P1‐193: suppression of kv1.1 channel activity by amyloid peptide fragments (1‐42) and the toxic core (25‐35)

Past studies have shown that Aβ’s role in the pathogenesis of Alzheimer's disease (AD) involves eventual disruption of Ca2+ homeostasis and synaptic communication, as well as impairment of long-term potentiation (LTP), but the underlying mechanism(s) is still unclear. Because Kv1.1 and related channels are activated during an action potential, regulate depolarization Ca2+ influx, and inhibition of Kv1 channels can be neurotoxic, we speculate that Aβ-suppression of Kv1 channels could contribute to AD pathology. Stage V and VI Xenopus laevis oocytes were commercially provided by Ecocyte Bioscience (Austin, TX) and injected with Kv1.1 cRNA. The effects of bath application of Aβ fragments (1-42) and (25-35) on macroscopic and microscopic Kv1.1 currents were assessed using standard two electrode voltage-clamp (TEVC) and patch clamp techniques, respectively. The direct effect of Aβ on the Kv1.1 channel was further investigated using artificial membrane techniques [“tip-dip” and black lipid membrane (BLM)]. Bath application of 1 μM Aβ(1-42) produced ∼50% suppression of macroscopic Kv1.1 current within 30 min. Aβ suppression of Kv1.1 was partially Ca2+- and PP2B-dependent, being reduced by ∼50% when cells were loaded with BAPTA-AM or exposed to the PP2B-inhibitor cyclosporine A (CsA). Patch-clamp results suggest that Aβ-suppression of Kv1.1 involves both PP2B-dephosphorylation and direct protein-protein interaction of Aβ with Kv1.1 channel subunits. Exposure of inside-out single Kv1.1 in ripped-off oocyte patches to application of purified, catalytically-active PP2B produced gradual reductions in p(open), followed by abrupt disappearance of Kv1.1 activity. Application of Aβ to the intracellular face of Kv1.1 channels also produced dramatic reductions in p(open). Additional results indicate that 2 μM of Aβ(25-35) suppressed Kv1.1 currents by ∼40%. Using “tip-dip” artificial membrane methods, 1 μM Aβ(25-35) exposure eliminated Kv1.1 channel activity when applied to the intracellular face. The toxic Aβ fragments (1-42) and (25-35) suppress the voltage-gated potassium channel, Kv1.1. Suppression of Kv1.1 and related K+ channels presynaptically could lead to larger and longer action potentials, allowing more influx of Ca2+, increased release of glutamate, and possibly the beginning of a disruption of Ca2+ homeostasis. Postsynaptically, the increased glutamate release, through activation of AMPA and NMDA receptors, may contribute to excitotoxicity.