Lysophosphatidic acid increases the electrophysiological instability of adult rabbit ventricular myocardium by augmenting L-type calcium current

Abstract

Lysophosphatidic acid (LPA) has diverse actions on the cardiovascular system and is widely reported to modulate multiple ion currents in some cell types. However, little is known about its electrophysiological effects on cardiac myocytes. This study investigated whether LPA has electrophysiological effects on isolated rabbit myocardial preparations. The results indicate that LPA prolongs action potential duration at 90% repolarization (APD(90)) in a concentration- and frequency-dependent manner in isolated rabbit ventricular myocytes. The application of extracellular LPA significantly increases the coefficient of APD(90) variability. LPA increased L-type calcium current (I(Ca,L)) density without altering its activation or deactivation properties. In contrast, LPA has no effect on two other ventricular repolarizing currents, the transient outward potassium current (I(to)) and the delayed rectifier potassium current (I(K)). In arterially perfused rabbit left ventricular wedge preparations, the monophasic action potential duration, QT interval, and Tpeak-end are prolonged by LPA. LPA treatment also significantly increases the incidence of ventricular tachycardia induced by S(1)S(2) stimulation. Notably, the effects of LPA on action potentials and I(Ca,L) are PTX-sensitive, suggesting LPA action requires a G(i)-type G protein. In conclusion, LPA prolongs APD and increases electrophysiological instability in isolated rabbit myocardial preparations by increasing I(Ca,L) in a G(i) protein-dependent manner.

Figure 1. Concentration and frequency-dependent prolongation of action potential duration (APD) induced by LPA in isolated rabbit ventricular myocytes.

(A) Action potentials were recorded at different frequencies from 0.25 to 4.0 Hz before and after perfusion with various concentrations of LPA. Closed circles: 1 µM, open circles: 5 µM, closed triangles: 10 µM, open triangles: 20 µM. Delta APD₉₀ is defined as the difference of APD₉₀ recorded before and after LPA addition at a constant frequency (n = 10). (B) Representative traces of action potential recorded at the frequency of 1.0 Hz in cardiac myocytes without (Control) and with different concentrations of LPA (1, 5, 10, 20 µM).

Figure 2. Time course of changes in action potential duration (APD) at 1.0 Hz in isolated rabbit ventricular myocytes after addition of LPA (10 µM).

(n = 10, ANOVA, P<0.01).

Figure 3. LPA-induced increase in action potential duration (APD) variability in isolated rabbit ventricular myocytes.

(A) Comparison of APD₉₀ variability subsequently recorded at 1.0 Hz before and after LPA treatment (10 µM). *P<0.01 in paired t-test (n = 10). (B) Representative action potentials recorded from 30 consecutive beats in one adult rabbit ventricular myocyte without (upper panel) and with (lower panel) LPA treatment. Coefficient of APD₉₀ variability = (SD/mean APD₉₀)×100%.

Figure 4. Effect of LPA on current-voltage relation of L-type calcium current (I_Ca,L) in isolated rabbit ventricular myocytes.

(A) Current amplitudes corrected for cell size in the presence (open circles) and absence (solid circles) of LPA (10 µM) plotted against the test potentials. Error bars are SDM. *P<0.01, two-way ANOVA followed by the Bonferroni test, comparing peak current density of I_Ca,L at the same potential recorded in myocytes without LPA (n = 10). (B) Representative traces of I_Ca,L in cardiac myocytes without (upper panel) and with (lower panel) LPA.

Figure 5. LPA increases L-type calcium current (I_Ca,L) density without altering activation or inactivation properties.

(A) Average current densities of I_Ca,L in the absence and presence of LPA (10 µM). The isolated rabbit ventricular myocytes were kept at a holding potential of −40 mV and depolarized to 0 mV for 200 ms. *P<0.05 compared with the control in an Independent Samples t-Test (n = 10). (B) Representative I_Ca,L in the control condition (upper panel) and LPA treatment (lower panel). Voltage dependence of I_Ca,L activation (C) and inactivation (D) in the control condition (closed circles, n = 10) and in the presence of LPA (open circles, n = 10).

Figure 6. LPA does not affect the current-voltage relation of transient outward potassium current (I_to) in isolated rabbit ventricular myocytes.

(A) Current amplitudes corrected for cell size observed in the presence (open circles) and in the absence (solid circles) of LPA were plotted against the test potentials. Error bars are SEM, n = 8. (B) Representative traces of I_to in cardiac myocytes without (upper panel) and with (lower panel) LPA.

Figure 7. LPA does not affect the current-voltage relation of delayed rectifier potassium current (I_K) in isolated rabbit ventricular myocytes.

(A) Current amplitudes corrected for cell size observed in the presence (open circles) and in the absence (solid circles) of LPA plotted against the test potentials. Error bars are SEM. No significant difference was observed, n = 7. (B) Representative traces of I_K in cardiac myocytes without (upper panel) and with (lower panel) LPA treatment.

Figure 8. LPA treatment increases electrophysiological instability in the heart.

LPA prolongs 90% of action potential duration (APD₉₀), Tpeak-end, and QT interval (A) and increased the incidence of ventricular tachycardia (B) induced by S₁S₂ stimulation in the arterially perfused rabbit left ventricular wedge preparations, n = 10. (C) Representative monophasic action potential in the endocardium and the transmural ECG before (left) and after (right) LPA treatment. (D) Representative ventricular tachycardia induced by S₁S₂ stimulation in the presence of LPA.

Figure 9. The effects of LPA on action potentials and I_Ca,L are abolished by pertussis toxin (PTX).

LPA increases APD₉₀ (A), APD₅₀?(B)?and I_Ca,L (D) above basal values, which does not occur after PTX pretreatment, * P<0.05, comparing the LPA treated sample with the control in the absence or presence of PTX), n = 10. I_Ca,L was recorded with the single-wave protocol at a holding potential of −40 mV and a test potential of 0 mV for 200 ms. (C) LPA has no effect on APD₂₀ without or with PTX pretreatment (n = 10).