Effective Perturbations by Phenobarbital on INa, IK(erg), IK(M) and IK(DR) during Pulse Train Stimulation in Neuroblastoma Neuro-2a Cells

Po-Ming Wu; Pei-Chun Lai; Hsin-Yen Cho; Tzu-Hsien Chuang; Sheng-Nan Wu; Yi-Fang Tu

doi:10.3390/biomedicines10081968

Effective Perturbations by Phenobarbital on I_Na, I_K(erg), I_K(M) and I_K(DR) during Pulse Train Stimulation in Neuroblastoma Neuro-2a Cells

Biomedicines. 2022 Aug 13;10(8):1968. doi: 10.3390/biomedicines10081968.

Authors

Po-Ming Wu^{1

2}, Pei-Chun Lai³, Hsin-Yen Cho⁴, Tzu-Hsien Chuang⁴, Sheng-Nan Wu^{4

5}, Yi-Fang Tu^{1

2}

Affiliations

¹ Department of Pediatrics, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 70101, Taiwan.
² Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan 70101, Taiwan.
³ Education Center, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 70101, Taiwan.
⁴ Department of Physiology, College of Medicine, National Cheng Kung University, Tainan 70101, Taiwan.
⁵ Institute of Basic Medical Sciences, College of Medicine, National Cheng Kung University, Tainan 70101, Taiwan.

Abstract

Phenobarbital (PHB, Luminal Sodium^®) is a medication of the barbiturate and has long been recognized to be an anticonvulsant and a hypnotic because it can facilitate synaptic inhibition in the central nervous system through acting on the γ-aminobutyric acid (GABA) type A (GABA_A) receptors. However, to what extent PHB could directly perturb the magnitude and gating of different plasmalemmal ionic currents is not thoroughly explored. In neuroblastoma Neuro-2a cells, we found that PHB effectively suppressed the magnitude of voltage-gated Na⁺ current (I_Na) in a concentration-dependent fashion, with an effective IC₅₀ value of 83 µM. The cumulative inhibition of I_Na, evoked by pulse train stimulation, was enhanced by PHB. However, tefluthrin, an activator of I_Na, could attenuate PHB-induced reduction in the decaying time constant of I_Na inhibition evoked by pulse train stimuli. In addition, the erg (ether-à-go-go-related gene)-mediated K⁺ current (I_K(erg)) was also blocked by PHB. The PHB-mediated inhibition on I_K(erg) could not be overcome by flumazenil (GABA antagonist) or chlorotoxin (chloride channel blocker). The PHB reduced the recovery of I_K(erg) by a two-step voltage protocol with a geometrics-based progression, but it increased the decaying rate of I_K(erg), evoked by the envelope-of-tail method. About the M-type K⁺ currents (I_K(M)), PHB caused a reduction of its amplitude, which could not be counteracted by flumazenil or chlorotoxin, and PHB could enhance its cumulative inhibition during pulse train stimulation. Moreover, the magnitude of delayed-rectifier K⁺ current (I_K(DR)) was inhibited by PHB, while the cumulative inhibition of I_K(DR) during 10 s of repetitive stimulation was enhanced. Multiple ionic currents during pulse train stimulation were subject to PHB, and neither GABA antagonist nor chloride channel blocker could counteract these PHB-induced reductions. It suggests that these actions might conceivably participate in different functional activities of excitable cells and be independent of GABA_A receptors.

Keywords: M-type K+ current; delayed-rectifier K+ current; erg-mediated K+ current; phenobarbital (phenobarbitone, Luminal Sodium®); pulse train stimulation; voltage-gated Na+ current; γ-aminobutyric acid type A receptors.

Abstract

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