Biomedical subjects
Publications and source records attributed to C W Balke.
Treatment of advanced heart failure is usually a challenge. The successful use of beta-blockers to alleviate mild to moderate symptoms of cardiac dysfunction suggests that even severe cases of heart failure may be reversible. Transplantation remains the only sure way to obtain a fully functional heart, but new medications, mechanical devices, and surgical procedures could eventually prove to be alternatives.
1. Whole-cell patch clamp currents from freshly isolated adult rat ventricular cells, recorded in external Ca2+ (Ca2+o) but no external Na+ (Na+o), displayed two inward current components: a smaller component that activated over more negative potentials and a larger component (L-type Ca2+ current) that activated at more positive potentials. The smaller component was not generated by Ca2+ channels. It was insensitive to 50 microM Ni2+ and 10 microM La3+ but suppressed by 10 microM tetrodotoxin (TTX). We refer to this component as ICa(TTX). 2. The conductance-voltage, g(V), relation in Ca2+o only was well described by a single Boltzmann function (half-maximum potential, V1/2, of -44.5; slope factor, k, of -4.49 mV, means of 3 cells). g(V) in Ca2+o plus Na+o was better described as the sum of two Boltzmann functions, one nearly identical to that in Ca2+o only (mean V1/2 of -45.1 and k of -3.90 mV), and one clearly distinct (mean V1/2 of -35.6 and k of -2.31 mV). Mean maximum conductance for ICa(TTX) channels increased 23.7% on adding 1 mM Na+o to 3 mM Ca2+o. ICa(TTX) channels are permeable to Na+ ions, insensitive to Ni2+ and La3+ and blocked by TTX. They are Na+ channels. 3. ICa(TTX) channels are distinct from classical cardiac Na+ channels. They activate and inactivate over a more negative range of potentials and have a slower time constant of inactivation than the classical Na+ channels. They are also distinct from yet another rat ventricular Na+ current component characterized by a much higher TTX sensitivity and by a persistent, non-fast-inactivating fraction. That ICa(TTX) channels activate over a more negative range of potentials than classical cardiac Na+ channels suggests that they may be critical for triggering the ventricular action potential and so of importance for cardiac arrhythmias.
The ryanodine receptor (RyR)/Ca2+ release channel is an essential component of excitation-contraction coupling in striated muscle cells. To study the function and regulation of the Ca2+ release channel, we tested the effect of caffeine on the full-length and carboxyl-terminal portion of skeletal muscle RyR expressed in a Chinese hamster ovary (CHO) cell line. Caffeine induced openings of the full length RyR channels in a concentration-dependent manner, but it had no effect on the carboxyl-terminal RyR channels. CHO cells expressing the carboxyl-terminal RyR proteins displayed spontaneous changes of intracellular [Ca2+]. Unlike the native RyR channels in muscle cells, which display localized Ca2+ release events (i.e., "Ca2+ sparks" in cardiac muscle and "local release events" in skeletal muscle), CHO cells expressing the full length RyR proteins did not exhibit detectable spontaneous or caffeine-induced local Ca2+ release events. Our data suggest that the binding site for caffeine is likely to reside within the amino-terminal portion of RyR, and the localized Ca2+ release events observed in muscle cells may involve gating of a group of Ca2+ release channels and/or interaction of RyR with muscle-specific proteins.
The [Ca2+]i transient in heart is now thought to involve the recruitment and summation of discrete and independent "units" of Ca2+ release (Ca2+ "sparks") from the sarcoplasmic reticulum, each of which is controlled locally by single coassociated L-type Ca2+ channels ("local control theory of excitation-contraction coupling"). All prior studies on Ca2+ sparks, however, have been performed in single enzymatically dissociated heart cells under nonphysiological conditions. In order to understand the possible significance of Ca2+ sparks to normal working cardiac muscle, we used confocal microscopy to record Ca2+ sparks, spatially averaged [Ca2+]i transients and Ca2+ waves in individual cells of intact rat right ventricular trabeculae (composed of < 15 cells in cross section) microinjected with the Ca2+ indicator fluo 3 under physiological conditions ([Ca2+]o, 1 mmol/L; temperature, 33 +/- 1 degree C). Twitch force was recorded simultaneously. When stretched to optimal length (sarcomere length, 2.2 microns) and stimulated at 0.2 Hz, the trabeculae generated approximately equal to 700 micrograms of force per cell. Spatially averaged [Ca2+]i transients recorded from individual cells within a trabecula were similar to those recorded previously from single cells. The amplitude distribution of the peak ratio of Ca2+ sparks was bimodal, with maxima at ratios of 1.8 +/- 0.3 and 2.7 +/- 0.2 (mean +/- SD), respectively. The amplitude of the peak of Ca2+ sparks was approximately equal to 170 nmol/L. Ca2+ sparks occurred at a frequency of 12.0 +/- 0.8/s (mean +/- SEM) in line scans covering 94 sarcomeres. Ca2+ waves occurred randomly at a frequency of 0.57 +/- 0.08/s and propagated with a velocity of 29.5 +/- 1.7 microns/s. The extent of Ca2+ wave propagation was 3.9 +/- 0.3 sarcomere lengths (sarcomere length, 2.2 microns). Ca2+ sparks could be identified along the leading edge of the waves at intervals of 1.30 +/- 0.11 sarcomere length. Our observations suggest that (1) Ca2+ sparks, similar to those recorded in single cells, occur in trabeculae under physiological conditions and (2) coupling of Ca2+ spark generation between neighboring sites occurs and may lead to (3) the development of Ca2+ waves, which propagate under physiological conditions at a low velocity over limited distances. The results suggest that concepts of excitation-contraction coupling recently derived from isolated myocytes are applicable to intact cardiac trabeculae [corrected].
The interpretation of confocal line-scan images of local [Ca2+]i transients (such as Ca2+ sparks in cardiac muscle) is complicated by uncertainties in the position of the origin of the Ca2+ spark (relative to the scan line) and by the dynamics of Ca(2+)-dye interactions. An investigation of the effects of these complications modeled the release, diffusion, binding, and uptake of Ca2+ in cardiac cells (producing a theoretical Ca2+ spark) and image formation in a confocal microscope (after measurement of its point-spread function) and simulated line-scan images of a theoretical Ca2+ spark (when it was viewed from all possible positions relative to the scan line). In line-scan images, Ca2+ sparks that arose in a different optical section or with the site of origin displaced laterally from the scan line appeared attenuated, whereas their rise times slowed down only slightly. These results indicate that even if all Ca2+ sparks are perfectly identical events, except for their site of origin, there will be an apparent variation in the amplitude and other characteristics of Ca2+ sparks as measured from confocal line-scan images. The frequency distributions of the kinetic parameters (i.e., peak amplitude, rise time, fall time) of Ca2+ sparks were calculated for repetitive registration of stereotyped Ca2+ sparks in two experimental situations: 1) random position of the scan line relative to possible SR Ca(2+)-release sites and 2) fixed position of the scan line going through a set of possible SR Ca(2+)-release sites. The effects of noise were incorporated into the model, and a visibility function was proposed to account for the subjective factors that may be involved in the evaluation of Ca(2+)-spark image parameters from noisy experimental recordings. The mean value of the resulting amplitude distributions underestimates the brightness of in-focus Ca2+ sparks because large numbers of out-of-focus Ca2+ sparks are detected (as small Ca2+ sparks). The distribution of peak amplitudes may split into more than one subpopulation even when one is viewing stereotyped Ca2+ sparks because of the discrete locations of possible SR Ca(2+)-release sites in mammalian ventricular heart cells.
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1. The origins of local [Ca2+]i transients (Ca2+ sparks) were studied using dual-channel confocal laser scanning microscopy. Line scan images showing [Ca2+]i (as fluo-3 fluorescence) and the transverse tubule membranes (as Di-8 fluorescence) were obtained simultaneously in single rat cardiac ventricular cells. 2. Line scan images of Di-8 fluorescence showed peaks regularly spaced at intervals of 1.83 +/- 0.30 microns (mean +/- S.D.). These peaks corresponded to the transverse tubules (T-tubules) in cross-section. 3. Line scan images of fluo-3 fluorescence showed local [Ca2+]i transients (LCTs or Ca2+ sparks) evoked by electrical stimulation. 4. Eighty-five per cent (85%) of all Ca2+ sparks evoked by electrical stimulation (n = 138, in 5 cells) occurred within 0.5 micron of a T-tubule. Thirty per cent (30%) occurred within 1 pixel (0.20 micron) of a T-tubule. 5. In some cells studied (3 out of 5), certain T-tubules had a higher probability of being sites of origin of Ca2+ sparks than others. 6. These results support local control theories of excitation-contraction coupling in which Ca2+ release from the sarcoplasmic reticulum (SR) is triggered by a high local [Ca2+]i established between the L-type Ca2+ channels in the T-tubules and associated ryanodine receptor(s) in the junctional SR.
Excitation-contraction coupling was studied in mammalian cardiac cells in which the opening probability of L-type calcium (Ca2+) channels was reduced. Confocal microscopy during voltage-clamp depolarization revealed distinct local transients in the concentration of intracellular calcium ions ([Ca2+]i). When voltage was varied, the latency to occurrence and the relative probability of occurrence of local [Ca2+]i transients varied as predicted if Ca2+ release from the sarcoplasmic reticulum (SR) was linked tightly to Ca2+ flux through L-type Ca2+ channels but not to that through the Na-Ca exchanger or to average [Ca2+]i. Voltage had no effect on the amplitude of local [Ca2+]i transients. Thus, the most efficacious "Ca2+ signal" for activating Ca2+ release from the SR may be a transient microdomain of high [Ca2+]i beneath an individual, open L-type Ca2+ channel.
In heart cells, several distinct kinds of transient spatial patterns of cytoplasmic calcium ion concentration ([Ca2+]i) can be observed: (1) [Ca2+]i waves, in which regions of spontaneously increased [Ca2+]i propagate at high velocity (100 microns/s) through the cell; (2) Ca2+ 'sparks', which are spontaneous, non-propagating changes in [Ca2+]i that are localized in small (approximately 2 microns) subcellular regions; and (3) evoked [Ca2+]i transients that are elicited by electrical depolarization, in association with normal excitation-contraction (E-C) coupling. In confocal [Ca2+]i images, evoked [Ca2+]i transients appear to be nearly spatially uniform throughout the cell, except during their rising phase or during small depolarizations. In contrast to [Ca2+]i waves and spontaneous Ca2+ sparks, evoked [Ca2+]i transients are triggered by L-type Ca2+ channel current and they are 'controlled', in the sense that stopping the L-type Ca2+ current stops them. Despite their different characteristics, all three types of Ca2+ transient involve Ca(2+)-induced release of Ca2+ from the sarcoplasmic reticulum. Here, we address the question of how the autocatalytic process of Ca(2+)-induced Ca2+ release, which can easily be understood to underlie spontaneous regenerative ('uncontrolled'), propagating [Ca2+]i waves, might be 'harnessed', under other circumstances, to produce controlled changes in [Ca2+]i, as during normal excitation-contraction coupling, or changes in [Ca2+]i that do not propagate. We discuss our observations of Ca2+ waves, Ca2+ sparks and normal Ca2+ transients in heart cells and review our results on the 'gain' of Ca(2+)-induced Ca2+ release. We discuss a model involving Ca2+ microdomains beneath L-type Ca2+ channels, and clusters of Ca(2+)-activated Ca2+ release channels in the sarcoplasmic reticulum which may form the basis of the answer to this question.
1. Confocal microscopy and the fluorescent Ca2+ indicator fluo-3 (K+ salt) were used to measure cytosolic free calcium ion concentration ([Ca2+]) during excitation-contraction (E-C) coupling in single, voltage-clamped, rat cardiac ventricular cells. 2. Local [Ca2+]i transients were measured nearly simultaneously in different, separate, subcellular volumes of approximately 2.0 microns 3. During depolarization, local [Ca2+]i transients were distinctly different from each other and from whole-cell [Ca2+]i transients. These differences were particularly apparent during small depolarizations, and were substantially reduced by ryanodine. 3. Components of the local [Ca2+]i transients, particularly those evoked by small depolarizations, were closely similar, in time course and amplitude, to spontaneous local [Ca2+]i transients, or 'sparks' (which have been shown previously to be Ca2+ released from sarcoplasmic reticulum). 4. Analysis of local [Ca2+]i transients in the spatial frequency domain (power spectrum) revealed that high power at spatial frequencies of 0.05-0.2 microns-1 was always associated with spontaneous calcium 'sparks' and with local [Ca2+]i transients evoked by small depolarizing pulses (e.g. to -31 mV). Evoked local [Ca2+]o transients in the presence of ryanodine, and those evoked by depolarization to very positive clamp-pulse potentials (+45 mV), were associated with considerably lower power at this frequency. 5. The results suggest that whole-cell [Ca2+]i transients evoked by voltage-clamp depolarization, and thus by L-type Ca2+ current, are comprised of local [Ca2+]i transients that are similar to the spontaneous calcium 'sparks'. At very positive clamp-pulse potentials, however, the electrically evoked local [Ca2+]i transients may be smaller, perhaps as a result of smaller unitary L-type Ca2+ current.
1. The processes that remove Ca2+ rapidly from the cytoplasm were studied in isolated rat ventricular myocytes subjected to whole-cell voltage clamp and internal perfusion with the Ca2+ indicator, indo-1. Na(+)-Ca2+ exchange was eliminated in most experiments by removing Na+ both internally and externally. 2. When the Ca(2+)-pumping ATPase of the sarcoplasmic reticulum (SR) was inhibited with cyclopiazonic acid and ryanodine interfered with the release of Ca2+ from the SR, [Ca2+]i transients rose slowly and declined extremely slowly. We concluded that transport of Ca2+ by mitochondria and the surface membrane Ca(2+)-pumping ATPase would be negligible over the time course of a single [Ca2+]i transient. 3. The influence of cytoplasmic Ca2+ ligands was characterized by internal perfusion with high concentrations of diffusible Ca2+ ligands (indo-1) or by superfusion with the membrane-permeant Ca2+ ligand, BAPTA AM. As the concentration of indo-1 in the cell increased from < 0.1 mM to at least 0.5 mM, the time constant of the decline of [Ca2+]i increased from about 0.15 s to nearly 3 s. 4. Calcium bound to endogenous Ca2+ ligands during depolarizing clamp pulses was characterized quantitatively as the difference between the total Ca2+ entering the cell via L-type Ca2+ channels and [Ca2+]i, in experiments in which SR function had been abolished. As total calcium increased during the entry of Ca2+, total calcium was found to agree reasonably well with that predicted by assuming that Ca2+ could bind to endogenous intracellular Ca2+ ligands and to indo-1. 5. The results indicate that, in the absence of Na+, the major factors determining the removal of cytoplasmic free Ca2+ are the Ca(2+)-pumping ATPase of the SR and the binding of Ca2+ to endogenous and exogenous Ca2+ ligands. 6. Several hypothetical 'Ca2+ removal functions' were fitted to the declining phase of [Ca2+]i transients. The best fit was one in which the flux of Ca2+ through the SR Ca(2+)-pumping ATPase was described by a Michaelis-Menten-type equation. The decline of the [Ca2+]i transient was thus described by a linear, first-order differential equation having terms giving the rate of Ca2+ transport by the SR Ca(2+)-pumping ATPase (Vmax and KM), the rates of complexation of Ca2+ with the various Ca2+ ligands (L), and a leak of Ca2+ into the cytoplasm from the SR (FSR,leak).(ABSTRACT TRUNCATED AT 400 WORDS)
1. Cytosolic free calcium ion concentration ([Ca2+]i) and whole-cell L-type Ca2+ channel currents were measured during excitation-contraction (E-C) coupling in single voltage-clamped rat cardiac ventricular cells. The measurements were used to compute the total cellular efflux of calcium ions through sarcoplasmic reticulum (SR) Ca2+ release channels (FSR,rel) and the influx of Ca2+ via L-type Ca2+ channels (FICa). 2. FSR,rel was elicited by depolarizing voltage-clamp pulses 200 ms in duration to membrane potentials from -30 to +80 mV. Over this range, peak FSR,rel had a bell-shaped dependence on clamp pulse potential. In all cells, the 'gain' of the system, measured as the ratio, FSR,rel(max)/FICa(max), declined from about 16, at 0 mV, to much lower values as clamp pulse voltage was made progressively more positive. We named this phenomenon of change in gain as a function of membrane potential, 'variable gain'. At clamp pulse potentials in the range -30 to 0 mV, the gain differed from cell to cell, being constant at about 16 in some cells, but decreasing from high values (approximately 65) at -20 mV in others. 3. At clamp pulse potentials at which Ca2+ influx (FICa) was maintained, FSR,rel also had a small maintained component. When macroscopic Ca2+ influx was brief (1-2 ms, during 'tails' of FICa), FSR,rel rose rapidly to a peak after repolarization and then declined with a half-time of about 9 ms (typically). 4. The rising phase of [Ca2+]i transients could be interrupted by stopping Ca2+ influx rapidly (by voltage clamp). We therefore termed this phenomenon 'interrupted SR Ca2+ release'.(ABSTRACT TRUNCATED AT 250 WORDS)
It is universally believed that the removal of external sodium ions is without effect on calcium current. We now report that in enzymatically isolated guinea pig ventricular cells, the replacement of external sodium ions with certain other cations causes a 3- to 6-fold increase in peak L-type calcium current. The increase in current is reversibly blocked by L-type calcium-channel antagonists, not mediated by changes in internal calcium, and is inhibited by intracellular 5'-adenylyl imidodiphosphate, a nonhydrolyzable ATP analogue. The effects of sodium removal (and isoproterenol) were almost completely blocked by intracellular application of a specific (peptide) inhibitor of cAMP-dependent protein kinase. These experiments demonstrate a previously unknown effect of sodium ions to modulate calcium-channel phosphorylation via cAMP-dependent protein kinase.
1. We sought to distinguish two types of Ca2+ channel in guinea-pig ventricular cells (T-type and L-type) and to characterize their respective gating and permeation properties when Ca2+ (1-10 mM) is the charge carrier, as is the case physiologically. 2. Na+ was removed from both the external and internal solutions to eliminate currents through Na+ channels and Na(+)-Ca2+ exchange. Major differences in the voltage dependence of steady-state inactivation were exploited to separate the two Ca2+ current components. 3. From a holding potential of -50 mV, only L-type channels were available to open with depolarization. When holding at -90 mV, T-type channels contributed an additional rapidly inactivating component superimposed upon the L-type current. Only the L-type channels thus identified were sensitive to the dihydropyridine Ca2+ channel blocker nitrendipine. 4. T-type currents, measured by taking the difference between the currents elicited from a holding potential of -90 mV and those elicited from -50 mV, peaked within 10 ms and decayed completely within 50-100 ms. 5. Macroscopic T-type currents were largest during depolarizing pulses between -40 and -30 mV (peak current density of 0.62 +/- 0.21 nA nF-1) and decreased at more positive potentials, becoming unmeasurably small above 0 mV. 6. Unitary currents recorded with similar ionic conditions and voltage protocols exhibited a single-channel conductance of 4-5 pS in 10 mM Ca2+. Ensemble average currents through a single channel reproduced accurately the time course of whole-cell T-type current. Permeation properties could not explain the absence of macroscopic T-type currents at positive test potentials, which must therefore be attributable to gating. 7. Convolution analysis was employed to clarify the single-channel basis of the rapidly decaying current waveform of T-type channels. The latencies to first opening and reopening, which reflect activation and deactivation, influenced the waveform most strikingly. Open times were sufficiently brief that they contributed little to shaping the average current. Thus, macroscopic inactivation largely reflects rate-limiting activation events. 8. The unitary current amplitudes and peak open probabilities measured for single T-type channels, when compared to the average macroscopic T-type current density, predict 10.6 functional channels per picofarad, or approximately 1700 T-type channels per typical ventricular myocyte.
1. We investigated the currents through L-type Ca2+ channels when Ca2+ (1-10 mM) was the charge carrier, as is the case physiologically. 2. Na+ was removed from both the external and internal solutions to eliminate currents through Na+ channels and Na(+)-Ca2+ exchange. 3. From a holding potential of -50 mV only L-type channels were available to open with depolarization. Macroscopic L-type currents were maximal during depolarizing pulses to +10 mV (peak current density of 4.7 +/- 0.3 nA nF-1). 4. During depolarizing steps as long as 180 ms, the decay of current through L-type channels was incomplete, in contrast to that of T-type current. 5. Unitary currents recorded with comparable ionic conditions and voltage protocols exhibited a single-channel conductance of 6.9 pS in 10 mM Ca2+. Ensemble average currents reproduced accurately the features of whole-cell L-type current, including the maintained component. 6. Convolution analysis was employed to clarify the single-channel basis of the complex current waveform of L-type channels. First openings underlie the peak, while the maintained pedestal is generated by multiple re-openings. As with T-type channels, single openings are brief and contribute little to the time course of the average current. 7. The prominent maintained component of macroscopic and ensemble average L-type current cannot be explained by simple Markov models in which current decay reflects the progressive entry of channels into an absorbing inactivated state. 8. We considered the possibility that the maintained component of current arises from the existence of multiple distinct gating patterns, one of which lacks inactivation. Individual sweeps were sorted among three patterns of gating (no openings, active-early and active-late). Patterns of activity are not randomly distributed; instead, they tend to cluster over time. 9. Most of the maintained current is attributable to the 'active-late' pattern of gating. Considered separately, this pattern can be well described by a simple Markov chain lacking an inactivated state. The 'active-early' gating pattern accounts entirely for the initial current transient, and for about one-third of the maintained component; thus, inactivation, even when present, must be reversible rather than absorbing. 10. The unitary current amplitudes and peak open probabilities measured for single L-type channels, when compared to the average macroscopic L-type current density, predict 170 functional channels per picofarad, or 28,000 L-type channels per typical ventricular myocyte.(ABSTRACT TRUNCATED AT 400 WORDS)
Calcium current in mammalian ventricular muscle is altered in the presence of ryanodine. Previous studies performed on rat ventricular cells have shown a slowing of Ca2+ current inactivation and suggest the hypothesis that ryanodine, by reducing the release of Ca2+ from the sarcoplasmic reticulum, reduces the availability of Ca2+ for inactivation of Ca2+ current (Ca(2+)-dependent inactivation). Another hypothesis is that the effects of ryanodine on Ca2+ current are due to a mechanical connection of the ryanodine receptor with the L-type Ca2+ channel. To further test these hypotheses we examined the effect of ryanodine on Ca2+ current in single voltage-clamped guinea pig ventricular myocytes that contained Ca2+ indicator and Ca2+ buffer. We used fura 2 (pentapotassium salt) to confirm that the ryanodine we used was capable of abolishing Ca2+ release from the sarcoplasmic reticulum during the period in which it was present. We perfused the cells with 10 mM EGTA to block changes in intracellular Ca2+ concentration. In the absence of internal EGTA, Ca2+ currents displayed biexponential inactivation and Ca(2+)-dependent inactivation (steady-state inactivation curves turned up at positive potentials). Inactivation was slowed by ryanodine at 10 microM. In cells perfused internally with EGTA, however, ryanodine had no effects, and steady-state inactivation curves were not shifted to the right. We conclude that, in guinea pig ventricular myocytes, the effects of ryanodine on Ca2+ current are mediated by Ca2+ and thus the effects of ryanodine do not provide a basis on which to postulate a physical connection between the L-type Ca2+ channel and the ryanodine receptor (sarcoplasmic reticulum Ca2+ release channel).
Experiments were performed in vitro on six normal thin ventricular epicardial tissue strips and 10 strips removed from the infarcted regions of dogs 21-60 days after experimental myocardial infarction. Conduction was evaluated by mapping activation sequences at 40-45 sites over an area of 1 x 2 cm during pacing at a basic cycle length of 2,000 msec. The amplitude and length of recorded electrograms were also determined at each site. After control recordings, heptanol, which increases gap junctional resistance, was added to the tissue bath at concentrations ranging between 0.2 and 1.0 mM. In contrast to its effect on normal tissues, heptanol caused 75 of 260 previously active sites in the infarcted tissues to become inactive. The affected sites were located in areas of very slow conduction and/or adjacent to areas of preexisting conduction block. In addition, heptanol decreased the length and degree of fractionation of electrograms recorded in slowly conducting regions of the infarcted tissues. The magnitude of the decrease in electrogram length following heptanol was related to the degree of electrogram abnormality during control as reflected in the ratio of electrogram length to amplitude. Heptanol shortened electrograms by causing local conduction block, which eliminated some components of the fractionated electrograms. In an additional eight epicardial strips removed from the infarcted region, 0.5 mM heptanol had only a slight effect (10.7% decrease) on the maximum rate of membrane depolarization. Thus, heptanol does not act primarily by way of depressing the fast inward current. We conclude from heptanol's effects on conduction and electrogram characteristics that slow and dissociated conduction in the infarcted region is due to an abnormality in gap junctional distribution between surviving cells and/or an abnormality in individual gap junctional function.
The maximum amplitude of vector loops formed by summing orthogonally recorded bipolar electrograms has been shown to reflect the direction of activation in cardiac muscle. To investigate whether components of vector loops could provide information about different activation directions in local areas of myocardium, we correlated "instantaneous vectors" with isochronal activation patterns in an in vitro preparation of experimental myocardial infarction. In thirteen 3 mm x 3 mm regions studied (from 11 tissues), at least 16 microelectrode impalements with a minimum density of 0.8 mm between sites were made. In seven situations in which notched, irregular, and prolonged duration electrograms were present, vector loops pointed in the same general direction throughout their entire time course. In these preparations, microelectrode impalements demonstrated only a single major direction of activation. In five of six areas in which multidirectional vector loops were present, two or more separate local directions of activation corresponded to the directions of the vector loop. Instantaneous vectors were then used to analyze propagation patterns in vivo in 10 animals with 2-4-week-old experimental myocardial infarction. Of 150 sites in the 10 animals, 13% contained more than one major local direction of activation. In 11 markedly abnormal sites, electrograms were recorded during pacing from four sites around the recording probe. When comparing electrogram characteristics from the four sites, a mean difference of 5.9 mV in electrogram amplitude and 19.1 msec in electrogram duration (coefficient of variation, 27% for amplitude and 22% for duration) was found. In only two of the 11 sites was it found that the same number of activation directions occurred from all pacing sites. We conclude: 1) Instantaneous components of vector loops accurately represent local directions of cardiac activation at differing times. 2) Most areas of experimental myocardial infarction have only one major direction of activation despite the presence of abnormal electrograms. 3) In some regions, however, two major local directions of activation can be identified within a relatively local area. 4) Geometric activation patterns in experimental myocardial infarction are markedly dependent on initial activation direction.