Effect of electroacupuncture on arginine vasopressin-induced endolymphatic hydrops.

OBJECTIVE: To investigate the influence of electroacupuncture (EA) on experimentally induced endolymphatic hydrops (EH) in guinea pigs, and elucidate the association between the dehydrating effect of EA and changes in stria vascularis ultrastructure and expression of vasopressin type 2 receptor (V2R), cyclic adenosine monophosphate (cAMP), and aquaporin 2 (AQP2) in the endolymphatic sac (ES). METHODS: The EH model was established by intraperitoneal injection of arginine vasopressin (AVP). As a treatment, EA was delivered to Baihui (GV 20) and Tinggong (SI 19) acupoints, once daily for 10 consecutive days. For histomorphological studies, degree of cochlear hydrops was evaluated by hematoxylin-eosin staining, and the ratio of scala media (SM) area to SM + scala vestibuli area was calculated. In mechanical studies, ultrastructural changes in stria vascularis tissue were examined by transmission electron microscopy. In addition, cAMP levels and mRNA expression levels of V2R and AQP2 in the ES were compared among groups. RESULTS: EA treatment significantly reduced cochlear hydrops compared with hydropic guinea pigs (P = 0.015). Furthermore, EA attenuated ultrastructural changes in the stria vascularis tissue following EH, significantly upregulated the expression of V2R (P = 0.016), and attenuated AVP-induced upregulation of both cAMP (P = 0.038) and AQP2 expression (P = 0.017) in the ES. CONCLUSION: Collectively, the results of the present study suggest that the dehydrating effect of EA is associated with improvement of stria vascularis ultrastructure and V2R-cAMP-AQP2 signaling pathway regulation in the ES.

[Optimization of delivering minimum Gd-DTPA at the posterior upper point on tympanic medial wall and hT2W-3D-FLAIR sequence for detecting endolymphatic hydrops].

Objective: To optimize delivery of gadolinium-diethylenetriamine pentaacetic acid(Gd-DTPA) at the posterior upper point on tympanic medial wall and heavily T2-weighted 3-dimensional fluid-attenuated inversion recovery (hT2W-3D-FLAIR) sequence, and to implement the technique of detecting endolymphatic hydrops using gadolinium-enhancement MRI. Methods: Thirteen patients with periphery vertigo, who visited Department of Otorhinolaryngology Head and Neck Surgery, Shanghai Changhai Hospital during June and December of 2017, were enrolled in the study.0.10-0.20 ml of Gd-DTPA in various dilutions (10, 20, and 40-fold) were delivered at the posterior upper point on tympanic medial wall using a soft-tipped tympanic suction and drug-spraying needle through an artificially perforated tympanic membrane. Inner ear MRI was performed at 8, 24 h after Gd-DTPA administration using a 3T MR machine in combination with a 20-channel Tim 4G head/neck coil and the sequence of hT2W-3D-FLAIR to detect the gadolinium-enhancement signal within the inner ear and possible endolymphatic hydrops. The scanning time was either 8 min 35 s or 15 min 11 s. Results: Efficient inner ear uptake of Gd-DTPA was detected and induced high signal to noise ratio of MRI in patients receiving targeted delivery of 0.15-0.20 ml of 10-fold diluted contrast agent at the posterior upper point on tympanic medial wall. At 8 h after delivery, significant uptake was detected in the scala tympani and vestibuli of hook region and basal turn of the cochlea, and perilymhatic compartment of the vestibule. At 24 h after delivery, the distribution of Gd-DTPA became homogenous in each turn of the cochlea and perilymphatic compartment of the vestibule. However, obvious individual variance existed in the inner ear uptake when 0.10 ml of 40-fold diluted Gd-DTPA was delivered. Efficient inner ear uptake and high quality images that generated in patients receiving 0.10, 0.15, and 0.20 ml of 20-fold Gd-DTPA demonstrated endolymphatic hydrops with minor individual variance. There was insignificant difference in the enhancement signal of inner ear between 0.15 and 0.10 ml groups when Gd-DTPA was diluted at 20-fold except for the signal of semicircular canal of 0.15 ml group (190.00±53.95 vs 165.50±42.13, t=2.61, P<0.05). There was insignificant difference in the image quality between 8 min 35 s and 15 min 11 s canning time. Various degrees of endolymphatic hydrops were detected in 7 cochleae and 11 vestibule, and both simultaneous cochlear and vestibular endolymphatic hydrops were detected in 4 ears. Cochlear endolymphatic hydrops was detected in all the 3 patients with definite Meniere's disease, and 2 of them had combined cochlear and vestibular endolymphatic hydrops. Endolymphatic hydrops was not detected in patients with possible Meniere's disease nor with symptoms of superior semicircular canal dehiscence. Conclusion: Targeted delivery of 0.10 ml with 20-fold diluted Gd-DTPA (total dosage of 5 µmol) at the posterior upper point on tympanic medial wall in combination with 8 min 35 s scanning time hT2W-3D-FLAIR sequence for inner ear MRI in a 3T MR machine is a clinically practical method to detect endolymphatic hydrops, and reduce the requirement for MRI hardware.

[Effects of intratympanic injection of dexamethasone on endolymphatic hydrops and changes in guinea pigs inner ear CFTR expression].

OBJECTIVE: To observe the effect of tympanic injection of dexamethasone in the guinea pig endolymphatic hydrops and the change CFTR expression, to explore the effect of glucocorticoid treatment endolymphatic and its possible mechanism. METHOD: Thirty guinea pigs were divided into three groups: hormone group, water group, control group. The animals(hormone group, water group) in study were injected DDAVP 4 µg/kg in the first 7 d, and will increase to 6 µg/kg in the second 3 d. The control group was given normal saline, continuous 10 d. After twelfth days, the hormone group transtympanic injection of dexamethasone (5 mg/ml, 0.5 ml), and water group, control group transtympanic given normal saline (0.5 ml), continuous injection 5 d. Using immuno- histochemistry and Western blot to detect the cystic fibrosis transmembrane conductance regulator cochlear factor (CFTR) expression. RESULT: The water group ABR thresholds was significantly higher than that before the experiment (P < 0.01), and the water group was significantly higher than the rest of the groups (P < 0.01); Hormone group compared with the control group increased threshold value (P < 0.05). The control group had no endolym- phatic hydrops, the water group showed varying degrees of endolymphatic hydrops, cochlear duct and vestibular plus cochlear duct area ratio compared with the control group, hormone group was significantly higher (P < 0.01). hormone group area ratio was higher than the control group (P < 0.05). CFTR was primarily expressed in the stria vascularis, Corti's, spiral ligament, basilar membrane, cochlear ganglion,etc . The expression of CFTR in the water group was increased than that in the control group, and the hormone group (P < 0.01); the expression of hormone group increased compared with the control group (P < 0.05). CONCLUSION: Tympanic injection of dexa- methasone can alleviate the desmopressin acetatein guinea pigs caused by membranous labyrinth, and the improve- ment of the hearing; Tympanic injection of dexamethasone can make the endolymphatic hydrops cochlea of guinea pigs decreased CFTR expression, indicating that the expression and possible mechanisms of CFTR intratympanic steroids reduce endolymphatic hydrops changes.

Plasma vasopressin and V2 receptor in the endolymphatic sac in patients with delayed endolymphatic hydrops.

OBJECTIVE: There are some kinds of sicknesses provoked by inadequate adaptation to physical and/or psychogenic stress in daily life. Delayed endolymphatic hydrops (DEH) is an inner ear disease like Ménière's disease (MD) characterized by episodic vertigo in the setting of preexisting unilateral deafness that especially occurs in civilized people with a stressful lifestyle. Its otopathologic finding was demonstrated to be inner ear endolymphatic hydrops through a temporal bone study in 1976, as in the case with MD in 1938. To elucidate the relationship between stress and the inner ear, we examined the plasma antidiuretic stress hormone vasopressin (pAVP) and its type 2 receptor (V2R) expression in the endolymphatic sac in patients with DEH. STUDY DESIGN: A prospective molecular biological study. METHODS: Between 1998 and 2007, we enrolled 20 patients with ipsilateral DEH to examine their pAVP during remission from vertigo attacks. Plasma vasopressin was also examined in 87 patients with unilateral MD and 30 control patients with chronic otitis media. Using the real-time polymerase chain reaction method with tissue samples obtained during surgery, we examined V2R mRNA expression in the endolymphatic sac in 6 patients with ipsilateral DEH, 9 patients with unilateral MD, and 6 control patients with acoustic neuroma. RESULTS: Plasma vasopressin (1.5 times versus controls; unpaired t test, p = 0.140) and V2R mRNA expression in the endolymphatic sac (35.8 times versus controls; unpaired t test, p = 0.002) were higher in patients with DEH compared with those with acoustic neuroma. There were no significant differences in pAVP or V2R expression in the endolymphatic sac between DEH and MD. Patients with DEH showed a significantly negative correlation between pAVP and V2R (Pearson test, r = -0.92, p = 0.009) as in those with MD (Pearson test, r = -0.68, p = 0.043). CONCLUSION: Civilized people are frequently exposed to stress in their daily life, and pAVP can easily become elevated at any time. Therefore, a negative feedback system between pAVP and V2R in the endolymphatic sac may function for inner ear fluid homeostasis against stress-induced increases in pAVP. For the pathogenesis of endolymphatic hydrops resulting in vertigo attacks in patients with DEH as well as MD, pAVP may represent a matter of consequence, but V2R overexpression in the endolymphatic sac could be much more essential as a basis for these diseases.

[Expression of aquaporin 1 in two types of animal model of endolymphatic hydrops].

OBJECTIVE: To study the expression of aquaporin 1 (AQP1) in cochlea and endolymphatic sac of guinea pig of two types of animal model of endolymphatic hydrops. METHODS: Thirty guinea pigs were divided into three groups at random: surgery group; aldosterone group and control group. Each group included 10 animals. Surgical endolymphatic sac obstruction or aldosterone intraperitoneal injection was used to build the animal models of endolymphatic hydrops. Immunohistochemistry and western blot were used to examine the expression of AQP1 in cochlea and endolymphatic sac of animal models. Image processing soft (Image Tool) was used to do the semiquantitative analysis. RESULTS: Middle to high grade endolymphatic hydrops were found in surgery group. It was most significant in the apical coil, and lessened from the apical coil to the basal coil. Low to middle expression grade in endolymphatic hydrops was found in aldosterone group. Most of them were found in the basal coil. The positions of AQP1 expression in two animal models of endolymphatic hydrops were identical with the control group. In cochlea, no difference of AQP1 expression was found in the surgery and control group (t = 0.718 , P > 0.05) , but the expression of AQP1 was down regulated in the cochlea of aldosterone group (t = 6.609, P < 0.01) while the expressions of AQP1 in endolymphatic sac of aldosterone and control group were no difference between them (t = 0.998, P > 0.05). The quantization of AQP1 protein in the lateral wall of cochlea of aldosterone group was lower than that of control group (t = 13.626 , P < 0.01). CONCLUSIONS: The expression of AQP1 is no change in surgery and control group, but decreased in aldosterone group. The expression of AQP1 may be regulated by the ionic concentration in inner ear of guinea pig.

Preliminary study of the role of endothelin-1 in the homeostasis of the inner ear.

Endothelin (ET), originally characterized as a 21-residue vasoconstrictor peptide from endothelial cells, has been reported to act as a local hormonal regulator of pressure, fluid, ions, and neurotransduction. Our previous studies suggested an important role of ET-1 in the inner ear. The present study investigated the time kinetics of ET-1 in the epithelium of the endolymphatic sac (ES) of guinea pigs and its relation to the development of endolymphatic hydrops (EH) following locally mounted secondary immune reaction. In the duration between 12 h and day 1, ET-1-like activity completely disappeared from the epithelium of the ES and was associated with the accumulation of inflammatory cells in the ES and a rapid development of EH. On day 7, ET-1-like activity recovered as a consequence of the decrease of inflammatory cells and reduction of EH. These findings suggest that ET-1 may play an important role as one of the regulators maintaining the fluid balance.

Changes in immunostaining of cochleas with experimentally induced endolymphatic hydrops.

Cochleas with experimentally induced endolymphatic hydrops were immunostained for Na+,K(+)-ATPase, intracellular Ca(++)-ATPase, carbonic anhydrase, aldehyde dehydrogenase, calcium-binding proteins, vimentin, and the gap junction protein, connexin 26. No changes in immunostaining of hydropic ears were observed 1 week after blockage of the endolymphatic duct. Two weeks to 1 month after the operation, immunostaining of type I fibrocytes in the spiral ligament, which are positive for all but Na+,K(+)-ATPase, was slightly decreased on the operated side. These changes became more pronounced 3 months after the operation. However, staining for Na+,K(+)-ATPase of the stria vascularis and of type II fibrocytes of the spiral ligament was not reduced until 6 months postoperative. The reduction of enzymes and other cell constituents that may be involved in ion balance of cochlear fluids indicates that cells in the spiral ligament play an important role in cochlear homeostasis and that they merit further study in animal and human otopathology.

Endolymph calcium increases with time after surgical induction of hydrops in guinea-pigs.

The ionized Ca2+ concentration in cochlear endolymph is normally extremely low. Previous studies have shown that endolymph Ca2+ levels become elevated when measured at long intervals after endolymphatic hydrops is surgically induced. The present study was designed to investigate how rapidly endolymph Ca2+ increases following endolymphatic duct ablation. Hydropic animals were tested at either 4 days, 4 weeks, 8 weeks or 16 weeks after surgery. In each animal endolymph Ca2+ and endocochlear potentials were measured in all four cochlear turns using double-barreled Ca(2+)-sensitive electrodes. Cochlear sensitivity was assessed using compound action potential thresholds. Our results confirm that hydropic animals show an elevation of endolymph Ca2+ and a reduction of EP which is initially small, but becomes more pronounced at longer times after surgery. At 16 weeks endolymph Ca2+ was increased by an average factor of 20 in the basal turn and 7.5 in the fourth turn. These findings suggest that endolymph Ca2+ changes may not be the primary factor responsible for hydrops generation, but probably contribute to cochlear dysfunction in later phases of hydrops. For some experimental groups, the elevation of AP threshold was more closely correlated with endolymph Ca2+ level than it was with endolymph volume. Endolymph Ca2+ changes must therefore be considered in order to account for dysfunction in the hydropic cochlea.