Suzuyo Okazaki

Expression and Translocation of Aquaporin‐2 in the Endolymphatic Sac in Patients with Meniere’s Disease

Meniere’s disease, characterised by episodic vertigo, fluctuating hearing loss and tinnitus, can occur under conditions of stress. Its pathology was first revealed to be inner ear hydrops through temporal bone studies in 1938. Although its pathogenesis has been proposed to be a disorder of water transport in the inner ear, subsequently, it remains unsolved, until now. A recent study revealed that both plasma stress hormone, vasopressin (pAVP) and its receptor, V2 (V2R) expression in the inner ear endolymphatic sac were significantly higher in Meniere’s patients. In the present study, to link V2R‐related molecules and inner ear hydrops, we examined V2R‐linked water channel molecule, aquaporin‐2 (AQP2) expression and translocation in human endolymphatic sac. AQP2 mRNA expression in the endolymphatic sac was significantly higher in Meniere’s patients by using real‐time polymerase chain reaction, as further confirmed by western blotting. AQP2‐like immunoreactivity (‐LIR) was translocated from luminal to basolateral side with endosomal trapping in the endolymphatic sac at the time of AVP exposure in human endolymphatic sac tissue culture. The similar AQP2‐LIR translocation was also demonstrated by forskolin and blocked by vasopressin/V2R specific antagonist, OPC31260 and protein kinase A (PKA) specific antagonists, H‐89 and KT‐5720. We concluded that in the pathogenesis of inner ear hydrops resulting in Meniere’s attacks, pAVP elevation as a result of stress and subsequent V2R‐cAMP‐PKA‐AQP2 activation and endosomal trapping of AQP2 in the endolymphatic sac, might be important as a basis of this disease. Further experimental and clinical studies are needed to better clarify the neuroscientific relationship between stress and Meniere’s disease.

Functional expression of TRPV1 and TRPA1 in rat vestibular ganglia

Both TRPV1 and TRPA1 are non-selective cation channels. They are co-expressed, and interact in sensory neurons such as dorsal root ganglia (DRG) and trigeminal ganglia (TG), and are involved in nociception, being activated by nociceptive stimuli. Immunohistological localization of TRPV1 in vestibular ganglion (VG) neurons has been reported. Although TRPA1 is co-expressed with TRPV1 in DRG and TG neurons, it is unclear whether TRPA1 channels are expressed in VG neurons. Moreover, it is unknown whether TRPV1 and TRPA1 channels are functional in VG neurons. We investigated the expression of TRPV1 and TRPA1 in rat VG neurons by RT-PCR, in situ hybridization, immunohistochemistry, and Ca(2+) imaging experiments. Both TRPV1 and TRPA1 RT-PCR products were amplified from the mRNA of rat VG neurons. In situ hybridization experiments showed TRPV1 and TRPA1 mRNA expression in the majority of VG neurons. Immunohistochemistry experiments confirmed TRPV1 protein expression. In Ca(2+) imaging experiments, capsaicin, a TRPV1 agonist, induced a significant increase in intracellular calcium ion concentration ([Ca(2+)]i) in rat primary cultured VG neurons, which was almost completely blocked by capsazepine, a TRPV1-specific antagonist. Cinnamaldehyde, a TRPA1 agonist, also caused an increase in [Ca(2+)]i, which was completely inhibited by HC030031, a TRPA1-specific antagonist. Moreover, in some VG neurons, a [Ca(2+)]i increase was evoked by both capsaicin and cinnamaldehyde in the same neuron. In summary, our histological and physiological studies reveal that TRPV1 and TRPA1 are expressed in VG neurons. It is suggested that TRPV1 and TRPA1 in VG neurons might participate in vestibular function and/or dysfunction such as vertigo.

Transient low-tone air-bone gaps during convalescence immediately after canal plugging surgery for BPPV

OBJECTIVES The aim of the present study was to elucidate the time course and frequency patterns of transient low-tone air-bone gaps (ABGs) after canal plugging for intractable BPPV. METHODS We investigated eight patients with intractable BPPV who underwent canal plugging. Four were cases with posterior type (pBPPV) and the other four were those with horizontal type (hBPPV). Pure-tone audiometries (PTAs) were performed before and 7 days, 1 month and 6 months after surgery. ABGs (+) were defined as the three-tone-average ≥20dB formulated by (a+b+c)/3, where a, b, and c are ABGs at 0.25, 0.5, and 1kHz, respectively. RESULTS The ratio of the number of patients with ABGs (+) at the post-operative 7th day and 1st month was 100.0% (8/8). The ratio at the post-operative 6th month was 0.0% (0/8). There were no significant differences in the time course or frequency patterns of the ABGs between pBPPV and hBPPV. CONCLUSIONS We clearly demonstrated eight cases with intractable BPPV showing transient low-tone ABGs during convalescence immediately after canal plugging. During that period, patients also complained of motion-evoked dizziness. All these findings suggest that, during such a convalescence period, the plugged area might not be fixed yet and could still induce the dizziness and low-tone ABGs, as enlarged vestibular aqueduct syndrome and superior semicircular canal deficiency syndrome exhibit low-tone ABGs due to the third mobile inner ear window. More than one month after surgery, both the ABGs and dizziness could disappear according to fixation of the plugged area.

Effects of repeated optic flow stimulation on gait termination in humans

Abstract Conclusions: Because the basic strategies to stop walking are stored as motor programs, visual stimulation may have little influence on body deviation during gait termination and its time course. Walking velocity, however, demonstrated dynamic flexible changes, which may subserve the stable process of gait termination under variable circumstantial changes such as optic flow. Objective: The aim of this study was to examine the effect of repeated optic flow on body deviation and walking velocity during gait termination, which may be more complicated than continuous standing or walking. Methods: Twenty-three healthy subjects were instructed to start walking upon an acoustic cue and to stop walking when the scenery changed in a virtual reality environment. Subjects underwent eight control trials without optic flow and three sets of optic flow conditions including four trials each of optic horizontal and rotational movement randomly. Results: Repeated optic flow caused no significant change of body deviation or the time course of the gait termination process in comparison with that in the control. The walking velocity at the start of the termination process showed short-term flexibility that denoted a gradual increase over the trial for within-set and long-term flexibility that denoted a gradual decrease for between-set.

Changes in beta-2 adrenergic receptor and AMP-activated protein kinase alpha-2 subunit in the rat vestibular nerve after labyrinthectomy.

In the present study, to elucidate the role of vestibular ganglion (VG) after the unilateral labyrinthine damage, we examined quantitative changes in mRNA expression of beta-adrenergic receptors (bARs) and AMP-activated protein kinase alpha catalytic subunits (aAMPKs) in VG after unilateral labyrinthectomy (UL) in rats. Using the real-time PCR method, beta2 AR mRNA expression in bilateral VG and AMPK alpha2 mRNA expression in the ipsilateral VG were significantly up-regulated with the maximum increase at the postoperative 7 day and 1 day, respectively. The up-regulation of beta2 AR in bilateral VG was long-lasting until 28 days after UL and that of AMPK alpha2 in the ipsilateral VG was just transient within 7 days after UL. These mRNA changes were supported by immunohistochemical data. According to previous reports, both of bARs and aAMPKs could regulate mitochondrial uncoupling protein (UCP) mRNA expression in several kinds of tissues and therefore might have thermogenic neurotransmission and antioxidant neuroprotective roles in neuronal tissues. UL requires not only long-lasting response of VG for central vestibular neuro-plasticity around 2-4 weeks but rapid response of VG against apoptosis of peripheral vestibular epithelia-neuronal synapses. The present findings suggest that beta2 AR in bilateral VG and AMPK alpha2 in the ipsilateral VG might play important signaling roles after the unilateral labyrinthine damage.

Unilateral posterior canal-plugging surgery for intractable bilateral posterior canal-type benign paroxysmal positional vertigo

OBJECTIVE To investigate the effectiveness of unilateral posterior semicircular canal (PSCC)-plugging surgery for patients with intractable bilateral PSCC-type benign paroxysmal positional vertigo (P-BPPV). METHODS From July 2011 to December 2015, we diagnosed 136 patients with P-BPPV. Of these, 3 patients had bilateral P-BPPV, and in 2 of the 3, the condition had been refractory to conservative treatment for more than 1 year. We planned a staged PSCC-plugging surgery for these 2 patients; initially one side was treated, and the contralateral side was treated 6 months later. RESULTS After the first surgery, both patients experienced improvement in symptoms of vertigo and nystagmus on the operated side and no change on the non-operated side. Patients underwent the Epley maneuver for the non-operated side. In one case, the non-operated side was cured. In the other case, although the P-BPPV was not completely resolved, the patient was satisfied with the result of unilateral surgery because he was now able to turn in bed to the operated side without vertigo. Before surgery, he had experienced vertigo when turning even slightly in bed. CONCLUSION We propose that even unilateral PSCC-plugging surgery is effective for some patients with intractable bilateral P-BPPV.

5-HT3 receptor expression in the mouse vestibular ganglion

The 5-hydroxytryptamine type 3 (5-HT3) receptor is a ligand-gated ion channel and a member of the Cys-loop family of receptors. Previous studies have shown 5-HT3 receptor expression in various neural cells of the central and peripheral nervous systems. Although the function and distribution of the 5-HT3 receptor has been well established, its role in the inner ear is still poorly understood. Moreover, no study has yet determined its localization and function in the peripheral vestibular nervous system. In the present study, we reveal mRNA expression of both 5-HT3A and 5-HT3B receptor subunits in the mouse vestibular ganglion (VG) by RT-PCR and in situ hybridization (ISH). We also show by ISH that 5-HT3 receptor mRNA is only expressed in the VG (superior and inferior division) in the peripheral vestibular nervous system. Moreover, we performed Ca(2+) imaging to determine whether functional 5-HT3 receptors are present in the mouse VG, using a selective 5-HT3 receptor agonist, SR57227A. In wild mice, 32% of VG neurons responded to the agonist, whereas there was no response in 5-HT3A receptor knockout mice. These results indicate that VG cells express functional 5-HT3 receptor channels and might play a modulatory role in the peripheral vestibular nervous system.

A high jugular bulb and poor development of perivestibular aqueductal air cells are not the cause of endolymphatic hydrops in patients with Ménière’s disease

OBJECTIVE The presence of endolymphatic hydrops in the inner ear, which can be detected with gadolinium-enhanced magnetic resonance imaging (Gd-MRI), is widely recognized as the main pathological cause of Ménières disease (MD). However, the precise mechanisms underlying the development of endolymphatic hydrops remains unclear. One hypothesis proposes a relationship between the presence of a high jugular bulb (HJB) and MD, which disrupts the vestibular aqueduct leading to the development of endolymphatic hydrops. This study sought to identify anatomical features in MD patients using computed tomography (CT) images of the temporal bone. METHODS Fifty-nine MD patients meeting the AAO-HNS diagnostic criteria and exhibiting endolymphatic hydrops in Gd-MRI were enrolled between July 2009 and December 2015. We only included MD patients who showed unilateral endolymphatic hydrops in Gd-MRI. Sixty-six patients with otosclerosis or facial palsy were also enrolled as control participants. In both groups, patients with other pathologies (e.g., chronic otitis media or cholesteatoma) and patients <16years old were excluded. HJB was defined as a JB that was observable in the axial CT image at the level where the round window could be visualized. JB surface area was measured on the axial image at the level where the foramen spinosum could be visualized. Finally, to investigate the relationship between the pneumatization of perivestibular aqueductal air cells and the existence of endolymphatic hydrops, the development of the air cells was rated using a three-grade evaluation system and the distance between the posterior semicircular canal (PSCC) and the posterior fossa dura was measured. RESULTS The presence of HJB was observed in 22 of 59 affected sides of MD patients and in 17 healthy sides. The likelihood that HJB was detected on an affected side (22/39) was not significantly above chance (50%). The HJB detection rate did not significantly differ between the three groups (MD affected side, MD healthy side, and control patients). Furthermore, there were no significant group differences in JB surface area, distance between the PSCC and posterior fossa dura, or the development of perivestibular aqueductal air cells. CONCLUSION We did not find any relationship between the anatomy of the temporal bones and the existence of endolymphatic hydrops. Moreover, we found no evidence suggesting that HJB or poor development of perivestibular aqueductal air cells were the cause of endolymphatic hydrops in MD patients.

Office-based differential diagnosis of transient and persistent geotropic positional nystagmus in patients with horizontal canal type of benign paroxysmal positional vertigo.

Abstract Conclusion: A 30 s observation of geotropic positional nystagmus is sufficient to distinguish persistent geotropic positional nystagmus (PGPN) from transient geotropic positional nystagmus (TGPN) in patients with horizontal canal type of benign paroxysmal positional vertigo (H-BPPV) in ENT office. Objective: As a canalith repositioning procedure effectively treats H-BPPV with TGPN, but not PGPN, the differentiation between patients with PGPN and with TGPN is essential. The purpose of this study is to determine the observation period enough to distinguish TGPN from PGPN. Methods: This study first analyzed positional nystagmus images recorded with an infrared CCD camera three-dimensionally in 47 patients with H-BPPV. PGPN is distinguished from TGPN in patients with H-BPPV precisely by means of time constant calculated form analysis of positional nystagmus. Ten-second and 30-s movies were made of positional nystagmus of the all 47 patients. Ten independent otolaryngologists were then asked to distinguish TGPN from PGPN after a 10 s or 30 s observation of the geotropic positional nystagmus images in 47 patients with H-BPPV. Results: The sensitivity and specificity to distinguish TGPN from PGPN was 100% and 97% after 30 s observation, but 100% and 40% after 10 s observation, respectively.

Stress Hormone Sensitivity and Endolymphatic Hydrops

Objective: Ménière’s disease is peculiar to humans, and attacks of this affliction can occur under conditions of stress. The present study aimed to assess the link between inner ear hydrops in Ménière’s disease and vasopressin, an anti-diuretic stress hormone with a potential role in inner ear fluid homeostasis. Method: We obtained blood samples from Ménière disease patients to examine plasma vasopressin (pAVP) and obtained inner ear tissue during endolymphatic sac surgery to examine vasopressin type-2 receptor (V2R) in the endolymphatic sac. We examined pAVP and the relative V2R mRNA expression in the endolymphatic sac using real-time PCR. Results: A significant negative correlation was revealed between pAVP and inner ear V2R (P = .014). We also examined relative cyclic AMP (cAMP) activity in the endolymphatic sac using tissue culture and cAMP assay. Both pAVP (1.6 times vs controls; P = .048) and inner ear V2R mRNA expression (41.5 times vs controls; P = .022) were significantly higher in Ménière patients. cAMP activity was basally upregulated (2.1 times vs controls) and cAMP sensitivity to vasopressin application was largely elevated (4.9 times vs controls) in Ménière patients. Conclusion: We concluded that in the pathogenesis of inner ear hydrops resulting in Ménière attacks, pAVP elevation probably as a result of stress may present a matter of consequence, but susceptibility of the V2R-overexpressed and cAMP-hypersensitized inner ear to pAVP elevation might be essential as a basis of this disease.