challenges, both technically and conceptually. Here, we review the most commonly used tools for silencing neuronal activity and provide an in-depth analysis of their mechanism of action and utility for particular experimental applications. We further discuss the considerations that need to be given to experimental design, analysis, and interpretation of collected data. Finally, we discuss future directions for the development of new silencing approaches in neuroscience., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Sodium pumping rhodopsins (NaRs) are a unique member of the microbial-type I rhodopsin family which actively transport Na+ and H+ depending on ionic condition. In this study, we surveyed 12 different NaRs from various sources of eubacteria for their electrophysiological as well as spectroscopic properties. In mammalian cells several of these NaRs exhibited a Na+ based pump photocurrent and four interesting candidates were chosen for further characterization. Voltage dependent photocurrent amplitudes revealed a membrane potential-sensitive turnover rate, indicating the presence of an electrically-charged intermediate(s) in the photocycle reaction. The NaR from Salinarimonas rosea DSM21201 exhibited a red-shifted absorption spectrum, and slower kinetics compared to the first described sodium pump, KR2. Although the ratio of Na+ to H+ ion transport varied among the NaRs we tested, the NaRs from Flagellimonas sp&#95;DIK and Nonlabens sp&#95;YIK&#95;SED-11 showed significantly higher Na+ selectivity when compared to KR2. All four further investigated NaRs showed a functional expression in dissociated hippocampal neuron culture and hyperpolarizing activity upon light-stimulation. Additionally, all four NaRs allowed optical inhibition of electrically-evoked neuronal spiking. Although efficiency of silencing was 3-5 times lower than silencing with the enhanced version of the proton pump AR3 from Halorubrum sodomense, our data outlines a new approach for hyperpolarization of excitable cells without affecting the intracellular and extracellular proton environment.

Key mitochondrial functions such as ATP production, Ca 2+ uptake and release, and substrate accumulation depend on the proton electrochemical gradient (ΔμH + ) across the inner membrane. Although several drugs can modulate ΔμH + , their effects are hardly reversible, and lack cellular specificity and spatial resolution. Although channelrhodopsins are widely used to modulate the plasma membrane potential of excitable cells, mitochondria have thus far eluded optogenetic control. Here we describe a toolkit of optometabolic constructs based on selective targeting of channelrhodopsins with distinct functional properties to the inner mitochondrial membrane of intact cells. We show that our strategy enables a light-dependent control of the mitochondrial membrane potential (Δψ m ) and coupled mitochondrial functions such as ATP synthesis by oxidative phosphorylation, Ca 2+ dynamics, and respiratory metabolism. By directly modulating Δψ m , the mitochondria-targeted opsins were used to control complex physiological processes such as spontaneous beats in cardiac myocytes and glucose-dependent ATP increase in pancreatic β-cells. Furthermore, our optometabolic tools allow modulation of mitochondrial functions in single cells and defined cell regions., Competing Interests: The authors declare no conflict of interest.

Fear-related disorders are thought to reflect strong and persistent fear memories. The basolateral amygdala (BLA) and the medial prefrontal cortex (mPFC) form strong reciprocal synaptic connections that play a key role in acquisition and extinction of fear memories. While synaptic contacts of BLA cells onto mPFC neurons are likely to play a crucial role in this process, the BLA connects with several additional nuclei within the fear circuit that could relay fear-associated information to the mPFC, and the contribution of direct monosynaptic BLA-mPFC inputs is not yet clear. Here we establish an optogenetic stimulation protocol that induces synaptic depression in BLA-mPFC synapses. In behaving mice, optogenetic high-frequency stimulation of BLA inputs to mPFC interfered with retention of cued associations, attenuated previously acquired cue-associated responses in mPFC neurons and facilitated extinction. Our findings demonstrate the contribution of BLA inputs to mPFC in forming and maintaining cued fear associations.

The complexity and heterogeneity of neuroimaging findings in individuals with autism spectrum disorder has suggested that many of the underlying alterations are subtle and involve many brain regions and networks. The ability to account for multivariate brain features and identify neuroimaging measures that can be used to characterize individual variation have thus become increasingly important for interpreting and understanding the neurobiological mechanisms of autism. In the present study, we utilize the Mahalanobis distance, a multidimensional counterpart of the Euclidean distance, as an informative index to characterize individual brain variation and deviation in autism. Longitudinal diffusion tensor imaging data from 149 participants (92 diagnosed with autism spectrum disorder and 57 typically developing controls) between 3.1 and 36.83 years of age were acquired over a roughly 10-year period and used to construct the Mahalanobis distance from regional measures of white matter microstructure. Mahalanobis distances were significantly greater and more variable in the autistic individuals as compared to control participants, demonstrating increased atypicalities and variation in the group of individuals diagnosed with autism spectrum disorder. Distributions of multivariate measures were also found to provide greater discrimination and more sensitive delineation between autistic and typically developing individuals than conventional univariate measures, while also being significantly associated with observed traits of the autism group. These results help substantiate autism as a truly heterogeneous neurodevelopmental disorder, while also suggesting that collectively considering neuroimaging measures from multiple brain regions provides improved insight into the diversity of brain measures in autism that is not observed when considering the same regions separately. Distinguishing multidimensional brain relationships may thus be informative for identifying neuroimaging-based phenotypes, as well as help elucidate underlying neural mechanisms of brain variation in autism spectrum disorders.

We investigated the efficacy of optogenetic inhibition at presynaptic terminals using halorhodopsin, archaerhodopsin and chloride-conducting channelrhodopsins. Precisely timed activation of both archaerhodopsin and halorhodpsin at presynaptic terminals attenuated evoked release. However, sustained archaerhodopsin activation was paradoxically associated with increased spontaneous release. Activation of chloride-conducting channelrhodopsins triggered neurotransmitter release upon light onset. Thus, the biophysical properties of presynaptic terminals dictate unique boundary conditions for optogenetic manipulation., Competing Interests: The authors declare no competing financial interests.

After the discovery of Channelrhodopsin, a light-gated ion channel, only a few people saw the diverse range of applications for such a protein. Now, more than 10 years later Channelrhodopsins have become widely accepted as the ultimate tool to control the membrane potential of excitable cells via illumination. The demand for more application-specific Channelrhodopsin variants started a race between protein engineers to design improved variants. Even though many engineered variants have undisputable advantages compared to wild-type variants, many users are alienated by the tremendous amount of new variants and their perplexing names. Here, we review new variants whose efficacy has already been proven in neurophysiological experiments, or variants which are likely to extend the optogenetic toolbox. Variants are described based on their mechanistic and operational properties in terms of expression, kinetics, ion selectivity, and wavelength responsivity.

It is commonly assumed, but has rarely been demonstrated, that sex differences in behaviour arise from sexual dimorphism in the underlying neural circuits. Parental care is a complex stereotypic behaviour towards offspring that is shared by numerous species. Mice display profound sex differences in offspring-directed behaviours. At their first encounter, virgin females behave maternally towards alien pups while males will usually ignore the pups or attack them. Here we show that tyrosine hydroxylase (TH)-expressing neurons in the anteroventral periventricular nucleus (AVPV) of the mouse hypothalamus are more numerous in mothers than in virgin females and males, and govern parental behaviours in a sex-specific manner. In females, ablating the AVPV TH(+) neurons impairs maternal behaviour whereas optogenetic stimulation or increased TH expression in these cells enhance maternal care. In males, however, this same neuronal cluster has no effect on parental care but rather suppresses inter-male aggression. Furthermore, optogenetic activation or increased TH expression in the AVPV TH(+) neurons of female mice increases circulating oxytocin, whereas their ablation reduces oxytocin levels. Finally, we show that AVPV TH(+) neurons relay a monosynaptic input to oxytocin-expressing neurons in the paraventricular nucleus. Our findings uncover a previously unknown role for this neuronal population in the control of maternal care and oxytocin secretion, and provide evidence for a causal relationship between sexual dimorphism in the adult brain and sex differences in parental behaviour.

Plant genomes encode large numbers of F-box proteins (FBPs), the substrate recognition subunit of SKP1-CULLIN-F-box (SCF) ubiquitin ligases. There are ~700 FBPs in Arabidopsis , most of which are uncharacterized. TIR1 is among the best-studied plant FBPs and functions as a receptor for the plant hormone auxin. Here we use a yeast two-hybrid system to identify novel TIR1 mutants with altered properties. The analysis of these mutants reveals that TIR1 associates with the CULLIN1 (CUL1) subunit of the SCF through the N-terminal H1 helix of the F-box domain. Mutations that untether TIR1 from CUL1 stabilize the FBP and cause auxin resistance and associated growth defects, probably by protecting TIR1 substrates from degradation. Based on these results we propose that TIR1 is subject to autocatalytic degradation when assembled into an SCF. Further, our results suggest a general method for determining the physiological function of uncharacterized FBPs. Finally, we show that a key amino acid variation in the F-box domain of auxin signalling F-box (AFB1), a closely related FBP, reduces its ability to form an SCF, resulting in an increase in AFB1 levels.

The brain is a large network of interconnected neurons where each cell functions as a nonlinear processing element. Unraveling the mysteries of information processing in the complex networks of the brain requires versatile neurostimulation and imaging techniques. Optogenetics is a new stimulation method which allows the activity of neurons to be modulated by light. For this purpose, the cell-types of interest are genetically targeted to produce light-sensitive proteins. Once these proteins are expressed, neural activity can be controlled by exposing the cells to light of appropriate wavelengths. Optogenetics provides a unique combination of features, including multimodal control over neural function and genetic targeting of specific cell-types. Together, these versatile features combine to a powerful experimental approach, suitable for the study of the circuitry of psychiatric and neurological disorders. The advent of optogenetics was followed by extensive research aimed to produce new lines of light-sensitive proteins and to develop new technologies: for example, to control the distribution of light inside the brain tissue or to combine optogenetics with other modalities including electrophysiology, electrocorticography, nonlinear microscopy, and functional magnetic resonance imaging. In this paper, the authors review some of the recent advances in the field of optogenetics and related technologies and provide their vision for the future of the field.

The phytohormone auxin regulates virtually every aspect of plant development. The hormone directly mediates the interaction between the two members of the auxin coreceptor complex, a TRANSPORT INHIBITOR RESPONSE (TIR1)/AUXIN SIGNALING F-BOX protein and an AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) transcriptional repressor. To learn more about the interaction between these proteins, a mutant screen was performed using the yeast (Saccharomyces cerevisiae) two-hybrid system in Arabidopsis (Arabidopsis thaliana). Two tir1 mutations were identified that increased interaction with Aux/IAAs. The D170E and M473L mutations increase affinity between TIR1 and the degron motif of Aux/IAAs and enhance the activity of the SCF(TIR1) complex. This resulted in faster degradation of Aux/IAAs and increased transcription of auxin-responsive genes in the plant. Plants carrying the pTIR1:tir1 D170E/M473L-Myc transgene exhibit diverse developmental defects during plant growth and display an auxin-hypersensitive phenotype. This work demonstrates that changes in the leucine-rich repeat domain of the TIR1 auxin coreceptor can alter the properties of SCF(TIR1).

Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Here we describe red, single-wavelength GECIs, "RCaMPs," engineered from circular permutation of the thermostable red fluorescent protein mRuby. High-resolution crystal structures of mRuby, the red sensor RCaMP, and the recently published red GECI R-GECO1 give insight into the chromophore environments of the Ca(2+)-bound state of the sensors and the engineered protein domain interfaces of the different indicators. We characterized the biophysical properties and performance of RCaMP sensors in vitro and in vivo in Caenorhabditis elegans, Drosophila larvae, and larval zebrafish. Further, we demonstrate 2-color calcium imaging both within the same cell (registering mitochondrial and somatic [Ca(2+)]) and between two populations of cells: neurons and astrocytes. Finally, we perform integrated optogenetics experiments, wherein neural activation via channelrhodopsin-2 (ChR2) or a red-shifted variant, and activity imaging via RCaMP or GCaMP, are conducted simultaneously, with the ChR2/RCaMP pair providing independently addressable spectral channels. Using this paradigm, we measure calcium responses of naturalistic and ChR2-evoked muscle contractions in vivo in crawling C. elegans. We systematically compare the RCaMP sensors to R-GECO1, in terms of action potential-evoked fluorescence increases in neurons, photobleaching, and photoswitching. R-GECO1 displays higher Ca(2+) affinity and larger dynamic range than RCaMP, but exhibits significant photoactivation with blue and green light, suggesting that integrated channelrhodopsin-based optogenetics using R-GECO1 may be subject to artifact. Finally, we create and test blue, cyan, and yellow variants engineered from GCaMP by rational design. This engineered set of chromatic variants facilitates new experiments in functional imaging and optogenetics.

Channelrhodopsin-2 is a light-gated ion channel and a major tool of optogenetics. It is used to control neuronal activity via blue light. Here we describe the construction of color-tuned high efficiency channelrhodopsins (ChRs), based on chimeras of Chlamydomonas channelrhodopsin-1 and Volvox channelrhodopsin-1. These variants show superb expression and plasma membrane integration, resulting in 3-fold larger photocurrents in HEK cells compared with channelrhodopsin-2. Further molecular engineering gave rise to chimeric variants with absorption maxima ranging from 526 to 545 nm, dovetailing well with maxima of channelrhodopsin-2 derivatives ranging from 461 to 492 nm. Additional kinetic fine-tuning led to derivatives in which the lifetimes of the open state range from 19 ms to 5 s. Finally, combining green- with blue-absorbing variants allowed independent activation of two distinct neural cell populations at 560 and 405 nm. This novel panel of channelrhodopsin variants may serve as an important toolkit element for dual-color cell stimulation in neural circuits.

The C. elegans nervous system is particularly well suited for optogenetic analyses of circuit function: Essentially all connections have been mapped, and light can be directed at the neuron of interest in the freely moving, transparent animals, while behavior is observed. Thus, different nodes of a neuronal network can be probed for their role in controlling a particular behavior, using different optogenetic tools for photo-activation or -inhibition, which respond to different colors of light. As neurons may act in concert or in opposing ways to affect a behavior, one would further like to excite these neurons concomitantly, yet independent of each other. In addition to the blue-light activated Channelrhodopsin-2 (ChR2), spectrally red-shifted ChR variants have been explored recently. Here, we establish the green-light activated ChR chimera C1V1 (from Chlamydomonas and Volvox ChR1's) for use in C. elegans. We surveyed a number of red-shifted ChRs, and found that C1V1-ET/ET (E122T; E162T) works most reliable in C. elegans, with 540-580 nm excitation, which leaves ChR2 silent. However, as C1V1-ET/ET is very light sensitive, it still becomes activated when ChR2 is stimulated, even at 400 nm. Thus, we generated a highly efficient blue ChR2, the H134R; T159C double mutant (ChR2-HR/TC). Both proteins can be used in the same animal, in different neurons, to independently control each cell type with light, enabling a further level of complexity in circuit analyses.

The capture and utilization of light is an exquisitely evolved process. The single-component microbial opsins, although more limited than multicomponent cascades in processing, display unparalleled compactness and speed. Recent advances in understanding microbial opsins have been driven by molecular engineering for optogenetics and by comparative genomics. Here we provide a Primer on these light-activated ion channels and pumps, describe a group of opsins bridging prior categories, and explore the convergence of molecular engineering and genomic discovery for the utilization and understanding of these remarkable molecular machines., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Severe behavioural deficits in psychiatric diseases such as autism and schizophrenia have been hypothesized to arise from elevations in the cellular balance of excitation and inhibition (E/I balance) within neural microcircuitry. This hypothesis could unify diverse streams of pathophysiological and genetic evidence, but has not been susceptible to direct testing. Here we design and use several novel optogenetic tools to causally investigate the cellular E/I balance hypothesis in freely moving mammals, and explore the associated circuit physiology. Elevation, but not reduction, of cellular E/I balance within the mouse medial prefrontal cortex was found to elicit a profound impairment in cellular information processing, associated with specific behavioural impairments and increased high-frequency power in the 30-80 Hz range, which have both been observed in clinical conditions in humans. Consistent with the E/I balance hypothesis, compensatory elevation of inhibitory cell excitability partially rescued social deficits caused by E/I balance elevation. These results provide support for the elevated cellular E/I balance hypothesis of severe neuropsychiatric disease-related symptoms.

Vascular plants appeared ~410 million years ago, then diverged into several lineages of which only two survive: the euphyllophytes (ferns and seed plants) and the lycophytes. We report here the genome sequence of the lycophyte Selaginella moellendorffii (Selaginella), the first nonseed vascular plant genome reported. By comparing gene content in evolutionarily diverse taxa, we found that the transition from a gametophyte- to a sporophyte-dominated life cycle required far fewer new genes than the transition from a nonseed vascular to a flowering plant, whereas secondary metabolic genes expanded extensively and in parallel in the lycophyte and angiosperm lineages. Selaginella differs in posttranscriptional gene regulation, including small RNA regulation of repetitive elements, an absence of the trans-acting small interfering RNA pathway, and extensive RNA editing of organellar genes.

Channelrhodopsin-2 (ChR2) is a light-gated ion channel that is successfully used in neurosciences to depolarize cells with blue light. In this regard control of membrane voltage with light opens new perspectives for the characterization of ion channels and the search for inhibitors or modulators. Here, we report a control of membrane potential with ChR2 and the potassium channel mTrek for the purpose of screening for ion channel specific drugs. To verify principle we have chosen the voltage gated calcium channel Ca(V)3.2 as potential drug target. For this purpose we transfected the ChR2 gene into a HEK293T-cell line that permanently expresses Ca(V)3.2 and the K-channel mTrek. The resting potential was adjusted with low concentration of extracellular potassium ions whereas transient depolarization was achieved by activation of ChR2 with short pulses of blue light. Calcium ion influx through Ca(V)3.2 was monitored by observing fura-2 fluorescence. This approach allowed a repetitive activation of Ca(V)3.2. The Ca(2+) influx was specifically blocked by the inhibitor mibefradil. Since this assay is genetically-encoded, it may be employed for a variety of voltage-gated calcium channels and should be applicable to multi-well reader formats for high-throughput screening.