The evolving capabilities of rhodopsin-based genetically encoded voltage indicators.

Protein engineering over the past four years has made rhodopsin-based genetically encoded voltage indicators a leading candidate to achieve the task of reporting action potentials from a population of genetically targeted neurons in vivo. Rational design and large-scale screening efforts have steadily improved the dynamic range and kinetics of the rhodopsin voltage-sensing domain, and coupling these rhodopsins to bright fluorescent proteins has supported bright fluorescence readout of the large and rapid rhodopsin voltage response. The rhodopsin-fluorescent protein fusions have the highest achieved signal-to-noise ratios for detecting action potentials in neuronal cultures to date, and have successfully reported single spike events in vivo. Given the rapid pace of current development, the genetically encoded voltage indicator class is nearing the goal of robust spike imaging during live-animal behavioral experiments.

A whole-cell and single-channel study of the voltage-dependent outward potassium current in avian hepatocytes.

Voltage-dependent membrane currents were studied in dissociated hepatocytes from chick, using the patch-clamp technique. All cells had voltage-dependent outward K+ currents; in 10% of the cells, a fast, transient, tetrodotoxin-sensitive Na+ current was identified. None of the cells had voltage-dependent inward Ca2+ currents. The K+ current activated at a membrane potential of about -10 mV, had a sigmoidal time course, and did not inactivate in 500 ms. The maximum outward conductance was 6.6 +/- 2.4 nS in 18 cells. The reversal potential, estimated from tail current measurements, shifted by 50 mV per 10-fold increase in the external K+ concentration. The current traces were fitted by n2 kinetics with voltage-dependent time constants. Omitting Ca2+ from the external bath or buffering the internal Ca2+ with EGTA did not alter the outward current, which shows that Ca2+-activated K+ currents were not present. 1-5 mM 4-aminopyridine, 0.5-2 mM BaCl2, and 0.1-1 mM CdCl2 reversibly inhibited the current. The block caused by Ba was voltage dependent. Single-channel currents were recorded in cell-attached and outside-out patches. The mean unitary conductance was 7 pS, and the channels displayed bursting kinetics. Thus, avian hepatocytes have a single type of K+ channel belonging to the delayed rectifier class of K+ channels.

Are the reference values of B cell subpopulations used in adults for classification of common variable immunodeficiencies appropriate for children?

Detailed phenotypic characterization of B cell subpopulations is of utmost importance for the diagnosis and management of humoral immunodeficiencies, as they are used for classification of common variable immunodeficiencies. Since age-specific reference values remain scarce in the literature, we analysed by flow cytometry the proportions and absolute values of total, memory, switched memory and CD21(-/low) B cells in blood samples from 168 healthy children (1 day to 18 years) with special attention to the different subpopulations of CD21(low) B cells. The percentages of total memory B cells and their subsets significantly increased up to 5-10 years. In contrast, the percentages of immature CD21(-) B cells and of immature transitional CD21(low)CD38(hi) B cells decreased progressively with age, whereas the percentage of CD21(low) CD38(low) B cells remained stable during childhood. Our data stress the importance of age-specific reference values for the correct interpretation of B cell subsets in children as a diagnostic tool in immunodeficiencies.

Hermeneutics, accreting receptions, hypermedia: A tool for reference versus a tool for instruction

We provide a select overview of tools supporting traditional Jewish learning. Then we go on to discuss our own HyperJoseph/HyperIsaac project in instructional hypermedia. Its application is to teaching, teacher training, and self-instruction in given Bible passages. The treatment of two narratives has been developed thus far. The tool enables an analysis of the text in several respects: linguistic, narratological, etc. Moreover, the Scriptures' focality throughout the cultural history makes this domain of application particularly challenging, in that there is a requirement for the tool to encompass the accretion of receptions in the cultural repertoire, i.e., several layers of textual traditions—either hermeneutic (i.e., interpretive), or appropriations—related to the given core passage, thus including "secondary" texts (i.e., such that are responding or derivative) from as disparate realms as Roman-age and later homiletics, Medieval and later commentaries or supercommentaries, literary appropriations, references to the arts and modern scholarship, etc. in particular, the Midrash (homiletic expansions) is adept at narrative gap filling, so the narratives mushroom at the interstices where the primary text is silent. The genealogy of the project is rooted in Weiss' index of novelist Agnon's writings, which was eventually upgraded into a hypertextual tool, including Agnon's full-text and ancillary materials. Those early tools being intended primarily for reference and research-support in literary studies, the Agnon hypertext system was initially emulated in the conception of HyperJoseph, which is applied to the Joseph story from Genesis. Then, the transition from a tool for reference to an instructional tool required a thorough reconception in an educational perspective, which led to HyperIsaac, on the sacrifice of Isaac, and to a redesign and upgrade of HyperJoseph as patterned after HyperIsaac.