Biomedical subjects

Vladimir E Bondarenko

Publications and source records attributed to Vladimir E Bondarenko.

10 recordsLinked to original sources

PubMed2007

W-7 modulates Kv4.3: pore block and Ca2+-calmodulin inhibition.

Ca(+)-calmodulin (Ca(2+)-CaM)-dependent protein kinase II (Ca(2+)/CaMKII) is an important regulator of cardiac ion channels, and its inhibition may be an approach for treatment of ventricular arrhythmias. Using the two-electrode voltage-clamp technique, we investigated the role of W-7, an inhibitor of Ca(2+)-occupied CaM, and KN-93, an inhibitor of Ca(2+)/CaMKII, on the K(v)4.3 channel in Xenopus laevis oocytes. W-7 caused a voltage- and concentration-dependent decrease in peak current, with IC(50) of 92.4 muM. The block was voltage dependent, with an effective electrical distance of 0.18 +/- 0.05, and use dependence was observed, suggesting that a component of W-7 inhibition of K(v)4.3 current was due to open-channel block. W-7 made recovery from open-state inactivation a biexponential process, also suggesting open-channel block. We compared the effects of W-7 with those of KN-93 after washout of 500 muM BAPTA-AM. KN-93 reduced peak current without evidence of voltage or use dependence. Both W-7 and KN-93 accelerated all components of inactivation. We used wild-type and mutated K(v)4.3 channels with mutant CaMKII consensus phosphorylation sites to examine the effects of W-7 and KN-93. In contrast to W-7, KN-93 at 35 muM selectively accelerated open-state inactivation in the wild-type vs. the mutant channel. W-7 had a significantly greater effect on recovery from inactivation in wild-type than in mutant channels. We conclude that, at certain concentrations, KN-93 selectively inhibits Ca(2+)/CaMKII activity in Xenopus oocytes and that the effects of W-7 are mediated by direct interaction with the channel pore and inhibition of Ca(2+)-CaM, as well as a change in activity of Ca(2+)-CaM-dependent enzymes, including Ca(2+)/CaMKII.

Animals↗

PubMed2006

DPP10 is an inactivation modulatory protein of Kv4.3 and Kv1.4.

Voltage-gated K(+) channels exist in vivo as multiprotein complexes made up of pore-forming and ancillary subunits. To further our understanding of the role of a dipeptidyl peptidase-related ancillary subunit, DPP10, we expressed it with Kv4.3 and Kv1.4, two channels responsible for fast-inactivating K(+) currents. Previously, DPP10 has been shown to effect Kv4 channels. However, Kv1.4, when expressed with DPP10, showed many of the same effects as Kv4.3, such as faster time to peak current and negative shifts in the half-inactivation potential of steady-state activation and inactivation. The exception was recovery from inactivation, which is slowed by DPP10. DPP10 expressed with Kv4.3 caused negative shifts in both steady-state activation and inactivation of Kv4.3, but no significant shifts were detected when DPP10 was expressed with Kv4.3 + KChIP2b (Kv channel interacting protein). DPP10 and KChIP2b had different effects on closed-state inactivation. At -60 mV, KChIP2b nearly abolishes closed-state inactivation in Kv4.3, whereas it developed to a much greater extent in the presence of DPP10. Finally, expression of a DPP10 mutant consisting of its transmembrane and cytoplasmic 58 amino acids resulted in effects on Kv4.3 gating that were nearly identical to those of wild-type DPP10. These data show that DPP10 and KChIP2b both modulate Kv4.3 inactivation but that their primary effects are on different inactivation states. Thus DPP10 may be a general modulator of voltage-gated K(+) channel inactivation; understanding its mechanism of action may lead to deeper understanding of the inactivation of a broad range of K(+) channels.

Animals↗

PubMed2005

Time- and voltage-dependent components of Kv4.3 inactivation.

Kv4.3 inactivation is a complex multiexponential process, which can occur from both closed and open states. The fast component of inactivation is modulated by the N-terminus, but the mechanisms mediating the other components of inactivation are controversial. We studied inactivation of Kv4.3 expressed in Xenopus laevis oocytes, using the two-electrode voltage-clamp technique. Inactivation during 2000 ms pulses at potentials positive to the activation threshold was described by three exponents (46 +/- 3, 152 +/- 13, and 930 +/- 50 ms at +50 mV, n = 7) whereas closed-state inactivation (at potentials below threshold) was described by two exponents (1079 +/- 119 and 3719 +/- 307 ms at -40 mV, n = 9). The fast component of open-state inactivation was dominant at potentials positive to -20 mV. Negative to -30 mV, the intermediate and slow components dominated inactivation. Inactivation properties were dependent on pulse duration. Recovery from inactivation was strongly dependent on voltage and pulse duration. We developed an 11-state Markov model of Kv4.3 gating that incorporated a direct transition from the open-inactivated state to the closed-inactivated state. Simulations with this model reproduced open- and closed-state inactivation, isochronal inactivation relationships, and reopening currents. Our data suggest that inactivation can proceed primarily from the open state and that multiple inactivation components can be identified.

Animals↗

PubMed2005

Heteropoda toxin 2 is a gating modifier toxin specific for voltage-gated K+ channels of the Kv4 family.

Kv4 voltage-gated K(+) channels are responsible for transient K(+) currents in the central nervous system and in the heart. HpTx2 is a peptide toxin that selectively inhibits these currents; making it a useful probe for understanding Kv4 channel structure and drug binding. Therefore, we developed a method to produce large amounts of recombinant HpTx2. Recombinant toxin inhibits all three Kv4 isoforms to the same degree; however, the voltage-dependence of inhibition is less apparent for Kv4.1 than for Kv4.3. Similarly, recombinant HpTx2(GS) effects gating characteristics of both channels, but Kv4.1 to a much lesser degree. The toxin lacks affinity for Kv1.4, Kv2.1, and Kv3.4. To locate the binding site, the amino acids linking the third and forth membrane spanning segments of Kv4.3 were replaced with analogous amino acids of Kv1.4. The chimeric K(+) channel was completely insensitive to block by rHpTx2, suggesting that its binding site is near the channel's voltage sensor. These data show that rHpTx2(GS) is a gating modifier toxin that binds to a site remote from the pore.

Amino Acid Sequence↗

PubMed2004

Computer model of action potential of mouse ventricular myocytes.

We have developed a mathematical model of the mouse ventricular myocyte action potential (AP) from voltage-clamp data of the underlying currents and Ca2+ transients. Wherever possible, we used Markov models to represent the molecular structure and function of ion channels. The model includes detailed intracellular Ca2+ dynamics, with simulations of localized events such as sarcoplasmic Ca2+ release into a small intracellular volume bounded by the sarcolemma and sarcoplasmic reticulum. Transporter-mediated Ca2+ fluxes from the bulk cytosol are closely matched to the experimentally reported values and predict stimulation rate-dependent changes in Ca2+ transients. Our model reproduces the properties of cardiac myocytes from two different regions of the heart: the apex and the septum. The septum has a relatively prolonged AP, which reflects a relatively small contribution from the rapid transient outward K+ current in the septum. The attribution of putative molecular bases for several of the component currents enables our mouse model to be used to simulate the behavior of genetically modified transgenic mice.

Action Potentials↗

PubMed2004

Activation properties of Kv4.3 channels: time, voltage and [K+]o dependence.

Rapidly inactivating, voltage-dependent K(+) currents play important roles in both neurones and cardiac myocytes. Kv4 channels form the basis of these currents in many neurones and cardiac myocytes and their mechanism of inactivation appears to differ significantly from that reported for Shaker and Kv1.4 channels. In most channel gating models, inactivation is coupled to the kinetics of activation. Hence, there is a need for a rigorous model based on comprehensive experimental data on Kv4.3 channel activation. To develop a gating model of Kv4.3 channel activation, we studied the properties of Kv4.3 channels in Xenopus oocytes, without endogenous KChIP2 ancillary subunits, using the perforated cut-open oocyte voltage clamp and two-electrode voltage clamp techniques. We obtained high-frequency resolution measurements of the activation and deactivation properties of Kv4.3 channels. Activation was sigmoid and well described by a fourth power exponential function. The voltage dependence of the activation time constants was best described by a biexponential function corresponding to at least two different equivalent charges for activation. Deactivation kinetics are voltage dependent and monoexponential. In contrast to other voltage-sensitive K(+) channels such as HERG and Shaker, we found that elevated extracellular [K(+)] modulated the activation process by slowing deactivation and stabilizing the open state. Using these data we developed a model with five closed states and voltage-dependent transitions between the first four closed states coupled to a voltage-insensitive transition between the final closed (partially activated) state and the open state. Our model closely simulates steady-state and kinetic activation and deactivation data.

Amino Acid Sequence↗

PubMed2004

Bifurcation of synchronous oscillations into torus in a system of two reciprocally inhibitory silicon neurons: experimental observation and modeling.

Oscillatory activity in the central nervous system is associated with various functions, like motor control, memory formation, binding, and attention. Quasiperiodic oscillations are rarely discussed in the neurophysiological literature yet they may play a role in the nervous system both during normal function and disease. Here we use a physical system and a model to explore scenarios for how quasiperiodic oscillations might arise in neuronal networks. An oscillatory system of two mutually inhibitory neuronal units is a ubiquitous network module found in nervous systems and is called a half-center oscillator. Previously we created a half-center oscillator of two identical oscillatory silicon (analog Very Large Scale Integration) neurons and developed a mathematical model describing its dynamics. In the mathematical model, we have shown that an in-phase limit cycle becomes unstable through a subcritical torus bifurcation. However, the existence of this torus bifurcation in experimental silicon two-neuron system was not rigorously demonstrated or investigated. Here we demonstrate the torus predicted by the model for the silicon implementation of a half-center oscillator using complex time series analysis, including bifurcation diagrams, mapping techniques, correlation functions, amplitude spectra, and correlation dimensions, and we investigate how the properties of the quasiperiodic oscillations depend on the strengths of coupling between the silicon neurons. The potential advantages and disadvantages of quasiperiodic oscillations (torus) for biological neural systems and artificial neural networks are discussed.

Action Potentials↗

PubMed2003

A model of graded calcium release and L-type Ca2+ channel inactivation in cardiac muscle.

We have developed a model of Ca(2+) handling in ferret ventricular myocytes. This model includes a novel L-type Ca(2+) channel, detailed intracellular Ca(2+) movements, and graded Ca(2+)-induced Ca(2+) release (CICR). The model successfully reproduces data from voltage-clamp experiments, including voltage- and time-dependent changes in intracellular Ca(2+) concentration ([Ca(2+)](i)), L-type Ca(2+) channel current (I(CaL)) inactivation and recovery kinetics, and Ca(2+) sparks. The development of graded CICR is critically dependent on spatial heterogeneity and the physical arrangement of calcium channels in opposition to ryanodine-sensitive release channels. The model contains spatially distinct subsystems representing the subsarcolemmal regions where the junctional sarcoplasmic reticulum (SR) abuts the T-tubular membrane and where the L-type Ca(2+) channels and SR ryanodine receptors (RyRs) are localized. There are eight different types of subsystems in our model, with between one and eight L-type Ca(2+) channels distributed binomially. This model exhibits graded CICR and provides a quantitative description of Ca(2+) dynamics not requiring Monte-Carlo simulations. Activation of RyRs and release of Ca(2+) from the SR depend critically on Ca(2+) entry through L-type Ca(2+) channels. In turn, Ca(2+) channel inactivation is critically dependent on the release of stored intracellular Ca(2+). Inactivation of I(CaL) depends on both transmembrane voltage and local [Ca(2+)](i) near the channel, which results in distinctive inactivation properties. The molecular mechanisms underlying many I(CaL) gating properties are unclear, but [Ca(2+)](i) dynamics clearly play a fundamental role.

Animals↗

PubMed2003

C-type inactivation involves a significant decrease in the intracellular aqueous pore volume of Kv1.4 K+ channels expressed in Xenopus oocytes.

Channels are water-filled membrane-spanning proteins, which undergo conformational changes as they gate, i.e. open or close. These conformational changes affect both the shape of the channel and the volume of the water-filled pore. We measured the changes in pore volume associated with activation, deactivation, C-type inactivation and recovery in an N-terminal-deleted mutant of the Kv1.4 K+ channel (Kv1.4DeltaN) expressed in Xenopus oocytes. We used giant-patch and cut-open oocyte voltage clamp techniques and applied solutes which are too large to enter the pore mouth to exert osmotic pressure and thus favour smaller pore volume conformations. Applied intracellular osmotic pressure (300 mM sucrose) sped inactivation (time constants (tauinactivation): control, 0.66 +/- 0.09 s; hyperosmotic solution, 0.29 +/- 0.04 s; n = 5, P < 0.01), sped deactivation (taudeactivation: control, 18.8 +/- 0.94 ms; hyperosmotic solution, 8.01 +/- 1.92 ms; n = 5, P < 0.01), and slowed activation (tauactivation: control, 1.04 +/- 0.05 ms; hyperosmotic solution, 1.96 +/- 0.31 ms; n = 5, P < 0.01). These effects were reversible and solute independent. We estimated the pore volume change on inactivation to be about 4500 A3. Osmotic pressure had no effect when applied extracellularly. These data suggest that the intracellular side of the pore closes during C-type inactivation and the volume change is similar to that associated with activation or deactivation. This is also similar to the pore volume estimated from the crystal structure of KcsA and MthK K+ channels. Intracellular osmotic pressure also strongly inhibited re-opening currents associated with recovery from inactivation, which is consistent with a physical similarity between the C-type inactivated and resting closed state.

Algorithms↗

PubMed2002

Regulation of N- and C-type inactivation of Kv1.4 by pHo and K+: evidence for transmembrane communication.

Kv1.4 encodes a slowly recovering transient outward current (I(to)), which inactivates by a fast N-type (intracellular ball and chain) mechanism but has slow recovery due to C-type inactivation. C-type inactivation of the NH(2)-terminal deletion mutant (fKv1.4DeltaN) was inhibited by 98 mM extracellular K(+) concentration ([K(+)](o)), whereas N-type was unaffected. In 98 mM [K(+)](o), removal of intracellular K(+) concentration ([K(+)](i)) speeded C-type inactivation but had no effect on N-type inactivation, suggesting that C-type inactivation is sensitive to K(+) binding to intracellular sites. C-type inactivation is thought to involve closure of the extracellular pore mouth. However, a valine to alanine mutation on the intracellular side of S6 (V561A) of fKv1.4DeltaN alters recovery and results in anomalous speeding of C-type inactivation with increasing [K(+)](o). Extracellular pH (pH(o)) modulated both N- and C-type inactivation through an S5-H5 linker histidine (H508) with acidosis speeding both N- and C-type inactivation. Mutation of an extracellular lysine to a tyrosine (K532Y) slowed C-type inactivation and inhibited the pH dependence of both N- and C-type inactivation. These results suggest that mutations, [K(+)], and pH modulate inactivation through membrane-spanning mechanisms involving S6.

Alanine↗