CD31 defines a subpopulation of human adipose-derived regenerative cells with potent angiogenic effects.

Cellular heterogeneity represents a major challenge for regenerative treatment using freshly isolated Adipose Derived Regenerative Cells (ADRCs). Emerging data suggest superior efficacy of ADRCs as compared to the ex vivo expanded and more homogeneous ADRCs (= ASCs) for indications involving (micro)vascular deficiency, however, it remains unknown which ADRC cell subtypes account for the improvement. Surprisingly, we found regarding erectile dysfunction (ED) that the number of injected CD31+ ADRCs correlated positively with erectile function 12 months after one bolus of autologous ADRCs. Comprehensive in vitro and ex vivo analyses confirmed superior pro-angiogenic and paracrine effects of human CD31+ enriched ADRCs compared to the corresponding CD31- and parent ADRCs. When CD31+, CD31- and ADRCs were co-cultured in aortic ring- and corpus cavernous tube formation assays, the CD31+ ADRCs induced significantly higher tube development. This effect was corroborated using conditioned medium (CM), while quantitative mass spectrometric analysis suggested that this is likely explained by secretory pro-angiogenic proteins including DKK3, ANGPT2, ANAX2 and VIM, all enriched in CD31+ ADRC CM. Single-cell RNA sequencing showed that transcripts of the upregulated and secreted proteins were present in 9 endothelial ADRC subsets including endothelial progenitor cells in the heterogenous non-cultured ADRCs. Our data suggest that the vascular benefit of using ADRCs in regenerative medicine is dictated by CD31+ ADRCs.

Summating potentials from the utricular macula of anaesthetized guinea pigs.

The Summating Potential (SP) was first recorded in the cochlea in the 1950s and represents an objective measure of cochlear hair cell function, in vivo. Despite being a regular tool in hearing research, a similar response has not yet been recorded from the vestibular system. This is mainly due to the lack of experimental techniques available to record electrical vestibular hair cell responses in isolation from the much larger cochlear potentials. Here we demonstrate the first recordings of the vestibular SP, evoked by Bone-Conducted Vibration (BCV) and Air-Conducted Sound (ACS) stimuli, in anaesthetized guinea pigs. Field potential measurements were taken from the basal surface of the utricular macula, and from the facial nerve canal following surgical or chemical ablation of the cochlea. SPs were evoked by stimuli with frequencies above ~200 Hz, and only with moderate to high intensity (~0.005-0.05 g) BCV and ACS (~120-140 dB SPL). Neural blockade abolished the Vestibular short-latency Evoked Potential (VsEP) and Vestibular Nerve Neurophonic (VNN) from the facial nerve canal recordings but did not abolish the vestibular SP nor the vestibular microphonic. Importantly, the vestibular SP was irreversibly abolished from the utricle and facial nerve canal recordings following local gentamicin application, highlighting its hair cell origin. This is the first study to record the Summating Potential from the mammalian vestibular system, in vivo, providing a novel research tool to assess vestibular hair cell function during experimental manipulations and animal models of disease.

Otopetrin-2 Immunolocalization in the Human Macula Utricle.

BACKGROUND: In the present study, we investigated the localization of otopetrin-2-a member of the otopetrin family that encodes proton-selective ion channels-in the human macula utricle using immunohistochemistry. METHODS: Macula utricle were acquired at surgery from patients who required transmastoid labyrinthectomy for intractable vertigo due to Meniere's disease (MD; n = 3) and/or vestibular drops attacks (VDA; n = 2) and from temporal bones (n = 2) acquired at autopsy from individuals with no balance disorders. Immunofluorescence staining with otopetrin-2 (rabbit affinity purified polyclonal antibody) and GFAP (mouse monoclonal antibody) to identify vestibular supporting cells was made in formalin fixed cryostat sections or whole microdissected utricle (for flat mount preparations). Secondary antibodies against rabbit and mouse were used for the identification of both proteins. Digital fluorescent images were obtained using a high-resolution laser confocal microscope. RESULTS: Using cryostat sections and flat mount preparations otopetrin-2 immunofluorescence was seen as punctated signal throughout the supporting cells cytoplasm. GFAP immunofluorescence was present in the supporting cell cytoplasm. The distribution of otopetrin-2 was similar in the macula utricle obtained from MD, VDA, or autopsy normative patients. CONCLUSIONS: Otopetrin-2 was localized in supporting cells in a similar fashion that otopetrin-1 previously reported in the mouse macula utricle. The differential expression of otopetrin-2 in the supporting cells of the human macula utricle suggest an important role in the vestibular sensory periphery homeostasis and otolith maintenance.

Dynamic response to sound and vibration of the guinea pig utricular macula, measured in vivo using Laser Doppler Vibrometry.

With the use of a commercially available Laser Doppler Vibrometer (LDV) we have measured the velocity of the surgically exposed utricular macula in the dorsoventral plane, in anaesthetized guinea pigs, during Air Conducted Sound (ACS) or Bone Conducted Vibration (BCV) stimulation. We have also performed simultaneous measurements of otolithic function in the form of the Utricular Microphonic (UM) and the Vestibular short-latency Evoked Potential (VsEP). Based on the level of macular vibration measured with the LDV, the UM was most sensitive to ACS and BCV between 100 and 200 Hz. The phase of the UM relative to the phase of the macular motion was relatively consistent across frequency for ACS stimulation, but varied by several cycles for BCV stimulation, suggesting a different macromechanical mode of utricular receptor activation. Moreover, unlike ACS, BCV evoked substantially distorted UM and macular vibration responses at certain frequencies, most likely due to complex resonances of the skull. Analogous to LDV studies of organ of Corti vibration, this method provides the means to study the dynamic response of the utricular macula whilst simultaneously measuring function.

The role of jab1, a putative downstream effector of the neurotrophic cytokine macrophage migration inhibitory factor (MIF) in zebrafish inner ear hair cell development.

Macrophage migration inhibitory factor (MIF) is a neurotrophic cytokine essential for inner ear hair cell (HC) development and statoacoustic ganglion (SAG) neurite outgrowth, and SAG survival in mouse, chick and zebrafish. Another neurotrophic cytokine, Monocyte chemoattractant protein 1 (MCP1) is known to synergize with MIF; but MCP1 alone is insufficient to support mouse/chick SAG neurite outgrowth or neuronal survival. Because of the relatively short time over which the zebrafish inner ear develops (~30hpf), the living zebrafish embryo is an ideal system to examine mif and mcp1 cytokine pathways and interactions. We used a novel technique: direct delivery of antisense oligonucleotide morpholinos (MOs) into the embryonic zebrafish otocyst to discover downstream effectors of mif as well as to clarify the relationship between mif and mcp1 in inner ear development. MOs for mif, mcp1 and the presumptive mif and mcp1 effector, c-Jun activation domain-binding protein-1 (jab1), were injected and then electroporated into the zebrafish otocyst 25-48hours post fertilization (hpf). We found that although mif is important at early stages (before 30hpf) for auditory macular HC development, jab1 is more critical for vestibular macular HC development before 30hpf. After 30hpf, mcp1 becomes important for HC development in both maculae.

[A new positioning maneuver for treating a subtype of postural unsteadiness - based on a modified model of the utricular macula]. / Ein neues Lagerungsmanöver zur Behandlung einer Form von Schwankschwindel auf der Basis eines modifizierten Modells der Macula utriculi

Objectives: A new method for the treatment of a subtype of postural unsteadiness frequently seen in the medical office is proposed. Ten years of successful practical application and continuous refining of this new treatment lead to the definition of symptom-criteria, which are useful to distinguish patients, who get benefit by this new procedure and to a modified model of the utricular macula. Materials and Methods: In a retrospective collection, the data of 15 patients with postural unsteadiness (mean duration of symptoms 61.7 months) successfully treated with the new maneuver are reported. Before treatment, all patients showed normal results in testing the caloric responses of the labyrinth, normal neurological investigation and no signs for cervical vertigo. Before and after treatment the DHI-score (Dizziness Handicap Index) was achieved by using the German version of the DHI-questionnaire. In all groups before and after treatment balance testing was performed using a posturography platform. Results: Application of the new treatment lead to a significant improvement of the DHI-scores and of the posturographic results (significant reduction of body sway). Discussion: The patients, who were successfully treated by the new maneuver have had presented the same symptoms like patients with a proven utricular dysfunction. This leads to a slightly modified model of the utricular macula. Beyond correctly positioned otoconia in the utricular macula and free otoconia in the endolymphatic utricular space, a third fraction of dysfunctional partially detached otoconia is postulated (causing the unsteadiness). Based on this theory, the single hair-cell with its accompanied otoconium might be the smallest receptor-unit of the macula-organ. This smallest receptor-unit might represent a biological mass-spring-system. According to technical mass-spring-systems used as accelerometers, the different sizes of the otoconia for a distinct direction of acceleration are necessary for covering different frequency-bands. Finally, a many years successfully tested modified variation of the new maneuver for treatment in the medical office and for self-treatment is proposed.

Ontogenetic development of the inner ear saccule and utricle in the Lusitanian toadfish: Potential implications for auditory sensitivity.

Studies addressing structure-function relationships of the fish auditory system during development are sparse compared to other taxa. The Batrachoididae has become an important group to investigate mechanisms of auditory plasticity and evolution of auditory-vocal systems. A recent study reported ontogenetic improvements in the inner ear saccule sensitivity of the Lusitanian toadfish, Halobatrachus didactylus, but whether this results from changes in the sensory morphology remains unknown. We investigated how the macula and organization of auditory receptors in the saccule and utricle change during growth in this species. Inner ear sensory epithelia were removed from the end organs of previously PFA-fixed specimens, from non-vocal posthatch fry (<1.4 cm, standard length) to adults (>23 cm). Epithelia were phalloidin-stained and analysed for area, shape, number and orientation patterns of hair cells (HC), and number and size of saccular supporting cells (SC). Saccular macula area expanded 41x in total, and significantly more (relative to body length) among vocal juveniles (2.3-2.9 cm). Saccular HC number increased 25x but HC density decreased, suggesting that HC addition is slower relative to epithelial growth. While SC density decreased, SC apical area increased, contributing to the epithelial expansion. The utricule revealed increased HC density (striolar region) and less epithelial expansion (5x) with growth, contrasting with the saccule that may have a different developmental pattern due to its larger size and main auditory functions. Both macula shape and HC orientation patterns were already established in the posthatch fry and retained throughout growth in both end organs. We suggest that previously reported ontogenetic improvements in saccular sensitivity might be associated with changes in HC number (not density), size and/or molecular mechanisms controlling HC sensitivity. This is one of the first studies investigating the ontogenetic development of the saccule and utricle in a vocal fish and how it potentially relates to auditory enhancement for acoustic communication.

The blood labyrinthine barrier in the human normal and Meniere's disease macula utricle.

The ultrastructural organization of the blood labyrinthine barrier (BLB) was investigated in the human vestibular endorgan, the utricular macula, using postmortem specimens from individuals with documented normal auditory and vestibular function and surgical specimens from patients with intractable Meniere's disease. Transmission electron microscopic analysis of capillaries located in the normal human utricular stroma showed vascular endothelial cells with few pinocytotic vesicles, covered by a smooth and uniform basement membrane surrounded by pericyte processes. Meniere's disease specimens revealed differential ultrastructural pathological changes in the cellular elements of the microvasculature. With moderate degeneration of the BLB, there were numerous vesicles within the vascular endothelial cells (VECs), with increased numbers at the abluminal face, pericyte process detachment and disruption of the perivascular basement membrane surrounding the VECs. With severe degeneration of the BLB, there was severe vacuolization or frank apparent necrosis of VECs and loss of subcellular organelles. A higher severity of BLB degenerative changes was associated with a higher degree of basement membrane thickening and edematous changes within the vestibular stroma. This study presents the first ultrastructural analysis of the capillaries constituting the BLB in the human vestibular macula utricle from normal and Meniere's disease.

[The ultrastructure of utricular maculae of mouse].

Objective: Using scanning electron microscope to observe the ultrastructure of utricular maculae of mouse. Methods: Ten young (6 to 8 weeks) and ten old (>12 months) mice were executed, and their utricles were harvested and the specimens were processed, using scanning electron microscope to observe the structures of the utricles from the surface of otoconia layer to the roots of hair cell cilia. Results: Under the scanning electron microscope, several ultrastructures were observed, including otoconia layer, unstructured gelatinous extracellular matrix layer, honeycomb-like gelatinous extracellular matrix layer, inter-cilia otoconia and hair cell cilia associated with these structures. When compared with young mouse, the otoconia surface of aged mouse was smoother, the gelatinous extracellular matrix between the adjacent otoconias was thinner. Conclusions: Using SEM, ultrastructures can be clearly observed from surface otoconia layer to the roots of hair cell cilia. By the analysis of the ultrastructure of utricular maculae, it is helpful for investigation of the pathological mechanisms of vestibular diseases, such as otolith diseases.

Deterioration of Vestibular Cells in Labyrinthitis.

OBJECTIVE: To quantitatively assess the effect of serous labyrinthitis, suppurative labyrinthitis, and labyrinthitis ossificans on vestibular hair cells, dark cells, and transitional cells. METHODS: We examined human temporal bone specimens with serous labyrinthitis, suppurative labyrinthitis, and labyrinthitis ossificans, then compared them with age-matched control groups without labyrinthitis. We evaluated the density of type I and II vestibular hair cells, dark cells, and transitional cells in the peripheral sensorial organs. RESULTS: The mean density of type I vestibular hair cells in the maculae of the saccule significantly differed between the serous labyrinthitis group and its control group. The loss of type I and II vestibular hair cells in all of the peripheral sensorial organs was significantly higher in the suppurative labyrinthitis group than in its control group. The mean density of dark cells in the lateral and posterior semicircular canals was significantly lower in the suppurative labyrinthitis group than in its control group. The mean density of type I and II vestibular hair cells, dark cells, and transitional cells was significantly lower in the labyrinthitis ossificans group than in its control group. CONCLUSION: The loss of vestibular hair cells and degenerative changes in dark cells and transitional cells could affect vestibular function in patients with labyrinthitis.

Superior canal dehiscence reveals concomitant unilateral utricular loss (UUL).

CONCLUSION: We report enhanced symmetrical cervical vestibular evoked myogenic potential (cVEMP) but asymmetrical ocular VEMP (oVEMP) responses in a patient with CT-verified bilateral superior semicircular canal dehiscence (SCD) but with acute vestibular syndrome. This implies that absence of unilateral utricular macula function alone is sufficient to cause symptoms of acute vertigo. Acute vertigo should not automatically be presumed to originate from semicircular canal dysfunction. OBJECTIVES: To identify the cause of an acute vertigo attack in a patient with bilateral SCD. METHODS: The functional state of all peripheral vestibular sense organs was tested using the video head impulse test (vHIT) for all semicircular canals and VEMPs to air-conducted sound (ACS) or bone-conducted vibration (BCV) to test all otolith organs. The cVEMP tested mainly saccular function and the oVEMP mainly utricular function. RESULTS: All semicircular canals showed normal function. The cVEMPs showed enhanced, but symmetrical saccular function. In contrast, oVEMPs showed an enhanced but asymmetric n10 component - it was greatly reduced beneath the left eye, implying decreased function in the right utricular macula. That result was confirmed using very high frequency stimuli which are effective in SCD: 4000 Hz BCV stimuli showed that oVEMP n10 was present beneath the right eye but absent beneath the left eye.

Hearing characteristics of cephalopods: modeling and environmental impact study.

Cephalopods (octopus, squid and cuttlefish) are some of the most intriguing molluscs, and they represent economically important commercial marine species for fisheries. Previous studies have shown that cephalopods are sensitive to underwater particle motion, especially at low frequencies in the order of 10 Hz. The present paper deals with quantitative modeling of the statocyst system in three cephalopod species: Octopus vulgaris, Sepia officinalis and Loligo vulgaris. The octopus's macula/statolith organ was modeled as a 2nd-order dynamic oscillator using parameter values estimated from scanning electron micrograph images. The modeling results agree reasonably well with experimental data (acceleration threshold) in the three cephalopod species. Insights made from quantitative modeling and simulating the particle motion sensing mechanism of cephalopods elucidated their underwater particle motion detection capabilities. Sensitivity to emerging environmental issues, such as low frequency noise caused by near-shore wind farms and increasing levels of carbon dioxide in the ocean, and sensitivity to sounds produced by impending landslides were investigated in octopus using the model.

ACF7 is a hair-bundle antecedent, positioned to integrate cuticular plate actin and somatic tubulin.

The precise morphology of the mechanosensitive hair bundle requires seamless integration of actin and microtubule networks. Here, we identify Acf7a (actin crosslinking family protein 7a) as a protein positioned to bridge these distinct cytoskeletal networks in hair cells. By imaging Acf7a-Citrine fusion protein in zebrafish and immunolabeling of vestibular and cochlear mouse hair cells, we show that Acf7a and ACF7 circumscribe, underlie, and are interwoven into the cuticular plate (CP), and they also encircle the basal body of the kinocilium. In cochlear hair cells, ACF7 localization is graded, with the highest concentration near each fonticulus--an area free of F-actin in the region of the CP that contains the basal body. During hair-cell development and regeneration, Acf7a precedes formation of the hair bundle and CP. Finally, electron tomography demonstrates that the ends of microtubules insert into the CP and are decorated with filamentous linkers connecting microtubules to the CP. These observations are consistent with ACF7 being a linker protein, which may shape the cytoskeleton of the hair cell early during hair-bundle genesis.

Temporal and spatial distribution of gentamicin in the peripheral vestibular system after transtympanic administration in guinea pigs.

BACKGROUND AND OBJECTIVE: Transtympanic administration of gentamicin is effective for treating patients with intractable vertigo. This study explored the spatial and temporal distribution of gentamicin in vestibular end-organs after transtympanic administration. METHODS: Thirty guinea pigs were transtympanically injected with gentamicin conjugated to Texas Red (GTTR) and their vestibular end-organs examined after various survival periods. Another 9 guinea pigs received GTTR at different doses. Nine animals received Texas Red only and served as controls. We used confocal microscopy to determine the cellular distribution of GTTR in semicircular canal cristae, as well as the utricular and saccular maculae. RESULTS: The most intense GTTR labeling was present in the saccule compared to other vestibular end-organs. GTTR fluorescence was detected predominantly in type I hair cells, type II hair cells and transitional cells after a single transtympanic dose of GTTR (0.1 mg/ml, 0.05 ml), while only weak fluorescence was observed in non-sensory cells such as supporting cells, dark cells and lumenal epithelial cells. Transitional cells displayed intense GTTR fluorescence in the supra-nuclear regions 24 h after transtympanic injection that was retained for at least 4 weeks. A decreasing spatial gradient of GTTR fluorescence was observed sensory epithelial regions containing central type I to peripheral type I and then type II hair cells in the crista ampullaris, and from striolar to extra-striolar hair cells within the vestibular macula. GTTR fluorescence extended from being restricted to the apical cytoplasm at lower doses to the entire cell body of type I hair cells with increasing dose. GTTR fluorescence reached peak intensities for individual regions of interest within the cristae and maculae between 3 and 7 days after transtympanic injection. CONCLUSION: The saccular uptake of GTTR is greater than other vestibular end-organs after transtympanic injection in the semicircular canals.

[Isolated functional loss of the lateral semicircular canal in vestibular neuritis]. / Isolierte Rezeptorfunktionsstörung des horizontalen Bogengangs.

Today, modern tools in vestibular testing are feasible to provide information about functional status of all five peripheral vestibular receptors. Isolated or combined loss of crista and macula receptor function can be determined in the diagnostic process. We describe a seldom case of isolated functional loss of lateral semicircular canal receptor function in a 55-year-old patient. Whereas there was no ispilateral caloric response and video head impulse test revealed a catch-up saccade, air-conducted (AC) cervical and ocular vestibular-evoked myogenic potentials (cVEMP, oVEMP), subjective visual vertical and MRI were normal.

Discromatosis hereditaria universal: relato de un caso / Dyschromatosis universal hereditary: case report

Comunicamos un caso de discromatosis hereditaria universal (DHU) en una paciente adolescente de 12 años de edad, que acude a la consulta por múltiples máculas hipo e hiperpigmentadas, asintomáticas y de distribución universal, con un patrón reticulado. Las lesiones aparecieron y fueron aumentando en número desde los 3 meses de edad, sin otro hallazgo patológico. Realizamos, además, una breve revisión de la literatura sobre el tema.

We report a case of dyschromatosis universalis hereditaria (DUH) in a 12 years old female with chief complain of asymptomatic, multiple hypopigmented and hyperpigmented macules, with reticulate pattern, involving almost the entire body, of many years of duration. The literature regarding this topic is also discussed.

Variations in shape-sensitive restriction points mirror differences in the regeneration capacities of avian and mammalian ears.

When inner ear hair cells die, humans and other mammals experience permanent hearing and balance deficits, but non-mammalian vertebrates quickly recover these senses after epithelial supporting cells give rise to replacement hair cells. A postnatal decline in cellular plasticity appears to limit regeneration in mammalian balance organs, where declining proliferation responses are correlated with decreased spreading of supporting cells on artificial and native substrates. By culturing balance epithelia on substrates that differed in flexibility, we assessed spreading effects independent of age, showing a strong correlation between shape change and supporting cell proliferation. Then we made excision wounds in utricles cultured from young and old chickens and mice and compared quantified levels of spreading and proliferation. In utricles from young mice, and both young and old chickens, wounds re-epithelialized in <24 hours, while those in utricles from mature mice took three times longer. More cells changed shape in the fastest healing wounds, which accounted for some differences in the levels of proliferation, but inter-species and age-related differences in shape-sensitive restriction points, i.e., the cellular thresholds for shape changes that promote S-phase, were evident and may be particularly influential in the responses to hair cell losses in vivo.

Mefloquine damage vestibular hair cells in organotypic cultures.

Mefloquine is an effective and widely used anti-malarial drug; however, some clinical reports suggest that it can cause dizziness, balance, and vestibular disturbances. To determine if mefloquine might be toxic to the vestibular system, we applied mefloquine to organotypic cultures of the macula of the utricle from postnatal day 3 rats. The macula of the utricle was micro-dissected out as a flat surface preparation and cultured with 10, 50, 100, or 200 µM mefloquine for 24 h. Specimens were stained with TRITC-conjugated phalloidin to label the actin in hair cell stereocilia and TO-PRO-3 to visualize cell nuclei. Some utricles were also labeled with fluorogenic caspase-3, -8, or -9 indicators to evaluate the mechanism of programmed cell death. Mefloquine treatment caused a dose-dependent loss of utricular hair cells. Treatment with 10 µM caused a slight reduction, 50 µM caused a significant reduction, and 200 µM destroyed nearly all the hair cells. Hair cell nuclei in mefloquine-treated utricles were condensed and fragmented, morphological features of apoptosis. Mefloquine-treated utricles were positive for the extrinsic initiator caspase-8 and intrinsic initiator caspase-9 and downstream executioner caspase-3. These results indicate that mefloquine can induce significant hair cell degeneration in the postnatal rat utricle and that mefloquine-induced hair cell death is initiated by both caspase-8 and caspase-9.

An adenylyl cyclase signaling pathway predicts direct dopaminergic input to vestibular hair cells.

Adenylyl cyclase (AC) signaling pathways have been identified in a model hair cell preparation from the trout saccule, for which the hair cell is the only intact cell type. The use of degenerate primers targeting cDNA sequence conserved across AC isoforms, and reverse transcription-polymerase chain reaction (RT-PCR), coupled with cloning of amplification products, indicated expression of AC9, AC7 and AC5/6, with cloning efficiencies of 11:5:2. AC9 and AC5/6 are inhibited by Ca(2+), the former in conjunction with calcineurin, and message for calcineurin has also been identified in the trout saccular hair cell layer. AC7 is independent of Ca(2+). Given the lack of detection of calcium/calmodulin-activated isoforms previously suggested to mediate AC activation in the absence of Gαs in mammalian cochlear hair cells, the issue of hair-cell Gαs mRNA expression was re-examined in the teleost vestibular hair cell model. Two full-length coding sequences were obtained for Gαs/olf in the vestibular type II-like hair cells of the trout saccule. Two messages for Gαi have also been detected in the hair cell layer, one with homology to Gαi1 and the second with homology to Gαi3 of higher vertebrates. Both Gαs/olf protein and Gαi1/Gαi3 protein were immunolocalized to stereocilia and to the base of the hair cell, the latter consistent with sites of efferent input. Although a signaling event coupling to Gαs/olf and Gαi1/Gαi3 in the stereocilia is currently unknown, signaling with Gαs/olf, Gαi3, and AC5/6 at the base of the hair cell would be consistent with transduction pathways activated by dopaminergic efferent input. mRNA for dopamine receptors D1A4 and five forms of dopamine D2 were found to be expressed in the teleost saccular hair cell layer, representing information on vestibular hair cell expression not directly available for higher vertebrates. Dopamine D1A receptor would couple to Gαolf and activation of AC5/6. Co-expression with dopamine D2 receptor, which itself couples to Gαi3 and AC5/6, will down-modulate levels of cAMP, thus fine-tuning and gradating the hair-cell response to dopamine D1A. As predicted by the trout saccular hair cell model, evidence has been obtained for the first time that hair cells of mammalian otolithic vestibular end organs (rat/mouse saccule/utricle) express dopamine D1A and D2L receptors, and each receptor co-localizes with AC5/6, with a marked presence of all three proteins in subcuticular regions of type I vestibular hair cells. A putative efferent, presynaptic source of dopamine was identified in tyrosine hydroxylase-positive nerve fibers which passed from underlying connective tissue to the sensory epithelia, ending on type I and type II vestibular hair cells and on afferent calyces.

Immunohistochemical localization and mRNA expression of aquaporins in the macula utriculi of patients with Meniere's disease and acoustic neuroma.

Meniere's disease is nearly invariably associated with endolymphatic hydrops (the net accumulation of water in the inner ear endolymphatic space). Vestibular maculae utriculi were acquired from patients undergoing surgery for Meniere's disease and acoustic neuroma and from autopsy (subjects with normal hearing and balance). Quantitative immunostaining was conducted with antibodies against aquaporins (AQPs) 1, 4, and 6, Na(+)K(+)ATPase, Na(+)K(+)2Cl co-transporter (NKCC1), and alpha-syntrophin. mRNA was extracted from the surgically acquired utricles from subjects with Meniere's disease and acoustic neuroma to conduct quantitative real-time reverse transcription with polymerase chain reaction for AQP1, AQP4, and AQP6. AQP1 immunoreactivity (-IR) was located in blood vessels and fibrocytes in the underlying stroma, without any apparent alteration in Meniere's specimens when compared with acoustic neuroma and autopsy specimens. AQP4-IR localized to the epithelial basolateral supporting cells in Meniere's disease, acoustic neuroma, and autopsy. In specimens from subjects with Meniere's disease, AQP4-IR was significantly decreased compared with autopsy and acoustic neuroma specimens. AQP6-IR occurred in the sub-apical vestibular supporting cells in acoustic neuroma and autopsy samples. However, in Meniere's disease specimens, AQP6-IR was significantly increased and diffusely redistributed throughout the supporting cell cytoplasm. Na(+)K(+)ATPase, NKCC1, and alpha-syntrophin were expressed within sensory epithelia and were unaltered in Meniere's disease specimens. Expression of AQP1, AQP4, or AQP6 mRNA did not differ in vestibular endorgans from patients with Meniere's disease. Changes in AQP4 (decreased) and AQP6 (increased) expression in Meniere's disease specimens suggest that the supporting cell might be a cellular target.