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Abstract Guinea pig hearts were stained with a voltage-sensitive dye and imaged on a photodiode array to record fluorescent action potentials (APs) from 124 sites. Activation and repolarization patterns were recorded from the epicardium during stimulation at different loci and correlated with the underlying fiber architecture. Endocardial APs were recorded by inserting a light guide into the ventricular cavity or by dissecting out the ventricular free wall to expose the endocardium. In hearts paced on the right atrium to simulate sinus rhythm, activation emerged synchronously over a large area of the ventricular epicardium and spread laterally in 5 to 7 ms. The apparent longitudinal and transverse velocities were 2.66±0.11 and 1.65±0.09 m/s (n=12). In contrast, repolarization began near the apex on the endocardium and spread transmurally in 6±1.3 ms (n=12) and then anisotropically along the epicardium in 25 to 30 ms with apparent maximum (0.53±0.11 m/s) and minimum (0.31±0.10 m/s) repolarization velocities that aligned with the longitudinal and transverse axes of epicardial fibers. When paced on the epicardium, activation of intact hearts (n=12) and perfused sheets (n=8) was anisotropic, with longitudinal (0.85±0.05 m/s) and transverse (0.44±0.04 m/s) conduction velocities that aligned with the epicardial fiber orientation. When activation was initiated at different sites on the epicardium, repolarization always began near the apex and exhibited patterns similar to those obtained under right atrial pacing, but with slower longitudinal (0.41±0.09 m/s) and transverse (0.23±0.07 m/s) repolarization velocities (n=18). In sheets stretched parallel to the longitudinal axis of surface fibers, AP durations (APDs) increased as a function of fiber length, from the length at zero developed tension to 120% of the length at maximum developed tension (L max ). Spatial distributions of APDs did not change during stretches along the rising phase of the length-tension curve. In sheets stretched to 50% of L max , APDs were shorter and more homogeneous on the endocardium (mean APD, 188 ms; ΔAPD, 195−186=9 ms) than on the epicardium (mean APD, 204 ms; ΔAPD, 212−186=26 ms; n=8). In guinea pig hearts, activation is rapid; therefore, repolarization depends primarily on intrinsic spatial heterogeneities of APDs. Consequently, repolarization begins at endocardial cells with the shortest APDs and spreads transmurally and then anisotropically on the surface according to the epicardial cell orientation.

Abstract Shear stress causes the vascular endothelium to release nitric oxide (NO), which is an important regulator of vascular tone. However, direct measurement of NO release after the imposition of laminar flow has not been previously accomplished because of chemical (oxidative degradation) and physical (diffusion, convection, and washout) complications. Consequently, the mechanism, time course, kinetics, and Ca 2+ dependence of NO release due to shear stress remain incompletely understood. In this study, we characterized these parameters by using fura 2 fluorescence and a polymeric porphyrin/Nafion-coated carbon fiber microsensor (detection limit, 5 nmol/L; response time, 1 millisecond) to directly measure changes in [Ca 2+ ] i and NO release due to shear stress or agonist (ATP or brominated Ca 2+ ionophore [Br-A23187]) from bovine aortic endothelial cells. The cells were grown to confluence on glass coverslips, loaded with fura 2-AM, and mounted in a parallel-plate flow chamber (volume, 25 μL). The microsensor was positioned ≈100 μm above the cells with its long axis parallel to the direction of flow. Laminar flow of perfusate was maintained from 0.04 to 1.90 mL/min, which produced shear stresses of 0.2 to 10 dyne/cm 2 . Shear stress caused transient NO release 3 to 5 seconds after the initiation of flow and 1 to 3 seconds after the rise in [Ca 2+ ] i , which reached a plateau after 35 to 70 seconds. Although the amount (peak rate) of NO release increased as a function of the shear stress (0.08 to 3.80 pmol/s), because of the concomitant increase in the flow rate, the peak NO concentration (133±9 nmol/L) remained constant. Maintenance of flow resulted in additional transient NO release, with peak-to-peak intervals of 15.5±2.5 minutes. During this 13- to 18-minute period, when the cells were unresponsive to shear stress, exogenous ATP (10 μmol/L) or Br-A23187 (10 μmol/L) evoked NO release. Prior incubation of the cells with exogenous NO or the removal and EGTA (100 μmol/L) chelation of extracellular Ca 2+ blocked shear stress but not ATP-dependent NO release. The kinetics of shear stress–induced NO release (2.23±0.07 nmol/L per second) closely resembled the kinetics of Ca 2+ flux but differed markedly from the kinetics of ATP-induced NO release (5.64±0.32 nmol/L per second). These data argue that shear stress causes a Ca 2+ -mediated ATP-independent transient release of NO, where the peak rate of release but not the peak concentration depends on the level of shear stress. The transient nature of this response may be due to NO-induced inhibition of Ca 2+ influx via a mechanism yet to be determined.

The effects of subthreshold stimulation (STS) delivered during right atrial pacing and ventricular tachycardia (VT) were investigated in Langendorff-perfused guinea pig hearts. The hearts were stained with a voltage-sensitive dye (RH 421) to map the propagation of optical action potentials. Sustained VT was reliably induced by 5-second trains (cycle length [CL], 25 to 50 milliseconds; duration, 0.5 to 10 milliseconds; and voltage, 2x threshold voltage) of impulses (n = 12 hearts) or a single premature beat (n = 6). The location of extrastimuli was not critical to the induction of VT, but the diameter of the heart had to be > or = 14.5 mm. During VT, heart rate increased from 200 to 600 beats per minute; action potential durations decreased from 112 to 175 milliseconds to 60 to 105 milliseconds, with no diastolic interval. Activation on the epicardium spread anisotropically, but VT decreased the "apparent" maximum conduction velocity (theta max) by 68% and altered the orientation of the major axis from beat to beat. Activation patterns and theta max measured during VT were similar to patterns recorded during direct pacing of the ventricle and indicated that Purkinje fibers no longer propelled ventricular excitation. STS (CL, 25 to 50 milliseconds; duration, 0.5 to 25 milliseconds; and voltage, 0.5x to 0.8x threshold; trains of 2.0 to 2.5 seconds) interrupted VT when applied to Purkinje fibers lining the endocardium (n = 6) but failed to interrupt VT when applied to the epicardium (n = 8). In atrial pacing, STS delivered to the endocardium increased theta max from 2.44 +/- 0.32 (mean +/- SEM) to 3.63 +/- 0.21 m/s in a local region surrounding the first activation sites (n = 4). Alternatively, VT could be terminated by reducing theta max (approximately 55%) with procainamide (10 mumol/L) (n = 6). STS terminates VT by synchronizing ventricular excitation most likely by increasing local conduction and/or improving the coupling between Purkinje and ventricular cells.