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Durham University

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Publication details for Dr Robert William Banks

Simon, Anna., Shenton, Fiona., Hunter, Irene., Banks, Robert W. & Bewick, Guy S. (2010). Amiloride-sensitive channels are a major contributor to mechanotransduction in mammalian muscle spindles. Journal of Physiology 588(1): 171-185.

Author(s) from Durham

Abstract

We investigated whether channels of the epithelial sodium/amiloride-sensitive degenerin(ENaC/DEG) family are a major contributor to mechanosensory transduction in primary mechanosensory afferents, using adult rat muscle spindles as a model system. Stretch-evoked afferent discharge was reduced in a dose-dependent manner by amiloride and three analogues – benzamil, 5-(N-ethyl-N-isopropyl) amiloride (EIPA) and hexamethyleneamiloride (HMA), reaching ≥85% inhibition at 1mm. Moreover, firing was slightly but significantly increased by ENaC δ subunit agonists (icilin and capsazepine). HMA’s profile of effects was distinct from that of the other drugs. Amiloride, benzamil and EIPA significantly decreased firing (P <0.01 each) at 1 μm, while 10 μm HMA was required for highly significant inhibition (P <0.0001). Conversely, amiloride, benzamil and EIPA rarely blocked firing entirely at 1mm, whereas 1mm HMA blocked 12 of 16 preparations. This pharmacology suggests low-affinity ENaCs are the important spindlemechanotransducer. In agreementwith this, immunoreactivity to ENaC α, β and γ subunits was detected both by Western blot and immunocytochemistry. Immunofluorescence intensity ratios for ENaC α, β or γ relative to the vesicle marker synaptophysin in the same spindle all significantly exceeded controls (P <0.001). Ratios for the relatedbrain sodiumchannelASIC2(BNaC1α)were also highly significantly greater (P <0.005). Analysis of confocal images showed strong colocalisation within the terminal of ENaC/ASIC2 subunits and synaptophysin. This study implicates ENaC and ASIC2 in mammalian mechanotransduction. Moreover, within the terminals they colocalise with synaptophysin,a marker for the synaptic-like vesicles which regulate afferent excitability in these mechanosensitive endings.