Characterisation of DRASIC in the mouse inner ear.
ABSTRACT Within the cochlea, the hair cells detect sound waves and transduce them into receptor potential. The molecular architecture of the highly specialised cochlea is complex and until recently little was known about the molecular interactions which underlie its function. It is now clear that the coordinated expression and interplay of hundreds of genes and the integrity of cochlear cells regulate this function. It was hypothesised that transcripts expressed highly or specifically in the cochlea are likely to have important roles in normal hearing. Microarray analyses of the Soares NMIE library, consisting of 1536 cDNA clones isolated from the mouse inner ear, suggested that the expression of the mechanoreceptor DRASIC was enriched in the cochlea compared to other tissues. This amiloride-sensitive ion channel is a member of the DEG/ENaC superfamily and a potential candidate for the unidentified mechanoelectrical transduction channel of the sensory hair cells of the cochlea. The cochlear-enriched expression of amiloride-sensitive cation channel 3 (ACCN3) was confirmed by quantitative real-time polymerase chain reaction. Using in situ hybridisation and immunofluorescence, DRASIC expression was localised to the cells and neural fibre region of the spiral ganglion. DRASIC protein was also detected in cells of the organ of Corti. DRASIC may be present in cochlear hair cells as the ACCN3 transcript was shown to be expressed in immortalised cell lines that exhibit characteristics of hair cells. The normal mouse ACCN3 cDNA and an alternatively spliced transcript were elucidated by reverse transcription polymerase chain reaction from mouse inner ear RNA. This transcript may represent a new protein isoform with an as yet unknown function. A DRASIC knockout mouse model was tested for a hearing loss phenotype and was found to have normal hearing at 2 months of age but appeared to develop hearing loss early in life. The human homologue of ACCN3, acid-sensing ion channel 3, maps to the same chromosomal region as the autosomal recessive hearing loss locus DFNB13. However, we did not detect mutations in this gene in a family with DFNB13 hearing loss.
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ABSTRACT: Sudden Infant Death Syndrome (SIDS) remains the leading cause of infant mortality in Western societies. A prior study identified an association between hearing suppression on the newborn hearing test and subsequent death from SIDS. This is the first finding of an abnormality in SIDS cases prior to death. A following study identified that inner ear dysfunction precipitates a marked suppression of the hypercapnic ventilatory response (HCVR). Failure of arousal has been proposed to be a key component in SIDS. The objective of the present study was to assess whether inner ear dysfunction not only weakens the hypercapnic response, but also plays a role in suppressing the arousal response to suffocating gas mixtures. Wild type mice (n=28) received intra-tympanic gentamicin (IT-Gent) injections bilaterally or unilaterally to precipitate inner ear hair cell dysfunction. Three control groups (n=22) received intra-tympanic saline (IT-Saline) bilaterally or unilaterally (right or left), or intra-peritoneal gentamicin (IP-Gent).The body movement arousal responses to severe hypoxia- hypercarbia combined (5% CO2 in nitrogen) were tested under light anesthesia eight days following the administration of gentamicin or saline. After injections, the bilateral and unilateral IT-Gent treated animals behaved similarly to controls, however the HCVR as well as the arousal movements in response to severe hypoxia-hypercarbia were suppressed in IT-Gent treated animals compared to control animals (P<0.05). Thus the HCVR was significantly decreased in the bilateral (n=9) and unilateral IT-Gent treated mice (n=19) compared to bilateral (n=7) and unilateral IT-Saline (n=9) control groups (p<0.05). Arousal movements were suppressed in the bilateral IT-Gent group (n=9) compared to bilateral IT-Saline controls (n=7, P<0.0001) and in the unilateral IT-Gent group (n=19) compared to unilateral IT-Saline controls (n=10, P<0.0001). The findings support the theory that inner ear dysfunction could be relevant in the pathophysiology of SIDS. The inner ear appears to play a key role in arousal from suffocating gas mixtures that has not been previously identified.Neuroscience 09/2013; · 3.33 Impact Factor
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ABSTRACT: BACKGROUND:: Understanding the mechanisms underlying deep tissue pain in the postoperative period is critical to improve therapies. Using the in vitro plantar flexor digitorum brevis muscle-nerve preparation and patch clamp recordings from cultured dorsal root ganglia neurons innervating incised and unincised muscle, the authors investigated responses to various pH changes. METHODS:: Incision including the plantar flexor digitorum brevis muscle or sham operation was made in the rat hind paw. On postoperative day 1, in vitro single-fiber recording was undertaken. On the basis of previous studies, the authors recorded from at least 40 fibers per group. Also DiI-labeled dorsal root ganglia innervating muscle from rats undergoing incision and a sham operation were cultured and tested for acid responses, using whole cell patch clamp recordings. RESULTS:: The prevalence of responsive group IV afferents to lactic acid pH 6.5 in the incision group (15 of 67; 22.3%) was greater than that in the control group (2 of 35; 5.7%; P= 0.022). In dorsal root ganglia neurons innervating muscle, incision increased mean current amplitudes of acid-evoked currents; the acid-sensing ion channel blocker, amiloride 300μM, inhibited more than 75% of the acid-evoked current, whereas, the transient receptor vanilloid receptor 1 blocker (AMG9810 1 μM) did not cause significant inhibition. CONCLUSION:: The authors' experiments demonstrated that incision increases the responses of flexor digitorum brevis muscle afferent fibers to weak acid solutions, and increased acid-evoked currents in dorsal root ganglia innervating muscle. The authors' data suggest that up-regulation of acid-sensing ion channels might underlie this increased chemosensitivity caused by surgery.Anesthesiology 05/2013; · 6.17 Impact Factor
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ABSTRACT: Advanced gene targeting technology and related tools in mice have been incorporated into studies of acid-sensing ion channels (ASICs). A single ASIC subtype can be knocked out specifically and screened thoroughly for expression in the nervous system at the cellular level. Mapping studies have further shed light on the initiation and identification of related behavioral phenotypes. Here we review studies involving genetically engineered mouse models used to investigate the physiological function of individual ASIC subtypes: ASIC1 (and ASIC1a), ASIC2, ASIC3 and ASIC4. We discuss the detailed expression studies and significant phenotypes revealed with gene knockout for most known Asic subtypes. Each strategy designed to manipulate mouse genetics has advantages and disadvantages. We discuss the limitations of these Asic-knockout models and propose future directions to solve the genetic issues. Copyright © 2014. Published by Elsevier Ltd.Neuropharmacology 01/2015; · 4.82 Impact Factor