RESUMO
Purpose/Aim: Besides as a cholinesterase (ChE) inhibitor, tacrine is able to act on multiple targets such as nicotinic receptors (nAChRs) and voltage-gated K+ (Kv) channels. Kv2.1, a Kv channel subunit underlying delayed rectifier currents with slow kinetics of inactivation, is highly expressed in the mammalian brain, especially in the hippocampus. Nevertheless, limited data are available concerning the relationship between tacrine and Kv2.1 channels. In the present study, we explore the possible effects of tacrine on Kv2.1 channels in heterologous expression systems and N2A cells.Materials and methods: The change of expression and currents of Kv2.1 after treatment with tacrine was detected by PCR and whole-cell recordings, respectively. WST-8 experiments were performed to reveal the effects of tacrine on cell proliferation.Results: Incubation with tacrine induced a significant reduction of the mRNA level of Kv2.1 channels in HEK293 cells. The decline of corresponding currents carried by Kv2.1 was also observed. Moreover, the proliferation rates of HEK293 cells with Kv2.1 channel were substantially enhanced after treatment with this chemical for 24 h. Similar results were also detected after exposure to tacrine in N2A cells with native expression of Kv2.1 channels.Conclusion: These lines of evidence indicate that application of tacrine downregulates the expression of Kv2.1 channels and increase cell proliferation. The effect of tacrine on Kv2.1 channels may provide an alternative explanation for its neuroprotective action.
Assuntos
Proliferação de Células/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Canais de Potássio Shab/efeitos dos fármacos , Tacrina/farmacologia , Animais , Linhagem Celular Tumoral , Regulação para Baixo , Células HEK293 , Humanos , Camundongos , Técnicas de Patch-Clamp , Reação em Cadeia da Polimerase , RNA Mensageiro , Sais de TetrazólioRESUMO
Previous findings indicated that microtubule-binding protein tau and voltage-gated K(+) (Kv) channels exhibit a regulatory role in cell proliferation. However, the possible interaction of tau with Kv channels remained obscure. In this report, transfection of tau plasmids into human neuroblastoma SK-N-SH cells caused a significant reduction in the messenger RNA (mRNA) levels of several Kv channels, including Kv2.1, Kv3.1, Kv5.1, Kv9.2, and KCNH4. Correspondingly, the Kv currents recorded using patch-clamp techniques were substantially declined in the tau-transfected SK-N-SH cells. Moreover, tau induction and treatment with the Kv channel blocker TEA (tetraethylammonium) were able to improve proliferation rates of SK-N-SH cells by 43.1 and 66.2%, respectively. These data suggested that the tau-mediated alteration of Kv channels could be involved in its action on neural proliferation.
Assuntos
Neurônios/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , RNA Mensageiro/metabolismo , Proteínas tau/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Neurônios/fisiologia , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , RNA Mensageiro/genética , Proteínas tau/genéticaRESUMO
It is well documented that nifedipine, a commonly used dihydropyridine Ca2+ channel blocker, has also significant interactions with voltage-gated K+ (Kv) channels. But to date, little is known whether nifedipine exerted an action on Kv2.1 channels, a member of the Shab subfamily with slow inactivation. In the present study, we explored the effects of nifedipine on rat Kv2.1 channels expressed in HEK293 cells. Data from whole-cell recording showed that nifedipine substantially reduced Kv2.1 currents with the IC50 value of 37.5 ± 5.7 µM and delayed the time course of activation without effects on the activation curve. Moreover, this drug also significantly shortened the duration of inactivation and deactivation of Kv2.1 currents in a voltage-dependent manner. Interestingly, the half-maximum inactivation potential (V1/2) of Kv2.1 currents was -11.4 ± 0.9 mV in control and became -38.5 ± 0.4 mV after application of 50 µM nifedipine. The large hyperpolarizing shift (27 mV) of the inactivation curve has not been reported previously and may result in more inactivation for outward delayed rectifier K+ currents mediated by Kv2.1 channels at repolarization phases. The Y380R mutant significantly increased the binding affinity of nifedipine to Kv2.1 channels, suggesting an interaction of nifedipine with the outer mouth region of this channel. The data present here will be helpful to understand the diverse effects exerted by nifedipine on various Kv channels.