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Amiloride‐sensitive channels are a major contributor to mechanotransduction in mammalian muscle spindles
Author(s) -
Simon Anna,
Shenton Fiona,
Hunter Irene,
Banks Robert W.,
Bewick Guy S.
Publication year - 2010
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2009.182683
Subject(s) - amiloride , epithelial sodium channel , acid sensing ion channel , sodium channel , chemistry , mechanotransduction , medicine , endocrinology , ion channel , microbiology and biotechnology , biology , sodium , receptor , biochemistry , organic chemistry
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 1 m m . 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 μ m , while 10 μ m HMA was required for highly significant inhibition ( P < 0.0001). Conversely, amiloride, benzamil and EIPA rarely blocked firing entirely at 1 m m , whereas 1 m m HMA blocked 12 of 16 preparations. This pharmacology suggests low‐affinity ENaCs are the important spindle mechanotransducer. In agreement with 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 related brain sodium channel ASIC2 (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.