Bimodal activation of different neuron classes with the spectrally red-shifted channelrhodopsin chimera C1V1 in Caenorhabditis elegans

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

A Cell Line for Detection of Botulinum Neurotoxin Type B

Botulinum neurotoxins (BoNTs) type A and type B are commonly used as biopharmaceutics for neurological diseases, uniquely allowing months-long paralysis of target muscles. Their exquisite neuronal specificity is conferred by a multistep process of binding, internalization, cytosolic escape and cleavage of the neuron-specific proteins, SNAP-25 and vesicle-associated membrane proteins (VAMPs), ultimately to inhibit secretion of neurotransmitters. Currently the mouse lethality bioassay is the only available method for quality control testing of VAMP-cleaving botulinum products. Refined assays for botulinum product testing are urgently needed. Specifically, in vitro replacement assays which can account for all steps of BoNT intoxication are in high demand. Here, we describe a novel SiMa cell-based approach where re-engineering of the VAMP molecule allows detection of all BoNT/B intoxication steps using a luminescent enzymatic reaction with sensitivity comparable to mouse LD50 bioassay. The presented one-step enzyme-linked immunosorbent assay meets 3Rs (replacement, reduction, and refinement of the use of animals) objectives, is user-friendly and will accelerate development of new botulinum drugs. The sensitive enzymatic reporter cell line could also be adapted for the detection of toxin activity during the manufacture of botulinum and tetanus vaccines

Raumordnung des Untergrundes

Neuartige und intensivierte Nutzungen des Untergrunds führen zu dem Erfordernis einer unterirdischen Raumordnung, die auf der Grundlage des geltenden Raumordnungsrechts grundsätzlich eröffnet ist. Ein weitgehender Ausfall der Zielbindung macht die Fortentwicklung des Fach-, insbesondere des Bergrechts unabdingbar

Maritime Raumordnung

Das Erfordernis raumordnerischen Handelns im Meeresbereich resultiert aus den zunehmenden Nutzungs- und Schutzkonflikten zu Wasser. Dem tragen das internationale, europäische und nationale Recht weitgehend Rechnung. Nach nationalem Recht findet Raumordnung sowohl in den Küstenmeeren als auch in der deutschen Ausschließlichen Wirtschaftszone (AWZ) statt

Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

Optogenetic tools have become indispensable in neuroscience to stimulate or inhibit excitable cells by light. Channelrhodopsin-2 (ChR2) variants have been established by mutating the opsin backbone or by mining related algal genomes. As an alternative strategy, we surveyed synthetic retinal analogues combined with microbial rhodopsins for functional and spectral properties, capitalizing on assays in C. elegans, HEK cells and larval Drosophila. Compared with all-trans retinal (ATR), Dimethylamino-retinal (DMAR) shifts the action spectra maxima of ChR2 variants H134R and H134R/T159C from 480 to 520 nm. Moreover, DMAR decelerates the photocycle of ChR2(H134R) and (H134R/T159C), thereby reducing the light intensity required for persistent channel activation. In hyperpolarizing archaerhodopsin-3 and Mac, naphthyl-retinal and thiophene-retinal support activity alike ATR, yet at altered peak wavelengths. Our experiments enable applications of retinal analogues in colour tuning and altering photocycle characteristics of optogenetic tools, thereby increasing the operational light sensitivity of existing cell lines or transgenic animals

Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

Optogenetic tools have become indispensable in neuroscience to stimulate or inhibit excitable cells by light. Channelrhodopsin-2 (ChR2) variants have been established by mutating the opsin backbone or by mining related algal genomes. As an alternative strategy, we surveyed synthetic retinal analogues combined with microbial rhodopsins for functional and spectral properties, capitalizing on assays in C. elegans, HEK cells and larval Drosophila. Compared with all-trans retinal (ATR), Dimethylamino-retinal (DMAR) shifts the action spectra maxima of ChR2 variants H134R and H134R/T159C from 480 to 520 nm. Moreover, DMAR decelerates the photocycle of ChR2(H134R) and (H134R/T159C), thereby reducing the light intensity required for persistent channel activation. In hyperpolarizing archaerhodopsin-3 and Mac, naphthyl-retinal and thiophene-retinal support activity alike ATR, yet at altered peak wavelengths. Our experiments enable applications of retinal analogues in colour tuning and altering photocycle characteristics of optogenetic tools, thereby increasing the operational light sensitivity of existing cell lines or transgenic animals

Feasibility of Certified Quality Management in a Comprehensive Stroke Care Network Using Telemedicine: STENO Project

Background Stroke care networks with and without telemedicine have been established in several countries over the last decade to provide specialized stroke expertise to patients in rural areas. Acute consultation is a first step in the management of stroke, but not the only one. Methods of standardization of care and treatment are much needed. So far, quality management systems have only been used for single stroke units. To the best of our knowledge, we are the first stroke network worldwide to aim for certification of a network-wide quality management system. Methods The Stroke Network Using Telemedicine in Northern Bavaria (STENO), currently with 20 associated medical institutions, is one of the world's largest stroke networks, caring for over 5000 stroke patients each year. In 2010, we initiated the implementation of a network-wide ‘total’ quality management system according to ISO standard 9001:2008 in cooperation with the German Stroke Society and a third-party certification organization (LGA InterCert). Results Certification according to ISO 9001:2008 was awarded in March 2011 and maintained over a complete certification cycle of 3 years without major deviation from the norm in three external third-party audits. Thrombolysis rate significantly increased from 8·2% (2009) to 12·8% (2012). Conclusions Certified quality management within a large stroke network using telemedicine is possible and might improve stroke care procedures and thrombolysis rates. Outcome studies comparing conventional stroke care and telestroke care are inevitable

First aid in acute stroke: Introducing a concept of first action to laypersons

OBJECTIVE: First aid training is well established to teach the public how to recognize a medical emergency and take appropriate action. Though it is now handled as a high priority emergency stroke is not among the main topics of first aid. We investigated if first aid training may be useful for enhancing stroke awareness. METHODS: We developed a 15–20 minute teaching session about stroke as an emergency including signs and symptoms and first hands-on measures. The session was integrated in standard first aid training of the St John Ambulance of Germany and participants were asked to fill out a questionnaire regarding their knowledge about stroke. Subjects were questioned before the stroke lesson and again at the end of the training. RESULTS: 532 participants of the training responded to the questionnaire (mean age 28.6 years, 53.6% male). There was a significant increase in proportion of subjects correctly defining what stroke is (28.4% vs. 69.9%, p < 0,001) and in the mean number of stroke symptoms listed (1.52 vs. 3.35, p < 0,001) by the participants. The number of participants unable to list at least 1 symptom decreased significantly (12.8 vs. 3.6%, p<0.001). CONCLUSIONS: In our study a teaching lesson integrated in first aid training was effective in improving stroke knowledge of participants. First aid training should be used for stroke information complementary to other activities like mass media campaigns as it is effective, could reach younger people that are not primarily interested in stroke and provides connections to other health topics