Induction Of Endolymphatic Hydrops Research Articles

Aldosterone inhibits Dot1l expression in guinea pig cochlea

BackgroundAldosterone relieves transcriptional repression of epithelial sodium channel (ENaC) by inhibiting Dot1a and Af9 expression and their interaction with ENaC promoter in various tissues. Expressions of ENaC and Af9 in inner ear have been identified. However, it is not known how Dot1l is regulated by aldosterone in inner ear.MethodsTwenty-eight adult guinea pigs were randomly divided into the control group and treatment group. Aldosterone 1 mg/kg/d was injected intraperitoneally in the treatment group and saline in the control group for 7 days. Animals were killed 1 month later following auditory brainstem response examination. Histomorphology of cochlea was detected with hematoxylin–eosin staining, and Dot1l expression was examined with immunohistochemistry and Western blot.ResultsThere was no significant difference in ABR thresholds before and after injection of aldosterone or saline in either group. Endolymphatic hydrops was found in 75% of animals in the treatment group. Dot1l was found in both groups in the stria vascularis, Reissner’s membrane, spiral limbus, organ of Corti and spiral ligament. Dot1l expression in the treatment group was decreased by aldosterone.ConclusionsDot1l in guinea pig cochlea is inhibited by aldosterone with induction of endolymphatic hydrops. Dot1l may be closely related to endolymph regulation by aldosterone and to pathogenesis of Meniere’s disease.

Surgical Induction of Endolymphatic Hydrops by Obliteration of the Endolymphatic Duct

Surgical induction of endolymphatic hydrops (ELH) in the guinea pig by obliteration and obstruction of the endolymphatic duct is a well-accepted animal model of the condition and an important correlate for human Meniere's disease. In 1965, Robert Kimura and Harold Schuknecht first described an intradural approach for obstruction of the endolymphatic duct (Kimura 1965). Although effective, this technique, which requires penetration of the brain's protective covering, incurred an undesirable level of morbidity and mortality in the animal subjects. Consequently, Andrews and Bohmer developed an extradural approach, which predictably produces fewer of the complications associated with central nervous system (CNS) penetration.(Andrews and Bohmer 1989) The extradural approach described here first requires a midline incision in the region of the occiput to expose the underlying muscular layer. We operate only on the right side. After appropriate retraction of the overlying tissue, a horizontal incision is made into the musculature of the right occiput to expose the right temporo-occipital suture line. The bone immediately inferio-lateral the suture line (Fig 1) is then drilled with an otologic drill until the sigmoid sinus becomes visible. Medial retraction of the sigmoid sinus reveals the operculum of the endolymphatic duct, which houses the endolymphatic sac. Drilling medial to the operculum into the area of the endolymphatic sac reveals the endolymphatic duct, which is then packed with bone wax to produce obstruction and ultimately ELH. In the following weeks, the animal will demonstrate the progressive, fluctuating hearing loss and histologic evidence of ELH.

Induction of endolymphatic hydrops by directly infused monoclonal antibody against type II collagen CB11 peptide.

Autoimmune ear disease induced by type II collagen has been investigated by Yoo et al. In the present study, we investigated the effects of direct chronic infusion of type II collagen-specific monoclonal antibodies into guinea pig cochlea. Type II collagen fragment CB11 peptide-specific monoclonal antibodies (anti-CB11 Mabs) were infused directly into the scala tympani of guinea pigs with an Alzet mini-osmotic pump (anti-CB11 Mab group). As a control, normal mouse serum was infused by the same method (control group). To evaluate the auditory function, we recorded brain stem auditory-evoked potentials (BAEPs). In the anti-CB11 Mab group, 80% of the animals showed an increased hearing threshold of more than 25 dB at 7 days after infusion. The hearing threshold shift observed in the guinea pigs of the control group was minimal (15 dB or less). To detect the structural changes, we performed histopathologic studies using hematoxylin and eosin staining. Inflammatory cell migration was detected mainly in the scala tympani of the guinea pigs of both groups. In the anti-CB11 Mab group, endolymphatic hydrops was also observed. The results of this experiment suggest that type II collagen autoimmunity is responsible for the production of hearing loss and endolymphatic hydrops.

内リンパ水腫とＮａ｀＋´‐Ｋ｀＋´‐２Ｃｌ｀−´共輸送体 モルモットか牛の免疫組織化学的検討

Na+-K+-2Cl- cotransporter 1 (NKCC1) and c-Jun N-terminal kinase (JNK) were immunohistochemically studied in guinea pig cochlea following the induction of endolymphatic hydrops by blocking the endolymphatic duct. Postoperative survivals were set to 1 day (n=3), 3 days (n=4), 1 week (n=3), 2 weeks (n=4), 1 month (n=4) and 3 months (n=4). The most striking finding was an increase in the immunoreactivity for NKCC1 at type 1 fibrocytes of the spiral ligament throughout the survival times. This suggests that the affected cells shrink as the result of surgical intervention and upregulate NKCC1 to restore the volume of the cells. The shrinkage of type 1 fibrocytes should reduce gap junctional connectivity among cells of the ligament, which could reduce K+ supply into the stria vascularis and disturb ionic homeostasis in the endolymph. On the other hand, JNK, which has been reported to activate NKCC1 by phosphorylation, showed a consistent decrease in immunoreactivity in the types 1 and 2 fibrocytes in hydropic ears, indicating that JNK does not play an important role in the activation of NKCC1 in the cochlea.The findings in the present study suggested that endolymphatic homeostasis depends on cell integrity within the spiral ligament and that NKCC1 in type 1 fibrocytes is crucial in maintaining cochlear fluid homeostasis.

Vestibular Neurectomy in the Guinea-pig: a Retrosigmoid Approach

The efferent cochlear system, composed of both ipsi- and contralateral bundles, runs along with the vestibular nerve and may serve as protection for the inner ear. Sectioning of the vestibular nerve could thus create an experimental model for further in-depth study of such a protective role. A personal surgical approach is described, involving a suboccipital (or retrosigmoid) route to the postero-medial aspect of the guinea-pig temporal bone, chosen because of its use for the induction of endolymphatic hydrops in the same species. The crucial step in this approach is the separation of the sigmoid sinus in its entirety, through an intra-dural access with a minimal removal/retraction of cerebellar tissue. This did not affect either the post-operative recovery of the animal or its hearing threshold level as assessed via auditory brainstem responses.

Induction of endolymphatic hydrops in the guinea pig by perisaccular deposition of sepharose beads carrying and not carrying immune complexes

We tried to induce endolymphatic hydrops in guinea pig cochleas by unilateral, perisaccular deposition of sepharose beads carrying immune complexes. Controls consisted of the deposition of sepharose beads without immune complexes and the contralateral, untreated ear. The effects of the treatment were studied by light microscopy and electrophysiological recordings of the gross cochlear potentials 1, 2, and 6 weeks after treatment. Each condition included six animals. Analysis of variance of the morphometric data concerning the ears treated with deposition of the beads showed a statistically significant difference ( P=0.04) between the degree of hydrops found for the beads with immune complexes and for those without. The difference between the treated ears and the contralateral untreated ears was significant ( P=0.01) for the beads with immune complexes and not significant ( P=0.8) for those without immune complexes while there was no significant effect of post-treatment time interval. Analysis of variance of the electrophysiological data, collected in response to tone bursts at the apex of the cochlea, showed no significant differences between the results for the beads with and without immune complexes. Therefore these results were pooled. One week after treatment the pooled results for the compound action potential showed a small decrease in amplitude, just significant at 2 kHz, but not at 4 and 8 kHz. This decrease disappeared completely after 6 weeks. The pooled results for the negative summating potential (SP) showed a significant increase in magnitude at all frequencies decreasing with post-treatment interval. The cochlear microphonics did not demonstrate any change in amplitude after treatment. The results indicate that deposition of sepharose beads with immune complexes induces endolymphatic hydrops. Also, deposition of the sepharose beads itself may have induced hydrops together with enhancement of the SP. SP enhancement may be related to the development of endolymphatic hydrops rather than to the presence of hydrops as such. This may be based on pressure build-up while hydrops develops.

Single cochlear fibre responses in guinea pigs with long-term endolymphatic hydrops.

Some cochlear fibre response properties have been measured in two GPs approximately one year after induction of endolymphatic hydrops (by surgical obliteration of the endolymphatic sac and duct). These animals are considered as models of the effects of hydrops in Menière's disease, and the purpose of the study was to examine any modifications of fibre response properties which may underly auditory symptoms of the disease in man. Neurones towards more apical cochlear regions (with low characteristic frequencies) showed the greatest deterioration in tuning properties; on average, in the 1-6 kHz range, Q10dB values were reduced by a factor of two compared with normal animals. Discharge rate versus intensity functions of such units were abnormally steep, with dynamic ranges reduced by 10-20 dB. Towards higher frequency regions neurone response properties showed less deterioration (contrasting with many other types of cochlear pathology where, in general, the high frequency basal region exhibits greatest vulnerability). We have also observed in a few units an abnormal bursting in both spontaneous and driven discharge. Interspike intervals during burst are less than 1 ms (within relative refractory period). These findings are related to the auditory symptoms of Menière's disease, in particular, poor frequency selectivity, loudness recruitment and tinnitus.