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

Fur in Magnetospirillum gryphiswaldense Influences Magnetosomes Formation and Directly Regulates the Genes Involved in Iron and Oxygen Metabolism

Magnetospirillum gryphiswaldense strain MSR-1 has the unique capability of taking up large amounts of iron and synthesizing magnetosomes (intracellular magnetic particles composed of Fe3O4). The unusual high iron content of MSR-1 makes it a useful model for studying biological mechanisms of iron uptake and homeostasis. The ferric uptake regulator (Fur) protein plays a key role in maintaining iron homeostasis in many bacteria. We identified and characterized a fur-homologous gene (MGR_1314) in MSR-1. MGR_1314 was able to complement a fur mutant of E. coli in iron-responsive manner in vivo. We constructed a fur mutant strain of MSR-1. In comparison to wild-type MSR-1, the mutant strain had lower magnetosome formation, and was more sensitive to hydrogen peroxide and streptonigrin, indicating higher intracellular free iron content. Quantitative real-time RT-PCR and chromatin immunoprecipitation analyses indicated that Fur protein directly regulates expression of several key genes involved in iron transport and oxygen metabolism, in addition it also functions in magnetosome formation in M. gryphiswaldense

Compatibility and stability of telavancin and vancomycin in heparin or sodium citrate lock solutions

Purpose. The compatibility and stability of telavancin and vancomycin in heparin or sodium citrate lock solutions were evaluated. Methods. Telavancin and vancomycin hydrochloride injection powder lyophilized for solution were reconstituted with 0.9% sodium chloride injection at room temperature according to the manufacturer\u27s instructions and then further diluted with (1) commercially available heparin sodium to reach a final heparin concentration of 2500 units/mL or (2) sodium citrate solution 2.2% or 4% to achieve final telavancin and vancomycin concentrations of 2 and 5 mg/mL. Physical stability, chemical compatibility, and biological anticoagulant stability were analyzed for each antibiotic-anticoagulant combination immediately after preparation and at 24, 48, and 72 hours. Changes in coagulation were measured at each time point and compared using two-way analysis of variance. Results. Both telavancin and vancomycin retained at least 90% of the initial concentration after incubation at 37°C over 72 hours. The biological stability of vancomycin 2 mg/mL and telavancin 2 mg/mL did not significantly alter prothrombin time when compared with that of 0.9% sodium chloride injection. However, telavancin 5 mg/mL and vancomycin 5 mg/mL significantly increased the activated partial thromboplastin time at 72 hours compared with the control solution. Visual precipitation only occurred with vancomycin-containing solutions; however, this dissipated after 10 minutes. Conclusion. Telavancin 2 and 5 mg/mL was physically compatible in combination with heparin 2500 units/mL and with sodium citrate 2.2% and 4% over 72 hours. Vancomycin 2 and 5 mg/mL initially precipitated in the sodium citrate 2.2% formulation, but no precipitation was noted after 10 minutes of incubation at 37°C. Telavancin and vancomycin 2 and 5 mg/mL retained over 90% of the initial concentration after incubation at 37°C over 72 hours