Biology subjects

Zahradnik, I.

Publications and source records attributed to Zahradnik, I..

4 recordsLinked to original sources

bioRxiv2024

Structure-based mechanism of RyR channel operation by calcium and magnesium ions

Ryanodine receptors (RyRs) serve for excitation-contraction coupling in skeletal and cardiac muscle cells in a noticeably different way, not fully understood at the molecular level. We addressed the structure of skeletal (RyR1) and cardiac (RyR2) isoforms relevant to gating by Ca2+ and Mg2+ ions (M2+). Bioinformatics analysis of RyR structures ascertained the EF-hand loops as the M2+ binding inhibition site and revealed its allosteric coupling to the channel gate. The intra-monomeric inactivation pathway interacts with the Ca2+-activation pathway in both RyR isoforms, and the inter-monomeric pathway, stronger in RyR1, couples to the gate through the S23*-loop of the neighbor monomer. These structural findings were implemented in the model of RyR operation based on statistical mechanics and the Monod-Wyman-Changeux theorem. The model, which defines closed, open, and inactivated macrostates allosterically coupled to M2+-binding activation and inhibition sites, approximated the open probability data for both RyR1 and RyR2 channels at a broad range of M2+ concentrations. The proposed mechanism of RyR operation provides a new interpretation of the structural and functional data of mammalian RyR channels on common grounds. This may provide a new platform for designing pharmacological interventions in the relevant diseases of skeletal and cardiac muscles. The synthetic approach developed in this work may find general use in deciphering mechanisms of ion channel functions.

molecular biology↗

bioRxiv2022

Analysis of noisy transient signals based on Gaussian process regression

Dynamic systems such as cells or tissues generate, either spontaneously or in response to stimuli, transient signals that carry information about the system. Characterization of recorded transients is often hampered by a low signal-to-noise ratio (SNR). Reduction of the noise by filtering has limited use due to partial signal distortion. Occasionally, transients can be approximated by a mathematical function, but such a function may not hold correctly if recording conditions change. We introduce here the model-independent approximation method for general noisy transient signals based on the Gaussian process regression (GPR). The method was implemented in the software TransientAnalyzer, which detects transients in a record, finds their best approximation by the Gaussian process, constructs a surrogate spline function, and estimates specified signal parameters. The method and software were tested on a cellular model of the calcium concentration transient corrupted by various SNR levels and recorded at a low sampling frequency. Statistical analysis of the model data sets provided the error of estimation <7.5% and the coefficient of variation of estimates <17% for peak SNR=5. The performance of GPR on signals of diverse experimental origin was even better than fitting by a function. The software and its description are available on GitHub. Statement of SignificanceTransient signals convey information on the state and function of the studied system. However, the estimation of their characteristic parameters is complicated by the noise present in the recordings. Methods used for noise reduction have various disadvantages, such as distortion of the time course by filtering, the difficult superposition of many transients for accurate averaging, or a lack of a model for data fitting. In this work, we present a general method for the automatic analysis of noisy transient signals based on Gaussian process regression and its implementation in Python. The method can analyze recorded transients reliably at peak SNR [≥] 2 with a precision equivalent to the model-fitting methods.

biophysics↗

bioRxiv2020

Synergy of calcium release site determinants in control of calcium release events in cardiac myocytes

Recent data on structure of dyads in cardiac myocytes indicate variable clustering of RyR calcium release channels. The question arises as to how geometric factors of RyR arrangement translate to their role in formation of calcium release events (CRE). Since this question is not experimentally testable in situ, we performed in silico experiments on a large set of calcium release site (CRS) models. The models covered the range of RyR spatial distributions observed in dyads, and included gating of RyRs with open probability dependent on Ca2+ and Mg2+ concentration. The RyR single-channel calcium current, varied in the range of previously reported values, was set constant in the course of CRE simulations. Other known features of dyads were omitted in the model formulation for clarity. CRE simulations initiated by a single random opening of one of the RyRs in a CRS produced spark-like responses with characteristics that varied with RyR vicinity, a newly defined parameter quantifying spatial distribution of RyRs in the CRSs, and with the RyR single-channel calcium current. The CRE characteristics followed the law of mass action with respect to a CRS state variable, defined as a weighed product of RyR vicinity and RyR single-channel calcium current. The results explained the structure-function relations among determinants of cardiac dyads on synergy principles and thus allowed to evolve the concept of CRS as a dynamic unit of cardiac dyad.

biophysics↗

bioRxiv2017

Deconvolution of membrane currents for low-crosstalk high-resolution membrane capacitance measurements

Many cellular activities involving cell membrane manifest through changes in membrane capacitance. The accuracy and precision of cell membrane capacitance measurement is limited by the measuring system, which distorts membrane current responses used for analysis of impedance parameters. We developed a practical deconvolution procedure for reconstruction of the time course of membrane current and applied it to current responses recorded at step voltage changes by the whole-cell patch-clamp technique. In contrast to the recorded current responses, the reconstructed current responses could be exactly described by adequate impedance models of the recorded circuit. Deconvolution of membrane currents improved the performance of the square-wave method of high-resolution membrane capacitance recording by providing higher accuracy in a wider range of cell sizes and by eliminating cross-talk errors in parameter estimates. This allowed resolving instabilities in the recording conditions arising from parasitic capacitance and seal resistance variation. Complex tests on hardware models, on simulated data sets, and on living cells confirmed reliability of the deconvolution method for membrane capacitance recording. The aptitude of the method was demonstrated in isolated cardiac myocytes by recording spontaneous vesicular events, by discerning formation of a fusion pore, and by revealing artefacts due to unstable seal resistance.

biophysics↗