Bicuculline block of small-conductance calcium-activated potassium channels

Abstract

Small-conductance calcium-activated potassium channels (SK channels) are gated solely by intracellular calcium ions and their activity is responsible for the slow afterhyperpolarization (AHP) that follows an action potential in many excitable cells. Brain slice studies commonly employ a methyl derivative of bicuculline (bicuculline-m), a GABAA (gamma-aminobutyric acid) receptor antagonist, to diminish the tonic inhibitory influences of GABAergic synapses, or to investigate the role of these synapses in specialized neural networks. However, recent evidence suggests that bicuculline-m may not be specific for GABAA receptors and may also block the slow AHP. Therefore, the effects of bicuculline-m on cloned apamin-sensitive SK2 and apamin-insensitive SK1 channels were examined following expression in Xenopus oocytes. The results show that at concentrations employed for slice recordings, bicuculline-m potently blocks both apamin-sensitive SK2 currents and apamin-insensitive SK1 currents when applied to outside-out patches. Apamin-insensitive SK1 currents run down in excised patches. The potency of bicuculline-m block also decreases with time after patch excision. Site-directed mutagenesis that changes two residues in the outer vestibule of the SK1 pore that confers apamin sensitivity also reduces run down of the current in patches, and endows stable sensitivity to bicuculline-m indistinguishable from SK2. Therefore, the use of bicuculline-m in slice recordings may mask apamin-sensitive slow AHPs that are important determinants of neuronal excitability. In addition, bicuculline-m-insensitive slow AHPs may indicate that the underlying channels have run down.

Small Conductance Ca2+-activated K+ Channels and Calmodulin

“…Fitting the data with single exponentials yielded time constants of 1.2 Ϯ 0.2, 1.7 Ϯ 0.3, and 1.8 Ϯ 0.4 min for Ca 2ϩ -CaM association and 0.7 Ϯ 0.1, 1.4 Ϯ 0.4, and 2.1 Ϯ 0.8 min for Ca 2ϩ -CaM dissociation (n ϭ 7). The maximum current amplitudes decreased over the course of the experiment, consistent with channel rundown (18).…”

Cerebellar ataxia and Purkinje cell dysfunction caused by Ca 2+ -activated K + channel deficiency

“…Saggital slices (350-400 m thick) from the cerebellar vermis of 4-to 5-week-old WT and BK Ϫ/Ϫ mice were prepared and kept at 34-36°C in artificial cerebrospinal fluid with 10 M bicuculline free base [a concentration that has been shown to not measurably affect currents of Ca 2ϩ -activated K ϩ channels with small conductance (SK) or afterhyperpolarizations (AHPs) in rat brain slices (14,15) and to cause only a Ϸ20% inhibition of SK channels expressed in frog oocytes (16)] and 10 M 6,7-dinitroquinoxaline-2,3-dione (DNQX) to block synaptic transmission. Whole-cell (Axoclamp 2A, Axon Instruments, Union City, CA) and extracellular (Multiclamp 700A, Axon Instruments) recordings were obtained from PC somata under visual control.…”

Neuronal Responses to Lemniscal Stimulation in Laminar Brain Slices of the Inferior Colliculus

“…The concentration of bicuculline used did not cause changes in resting membrane potentials. Bicuculline also affects apamin-sensitive, calcium-activated potassium channels (Khawaled et al 1999). Because these channels are found in the ICC (Sivaramakrishnan and Oliver 2001), we tested the effect of bicuculline on firing patterns across cell types.…”