Stimulatory actions of a novel thiourea derivative on large-conductance, calcium-activated potassium channels

In this study, we examine whether an anti-inflammatory thiourea derivative, compound #326, actions on ion channels. The effects of compound #326 on Ca²⁺ -activated K⁺ channels were evaluated by patch-clamp recordings obtained in cell-attached, inside-out or whole-cell configuration. In pituitary GH₃ cells, compound #326 increased the amplitude of Ca²⁺ -activated K⁺ currents (I_K(Ca) ) with an EC₅₀ value of 11.6 μM, which was reversed by verruculogen, but not tolbutamide or TRAM-34. Under inside-out configuration, a bath application of compound #326 raised the probability of large-conductance Ca²⁺ -activated K⁺ (BK_Ca ) channels. The activation curve of BK_Ca channels was shifted to less depolarised potential with no modification of the gating charge of the curve; consequently, the difference of free energy was reduced in the presence of this compound. Compound #326-stimulated activity of BK_Ca channels is explained by a shortening of mean closed time, despite its inability to alter single-channel conductance. Neither delayed-rectifier nor erg-mediated K⁺ currents was modified. Compound #326 decreased the peak amplitude of voltage-gated Na⁺ current with no clear change in the overall current-voltage relationship of this current. In HEK293T cells expressing α-hSlo, compound #326 enhanced BK_Ca channels effectively. Intriguingly, the inhibitory actions of compound #326 on interleukin 1β in lipopolysaccharide-activated microglia were significantly reversed by verruculogen, whereas BK_Ca channel inhibitors suppressed the expressions of inducible nitric oxide synthase. The BK_Ca channels could be an important target for compound #326 if similar in vivo results occur, and the multi-functionality of BK_Ca channels in modulating microglial immunity merit further investigation.