Hypericin prolongs action potential duration in hippocampal neurons by acting on K+ channels

Abstract

Background and purpose: Synaptic deficiency is generally accepted to be involved in major depression, and accordingly classic antidepressants exert their effects through enhancing synaptic efficiency. Hypericin is one of the major active constituents of extracts of St. John's Wort (Hypericum perforatum L.) with antidepressive actions, but little is known about its therapeutic mechanisms. Our aim was to explore whether hypericin has a modulatory effect on neuronal action potential (AP) duration by acting on voltage-gated ion channels.

Experimental approach: We used voltage-clamp and current-clamp techniques in a whole-cell configuration to study primary cultures of neonatal rat hippocampal neurones. We measured the effects of extracellularly applied hypericin on AP duration as well as on voltage-gated Na(+), I(A) and I(K) currents.

Key results: Extracellularly applied hypericin dose-dependently increased AP duration but barely affected its amplitude. Further analysis revealed that hypericin inhibited both transient I(A) and delayed rectifier I(K) potassium currents. In contrast, hypericin exerted no significant effect on both Na(+) peak current and its decay kinetics.

Conclusions and implications: Extracellularly applied hypericin increased AP duration, which might be ascribed to its effect on I(A) and I(K) currents. As a small increase in AP duration could lead to a dramatic increase in synaptic efficiency, our results imply that hypericin might exert its antidepressant effects by enhancing presynaptic efficiency.

Figure 1

Effect of hypericin on action potential waveforms. (A) Action potential traces showing effect of 1.0 µM hypericin on AP waveform (representative of six cells). The AP was evoked from the same cell by a 100 ms depolarizing step of 130 pA. AP duration (B) and AP amplitude (C) relative to control values (=1.0) after different concentrations of hypericin. AP amplitude was measured from the holding potential and AP duration was defined as the width at half of the AP amplitude. **P < 0.01, ***P < 0.001, significantly different from control; one-way anova). AP, action potential.

Figure 2

Effects of hypericin on the whole-cell peak Na⁺ currents. (A) Current traces showing effect of extracellularly applied 1.0 µM hypericin on whole-cell Na⁺ currents (representative of seven cells). The currents was evoked by voltage steps from −100 mV to +20 mV in 10 mV increments for 12 ms. (B) Summary data of the ratio (I/I₀) of the whole-cell Na⁺ peak currents before (I₀) and after (I) application of different concentrations of hypericin. (C) Summary data of the ratio (τ/τ_o) of the decay time constant of Na⁺ current evoked at −30 mV before (τ₀) and after (τ) application of different concentrations of hypericin. The decay time constant of Na⁺ current was derived by fitting a single exponential. There are no significant differences between control and different hypericin concentration groups in both (B) and (C) (P > 0.05, one-way anova).

Figure 3

Effects of hypericin on peak I_A current and its decay kinetics. (A) Whole-cell I_A current traces(representative of seven cells) for the control (left panel) and in the presence of 1.0 µM hypericin (middle panel). The currents were evoked by voltage steps from −110 to +70 mV in 20 mV increments for 200 ms and derived from subtraction (see Methods). Right panel: Normalized I_A current traces evoked at +30 mV showing effects of 1.0 µM hypericin on its decay kinetics. (B) Ratios (I/I₀) of the whole-cell I_A peak current evoked at −30 mV before (I₀) and after (I) application of different concentrations of hypericin. (C) Summary data of the ratio (τ/τ_o) of the decay time constant of the I_A current evoked at +30 mV before (τ₀) and after (τ) application of different concentrations of hypericin. The decay time constant of I_A current was derived by fitting a single exponential function. *P < 0.05, **P < 0.01, ***P < 0.001, significantly different from control; one-way anova.

Figure 4

Effects of hypericin on I_K currents. (A) Current traces showing effects of 1.0 µM hypericin on whole-cell I_K currents (representative of seven cells). The currents were evoked by voltage steps from −110 to +70 mV in 20 mV increments for 150 ms following a prepulse to −50 mV for 50 ms to inactivate the I_A current. (B) Summary data of the ratio (I/I_o) of the whole-cell I_K current evoked at +30 mV before (I₀) and after (I) application of different concentrations of hypericin. ***P < 0.001, significantly different from control; one-way anova. The steady-state outward I_K current (+30 mV) was measured as mean value in a range from 85% to 95% of the current trace.