title: In vitro and intrathecal siRNA mediated K(V)1.1 knock-down in primary sensory neurons.
abstract: K(V)1.1 is a Shaker homologue K(+) channel that contributes to the juxta-paranodal membrane conductance in myelinated axons, and is blocked by fampridine (4-aminopyridine), used to treat the symptoms of multiple sclerosis. The present experiments investigate K(V)1.1 function in primary sensory neurons and A-fibres, and help define its characteristics as a drug-target using sequence specific small-interfering RNAs (siRNAs). siRNA (71nM) was used to knock-down functional expression of K(V)1.1 in sensory neurons (>25μm in apparent diameter) in culture, and was also delivered intrathecally in vivo (9.3μg). K(+) channel knock-down in sensory neurons was found to make the voltage-threshold for action potential generation significantly more negative than in control (p=0.02), led to the breakdown of accommodation and promoted spontaneous action potential firing. Exposure to dendrotoxin-K (DTX-K, 10-100nM) also selectively abolished K(+) currents at negative potentials and made voltage-threshold more negative, consistent with K(V)1.1 controlling excitability close to the nominal resting potential of the neuron cell body, near -60mV. Introduction of one working siRNA sequence into the intrathecal space in vivo was associated with a small increase in the amplitude of the depolarising after-potential in sacral spinal roots (p<0.02), suggesting a reduction in the number of working K(+) channels in internodal axon membrane. Our study provides evidence that K(V)1.1 contributes to the control of peripheral sensory nerve excitability, and suggests that its characteristics as a putative drug target can be assessed by siRNA transfection in primary sensory neurons in vitro and in vivo.
authors: Queen Mary University of London, Barts and The London School of Medicine and Dentistry, Neuroscience and Trauma Centre, Blizard Institute of Cell and Molecular Science, 4 Newark Street, London E1 2AT, UK. m.d.baker@qmul.ac.uk
authors: Mark D
authors: MD
authors: Baker
authors: Ya-Chun
authors: YC
authors: Chen
authors: Syed U
authors: SU
authors: Shah
authors: Kenji
authors: K
authors: Okuse
mesh_terms: D000200:Action Potentials / Q000502:physiology*; D000818:Animals; D005727:Ganglia, Spinal / Q000166:cytology / Q000502:physiology; D051662:Kv1.1 Potassium Channel / Q000235:genetics* / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology; D034622:RNA Interference; D034741:RNA, Small Interfering; D051381:Rats; D017208:Rats, Wistar; D011984:Sensory Receptor Cells / Q000502:physiology*
chemical_list: D034741:RNA, Small Interfering; D051662:Kv1.1 Potassium Channel