Limits...
Effects of neferine on Kv4.3 channels expressed in HEK293 cells and ex vivo electrophysiology of rabbit hearts.

Wang C, Chen YF, Quan XQ, Wang H, Zhang R, Xiao JH, Wang JL, Zhang CT, Xiang JZ, Tang Q - Acta Pharmacol. Sin. (2015)

Bottom Line: Furthermore, neferine (10 μmol/L) accelerated the inactivation but not the activation of Kv4.3 currents, and markedly slowed the recovery of Kv4.3 currents from inactivation.Neferine-induced blocking of Kv4.3 currents was frequency-dependent.Neferine inhibits Kv4.3 channels likely by blocking the open state and inactivating state channels, which contributes to neferine-induced dramatic increase of APD10 at Epi sites of rabbit heart.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.

ABSTRACT

Aim: Neferine is an isoquinoline alkaloid isolated from seed embryos of Nelumbo nucifera (Gaertn), which has a variety of biological activities. In this study we examined the effects of neferine on Kv4.3 channels, a major contributor to the transient outward current (I(to)) in rabbit heart, and on ex vivo electrophysiology of rabbit hearts.

Methods: Whole-cell Kv4.3 currents were recorded in HEK293 cells expressing human cardiac Kv4.3 channels using patch-clamp technique. Arterially perfused wedges of rabbit left ventricles (LV) were prepared, and transmembrane action potentials were simultaneously recorded from epicardial (Epi) and endocardial (Endo) sites with floating microelectrodes together with transmural electrocardiography (ECG).

Results: Neferine (0.1-100 μmol/L) dose-dependently and reversibly inhibited Kv4.3 currents (the IC50 value was 8.437 μmol/L, and the maximal inhibition at 100 μmol/L was 44.12%). Neferine (10 μmol/L) caused a positive shift of the steady-state activation curve of Kv4.3 currents, and a negative shift of the steady-state inactivation curve. Furthermore, neferine (10 μmol/L) accelerated the inactivation but not the activation of Kv4.3 currents, and markedly slowed the recovery of Kv4.3 currents from inactivation. Neferine-induced blocking of Kv4.3 currents was frequency-dependent. In arterially perfused wedges of rabbit LV, neferine (1, 3, and 10 μmol/L) dose-dependently prolonged the QT intervals and action potential durations (APD) at both Epi and Endo sites, and caused dramatic increase of APD10 at Epi sites.

Conclusion: Neferine inhibits Kv4.3 channels likely by blocking the open state and inactivating state channels, which contributes to neferine-induced dramatic increase of APD10 at Epi sites of rabbit heart.

Show MeSH

Related in: MedlinePlus

Effects of neferine on steady-state inactivation of Kv4.3 channels. (A) Representative traces show the steady-state inactivation of Kv4.3 currents, which were evoked using a two-pulse protocol. The membrane potential was held at −80 mV and then stepped from −120 to +50 mV in 500-ms in increments of 10 mV followed by a 500-ms depolarizing pulse to +50 mV in the absence or presence of 10 μmol/L neferine. (B) Steady-state inactivation curves of Kv4.3 currents were fitted to the Boltzmann equation; the plots represent the normalized peak currents during the test pulse as a function of the conditioning potential (n=5, P<0.05). The data are expressed as the mean±SEM.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4816235&req=5

fig3: Effects of neferine on steady-state inactivation of Kv4.3 channels. (A) Representative traces show the steady-state inactivation of Kv4.3 currents, which were evoked using a two-pulse protocol. The membrane potential was held at −80 mV and then stepped from −120 to +50 mV in 500-ms in increments of 10 mV followed by a 500-ms depolarizing pulse to +50 mV in the absence or presence of 10 μmol/L neferine. (B) Steady-state inactivation curves of Kv4.3 currents were fitted to the Boltzmann equation; the plots represent the normalized peak currents during the test pulse as a function of the conditioning potential (n=5, P<0.05). The data are expressed as the mean±SEM.

Mentions: The voltage dependence of the steady-state inactivation of Kv4.3 currents was determined using a conventional double-pulse protocol: holding the membrane potential at −80 mV and then stepping from −120 mV to +50 mV for 500 ms with increments of 10 mV followed by a 500-ms depolarizing pulse to +50 mV at 5 s intervals (Figure 3A). The Boltzmann equation was fitted to the steady-state inactivation curve: (I-I0)/(Imax-I0)=a/[1+exp(V-V1/2)/k], where Imax represents the current measured at the most hyperpolarized preconditioning pulse, I0 represents a non-inactivating current at the most depolarized preconditioning pulse, and V1/2 represents the point at which channels were half-inactivated. The steady-state inactivation curve was negatively shifted by 10 μmol/L neferine (Figure 3B): the V1/2 value of the control was −30.18±7.13 mV, whereas that of 10 μmol/L neferine was −37.85±7.92 mV (n=5, P<0.05); the slope factor was 11.67±2.33 for the control and 9.36±0.93 for 10 μmol/L neferine (n=5, P>0.05).


Effects of neferine on Kv4.3 channels expressed in HEK293 cells and ex vivo electrophysiology of rabbit hearts.

Wang C, Chen YF, Quan XQ, Wang H, Zhang R, Xiao JH, Wang JL, Zhang CT, Xiang JZ, Tang Q - Acta Pharmacol. Sin. (2015)

Effects of neferine on steady-state inactivation of Kv4.3 channels. (A) Representative traces show the steady-state inactivation of Kv4.3 currents, which were evoked using a two-pulse protocol. The membrane potential was held at −80 mV and then stepped from −120 to +50 mV in 500-ms in increments of 10 mV followed by a 500-ms depolarizing pulse to +50 mV in the absence or presence of 10 μmol/L neferine. (B) Steady-state inactivation curves of Kv4.3 currents were fitted to the Boltzmann equation; the plots represent the normalized peak currents during the test pulse as a function of the conditioning potential (n=5, P<0.05). The data are expressed as the mean±SEM.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4816235&req=5

fig3: Effects of neferine on steady-state inactivation of Kv4.3 channels. (A) Representative traces show the steady-state inactivation of Kv4.3 currents, which were evoked using a two-pulse protocol. The membrane potential was held at −80 mV and then stepped from −120 to +50 mV in 500-ms in increments of 10 mV followed by a 500-ms depolarizing pulse to +50 mV in the absence or presence of 10 μmol/L neferine. (B) Steady-state inactivation curves of Kv4.3 currents were fitted to the Boltzmann equation; the plots represent the normalized peak currents during the test pulse as a function of the conditioning potential (n=5, P<0.05). The data are expressed as the mean±SEM.
Mentions: The voltage dependence of the steady-state inactivation of Kv4.3 currents was determined using a conventional double-pulse protocol: holding the membrane potential at −80 mV and then stepping from −120 mV to +50 mV for 500 ms with increments of 10 mV followed by a 500-ms depolarizing pulse to +50 mV at 5 s intervals (Figure 3A). The Boltzmann equation was fitted to the steady-state inactivation curve: (I-I0)/(Imax-I0)=a/[1+exp(V-V1/2)/k], where Imax represents the current measured at the most hyperpolarized preconditioning pulse, I0 represents a non-inactivating current at the most depolarized preconditioning pulse, and V1/2 represents the point at which channels were half-inactivated. The steady-state inactivation curve was negatively shifted by 10 μmol/L neferine (Figure 3B): the V1/2 value of the control was −30.18±7.13 mV, whereas that of 10 μmol/L neferine was −37.85±7.92 mV (n=5, P<0.05); the slope factor was 11.67±2.33 for the control and 9.36±0.93 for 10 μmol/L neferine (n=5, P>0.05).

Bottom Line: Furthermore, neferine (10 μmol/L) accelerated the inactivation but not the activation of Kv4.3 currents, and markedly slowed the recovery of Kv4.3 currents from inactivation.Neferine-induced blocking of Kv4.3 currents was frequency-dependent.Neferine inhibits Kv4.3 channels likely by blocking the open state and inactivating state channels, which contributes to neferine-induced dramatic increase of APD10 at Epi sites of rabbit heart.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.

ABSTRACT

Aim: Neferine is an isoquinoline alkaloid isolated from seed embryos of Nelumbo nucifera (Gaertn), which has a variety of biological activities. In this study we examined the effects of neferine on Kv4.3 channels, a major contributor to the transient outward current (I(to)) in rabbit heart, and on ex vivo electrophysiology of rabbit hearts.

Methods: Whole-cell Kv4.3 currents were recorded in HEK293 cells expressing human cardiac Kv4.3 channels using patch-clamp technique. Arterially perfused wedges of rabbit left ventricles (LV) were prepared, and transmembrane action potentials were simultaneously recorded from epicardial (Epi) and endocardial (Endo) sites with floating microelectrodes together with transmural electrocardiography (ECG).

Results: Neferine (0.1-100 μmol/L) dose-dependently and reversibly inhibited Kv4.3 currents (the IC50 value was 8.437 μmol/L, and the maximal inhibition at 100 μmol/L was 44.12%). Neferine (10 μmol/L) caused a positive shift of the steady-state activation curve of Kv4.3 currents, and a negative shift of the steady-state inactivation curve. Furthermore, neferine (10 μmol/L) accelerated the inactivation but not the activation of Kv4.3 currents, and markedly slowed the recovery of Kv4.3 currents from inactivation. Neferine-induced blocking of Kv4.3 currents was frequency-dependent. In arterially perfused wedges of rabbit LV, neferine (1, 3, and 10 μmol/L) dose-dependently prolonged the QT intervals and action potential durations (APD) at both Epi and Endo sites, and caused dramatic increase of APD10 at Epi sites.

Conclusion: Neferine inhibits Kv4.3 channels likely by blocking the open state and inactivating state channels, which contributes to neferine-induced dramatic increase of APD10 at Epi sites of rabbit heart.

Show MeSH
Related in: MedlinePlus