Biomedical subjects

C F Starmer

Publications and source records attributed to C F Starmer.

At least 19 recordsLinked to original sources

PubMed1992

Na channel kinetics remain stable during perforated-patch recordings.

The results of studies on modulation of Na channel function are often difficult to interpret due to time-dependent changes in channel kinetics. Although the "tight-seal" whole cell voltage-clamp technique has proved very useful in studying the properties of the cardiac Na current, the spontaneous shift of parameters of inactivation and activation gating to more negative potential is a serious limitation to the use of the technique. The shifts are believed to result from changes in the intracellular milieu effected by dialysis; moreover, use of a variety of different anions and cations in the internal micropipette solution has not obviated the problem. The perforated-patch technique permits low-resistance intracellular access without free dialysis between the intracellular solution and the recording micropipette. We have compared steady-state inactivation and peak current-voltage relationship of whole cell Na currents measured with the conventional whole cell and perforated-patch techniques in rabbit atrial myocytes at 17 degrees C. Although gating parameters shifted to more negative potentials when recorded with the conventional technique, stable kinetics could be observed for up to 150 min with the perforated-patch technique. The potential for one-half Na channel inactivation was -73 +/- 5.1 mV and is consistent with measurements made using indirect techniques such as upstroke velocity measurements. The fact that the intracellular milieu is left relatively intact makes the approach attractive for studying modulation of the Na current by neurotransmitters and hormones.

Animals↗

PubMed1992

Cardiac instability amplified by use-dependent Na channel blockade.

Drugs that exhibit use-dependent Na channel blockade, including antiarrhythmic agents, tricyclic antidepressants, opiate-like analgesics, and cocaine, are linked with an increased susceptibility to cardiac arrhythmias and sudden death. Computer simulations indicate that Na channel blockade retards recovery of excitability, thereby increasing the spatial dispersion of refractoriness, a precursor of many cardiac arrhythmias. In isolated rabbit left atria, stimuli timed to occur at increasing intervals following conditioning stimuli reveal an unstable interval (vulnerable period) during which single stimuli initiate trains of responses. The vulnerable period is extended by use-dependent Na channel blockade and provides a model for assaying proarrhythmic potential and probing cardiac instability.

Animals↗

PubMed1992

Cellular Vmax reflects both membrane properties and the load presented by adjoining cells.

This study was designed to test the hypothesis that the electrical load seen at a microelectrode impalement site is sensitive to the direction of propagation of the approaching wavefront as a reflection of an altered spatial relationship between the impalement site and the surrounding microscopic electrical boundaries located up- and downstream. These boundaries correspond to the different sizes and shapes of the impaled and surrounding cells as well as to the distribution of the associated electrical connections between the cells. The effects of changes in these geometric relationships on maximum rate of rise of transmembrane potential (Vmax) were investigated in canine ventricular muscle by measuring Vmax in different cells while the direction of propagation was changed from along the longitudinal axis to the transverse axis of the fibers or the direction of conduction was reversed along either of these axes. Comparison of the Vmax values for longitudinal propagation (LP) and transverse propagation (TP), each in one direction, showed that TP Vmax was significantly greater than LP Vmax (P < 0.001). However, the values of Vmax were different from cell to cell during LP (93-139 V/s) and TP (110-181 V/s). The absolute values of LP Vmax and TP Vmax at the same site varied independently of each other, e.g., some of the lowest LP Vmax values occurred at the same site as the highest TP Vmax values. Furthermore, at the same site, Vmax changed considerably when propagation was maintained along the longitudinal axis but the direction of conduction was reversed. Similar prominent changes in Vmax occurred when the direction was reversed along the transverse axis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Kinetics of interaction of the lidocaine metabolite glycylxylidide with the cardiac sodium channel. Additive blockade with lidocaine.

The recovery of the sodium channel from blockade by local anesthetic antiarrhythmic drugs is voltage dependent. Recovery from lidocaine-induced blockade is accelerated by hyperpolarization, whereas that from glycylxylidide (GX) blockade has been reported to be slowed by hyperpolarization. This striking difference occurs despite similarities in chemical structure. The fast recovery from GX block at depolarized potentials may lead to a partial reversal of lidocaine blockade when the two drugs are combined. We have examined the kinetics of interaction of GX with the cardiac sodium channel over a range of membrane potentials by measuring whole-cell currents in isolated rabbit myocytes under voltage clamp at 15 degrees C. In the absence of drug, slow inactivation developed with a time constant of 10.7 +/- 5.1 seconds (n = 6). During exposure to 74 mumol/l GX, block developed with a time constant of 7.0 +/- 3 seconds (n = 6). Because of the similar time course of slow inactivation and block, we used a high concentration of GX to induce a level of block sufficient for analysis. The onset of block was slower than that induced by lidocaine and was unaffected by variation of external sodium from 20 to 75 mmol/l. Use-dependent blockade of sodium channels was greater when pulse trains were applied from a holding potential of -100 than -140 mV. This suggested that recovery from GX block might be slower at -100 than -140 mV. Direct measurements gave time constants of recovery of 10.3 +/- 4.2 seconds at -100 mV (n = 6) and 4.1 +/- 0.4 seconds at -140 mV (n = 4). The combination of GX with lidocaine produced only additive blocking effects when pulse trains were applied from both holding potentials. Computer simulations of the requirements for the competitive displacement of a sodium channel blocker with slow kinetics by one with fast kinetics suggest that the recovery time constant of the fast drug must be 10-100-fold smaller than that of the slow drug. Rapid association kinetics effected by a large binding rate constant or a higher concentration of the fast blocking drug is also important. The simulations suggest that, for the interaction of GX and lidocaine, only additive blocking action should be observed over the range of stimulus frequencies used in these experiments.

Animals↗

PubMed1992

Blockade of cardiac sodium channels. Competition between the permeant ion and antiarrhythmic drugs.

A number of basic and clinical studies suggest that elevation of external sodium concentrations, [Na]o, may reverse the cardiotoxic effect of local anesthetic-class drugs. The mechanisms of reversal are uncertain. The blocking action of lidocaine and disopyramide were studied over a range of [Na]o. Both whole-cell voltage clamp and single-channel recordings were performed on isolated rabbit myocytes at 17 and 22 degrees C, respectively. In the presence of lidocaine, an inactivated channel blocker, the level of steady-state block in response to pulse train stimulation was not affected by variations in [Na]o from 20 to 150 mM. Estimates of the rate of dissociation of drug from the channel also were unaffected. In contrast, steady-state block by disopyramide, a drug that blocks open channels, was decreased as [Na]o was increased. Single-channel measurements suggest that the influence of [Na]o on channel current amplitude was small, 12% for a 25 mM increase in [Na]o. This increase in single-channel current amplitude would affect drug-free channels only, in that our studies suggest that drug-associated channels do not conduct. The association rate constant of disopyramide with open single sodium channels was decreased from 10 x 10(6) to 5 x 10(6)/M per s by an increase in [Na]o from 120 to 180 mM. Elevation of [Na]o may reverse the blocking action of local anesthetic-class drugs by an increase in single-channel current amplitude or by a decrease in drug association rate with the sodium channel. The occurrence of the latter action depends on the mode of block of the specific agent.

Animals↗

PubMed1992

A proarrhythmic response to sodium channel blockade: modulation of the vulnerable period in guinea pig ventricular myocardium.

The vulnerable period (VP) is an interval of time during the cardiac cycle within which premature stimulation may lead to trains of responses (one: many stimulus-response coupling). Although the VP parallels the recovery of sodium channel availability, modulators of its boundaries remain unclear. Numerical studies of a uniform cable demonstrated that reduction in sodium channel availability increased the range of premature stimuli, resulting in unidirectional block, a precursor of reentrant activation. Consequently, we hypothesized that the kinetics of use-dependent sodium channel blockade could reflect one dimension of a drug's proarrhythmic potential. In strips from guinea pig right ventricle, we probed the boundaries of the VP in the presence of use-dependent sodium channel antagonists utilizing a train of stimuli followed by a premature stimulus. Under drug-free conditions when the sites of drive and premature stimulation were the same, the VP was less than 4 ms in duration. When the drive and premature sites were different, the drug-free VP was greater than 5 ms in 22 of 24 preparations and 0 in the other two, with an average VP duration of 16 +/- 10 ms (mean +/- SD). In the presence of 1 microM moricizine, VP = 17 +/- 4 ms; 12 microM moricizine, VP = 35 +/- 4 ms; 3 microM flecainide, VP = 50 +/- 17 ms; and 4 microM quinidine, VP = 2 +/- 1 ms. These results suggest that residual unsuppressed premature ventricular contractions (PVCs) in the presence of some class 1 drugs have a greater potential for initiating a proarrhythmic response than PVCs in the absence of a class 1 drug.

Action Potentials↗

PubMed1991

The quantitative investigation of the binding process of calcium blocking drugs in a sinoatrial node.

Application of existing models of sinoatrial node pacemaker activity and of channel-drug interaction allow us to reproduce action potential changes as a result of the blocking effect of drugs. Two calcium antagonistic drugs, nifedipine and mesudipine, were investigated and as a result averaged rate constants of binding and unbinding were evaluated. The procedure applied which is based on experimental results and on computer simulations, can be used as an initial step for comparison of different drugs.

Action Potentials↗

PubMed1991

A source of bias in the analysis of single channel data: assessing the apparent interaction between channel proteins.

A recent study of single sodium channel currents in neuroblastoma cells suggested interaction between ion channels in close proximity to one another (T. Kiss and K. Nagy, Eur. Biophys. J. 12, 13, 1985). The opening of one channel appeared to affect the likelihood that neighboring channels might open. Some of the conclusions were based on the analysis of observed channel openings that were segregated depending on whether one channel or more than one channel was open at the same time. We hypothesized that the longer one channel remained open, the more likely another channel operating independently, would open, thereby creating the impression of an apparent coupling of channel behavior. We performed simulations and measurements of single sodium channel currents to determine whether the technique of event segregation could account for apparent channel interactions. The simulations showed that the segregation of overlapping (more than one channel open at the same time) and nonoverlapping events led to a bias in the estimated open time and the derived closing rate. To avoid the bias, we found that random pairing of opening and closing events provided an unbiased estimate of the mean closing rate. Using this random assignment approach, we showed that the mean closing rate of single sodium channels in neonatal rat myocytes decreased with depolarization over a limited range of membrane potential. This suggested that the underlying closure mechanism(s) was voltage dependent. From the analysis of open times, we found no evidence for channel interaction in the time scale of tens of milliseconds. Depolarizing steps without events occurred in runs suggesting the existence of long-lived shut state(s). Double pulse experiments with the prepulse and test pulse above threshold showed significant inactivation of channels that did not open. The rate of inactivation of shut channels was substantially slower than the closure rate of open channels. The rate of inactivation of cardiac sodium channels appeared to be strongly dependent on the initial channel state.

Animals↗

PubMed1991

Lidocaine blockade of continuously and transiently accessible sites in cardiac sodium channels.

Lidocaine binds to sodium channels in a voltage dependent manner where depolarization enhances block and hyperpolarization relieves block. Voltage--clamp studies demonstrate that there are two components of block: one involving interaction with a binding site that is accessible for the duration of a depolarizing clamp (continuous access or availability) and one involving interaction with a site that is transiently available or accessible during transitions between polarized and depolarized potentials. Here we report results demonstrating two distinct voltage dependencies of blockade. The voltage dependence of block of the transiently accessible site is similar to that of channel activation and exhibits a maximal binding rate of 1.37 x 10(6)/M/S and an unbinding rate of 39.5/s at -30 mV. Blockade of the sustained site exhibits a voltage dependence similar to inactivation with a maximal binding rate of 3.59 x 10(4)/M/S and an unbinding rate of 0.678/s at -30 mV. Recovery from blockade acquired by either process is voltage dependent and proportional to exp(-0.037 Vm). Drug induced shifts in channel availability and transient site block are accurately predicted from kinetic rates estimated from frequency dependent protocols.

Animals↗

PubMed1991

Modulating intraventricular conduction through competition of two class 1 antiarrhythmic agents: experience with ethacizin and lidocaine in canine heart.

The frequency-dependent effects on the intraventricular conduction through the dog heart in situ produced by two class 1 antiarrhythmic drugs, ethacizin and lidocaine, with different kinetic properties were investigated. Conduction delay was measured using stimulation of the His-bundle after pharmacologically induced atrioventricular (AV) block. Electrical events were derived from local epicardial bipolar electrograms at the base of the right ventricle. The stimulation program consisted of several 50-pulse trains with progressively shorter interstimulus intervals (ISI) separated by a l-s pause. Ethacizin (1.5 mg/kg) increased conduction delay by 30% at ISI of 1000 ms, and the effect was enhanced when ISI was shortened to 200 ms; l-s pauses did not significantly increase conduction velocity. Addition of lidocaine (12 mg/kg) strongly potentiated the ethacizin effect at ISI shorter than 300 ms without any noticeable increase in conduction delay at longer intervals. The major result was dramatic acceleration of conduction during the l-s pauses while both drugs were infused. With this combination, conduction delay after pause was shorter than with ethacizin alone, which is consistent with the competition of the drugs for the same binding site inside the sodium channel. Combination of two class 1 compounds in clinical practice may enhance their antiarrhythmic effects without adversely inhibiting normal impulse conduction in the heart. Computer-predicted data were in reasonable agreement with experimental results. The "guarded receptor" model, thus, can provide a simple method for predicting local anesthetic drug interactions in man.

Animals↗

PubMed1991

Proarrhythmic response to sodium channel blockade. Theoretical model and numerical experiments.

BACKGROUND: The use of flecainide and encainide was terminated in the Cardiac Arrhythmia Suppression Trial because of an excess of sudden cardiac deaths in the active treatment group. Such events might arise from reentrant rhythms initiated by premature stimulation in the presence of anisotropic sodium channel availability. Drugs that bind to sodium channels increase the functional dispersion of refractoriness by slowing (a result of the drug-unbinding process) the transition from an inexcitable state to an excitable state. It is interesting that encainide and flecainide unbind slowly (15-20 seconds), whereas lidocaine and moricizine unbind rapidly (0.2-1.3 seconds). METHODS AND RESULTS: With a computer representation of a cable with Beeler-Reuter membrane properties, we found a small (6 msec) vulnerable window that occurred 338 msec after the last drive stimulus. Premature stimuli falling within the vulnerable window resulted in unidirectional block and reentrant activation. In the presence of a slowly unbinding drug, the window was delayed an additional 341 msec, and its duration was extended to 38 msec. The delay (antiarrhythmic effect) before the onset of the vulnerable window and its duration (proarrhythmic effect) were both dependent on the sodium channel availability and the recovery process. Both effects were also prolonged when sodium channel availability was reduced by membrane depolarization. Defining the proarrhythmic potential as the duration of the vulnerable window, we found that hypothetical use-dependent class I drugs have a greater proarrhythmic potential than non-use-dependent drugs. CONCLUSIONS: The antiarrhythmic and proarrhythmic properties of pure sodium channel antagonists are both dependent on sodium channel availability. Consequently, the price for increased antiarrhythmic efficacy (suppressed premature ventricular contractions) is an increased proarrhythmic vulnerability to unsuppressed premature ventricular contractions.

Anti-Arrhythmia Agents↗

PubMed1991

Blockade of cardiac sodium channels by amitriptyline and diphenylhydantoin. Evidence for two use-dependent binding sites.

Cardiac toxicity is a frequent manifestation in amitriptyline overdose and is felt to be due, in part, to sodium channel blockade by the drug. Another agent with sodium channel blocking properties, diphenylhydantoin, has been used clinically to reverse cardiac conduction abnormalities induced by amitriptyline. This reversal of toxicity is believed to occur secondary to competition for the sodium channel binding site. We evaluated individually and in combination the effects of amitriptyline (0.4 microM) and diphenylhydantoin (10-80 microM) on the sodium current in isolated rabbit atrial and ventricular myocytes at 17 degrees C. Using the whole-cell variant of the patch-clamp technique, we found that both amitriptyline and diphenylhydantoin reduced the sodium current in a use-dependent fashion. The time constant of recovery (tau r) from block by amitriptyline at -130 mV was very slow (13.6 +/- 3.2 seconds), whereas tau r during diphenylhydantoin exposure was fast (0.71 +/- 0.21 seconds, p less than 0.0001 compared with amitriptyline). During exposure of cells to a mixture of the two drugs, tau r was found to be 6.6 +/- 1.8 seconds, but no evidence of direct competition between amitriptyline and diphenylhydantoin was seen. Attempts to fit the recovery data of the mixture to two exponentials resulted in no significant improvement in the fit when compared with that using a single exponential. Use of the sodium channel blocking agent lidocaine (similar kinetics to diphenylhydantoin) in competition with amitriptyline resulted in findings consistent with direct competition of these two drugs for a single binding site. These observations prompted us to evaluate the possibility that diphenylhydantoin was not acting at (and therefore not competing for) the same channel binding site as amitriptyline. Experiments altering pHi and pHo revealed dramatic differences between amitriptyline and diphenylhydantoin. When pHo was increased from 7.4 to 8.0, tau r was reduced approximately threefold (from 13.6 +/- 3.2 to 4.2 +/- 0.1 seconds, p less than 0.0001) during exposure to amitriptyline, but no effect was seen on tau r after exposure to diphenylhydantoin. Conversely, when pHi was increased from 7.3 to 8.0, tau r after amitriptyline was unaffected, but tau r after diphenylhydantoin markedly increased (from 0.71 +/- 0.21 to 2.60 +/- 1.30 seconds, p less than 0.001). Additionally, diphenylhydantoin block demonstrated profound voltage dependence across the range of -130 to -90 mV, whereas amitriptyline block appeared less voltage sensitive. Single-channel studies using patch-clamp techniques in isolated ventricular myocytes supported these data.(ABSTRACT TRUNCATED AT 400 WORDS)

Amitriptyline↗

PubMed1990

An object oriented user interface for analysis of biological data.

In a previous paper we described a self-documented file and a collection of general purpose programs or tools that facilitates the management and analysis of biological data. The tools can be specified in a pipeline to accomplish a specific analysis task. However, we found that it was difficult for investigators to learn the UNIX command language for specifying pipelines, specify selection tasks through a command language, and visualize the data as they were transformed and rearranged. To alleviate these problems we developed an object-oriented user interface for the pipeline programs. The system consists of four major programs for visualization: Vedit, Vgraf, Vscan, and V spread. Vedit is a simple text editor, Vgraf is a flexible graphics program, Vscan facilitates scanning graphically through large files, and Vspread provides spreadsheet-like capabilities. To demonstrate how the visualization programs are used together to accomplish the needed analysis we describe two case studies and then discuss how well the system accomplished the goals of visualization, short learning curve, and user adaptability.

Biology↗

PubMed1990

Use of ionic currents to identify and estimate parameters in models of channel blockade.

Models of ion channel blockade are frequently validated with observations of ionic currents resulting from electrical or chemical stimulation. Model parameters for some models (modulated receptor hypothesis) cannot be uniquely determined from ionic currents. The time course of ionic currents reflects the activation (fraction of available channels that conduct in the presence of excitation) and availability of channels (the ability of the protein to make a transition to a conducting conformation and where this conformation is not complexed with a drug). In the presence of a channel blocking agent, the voltage dependence of availability appears modified and has been interpreted as evidence that drug-complexed channels exhibit modified transition rates between channel protein conformations. Because blockade and availability both modify ionic currents, their individual contributions to macroscopic conductance cannot be resolved from ionic currents except when constant affinity binding to a bindable site is assumed. Experimental studies of nimodipine block of calcium channels and lidocaine block of sodium channels illustrate these concepts.

Animals↗

PubMed1990

Extracellular pH modulates block of both sodium and calcium channels by nicardipine.

Slowing of the recovery of the Na channel from local anesthetic block at low external pH has been well described. In contrast the Ca channel has been reported not to show such an effect. Because of the absence of this response, a two-site model for the interaction of Ca antagonists with the Ca channel has been proposed. We sought to determine whether these results were a consequence of utilizing the poorly lipid-soluble Ca antagonist diltiazem as the test drug. We have measured the time constants of recovery (tau r) of Na (INa) and Ca currents (ICa) recorded from rabbit atrial myocytes during control and exposure to the lipid-soluble Ca antagonist nicardipine as external pH was reduced from 7.8 to 6.9. In the absence of drug, tau r for INa decreased from 19 +/- 1.3 to 12.1 +/- 0.6 ms for the pH reduction. Similarly, tau r for ICa decreased from 53 +/- 7 to 33.6 +/- 7 ms. During exposure to nicardipine tau r for INa increased from 2.24 +/- 0.8 to 5.96 +/- 0.8 s as external pH was reduced. For ICa, tau r also increased from 1.43 +/- 0.55 to 2 +/- 0.32 s. For each channel type, the results are well explained by a single blocking site model with hydrophobic and hydrophilic pathways to a single membrane binding site as proposed by Hille (J. Gen. Physiol. 69: 497-517, 1977).

Animals↗

PubMed1990

Quinidine blocks adenosine 5'-triphosphate-sensitive potassium channels in heart.

The ATP-sensitive potassium channel current (IK-ATP) was studied in excised inside-out patches from rat ventricular cells at 20-23 degrees C. The bath solution contained 140 mM KF, and the pipette solution contained 140 mM KCl and 1.2 mM MgCl2. ATP (0.5 mM) in the bath inhibited IK-ATP. In the absence of ATP, 10 microM quinidine decreased open probability 67 +/- 1% (n = 6) at -50 mV and 28 +/- 12% at -130 mV (n = 5) without affecting single channel conductance (48-52 pS). The block increased with 25 and 50 microM quinidine and could be reversed on washing quinidine for several minutes. Interburst (closed) intervals were increased by quinidine, whereas open and closed time distributions within bursts were not changed. We conclude that quinidine blocks IK-ATP in a "slow" and voltage-dependent manner in clinically relevant concentrations. Because of the postulated role for IK-ATP in cardiac ischemia, quinidine block of this channel may play a role in ischemic arrhythmias.

Adenosine Triphosphate↗

PubMed1990

Channel specificity in antiarrhythmic drug action. Mechanism of potassium channel block and its role in suppressing and aggravating cardiac arrhythmias.

Although work on class III antiarrhythmics remains at an early stage, these agents still appear to possess greater efficacy and less proarrhythmia than conventional class I agents in those experimental arrhythmia models considered to be most representative of the clinical situation. Although prolongation of repolarization carries with its own tendency for pause-dependent arrhythmogenesis (i.e., torsade de pointes), available data suggest that this may be a function of nonspecificity in potassium channel block rather than a general characteristic of class III activity. The availability of new and more selective blockers of specific cardiac potassium channels under development as class III agents have already helped to clarify basic questions about the ionic mechanism of repolarization in the heart, and one hopes that a growing clinical data base will eventually determine the relative safety and efficacy of these agents in preventing symptomatic and life-threatening arrhythmias.

Animals↗

PubMed1990

Managing clinical research data: software tools for hypothesis exploration.

Data representation, data file specification, and the communication of data between software systems are playing increasingly important roles in clinical data management. This paper describes the concept of a self-documenting file that contains annotations or comments that aid visual inspection of the data file. We describe access of data from annotated files and illustrate data analysis with a few examples derived from the UNIX operating environment. Use of annotated files provides the investigator with both a useful representation of the primary data and a repository of comments that describe some of the context surrounding data capture.

Data Interpretation, Statistical↗