Daizo Taguchi

Expressions of Aquaporin-2, Vasopressin Type 2 Receptor, Transient Receptor Potential Channel Vanilloid (TRPV)1, and TRPV4 in the Human Endolymphatic Sac†

Objective: To localize aquaporin (AQP)2, vasopressin type 2 receptor (V2‐R), and transient receptor potential channel vanilloid subfamily 1, 4 (TRPV1, TRPV4) in the human endolymphatic sac (ES).

Hormonal Aspects of Ménière’s Disease on the Basis of Clinical and Experimental Studies

Conclusions: Endolymph homeostasis is thought to be mediated by the vasopressin-aquaporin-2 (VP-AQP2) system in the inner ear. Endolymphatic hydrops, the morphological characteristics of Ménière’s disease (MD), seems to reflect the malregulation of the VP-AQP2 system in inner ear fluid. The elevation of plasma vasopressin (p-VP) level, which is often observed in MD and its related diseases, might be one of the causative factors underlying these diseases. Purpose of Review: Review of the role of the VP-AQP2 system in the inner ear fluid homeostasis and in the formation and development of endolymphatic hydrops. Recent Clinical and Experimental Findings: A clinical survey has revealed that the p-VP level is often elevated in MD and its related diseases and that the increase in the p-VP level was closely linked to vertigo attacks in MD. Experimental studies have revealed that proteins and mRNAs of aquaporin-2 and vasopressin type 2 receptor were expressed in the stria vascularis of the cochlea and the epithelium of the endolymphatic sac, and that the volume of the endolymphatic compartment was mediated by the activity of the VP-AQP2 system in the inner ear.

Expression of aquaporin1, 3, and 4, NKCC1, and NKCC2 in the human endolymphatic sac

OBJECTIVE To locate aquaporin (AQP) 1, 3, and 4, Na-K-2Cl cotransporter (NKCC) 1 and 2 in the human endolymphatic sac (ES). METHODS A sample of human ES was harvested during the removal of vestibular schwannoma via the translabyrinthine approach. The sample was immediately fixed in 4% paraformaldehyde and embedded in OCT compound. Immunohistochemistry was performed with AQP1, 3, and 4, NKCC1, and NKCC2 polyclonal antibodies. RESULTS AQP1, AQP3, and NKCC2 were strongly expressed in the epithelial layer of the ES. AQP4 and NKCC1 were weakly expressed in the epithelial layer of the ES. CONCLUSIONS As it is impossible to perform quantitative analysis based on the fluorescence intensity of each immunoreactivity, we have presented the existence of AQP1, 3, and 4, NKCC1, and NKCC2 in the ES. The expression of NKCC1 and 2 indicated that the ES may have both secretory and adsorptive functions to maintain the homeostasis of endolymph.

Aquaporins as potential drug targets for Meniere's disease and its related diseases.

The homeostasis of water in the inner ear is essential for maintaining function of hearing and equilibrium. Since the discovery of aquaporin water channels, it has become clear that these channels play a crucial role in inner ear fluid homeostasis. Indeed, proteins or mRNAs of AQP1, AQP2, AQP3, AQP4, AQP5, AQP6, AQP7 and AQP9 are expressed in the inner ear. Many of them are expressed mainly in the stria vascularis and the endolymphatic sac, which are the main sites of secretion and/or absorption of endolymph. Vasopressin type2 receptor is also expressed there. Water homeostasis of the inner ear is regulated in part via the arginine vasopressin-AQP2 system in the same fashion as in the kidney, and endolymphatic hydrops, a morphological characteristic of Menieres disease, is thought to be caused by mal-regulation of this system. Therefore, aquaporins appear to be important for the development of novel drug therapies for Menieres disease and related disorders.

A comparison of dehydration effects of V2-antagonist (OPC-31260) on the inner ear between systemic and round window applications

V2-antagonist (OPC-31260 (OPC)) application to the scala tympani reduced endolymphatic hydrops. In the present study, we investigated whether systemic administration or local infusion via the round window (RW application) of OPC would be more suitable for clinical use. In Experiment 1, the increase ratios of the cross-sectional area of the scala media of experimentally induced endolymphatic hydrops were quantitatively assessed among four groups of non-OPC application, RW application of xanthan gum, systemic application of OPC and RW application of OPC. In Experiment 2, the effects of systemic and RW applications of OPC on plasma vasopressin (p-VP) concentrations and plasma osmolality (p-OSM) were investigated. In Experiment 3, endocochlear DC potential (EP) was measured in guinea pigs with the RW application of OPC. Electron microscopic observations of the stria vascularis and the hair cells were also made. Both systemic and RW applications of OPC significantly reduced endolymphatic hydrops. However, systemic application resulted in the distension of the Reissners membrane in the non-operated ear, which seemed to be caused by elevated p-VP levels resulting from the systemic application of OPC. In contrast, RW application of OPC produced no apparent toxic effects in the inner ear, as indicated electrophysiological or morphological changes. Thus, drug delivery via the round window is more useful for the clinical application of OPC for medical decompression.

Expression and immunolocalization of aquaporin-6 (Aqp6) in the rat inner ear

Conclusion. Since aquaporin-6 (Aqp6) protein was located in the membrane of intracellular vesicles of the stria vascularis, endolymphatic sac, and vestibule, Aqp6 might be involved in some distinct physiological function of acid–base metabolism and water balance in endolymphatic fluid homeostasis. However, its lack of expression on the plasma membrane indicates that Aqp6 does not have a direct role in water flux via the plasma membrane. Objective. To evaluate the expression and immunolocalization of Aqp6 in the rat inner ear. Materials and methods. Wistar rats were used. Aqp6 mRNA expression in the rat inner ear was investigated in the vestibulum as well as in the cochlea and endolymphatic sac using the reverse transcription-polymerase chain reaction (RT-PCR) method, and detailed immunolocalization of Aqp6 in the rat inner ear was investigated using immunohistochemical methods including immunofluorescence microscopy and immunoelectron microscopy. Results. We obtained novel data showing that not just Aqp6 mRNA but also Aqp6 protein is expressed in the cochlea, endolymphatic sac, and vestibule. Immunoelectron microscopic studies revealed that the immunolabelled gold was diffusely seen in the intracellular area of the stria vascularis, endolymphatic sac, and vestibule, but never in the plasma membranes.

Presence and regulation of epithelial sodium channels in the marginal cells of stria vascularis

Conclusion. This study indicates that epithelial Na+-selective channels (ENaC) recycle Na+ via clathrin-mediated endocytosis in the marginal cells of the stria vascularis and that clathrin-independent endocytosis appeared to be modulated by the amount of Na+ transported. These results suggest the presence of ENaC in the luminal membrane of marginal cells and that ENaC are an efficient pathway for the uptake of Na+ from the endolymph. Objective. The ENaC found in many transporting epithelia play a key role in the regulation of salts and water homeostasis, cellular pH, cell volume, and cell function. Both biochemical and physiological approaches have been used to identify, characterize, and quantify this important channel, but its location in the marginal cells of the stria vascularis has not been fully clarified. The aim of this study was to determine the localization and regulation of ENaC. Materials and methods. Forty healthy female guinea pigs were used: 20 for the control experiment, 10 for the amiloride experiment, and 10 for the aldosterone experiment. We perfused cationized ferritin (CF) and microperoxidase (MPO) as tracers for clathrin-mediated and clathrin-independent endocytosis, respectively, into the cochlear duct. After 30 min of endolymphatic perfusion, the tissues were fixed and CF- and MPO-loaded endosomes within the marginal cell were observed by transmission electron microscopy. The numbers of CF- and MPO-loaded endosomes were compared between the three groups. Results. In the amiloride group, the numbers of CF- and MPO-loaded endosomes decreased in comparison with the control. In the aldosterone group, the numbers of CF- and MPO-loaded endosomes decreased and increased, respectively. Recently, it has been reported that ENaC are endocytosed via clathrin-mediated endosomes and aldosterone decreases the rate of endocytosis of ENaC. In this study, the results of the aldosterone experiment were consistent with those of recent studies.

Mastoid obliteration using calcium phosphate bone paste with an artificial dermis soaked with basic fibroblast growth factor: Preliminary clinical report

OBJECTIVES To describe mastoid obliteration using a calcium phosphate paste with an artificial dermis soaked with basic fibroblast growth factor (b-FGF) and the usefulness of this procedure. METHODS Ten ears that underwent a canal wall-down mastoidectomy were treated with calcium phosphate paste. The cases consisted of four patients with mastoid cavity problem, four with middle ear cholesteatoma, one with adhesive otitis media, and one with external auditory canal cholesteatoma. The post-operative follow-up period was 19.9 months on average (ranging from 13 to 23 months). First, the canal wall-down mastoidectomy was performed. Then, the calcium phosphate paste was used to fill up to the facial ridge posteriorly and the horizontal segment of the facial canal superiorly. The surface of the paste was covered with an artificial dermis soaked with b-FGF. RESULTS Following calcium phosphate paste treatment, all open cavities were decreased in volume and epithelized within 2 months. CONCLUSION Calcium phosphate paste appears effective for mastoid obliteration. Using an artificial dermis soaked with b-FGF appears to prevent exposure of the paste to the skin of the external auditory meatus. However, the number of the patients is too small to draw any conclusion.

Plasma antidiuretic hormone in cases with the early onset of profound unilateral deafness

OBJECTIVE The p-ADH level in cases of juvenile unilateral profound deafness (JUPD) and the timecourse of the level were examined to investigate whether or not an increase of p-ADH is involved in the development of delayed endolymphatic hydrops (DEH) in JUPD. MATERIALS AND METHODS In 90 consecutive patients with unilateral profound or total sensorineural deafness with the onset in early childhood, pure-tone audiometric examination and the measurement of p-ADH and plasma osmolality (p-OSM) were followed up once or twice a year as far as possible. At every testing, we performed careful history-taking about episodic vertigo/dizziness, fluctuant hearing loss, and tinnitus in order to find out whether patients had experienced these clinical signs of the development of DEH. RESULTS Means and standard deviation (S.D.) of p-ADH level and osmolality in all samples tested (n=368) were 7.3+/-7.0 pg/mL (0.7-52.0 pg/mL), and 288.6+/-4.4 mOsm/L (273-306 mOsm/L), respectively. The mean of p-ADH level was much higher than those previously reported in children and adolescents. High levels of p-ADH (over 5.0 pg/mL) were often observed in subjects between 6 and 19 years of age, but not so frequently in subjects of 20 years of age or older. Long-term follow-up of p-ADH levels revealed that DEH frequently developed in cases with persistent elevation of p-ADH. CONCLUSIONS The elevation of p-ADH is likely to promote the development of DEH in cases of JUPD, although the underlying mechanism remains to be elucidated.

Antidiuretic hormone and osmolality in isosorbide therapy and glycerol test.

Changes in plasma antidiuretic hormone (p-ADH) and plasma osmolality (p-Osm) levels were studied in 63 patients with Ménière’s disease before and after isosorbide administration and a glycerol test. Increments in both p-ADH and p-Osm levels were observed after isosorbide administration and the glycerol test. The p-ADH secretion appeared to be secondarily stimulated by an increase in the p-Osm level. This p-ADH level increase affects cochlear fluid homeostasis. Based on these results, we must consider both the p-Osm and p-ADH levels when treating patients with Ménière’s disease by osmotic diuretics.