Chronic Ca2+influx through voltage-dependent Ca2+channels enhance delayed rectifier K+currents via activating Src family tyrosine kinase in rat hippocampal neurons

Figures

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  Fig. 1. IDR is enhanced under high Ca2+ condition in cultured hippocampal neurons, showing the dependence on VDCCs but not NMDARs. (A) The experimental protocol of voltage clamping to record IDR in hippocampal neurons and example traces of IDR. High Ca2+ condition was induced by doubling Ca2+ concentration (CaCl2, 3.6 mM) of culture media for 24 hours (High Ca2+) and then neurons were washed in normal ACSF during recording. Either APV (100 µM, APV+High Ca2+) or nimodipine (10 µM, Nimodipine + High Ca2+) was added to culture media for 24 hours under high Ca2+ condition. Scale bars 500 pA, 100 ms. (B) Individuals (circles) and averaged (square bars) densities of IDR. Currents were adjusted with whole cell capacitance for measuring current densities of each group. Error bars represent SEM. *p<0.05.

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  Fig. 2. The enhancement of IDR under high Ca2+ condition does not involve the significant alteration of activation properties of IDR channels. The activation property was measured at −60 to 80 mV with 20 steps after prepulse injection (−20 mV, 200 ms duration). (A) The experimental protocol of voltage clamping to measure the activation kinetics of IDR channels and example traces of IDR. The amplitude of IDR is increased by steps of voltage clamping. However, the constant rate of increment indicates no changes of IDR activating kinetics. Scale bars 500 pA, 100 ms. (B) Averaged densities of IDR at each clamping voltage (−40 to 80 mV). (C) and (D) Boltzmann fitted curves of gating kinetics of IDR activation and averaged values of voltage half (Vh, dotted line in C). The significant change of activation curves was not observed. Error bars represent SEM.

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  Fig. 3. SFKs are participated in IDR enhancement under high Ca2+ condition. (A) Example traces of IDR in each group. To confirm the contribution of SK channels, apamin (100 nM, Ca2++Apamin) was added to culture media. PP2 (1 µM) abolished the Ca2+-induced enhancement of IDR, indicating the involvement of SFK in IDR upregulation. (B) Individual (open circles) and averaged values (square bars) of IDR densities. Error bars represent SEM. *p<0.05 compared with control and †p<0.05 compared with High Ca2+. (C) Averaged densities of IDR at each clamping voltage (−40 to 80 mV). Error bars represent SEM.