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Objective: Catecholamines that accompany acute physiological stress are also involved in mediating the development of hypertrophy and failure. However, the cellular mechanisms involved in catecholamine-induced cardiac hypertrophy, particularly Ca2+ handling, are largely unknown. We therefore investigated the effects of cardiac hypertrophy, produced by isoprenaline, on INa/Ca and sarcoplasmic reticulum (SR) function in isolated myocytes. Methods: INa/Ca was studied in myocytes from Wistar rats, using descending (+80 to −110 mV) voltage ramps under steady state conditions. Myocytes were also loaded with fura-2 and either field stimulated or voltage clamped to assess [Ca2+]i and SR Ca2+ content. Results: Ca2+-dependent, steady state INa/Ca density was increased in hypertrophied myocytes (P<0.05). Ca2+ release from the SR was also increased, whereas resting [Ca2+]i and the rate of decline of [Ca2+]i to control levels were unchanged. SR Ca2+ content, estimated by using 10.0 mmol/l caffeine, was also significantly increased in hypertrophied myocytes, but only when myocytes were held and stimulated from their normal resting potential (−80 mV) but not from −40 mV. However, the rate of decline of caffeine-induced Ca2+ transients or INa/Ca was not significantly different between control and hypertrophied myocytes. Ca2+-dependence of INa/Ca, examined by comparing the slope of the descending phase of the hysteresis plots of INa/Ca vs. [Ca2+]i, was also similar in the two groups of cells. Conclusion: Data show that SR Ca2+ release and SR Ca2+ content were increased in hypertrophied myocytes, despite an increase in the steady state INa/Ca density. The observation that increased SR function occurred only when myocytes were stimulated from −80 mV suggests that Na+ influx may play a role in altering Ca2+ homeostasis in hypertrophied cardiac muscle, possibly through increased reverse Na+/Ca2+ exchange, particularly at low stimulation frequencies. [Copyright &y& Elsevier]

Objective: To test the hypothesis that changes in the normal regional distribution of potassium and calcium currents contribute to the different regional changes in action potential duration in isoprenaline-induced hypertrophy in rats. Methods: Hypertrophy was elicited in rats by seven daily injections of isoprenaline. Left ventricular myocytes were isolated from basal sub-endocardial, basal mid-myocardial and apical sub-epicardial tissue. Membrane currents were measured using the whole-cell patch-clamp technique at 35±1°C. Results: Cell membrane capacitance was similar in all three groups and was increased by 17% in hypertrophy (P<0.001, t-test). Changes in the calcium-independent transient outward current (Ito1) density in hypertrophy were different in the three regions (P<0.05, ANOVA). Ito1 was reduced in sub-epicardial (control, 23.4±2.0 pA pF−1; hypertrophy, 15.8±1.5 pA pF−1, P<0.01 ANOVA) and in mid-myocardial myocytes (control, 24.0±2.8 pA pF−1; hypertrophy, 13.8±1.3 pA pF−1, P<0.01 ANOVA) and was not significantly altered in sub-endocardial myocytes (control, 8.5±0.7 pA pF−1; hypertrophy, 7.4±1.8 pA pF−1). Steady-state background current density was reduced in hypertrophy (P<0.05, ANOVA). The regional difference in steady-state background current in control hearts (P<0.05, ANOVA) was altered in hypertrophy. Calcium current (ICa) density was similar in the three regions studied in both control and hypertrophied hearts. ICa was reduced in hypertrophy (P<0.05, ANOVA). Conclusion: The normal regional differences in Ito1 are reduced, in steady-state background current are altered and in ICa are unchanged in catecholamine-induced hypertrophy in the rat left ventricle. These data may in part explain the reduction in the normal regional differences in APD observed in hypertrophy. [ABSTRACT FROM PUBLISHER]

Objective: To compare the properties of single myocytes isolated from different layers of the basal region of the left ventricle and to test the hypothesis that differences in the delayed rectifier current (IK) contribute to regional differences in action potential duration. Methods: Myocytes were isolated from basal sub-endocardial, mid-myocardial and sub-epicardial layers of the guinea-pig left ventricle. Membrane voltage and current were measured using the switch-clamp technique. Results: Mean action potential duration measured at 90% repolarisation (APD90) was longer in sub-endocardial myocytes than in mid-myocardial and sub-epicardial myocytes [APD90 ms at 0.2 Hz: sub-endocardial 292±12 (n=40), mid-myocardial 243±8 (n=42) and sub-epicardial 227±9 (n=36), P<0.001, analysis of variance (ANOVA)]. The APD–rate relationship (stimulation frequencies 2, 1, 0.2 and 0.017 Hz) was steeper in sub-endocardial than in mid-myocardial or sub-epicardial myocytes (P<0.001, ANOVA). The density of IK was greater in mid-myocardial (4.05±0.09 pA pF−1) and sub-epicardial (3.90±0.41 pA pF−1) than in sub-endocardial myocytes (2.74±0.27 pA pF−1, P<0.01 ANOVA). The rapidly-activating (IKr) and slowly-activating (IKs) components of IK were significantly smaller in sub-endocardial than in mid-myocardial or sub-epicardial myocytes. d,l-Sotalol-induced prolongation of APD90 was similar in the three regions studied. Conclusions: There are significant transmural gradients in the electrophysiological properties of myocytes isolated from the base of the left ventricular free wall in guinea-pig. Sub-endocardial myocytes had a longer APD90 attributable in part to a significantly smaller IK density. We have been unable to identify M cells in the guinea-pig left ventricular free wall. [ABSTRACT FROM PUBLISHER]

Objective: To investigate electrical and mechanical properties of single myocytes isolated from different regions of the left ventricle in control and hypertrophied hearts. Methods: Mild cardiac hypertrophy was induced in guinea-pigs by aortic constriction. Myocytes were isolated from basal sub-endocardial, basal mid-myocardial and apical sub-epicardial layers of the left ventricle. Action potentials were stimulated at 1 Hz. Membrane currents were measured using the switch-clamp technique. Cell shortening was measured using a photodiode array. Results: In control hearts mean action potential duration (APD) was longer in sub-endocardial myocytes than in sub-epicardial myocytes. In hypertrophy APD was prolonged in sub-epicardial and mid-myocardial myocytes and unchanged in sub-endocardial myocytes (APD90 ms, control: sub-endocardial 273±12, mid-myocardial 254±14, sub-epicardial 229±9; hypertrophy: sub-endocardial 259±13, mid-myocardial 291±9, sub-epicardial 268±11, P<0.05, ANOVA). There was no significant regional difference in APD in hypertrophied hearts. In control hearts L-type calcium current (ICa) was similar in all regions. In hypertrophy ICa was increased in sub-epicardial and mid-myocardial myocytes and reduced in sub-endocardial myocytes. Calcium-activated tail currents were not regionally different in control or hypertrophied hearts, but were increased in hypertrophy. Conclusions: Changes in electrical and mechanical properties associated with hypertrophy are not homogeneous throughout the left ventricle. The difference in APD between sub-endocardial and sub-epicardial myocytes seen in control hearts is lost in hypertrophy. These results may favour the propagation of re-entry arrhythmias in hypertrophied hearts. [ABSTRACT FROM PUBLISHER]

Objective: Tissue-type plasminogen activator (t-PA) is an important component of the blood fibrinolytic system responsible for thrombus dissolution. It is often required to function under oxidative stress, and exogenous t-PA is used clinically in the treatment of acute myocardial infarction (AMI). The aim of this study was to examine alterations in the residual activity of t-PA pre-treated with certain oxidants. Methods: Recombinant t-PA (rt-PA) and native t-PA (nt-PA) were pre-treated with freshly generated hypochlorous acid (HOCl) and chloramine T at varying concentrations. The amidolytic activity, the plasminogenolytic activity and the fibrin-binding affinity were then examined using chromogenic assays based on S-2288 and S-2251. Results: The amidolytic activity of t-PA was surprisingly found to be rather sensitive (IC50 1 and 12 μmol/l, respectively), and the plasminogenolytic activity rather resistant to pre-treatment with HOCl and chloramine T. The fibrin binding study of treated t-PA revealed substantial loss of binding to CNBr-digested fibrinogen (FDP-CNBr). The velocity of t-PA in reaction with plasminogen remained the same as non-treated t-PA. The possible mechanisms of this asymmetrical oxidative modification of the biological functions are also discussed. Conclusions: (1) The catalytic activity of t-PA and the binding affinity for its large-molecule substrate plasminogen, rather than the small-molecule substrate S-2288, are highly resistant to oxidative damage; (2) the fibrin-binding affinity of t-PA can be selectively and asymmetrically damaged by exposure to these oxidants. Thus it is possible that the characteristic advantage of thrombus selectivity of t-PA in both spontaneous thrombolysis and thrombolytic therapy may be diluted in circumstances where toxic and reactive oxidants exist. [ABSTRACT FROM PUBLISHER]

Objective: Catecholamines that accompany acute physiological stress are also involved in mediating the development of hypertrophy and failure. However, the cellular mechanisms involved in catecholamine-induced cardiac hypertrophy, particularly Ca2+ handling, are largely unknown. We therefore investigated the effects of cardiac hypertrophy, produced by isoprenaline, on I(Na/Ca) and sarcoplasmic reticulum (SR) function in isolated myocytes., Methods: I(Na/Ca) was studied in myocytes from Wistar rats, using descending (+80 to -110 mV) voltage ramps under steady state conditions. Myocytes were also loaded with fura-2 and either field stimulated or voltage clamped to assess [Ca2+]i and SR Ca2+ content., Results: Ca2+-dependent, steady state I(Na/Ca) density was increased in hypertrophied myocytes (P<0.05). Ca2+ release from the SR was also increased, whereas resting [Ca2+]i and the rate of decline of [Ca2+]i to control levels were unchanged. SR Ca2+ content, estimated by using 10.0 mmol/l caffeine, was also significantly increased in hypertrophied myocytes, but only when myocytes were held and stimulated from their normal resting potential (-80 mV) but not from -40 mV. However, the rate of decline of caffeine-induced Ca2+ transients or I(Na/Ca) was not significantly different between control and hypertrophied myocytes. Ca2+-dependence of I(Na/Ca), examined by comparing the slope of the descending phase of the hysteresis plots of I(Na/Ca) vs. [Ca2+]i, was also similar in the two groups of cells., Conclusion: Data show that SR Ca2+ release and SR Ca2+ content were increased in hypertrophied myocytes, despite an increase in the steady state I(Na/Ca) density. The observation that increased SR function occurred only when myocytes were stimulated from -80 mV suggests that Na+ influx may play a role in altering Ca2+ homeostasis in hypertrophied cardiac muscle, possibly through increased reverse Na+/Ca2+ exchange, particularly at low stimulation frequencies.