Elementary Functional Properties of Single HCN2 Channels

AbstractHyperpolarization-activated cyclic-nucleotide-gated (HCN) channels are tetramers that evoke rhythmic electrical activity in specialized neurons and cardiac cells. These channels are activated by hyperpolarizing voltage, and the second messenger cAMP can further enhance the activation. Despite the physiological importance of HCN channels, their elementary functional properties are still unclear. In this study, we expressed homotetrameric HCN2 channels in Xenopus oocytes and performed single-channel experiments in patches containing either one or multiple channels. We show that the single-channel conductance is as low as 1.67 pS and that channel activation is a one-step process. We also observed that the time between the hyperpolarizing stimulus and the first channel opening, the first latency, determines the activation process alone. Notably, at maximum hyperpolarization, saturating cAMP drives the channel to open for unusually long periods. In particular, at maximum activation by hyperpolarization and saturating cAMP, the open probability approaches unity. In contrast to other reports, no evidence of interchannel cooperativity was observed. In conclusion, single HCN2 channels operate only with an exceptionally low conductance, and both activating stimuli, voltage and cAMP, exclusively control the open probability

A Robust and Universal Metaproteomics Workflow for Research Studies and Routine Diagnostics Within 24 h Using Phenol Extraction, FASP Digest, and the MetaProteomeAnalyzer

The investigation of microbial proteins by mass spectrometry (metaproteomics) is a key technology for simultaneously assessing the taxonomic composition and the functionality of microbial communities in medical, environmental, and biotechnological applications. We present an improved metaproteomics workflow using an updated sample preparation and a new version of the MetaProteomeAnalyzer software for data analysis. High resolution by multidimensional separation (GeLC, MudPIT) was sacrificed to aim at fast analysis of a broad range of different samples in less than 24 h. The improved workflow generated at least two times as many protein identifications than our previous workflow, and a drastic increase of taxonomic and functional annotations. Improvements of all aspects of the workflow, particularly the speed, are first steps toward potential routine clinical diagnostics (i.e., fecal samples) and analysis of technical and environmental samples. The MetaProteomeAnalyzer is provided to the scientific community as a central remote server solution at www.mpa.ovgu.de.Peer Reviewe

From Hubble to Snap Parameters: A Gaussian Process Reconstruction

By using recent $H(z)$ and SNe Ia data, we reconstruct the evolution of kinematic parameters $H(z)$, $q(z)$, jerk and snap, using a model-independent, non-parametric method, namely, the Gaussian Processes. Throughout the present analysis, we have allowed for a spatial curvature prior, based on Planck 18 [1] constraints. In the case of SNe Ia, we modify a python package (GaPP) [2] in order to obtain the reconstruction of the fourth derivative of a function, thereby allowing us to obtain the snap from comoving distances. Furthermore, using a method of importance sampling, we combine $H(z)$ and SNe Ia reconstructions in order to find joint constraints for the kinematic parameters. We find for the current values of the parameters: $H_0 =67.2 \pm 6.2$ km/s/Mpc, $q_0 = -0.60^{+0.21}_{-0.18}$, $j_0=0.90^{+0.75}_{-0.65}$, $s_0=-0.57^{+0.52}_{-0.31}$ at 1$\sigma$ c.l. We find that these reconstructions are compatible with the predictions from flat $\Lambda$CDM model, at least for 2$\sigma$ confidence intervals.Comment: 18 pages, 6 figure

New strategies to measure intracellular sodium concentrations

Fluorescent ion indicators are widely used to measure ion concentrations in living cells. However, despite considerable efforts in synthesizing new compounds, no ratiometric sodium indicator is available that can be excited at visible wavelengths. Ratiometric indicators have an advantage in that measured fluorescence intensities can be corrected for fluctuations of the indicator concentration and the illumination intensity, which is not possible when non-ratiometric indicators are used. One way to circumvent this problem is to measure fluorescence lifetimes, which are independent of these factors. Another way to overcome the disadvantages of a non-ratiometric indicator dye is to embed it, together with a reference dye, into nanoparticles. By relating the indicator fluorescence to the fluorescence of the reference dye, inhomogeneities in the nanosensor concentration or the illumination intensity can be cancelled out reliably. In this study we compare the benefits and drawbacks of these approaches. © 2010 Copyright SPIE - The International Society for Optical Engineering

Structural characterization of a-plane Zn1−xCdxO (0 < x <0.085) thin films grown by metal-organic vapor phase epitaxy.

Zn1−xCdxO(11math0) films have been grown on (01math2) sapphire (r–plane) substrates by metal-organic vapor phase epitaxy. A 800-nm-thick ZnO buffer, deposited prior to the alloy growth, helps to prevent the formation of pure CdO. A maximum uniform Cd incorporation of 8.5 at. % has been determined by Rutherford backscattering spectrometry. Higher Cd contents lead to the coexistence of Zn1−xCdxO alloys of different compositions within the same film. The near band-edge photoluminescence emission shifts gradually to lower energies as Cd is incorporated and reaches 2.93 eV for the highest Cd concentration (8.5 at. %). The lattice deformation, due to Cd incorporation, has been described using a new reference frame in which the lattice distortions are directly related to the a-plane surface structure. Cd introduction does not affect the c lattice parameter but expands the lattice along the two perpendicular directions, [11math0] and [math100], resulting in a quadratic volume [email protected] [email protected]

Fecal Metaproteomics Reveals Reduced Gut Inflammation and Changed Microbial Metabolism Following Lifestyle-Induced Weight Loss

Gut microbiota-mediated inflammation promotes obesity-associated low-grade inflammation, which represents a hallmark of metabolic syndrome. To investigate if lifestyle-induced weight loss (WL) may modulate the gut microbiome composition and its interaction with the host on a functional level, we analyzed the fecal metaproteome of 33 individuals with metabolic syndrome in a longitudinal study before and after lifestyle-induced WL in a well-defined cohort. The 6-month WL intervention resulted in reduced BMI (−13.7%), improved insulin sensitivity (HOMA-IR, −46.1%), and reduced levels of circulating hsCRP (−39.9%), indicating metabolic syndrome reversal. The metaprotein spectra revealed a decrease of human proteins associated with gut inflammation. Taxonomic analysis revealed only minor changes in the bacterial composition with an increase of the families Desulfovibrionaceae, Leptospiraceae, Syntrophomonadaceae, Thermotogaceae and Verrucomicrobiaceae. Yet we detected an increased abundance of microbial metaprotein spectra that suggest an enhanced hydrolysis of complex carbohydrates. Hence, lifestyle-induced WL was associated with reduced gut inflammation and functional changes of human and microbial enzymes for carbohydrate hydrolysis while the taxonomic composition of the gut microbiome remained almost stable. The metaproteomics workflow has proven to be a suitable method for monitoring inflammatory changes in the fecal metaproteome