Biomedical subjects

T F Wiesner

Publications and source records attributed to T F Wiesner.

11 recordsLinked to original sources

PubMed2001

Potentiation of Ca(2+) release by cADP-ribose in the heart is mediated by enhanced SR Ca(2+) uptake into the sarcoplasmic reticulum.

cADP-Ribose (cADPR) is a novel endogenous messenger that is believed to mobilize Ca(2+) from ryanodine-sensitive Ca(2+) stores. Despite intense research, the precise mechanism of action of cADPR remains uncertain, and experimental findings are contradictory. To elucidate the mechanism of cADPR action, we performed confocal Ca(2+) imaging in saponin-permeabilized rat ventricular myocytes. Exposure of the cells to cADPR resulted in a slow (>2 minutes) and steady increase in the frequency of Ca(2+) sparks. These effects on local release events were accompanied by a significant increase in sarcoplasmic reticulum (SR) Ca(2+) content. In comparison, sensitization of ryanodine receptors (RyRs) by caffeine, a true RyR agonist, caused a rapid (<1 second) and transient potentiation of Ca(2+) sparks followed by a decrease in SR Ca(2+) content. When the increase in the SR load was prevented by partial inhibition of the SR Ca(2+) with thapsigargin, cADPR failed to produce any increase in sparking activity. cADPR had no significant impact on activity of single cardiac RyRs incorporated into lipid bilayers. However, it caused a significant increase in the rate of Ca(2+) uptake by cardiac SR microsomes. Our results suggest that the primary target of cADPR is the SR Ca(2+) uptake mechanism. Potentiation of Ca(2+) release by cADPR is mediated by increased accumulation of Ca(2+) in the SR and subsequent luminal Ca(2+)-dependent activation of RyRs.

Adenosine Diphosphate Ribose↗

PubMed2001

Dynamic regulation of sarcoplasmic reticulum Ca(2+) content and release by luminal Ca(2+)-sensitive leak in rat ventricular myocytes.

In cardiac muscle, excitation-contraction (E-C) coupling is determined by the ability of the sarcoplasmic reticulum (SR) to store and release Ca(2+). It has been hypothesized that the Ca(2+) sequestration and release mechanisms might be functionally linked to optimize the E-C coupling process. To explore the relationships between the loading status of the SR and functional state of the Ca(2+) release mechanism, we examined the effects of changes in SR Ca(2+) content on spontaneous Ca(2+) sparks in saponin-permeabilized and patch-clamped rat ventricular myocytes. SR Ca(2+) content was manipulated by pharmacologically altering the capacities of either Ca(2+) uptake or leak. Ca(2+) sparks were recorded using a confocal microscope and Fluo-3 and were quantified considering missed events. SR Ca(2+) content was assessed by application of caffeine. Exposure of permeabilized cells to anti-phospholamban antibodies elevated the SR Ca(2+) content and increased the frequency of sparks. Suppression of the SR Ca(2+) pump by thapsigargin lowered [Ca(2+)](SR) and reduced the frequency of sparks. The ryanodine receptor (RyR) blockers tetracaine and Mg(2+) transiently suppressed the frequency of sparks. Upon washout of the drugs, sparking activity transiently overshot control levels. Low doses of caffeine transiently potentiated sparking activity upon application and transiently depressed the sparks upon removal. In patch-clamped cardiac myocytes, exposure to caffeine produced only a transient increase in the probability of sparks induced by depolarization. We interpret these results in terms of a novel dynamic control scheme for SR Ca(2+) cycling. A central element of this scheme is a luminal Ca(2+) sensor that links the functional activity of RyRs to the loading state of the SR, allowing cells to auto-regulate the size and functional state of their SR Ca(2+) pool. These results are important for understanding the regulation of intracellular Ca(2+) release and contractility in cardiac muscle.

Animals↗

PubMed2001

Underlying mechanisms of symmetric calcium wave propagation in rat ventricular myocytes.

Calcium waves in heart cells are mediated by diffusion-coupled calcium-induced calcium release. The waves propagate in circular fashion. This is counterintuitive in view of the accepted ultrastructure of the cardiac myocyte. The density of calcium release sites in the transverse direction is four times higher than in the longitudinal direction. Simulations with release sites localized along Z-lines and isotropic diffusion yielded highly elliptical, nonphysiological waves. We hypothesized that subcellular organelles counteracted the higher release site density along the Z-lines by acting as transverse diffusion barriers and sites of active calcium uptake. We quantified the reduction of transverse diffusion by microinjecting cells with the nonreactive dye fluorescein. The ratio of the radial diffusion coefficient to the longitudinal coefficient was 0.39. Inhibition of mitochondrial uptake by rotenone accelerated the wave in the transverse direction. Simulations with release sites clustered at the Z-lines and a transverse diffusion coefficient 50% of the longitudinal coefficient generated waves of ellipticity 2/1 (major axis along the Z-line). Introducing additional release sites between the Z-lines at a density 20% of that on the Z-lines produced circular waves. The experiments and simulations support the presence of transverse diffusion barriers, additional uptake sites, and possibly intermediate release sites as well.

Animals↗

PubMed2000

Inhibition of Ca(2+) sparks by ruthenium red in permeabilized rat ventricular myocytes.

We have compared the effects of the sarcoplasmic reticulum (SR) Ca(2+) release inhibitor, ruthenium red (RR), on single ryanodine receptor (RyR) channels in lipid bilayers, and on Ca(2+) sparks in permeabilized rat ventricular myocytes. Ruthenium red at 5 microM inhibited the open probability (P(o)) of RyRs approximately 20-50-fold, without significantly affecting the conductance or mean open time of the channel. At the same concentration, RR inhibited the frequency of Ca(2+) sparks in permeabilized myocytes by approximately 10-fold, and reduced the amplitude of large amplitude events (with most probable localization on the line scan) by approximately 3-fold. According to our theoretical simulations, performed with a numerical model of Ca(2+) spark formation, this reduction in Ca(2+) spark amplitude corresponds to an approximately 4-fold decrease in Ca(2+) release flux underlying Ca(2+) sparks. Ruthenium red (5 microM) increased the SR Ca(2+) content by approximately 2-fold (from 151 to 312 micromol/l cytosol). Considering the degree of inhibition of local Ca(2+) release events, the increase in SR Ca(2+) load by RR, and the lack of effects of RR on single RyR open time and conductance, we have estimated that Ca(2+) sparks under normal conditions are generated by openings of at least 10 single RyRs.

Animals↗

PubMed1999

The role of luminal Ca2+ in the generation of Ca2+ waves in rat ventricular myocytes.

1. We used confocal Ca2+ imaging and fluo-3 to investigate the transition of localized Ca2+ releases induced by focal caffeine stimulation into propagating Ca2+ waves in isolated rat ventricular myocytes. 2. Self-sustaining Ca2+ waves could be initiated when the cellular Ca2+ load was increased by elevating the extracellular [Ca2+] ([Ca2+]o) and they could also be initiated at normal Ca2+ loads when the sensitivity of the release sites to cytosolic Ca2+ was enhanced by low doses of caffeine. When we prevented the accumulation of extra Ca2+ in the luminal compartment of the sarcoplasmic reticulum (SR) with thapsigargin, focal caffeine pulses failed to trigger self-sustaining Ca2+ waves on elevation of [Ca2+]o. Inhibition of SR Ca2+ uptake by thapsigargin in cells already preloaded with Ca2+ above normal levels did not prevent local Ca2+ elevations from triggering propagating waves. Moreover, wave velocity increased by 20 %. Tetracaine (0.75 mM) caused transient complete inhibition of both local and propagating Ca2+ signals, followed by full recovery of the responses due to increased SR Ca2+ accumulation. 3. Computer simulations using a numerical model with spatially distinct Ca2+ release sites suggested that increased amounts of releasable Ca2+ might not be sufficient to generate self-sustaining Ca2+ waves under conditions of Ca2+ overload unless the threshold of release site Ca2+ activation was set at relatively low levels (< 1.5 microM). 4. We conclude that the potentiation of SR Ca2+ release channels by luminal Ca2+ is an important factor in Ca2+ wave generation. Wave propagation does not require the translocation of Ca2+ from the spreading wave front into the SR. Instead, it relies on luminal Ca2+ sensitizing Ca2+ release channels to cytosolic Ca2+.

Algorithms↗

PubMed1999

Dynamic reoptimization of a fed-batch fermentor.

Traditionally, fed-batch biochemical process optimization and control use complicated models and off-line optimizers with no on-line model adaptation and re-optimization. This work demonstrates the applicability, effectiveness, and economic potential, of a simple phenomenological model for modeling, and a novel optimizer for on-line re-optimization and control of an aerobic fed-batch fermentor.

Animals↗

PubMed1998

Termination of Ca2+ release during Ca2+ sparks in rat ventricular myocytes.

1. Confocal Ca2+ imaging was used to measure spontaneous release events (Ca2+ sparks) in fluo-3-loaded isolated rat ventricular myocytes. 2. The microscopic Ca2+ release flux underlying Ca2+ sparks was derived by adapting the methods used previously to describe macroscopic Ca2+ release from cell-averaged Ca2+ transients. 3. The magnitude of the local release fluxes varied from 2 to 5 microM ms-1, depending on SR Ca2+ loading conditions. Following spontaneous activation, the release flux rapidly decayed (tau = 6-12 ms). The rate of termination of release flux was found to be directly related to the magnitude of the flux (r2 = 0.88). 4. The rate of termination of local release flux was slowed in the presence of FK506, a compound that is known to reduce inactivation of SR Ca2+ channels in vitro. 5. These results suggest that termination of release flux during sparks is not due to a spontaneous stochastic decay process or local depletion of Ca2+ from the SR, but rather involves an active extinguishing mechanism such as Ca2+-dependent inactivation or adaptation.

Algorithms↗

PubMed1997

A mathematical model of the cytosolic-free calcium response in endothelial cells to fluid shear stress.

Important among the responses of endothelial cells to flow stimuli are cytosolic-free calcium transients. These transients are mediated by several factors, including blood-borne agonists, extracellular calcium, and fluid-imposed shear forces. A mathematical model has been developed describing the recognition and transduction of shear stress to the second messenger cytosolic calcium. Shear stress modulates the calcium response via at least two modalities. First, mass transfer of agonist to the cell surface is enhanced by perfusion and is thus related to shear stress. Second, the permeability of the cell membrane to extracellular calcium increases upon exposure to shear stress. A mass balance for agonist in the perfusate is coupled to a previously published calcium dynamics model. Computations indicate a flow region where the transient moves from transport limited to kinetically limited. Parametric studies indicate distinct contributions to the time course by each step in the process. These steps include the time to develop the concentration boundary layer of agonist, receptor activation, and the mobilization of calcium from intracellular stores. Exogenous calcium is presumed to enter the cell via shear stress-gated ion channels. The model predicts a sigmoidal dependence of calcium influx upon shear stress. The peak value of the transient is determined largely by the agonist pathway, whereas the plateau level is governed by calcium influx. The model predicts the modulation of the calcium transient in the physiologically relevant range of flow and the associated shear stress. This implies that hemodynamics is important in regulating endothelial biology.

Animals↗

PubMed1996

A mathematical model of cytosolic calcium dynamics in human umbilical vein endothelial cells.

Important among the responses of endothelial cells are cytosolic free calcium transients. These transients are mediated by several factors, including blood-borne agonists, extracellular calcium, and fluid-imposed shear forces. The transients are characterized by a rapid rise followed by a plateau phase. A base mathematical model is presented that reasonably reproduces the measured calcium transient in cultured human umbilical vein endothelial cells responding to thrombin. Kinetic equations for receptor activation and calcium mobilization comprise the model. A graded response of intracellular free calcium to increasing concentrations of agonist is predicted. Also predicted is the elevation of the peak value and the plateau level by steady nonspecific leak of calcium across the plasma membrane. The influences of capacitative calcium entry, calcium-induced calcium release, and buffering by cytosolic proteins are investigated parametrically. The model predicts significant depletion of cellular calcium in response to agonist stimulation.

Buffers↗

PubMed1988

Epicardial coronary blood flow including the presence of stenoses and aorto-coronary bypasses--II: Experimental comparison and parametric investigations.

This article is the second in a series which presents a computer model of the left coronary arteries. The first article discussed the geometry, the governing equations, and the numerical method employed. This paper details an acute canine experiment used to validate the approach as well as the systematic investigation of several important parameters governing the left coronary circulation. These parameters include peripheral resistance, wall properties, and altered geometric properties through various stenosis/bypass configurations. With appropriate selection of parameters, the model reproduces an in vivo waveform very closely. The model also predicts many clinical phenomena, such as the "critical" value of stenosis, the dramatic reduction in flow through a stenosis when bypassed, and the restorative effect of the bypass upon flow to the distal bed. The model also is used to show that the autonomic state of the animal profoundly affects the influence of various factors, e.g., the critical value of a stenosis is much higher under resting conditions than under hyperemic conditions.

Algorithms↗

PubMed1985

Epicardial coronary blood flow including the presence of stenoses and aorto-coronary bypasses--I: Model and numerical method.

A computer model and numerical method for calculating left epicardial coronary blood flow has been developed. This model employs a finite-branching geometry of the coronary vasculature and the one-dimensional, unsteady equations for flow with friction. The epicardial coronary geometry includes the left main and its bifurcation, the left anterior descending and left circumflex coronary arteries, and a selected number of small branches. Each of the latter terminate in an impedance, whose resistive component is related to intramyocardial compression through a linear dependence on left ventricular pressure. The elastic properties of the epicardial arteries are taken to be non-linear and are prescribed by specifying the local small-disturbance wave speed. The model allows for the incorporation of multiple stenoses as well as aorto-coronary bypasses. Calculations using this model predict pressure and flow waveform development and allow for the systematic investigation of the dependence of coronary flow on various parameters, e.g., peripheral resistance, wall properties, and branching pattern, as well as the presence of stenoses and bypass grafts. Reasonable comparison between calculations and earlier experiments in horses has been obtained.

Animals↗