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James N Weiss

Publications and source records attributed to James N Weiss.

At least 37 records · Page 2Linked to original sources
PubMed2005

ATP-sensitive K+ channels: regulation of bursting by the sulphonylurea receptor, PIP2 and regions of Kir6.2.

ATP-sensitive K+ channels composed of the pore-forming protein Kir6.2 and the sulphonylurea receptor SUR1 are inhibited by ATP and activated by Phosphatidylinositol Bisphosphate (PIP2). Residues involved in binding of these ligands to the Kir6.2 cytoplasmic domain have been identified, and it has been hypothesized that gating mechanisms involve conformational changes in the regions of the bundle crossing and/or the selectivity filter of Kir6.2. Regulation of Kir6.2 by SUR1, however, is not well-understood, even though this process is ATP and PIP2 dependent. In this study, we investigated the relationship between channel regulation by SUR1 and PIP2 by comparing a number of single and double mutants known to affect open probability (P(o)), PIP2 affinity, and sulphonylurea and MgADP sensitivity. When coexpressed with SUR1, the Kir6.2 mutant C166A, which is characterized by a P(o) value close to 0.8, exhibits no sulphonylurea or MgADP sensitivity. However, when P(o) was reduced by combining mutations at the PIP2-sensitive residues R176 and R177 with C166A, sulphonylurea and MgADP sensitivities were restored. These effects correlated with a dramatic decrease in PIP2 affinity, as assessed by PIP2-induced channel reactivation and inhibition by neomycin, an antagonist of PIP2 binding. Based on macroscopic and single-channel data, we propose a model in which entry into the high-P(o) bursting state by the C166A mutation or by SUR1 depends on the interaction of PIP2 with R176 and R177 and, to a lesser extent, R54. In conjunction with this PIP2-dependent process, SUR1 also regulates channel activity via a PIP2-independent, but MgADP-dependent process.

ATP-Binding Cassette Transporters↗
PubMed2005

ATP sensitivity of ATP-sensitive K+ channels: role of the gamma phosphate group of ATP and the R50 residue of mouse Kir6.2.

ATP-sensitive K (K(ATP)) channels are composed of Kir6, the pore-forming protein, and the sulphonylurea receptor SUR, a regulatory protein. We and others have previously shown that positively charged residues in the C terminus of Kir6.2, including R201 and K185, interact with the alpha and beta phosphate groups of ATP, respectively, to induce channel closure. A positively charged residue in the N terminus, R50, is also important, and has been proposed to interact with either the gamma or beta phosphate group of ATP. To examine this issue, we systematically mutated R50 to residues of different size, charge and hydropathy, and examined the effects on adenine nucleotide sensitivity in the absence and presence of SUR1. In the absence of SUR1, only the size of residue 50 significantly altered ATP sensitivity, with smaller side chains decreasing ATP sensitivity. In the presence of SUR1, however, hydrophathy and charge also played a role. Hydrophilic residues decreased ATP sensitivity more than hydrophobic residues for small size residues, and, surprisingly, negatively charged residues E and D preserved ATP sensitivity and increased ADP sensitivity relative to the wild-type residue R. These observations suggest that a negative charge near position 50, due to either mutation of R50 or the interaction of the gamma phosphate group of ATP with R50, facilitates closure of the ATP-dependent gate. Mutation of the nearby positively charged residue R54, known to be involved in stabilizing channel opening via electrostatic interactions with phosphatidylinositol 4,5-bisphosphate (PIP2), also caused increased ADP sensitivity as compared with ATP, suggesting a loss of function of ATP's gamma phosphate. Based on these results, we propose that a phosphate group or a negative charge at position 50 initiates channel closure by destabilizing the electrostatic interactions between negative phosphate groups of PIP2 and residues such as R54.

Adenosine Triphosphate↗
PubMed2005

The dynamics of cardiac fibrillation.

Reentry occurs when the electrical wave propagating through the atria or ventricles breaks locally and forms a rotor (also called a scroll wave or functional reentry). If the waves propagating outward from a rotor develop additional wavebreaks (which may form new rotors), fibrillation results. Tissue heterogeneity, exacerbated by electrical and structural remodeling from cardiac disease, has traditionally been considered the major factor promoting wavebreak and its degeneration to fibrillation. Recently, however, dynamic factors have also been recognized to play a key role. Dynamic factors refer to cellular properties of the cardiac action potential and Ca(i) cycling, which dynamically generate wave instability and wavebreak, even in tissue that is initially completely homogeneous. Although the latter situation can only be created in computer simulations, its relevance to real (heterogeneous) cardiac tissue has been unequivocally demonstrated. Dynamic factors are related to membrane voltage (Vm) and Ca(i). Vm factors include electrical restitution of action potential duration and conduction velocity, short-term cardiac memory, and electrotonic currents. Ca(i) factors are related to dynamic Ca(i) cycling properties. They act synergistically, as well as with tissue heterogeneity, to promote wavebreak and fibrillation. As global properties, rather than local electrophysiological characteristics, dynamic factors represent an attractive target for novel therapies to prevent ventricular fibrillation.

Action Potentials↗
PubMed2005

Effects of Na(+) and K(+) channel blockade on vulnerability to and termination of fibrillation in simulated normal cardiac tissue.

Na(+) and K(+) channel-blocking drugs have anti- and proarrhythmic effects. Their effects during fibrillation, however, remain poorly understood. We used computer simulation of a two-dimensional (2-D) structurally normal tissue model with phase I of the Luo-Rudy action potential model to study the effects of Na(+) and K(+) channel blockade on vulnerability to and termination of reentry in simulated multiple-wavelet and mother rotor fibrillation. Our main findings are as follows: 1) Na(+) channel blockade decreased, whereas K(+) channel blockade increased, the vulnerable window of reentry in heterogeneous 2-D tissue because of opposing effects on dynamical wave instability. 2) Na(+) channel blockade increased the cycle length of reentry more than it increased refractoriness. In multiple-wavelet fibrillation, Na(+) channel blockade first increased and then decreased the average duration or transient time ( ) of fibrillation. In mother rotor fibrillation, Na(+) channel blockade caused peripheral fibrillatory conduction block to resolve and the mother rotor to drift, leading to self-termination or sustained tachycardia. 3) K(+) channel blockade increased dynamical instability by steepening action potential duration restitution. In multiple-wavelet fibrillation, this effect shortened because of enhanced wave instability. In mother rotor fibrillation, this effect converted mother rotor fibrillation to multiple-wavelet fibrillation, which then could self-terminate. Our findings help illuminate, from a theoretical perspective, the possible underlying mechanisms of termination of different types of fibrillation by antiarrhythmic drugs.

Anti-Arrhythmia Agents↗
PubMed2005

K+-dependent regulation of matrix volume improves mitochondrial function under conditions mimicking ischemia-reperfusion.

To delineate the role of mitochondrial K+ fluxes in cardioprotection, we investigated the effect of extramitochondrial K+ on the ability of mitochondria to support membrane potential (DeltaPsi), regulate matrix volume, consume oxygen, and phosphorylate ADP under conditions mimicking key elements of ischemia-reperfusion. Isolated energized mitochondria responded to ADP addition with depolarization, increased O2 consumption, and matrix shrinkage. The time required for full recovery of DeltaPsi, signaling the completion of ADP phosphorylation, was used to evaluate the rate of ATP synthesis during repeated ADP pulses. In mitochondria with a decreased ability to support DeltaPsi, the rate of ADP phosphorylation was significantly improved by extramitochondrial K+ > Na+ > Li+, especially at higher buffer osmolarity, which promotes matrix shrinkage. K+-induced improvement in DeltaPsi recovery after ADP pulses was accompanied by more rapid and complete matrix volume recovery and enhanced O2 consumption. Manipulations expected to affect matrix swelling by regulating K+ fluxes or water distribution indicate that matrix volume regulation by external factors becomes increasingly important in mitochondria with decreased ability to support DeltaPsi in the face of a high ADP load. Under these conditions, opening of K+ influx pathways improved mitochondrial function and delayed failure. This may be an important factor in the mechanism of diaxozide-induced cardioprotection.

Adenosine Diphosphate↗
PubMed2005

Action potential duration restitution and alternans in rabbit ventricular myocytes: the key role of intracellular calcium cycling.

Action potential duration (APD) restitution properties and repolarization alternans are thought to be important arrhythmogenic factors. We investigated the role of intracellular calcium (Ca2+i) cycling in regulating APD restitution slope and repolarization (APD) alternans in patch-clamped rabbit ventricular myocytes at 34 to 36 degrees C, using the perforated or ruptured patch clamp techniques with Fura-2-AM to record Ca2+i. When APD restitution was measured by either the standard extrastimulus (S1S2) method or the dynamic rapid pacing method, the maximum APD restitution slope exceeded 1 by both methods, but was more shallow with the dynamic method. These differences were associated with greater Ca2+i accumulation during dynamic pacing. The onset of APD alternans occurred at diastolic intervals at which the APD restitution slope was significantly <1 and was abolished by suppressing sarcoplasmic reticulum (SR) Ca2+i cycling with thapsigargin and ryanodine, or buffering the global Ca2+i transient with BAPTA-AM or BAPTA. Thapsigargin and ryanodine flattened APD restitution slope to <1 when measured by the dynamic method, but not by the S1S2 method. BAPTA-AM or BAPTA failed to flatten APD restitution slope to <1 by either method. In conclusion, APD alternans requires intact Ca2+i cycling and is not reliably predicted by APD restitution slope when Ca2+i cycling is suppressed. Ca2+i cycling may contribute to differences between APD restitution curves measured by S1S2 versus dynamic pacing protocols by inducing short-term memory effects related to pacing-dependent Ca2+i accumulation.

Action Potentials↗
PubMed2005

Regulation of the ATP-sensitive K channel Kir6.2 by ATP and PIP(2).

ATP-sensitive K (K(ATP)) channels are blocked by ATP and activated by PIP(2). Both negatively-charged ligands are presumed to bind to positively-charged residues on the N-and C-termini of the channel's cytoplasmic domain. Evidence summarized here suggests that the channel's interaction with ATP and PIP(2) is regulated by separate groups of residues, involving both direct charge-charge interactions and allosteric effects. ATP interaction is regulated by R50 in the N-terminus and by K185, R192 and R201 in the C-terminus. R192 and R201 mutations decrease channel sensitivity to ATP, ADP and AMP to a similar extent, implying that they regulate interaction with either the alpha phosphate group, common to all three adenine nucleotides, or the adenosine moiety. K185 mutations, and to a lesser extent R50 mutations, decrease ATP and ADP sensitivity without markedly affecting AMP sensitivity, implying that they regulate interaction with the beta phosphate of ATP and ADP. In addition, when open probability decreases due to rundown, ATP sensitivity increases in R50, K185 and R192, but not in R201 mutants. Combining these observations with recent structural data, we hypothesize the following scenario: 1) the ATP binding site is located at the outside of the channel's cytoplasmic domain away from the pore. 2) When the channel is open, R50 and K185 interact directly with the beta phosphate of ATP, whereas R192, which appears to be removed from the ATP binding site, modulates the initial interaction with ATP allosterically. 3) When the channel closes, R201 is in position to interact with the alpha phosphate of ATP to stabilize the closed state. 4) PIP(2) also interacts with the channel's cytoplasmic domain, but at distinct positively-charged residues located above the ATP binding site and near to the plasma membrane. These residues include R54 in the N-terminus and R176, R177 and R206 in the C-terminus. Thus, the binding domains of ATP and PIP(2) in the N- and C-termini do not appear to overlap.

Adenosine Triphosphate↗
PubMed2005

Dissociation of membrane potential and intracellular calcium during ventricular fibrillation.

UNLABELLED: Membrane potential and intracellular calcium during VF. INTRODUCTION: The cardiac action potential (AP) and the intracellular Ca transient (CaT) are closely associated under normal physiological conditions, but not during ventricular fibrillation (VF). The purpose of this study was to determine whether this dissociation is directly related to the fast activation rate during VF. METHODS AND RESULTS: We optically mapped AP and CaT simultaneously in nine isolated rabbit hearts. Pinacidil, a K(ATP) channel opener, was used to shorten the action potential duration (APD) in order to capture tissue at fast pacing rates or to induce ventricular tachycardia (VT) comparable to VF activation rates. Mutual information (MI) was used to calculate the degree of AP and CaT coupling. Pinacidil (40 microM) infusion significantly shortened APD. The CL of VF without pinacidil averaged 77+/-13 ms, whereas the shortest CL achieved during VT under pinacidil infusion was 76 ms. MIs during fast pacing (1.13+/-0.15 bits) and fast VT (0.88+/-0.18 bits) were higher than those during baseline VF (0.39+/-0.11 bits), VF with pinacidil infusion (0.21+/-0.07 bits) and VF after pinacidil washout (0.36+/-0.15 bits). MIs during fast pacing or fast VT were higher than that of VFs at comparable dominant frequencies. CONCLUSIONS: CaT is closely associated with the AP during fast pacing and fast VT, but not during VF. The reduced MI during VF is not secondary to the fast rate of activation.

Animals↗
PubMed2005

Long polyamines act as cofactors in PIP2 activation of inward rectifier potassium (Kir2.1) channels.

Phosphatidylinosital-4,5-bisphosphate (PIP2) acts as an essential factor regulating the activity of all Kir channels. In most Kir members, the dependence on PIP2 is modulated by other factors, such as protein kinases (in Kir1), G(betagamma) (in Kir3), and the sulfonylurea receptor (in Kir6). So far, however, no regulator has been identified in Kir2 channels. Here we show that polyamines, which cause inward rectification by selectively blocking outward current, also regulate the interaction of PIP2 with Kir2.1 channels to maintain channel availability. Using spermine and diamines as polyamine analogs, we demonstrate that both spontaneous and PIP2 antibody-induced rundown of Kir2.1 channels in excised inside-out patches was markedly slowed by long polyamines; in contrast, polyamines with shorter chain length were ineffective. In K188Q mutant channels, which have a low PIP2 affinity, application PIP2 (10 microM) was unable to activate channel activity in the absence of polyamines, but markedly activated channels in the presence of long diamines. Using neomycin as a measure of PIP2 affinity, we found that long polyamines were capable of strengthening either the wild type or K188Q channels' interaction with PIP2. The negatively charged D172 residue inside the transmembrane pore region was critical for the shift of channel-PIP2 binding affinity by long polyamines. Sustained pore block by polyamines was neither sufficient nor necessary for this effect. We conclude that long polyamines serve a dual role as both blockers and coactivators (with PIP2) of Kir2.1 channels.

Animals↗
PubMed2005

Mitochondria and ischemia/reperfusion injury.

Cardiac ischemia/reperfusion injury results in a variable mixture of apoptotic, necrotic, and normal tissue that depends on both the duration and severity of ischemia. Injury can be abrogated by activation of protective pathways via ischemic and pharmacologic preconditioning. Mitochondria serve as final arbiters of life and death of the cell as these organelles not only are required to generate ATP but also can trigger apoptosis or necrosis. A key mechanism of mitochondrial injury is by the mitochondrial permeability transition (MPT) that has been shown to occur at reperfusion. The article hypothesizes that ischemia/reperfusion promotes MPT in two phases: (1) MPT priming during ischemia occurs as progressive inner mitochondrial membrane leak is accompanied by depressed electron transport in the setting of fatty acid accumulation and loss of cytochrome c and antioxidants; and (2) Triggering of MPT at reperfusion is determined by the interplay of mitochondrial membrane potential (DeltaPsi(m)) with mitochondrial matrix Ca, reactive oxygen species, and pH. It has been found that strategies that promote mitochondrial recovery such as pharmacologic preconditioning by diazoxide are mediated by K(+)-dependent regulation of matrix volume and DeltaPsi(m), resulting in improved efficiency of ATP synthesis as well as prevention of cytochrome c loss. If mitochondria fail to recover, then MPT and hypercontracture can result as DeltaPsi(m) depolarization waves regeneratively cross the cell (0.1 to 0.2 microm/s).

Adenosine Triphosphate↗
PubMed2004

Hysteresis and cell cycle transitions: how crucial is it?

Recently, experiments have shown that cyclin-dependent kinase (CDK) activity exhibits hysteresis in its response to total cyclin when cyclin is made nondegradable and controlled externally. This observation was taken to support mathematical modeling predictions regarding the underlying dynamics of the cell cycle. However, cell cycle dynamics can also be generated by other nonhysteretic mechanisms. To examine the robustness of the hysteretic response of CDK activity to total cyclin, we simulated various cell cycle signal transduction networks, and correlated the dynamics to the response function of CDK activity versus total cyclin. By randomly searching the parameter space, we assessed robustness by estimating the frequency of hysteretic versus nonhysteretic dynamical mechanisms. When the dynamical instabilities were caused by feedback loops in CDK phosphorylation and dephosphorylation or by feedback between cyclin and the CDK inhibitor, the response function of CDK activity versus total cyclin correlated well with the dynamical instabilities. However, when the dynamical instabilities originated from feedback between cyclin and APC-CDH1 or RB-E2F, the response function did not correlate with dynamical instabilities. Thus, although a hysteretic response is neither necessary nor sufficient, it is in general a much more robust mechanism for generating cell cycle dynamics than nonhysteretic mechanisms.

Biological Clocks↗
PubMed2004

Nitric oxide donors protect murine myocardium against infarction via modulation of mitochondrial permeability transition.

Mitochondrial permeability transition (MPT) pores have recently been implicated as a potential mediator of myocardial ischemic injury. Nitric oxide (NO) donors induce a powerful late phase of cardioprotection against ischemia-reperfusion injury; however, the cellular mechanisms involved are poorly understood. The role of MPT pores as a target of cardioprotective signaling pathways activated by NO has never been explored in detail. Thus mice were administered the NO donor diethylenetriamine (DETA)/NO (4 doses of 0.1 mg/kg i.v. each) 24 h before 30 min of coronary artery occlusion followed by 24 h of reperfusion. Infarct size was significantly reduced in DETA/NO-treated mice (30 +/- 2% of risk region in treated mice vs. 50 +/- 2% in control mice; P < 0.05), which demonstrates powerful cardioprotection. To examine the role of MPT pores, mice were administered atractyloside (Atr; 25 mg/kg i.v.), which induces adenine nucleotide translocase-dependent MPT, 20 min before ischemia. Atr blocked the infarct-sparing effects of DETA/NO (infarct size, 58 +/- 1 vs. 30 +/- 2% of risk region in DETA/NO; P < 0.05), whereas Atr alone had no effect. Mitochondria isolated from DETA/NO-treated mice exhibited increased resistance to Ca(2+)-induced swelling by 20 micromol/l CaCl(2) or by the higher concentration of 200 micromol/l, which suggests that cardioprotection involves decreased propensity for MPT. Preincubation of mitochondria from control hearts with 30 nmol/l of the pore inhibitor cyclosporin A prevented swelling by 200 micromol/l CaCl(2), thereby confirming that Ca(2+) induces mitochondrial swelling via MPT. In accordance with the effects on infarct size, administration of Atr to the mice significantly abrogated DETA/NO-induced protection against Ca(2+)-induced mitochondrial swelling. These phenotypic alterations were associated with an increase in the antiapoptotic protein Bcl-2, which suggests that the underlying mechanisms may involve inhibition of cell death by Bcl-2. These data suggest that a critical process during NO donor-induced cardioprotection is to prevent MPT pore opening potentially via targeting of the adenine nucleotide translocator.

Animals↗
PubMed2004

Mother rotors and the mechanisms of D600-induced type 2 ventricular fibrillation.

BACKGROUND: Two types of ventricular fibrillation (VF) have been demonstrated in isolated rabbit hearts during D600 infusion. Type 1 VF is characterized by the presence of multiple, wandering wavelets, whereas type 2 VF shows local spatiotemporal periodicity. We hypothesized that a single mother rotor underlies type 2 VF. METHODS AND RESULTS: One (protocol I) or 2 (protocol II) cameras were used to map the epicardial ventricular activations in Langendorff-perfused rabbit hearts. Multiple episodes of type 2 VF were induced in 22 hearts by high-concentration (> or =2.5 mg/L) D600 (protocol I). During type 2 VF, a single spiral wave (n=19) and/or an epicardial breakthrough pattern (n=11) was present in 14 hearts. These spiral waves either slowly drifted or intermittently anchored on the papillary muscle (PM) of the left ventricle. Dominant-frequency (DF) analyses showed that the highest local DF was near the PM (12.5+/-1.1 Hz). There was an excellent correlation between the highest local DF of these spiral waves and breakthroughs (11.8+/-1.7 Hz) and the DF of simultaneously obtained global pseudo-ECG (11.2+/-1.8 Hz, r=0.97, P<0.0001) during type 2 VF. We also successfully reproduced the major features of type 2 VF by using the Luo-Rudy action-potential model in a simulated, 3-dimensional tissue slab, under conditions of reduced excitability and flat action-potential duration restitution. CONCLUSIONS: Either a stationary or a slowly drifting mother rotor can result in type 2 VF. Colocalization of the stationary mother rotors with the PM suggests the importance of underlying anatomic structures in mother rotor formation.

Action Potentials↗
PubMed2004

Regulation of gating by negative charges in the cytoplasmic pore in the Kir2.1 channel.

Inward rectifier K(+) channels commonly exhibit long openings (slow gating) punctuated by rapid open-close transitions (fast gating), suggesting that two separate gates may control channel open-closed transitions. Previous studies have suggested possible gate locations at the selectivity filter and at the 'bundle crossing', where the two transmembrane segments (M1 and M2) cross near the cytoplasmic end of the pore. Wild-type Kir2.1 channels exhibit only slow gating, but mutations in the cytoplasmic pore domain at E224 and E299 have been shown to induce fast flickery gating. Since these mutations also affect polyamine affinity, we conjectured that the fast gating mechanism might affect the kinetics of polyamine block/unblock if located more intracellularly than the polyamine blocking site in the pore. Neutralization of either E224 or E299 induced fast gating and slowed both block and unblock rates by the polyamine diamine 10. The slowing of polyamine block/unblock was partly relieved by raising pH from 7.2 to 9.0, which also slowed fast gating kinetics. These findings indicate that the fast flickery gate is located intracellularly with respect to the polyamine pore-plugging site near D172, thereby excluding the selectivity filter, and implicating the bundle crossing or more intracellular site as the gate. As additional proof, fast gating induced at the selectivity filter by disrupting P loop salt bridges in WT-E138D-E138D-WT tandem had no effect on polyamine block and unblock rates. The pH sensitivity of fast gating in E224 and E299 mutants was attributed to the protonation state of H226, since the double mutant E224Q/H226K induced fast gating which was pH insensitive. Moreover, introducing a negative charge in the 224-226 region was sufficient to prevent fast gating, since the double mutant E224Q/H226E rescued wild-type Kir2.1 slow gating. These observations implicate E224 and E299 as allosteric modulators of a fast gate, located at the bundle crossing or below in Kir2.1 channels. By suppressing fast gating, these negative charges facilitate polyamine block and unblock, which may be their physiologically important role.

Animals↗
PubMed2004

Coordination of cell growth and cell division: a mathematical modeling study.

Although there is general agreement that cell growth and division are functionally coordinated, the mechanisms that link the two processes are poorly understood. In this study, we have developed a mathematical model based on current biological concepts of the signaling transduction pathways involved in cell growth, which predicts that cell growth rate is proportional to cell surface area at birth. To investigate the relationship between growth control and cell division, we then applied our mathematical model to three classic experiments measuring cycle time versus cell birth size in fission yeast and Xenopus laevis, and the cell cycle delay in mammalian cells after serum withdrawal. When coupled to a cell cycle exhibiting 'sizer' and 'timer' phases, we show that a simple model in which growth rate is proportional to the cell surface area immediately after division reproduces the experimental observations including the relationship between cycle time and birth size in fission yeast and Xenopus laevis. The model also accounts for the cell cycle delay seen in restriction point experiments performed in HeLa cells.

Animals↗