Mammalian cochlea undergoes morphological and functional changes during the postnatal period, around the hearing onset. Major changes during the initial 2 postnatal weeks of mouse include maturation of sensory hair cells and supporting cells, and acquisition of afferent and efferent innervations. During this period, supporting cells in the greater epithelial ridge (GER) of the cochlea exhibit spontaneous and periodic activities which involves ATP, increase in intracellular Ca2+, and cell volume change. This Ca2+-dependent volume change has been proposed to involve chloride channels or transporters. We found that the spontaneous volume changes were eliminated by anion channel blocker, 100 microM NPPB. Among candidates, expression of Anoctamin-1 (Ano1 or TMEM16A), bestriphin-1 and NKCC1 were investigated in whole-mount cochlea of P9-10 mice. Immunolabeling indicated high level of Ano1 expression in the GER, but not of betrophin-1 or NKCC1. Double-labeling with calretinin and confocal image analysis further elucidated the cellular localization of Ano1 immunoreactivity in supporting cells. It was tested if the Ano1 expression exhibits similar time course to the spontaneous activities in postnatal cochlear supporting cells. Cochlear preparations from P2-3, P5-6, P9-10, P15-16 mice were subjected to immunolabeling. High level of Ano1 immunoreactivity was observed in the GER of P2-3, P5-6, P9-10 cochleae, but not of P15-17 cochleae. Taken together, the localization and time course in Ano1 expression pattern correlates with the spontaneous, periodic volume changes recorded in postnatal cochlear supporting cells. From these results we propose that Ano1 is the pacemaker of spontaneous activities in postnatal cochlea.
Adenosine Triphosphate ; Animals ; Calbindin 2 ; Cell Size ; Chloride Channels ; Cochlea* ; Hair ; Hearing ; Mice*

PURPOSE: The smoking rates among teenagers in Korea grow gradually since 1980s despite of the numerous programs for youth smoking prevention and cessation have been developed and implemented. The purpose of this study was to explore and describe the processes of habituating to smoking among teenagers using the grounded theory methodology. METHOD: Qualitative data was collected via six focus group interviews. A total of 38 people, twelve teachers and 24 middle school students participated in this study. All focus group interviews were tape-recorded, transcribed, and analyzed according to the grounded theory methodology. RESULTS: The overriding theme of the elicited grounded theory was "stepping into a quagmire by a merest chance". The student participants began smoking by a simple chance. The contingent factors to starting smoking were "discord within the family", "family member's smoking", "schoolwork stresses", or "a rebellious spirit". The conditions of smoking included accessibility, going around in group, and the lack of discipline. "Stigmatizing", "involved in mob violence", and "making a poor academic record" coexisted as the covariance of the smoking habituation. CONCLUSION: The findings of this study illustrated the comprehensive and insightful picture of the phenomena under investigation. Nursing implications and further directions for research were discussed.
Adolescent* ; Focus Groups ; Humans ; Korea ; Nursing ; Smoke* ; Smoking* ; Tobacco Products

The vast majority of malignant tumors involving the vagina are secondary spread from primary malignant lesion of the cervix uteri, the sigmoid colon, the bladder and the vulva. Primary invasive carcinoma of the vagina remains among the rare gynecologic malignant tumor. The diagnosis of primary carcinoma of the vagina requires that the cervix and the vulva be intact and no clinical evidence of other primary tumors exist. Greater than 80-90% of all vaginal tumors are squamous cell type. We experienced two cases of primary vaginal cancer of 68 years old woman without any other gynecologic disease and 67 years old woman after hysterectomy for benign desease. We presented these cases with a brief review of related literatures.
Aged ; Cervix Uteri ; Colon, Sigmoid ; Diagnosis ; Female ; Genital Diseases, Female ; Humans ; Hysterectomy ; Neoplasms, Squamous Cell ; Urinary Bladder ; Vagina ; Vaginal Neoplasms* ; Vulva

The synaptic contacts of cochlear afferent fibers (CAFs) with inner hair cells (IHCs) are spatially segregated according to their firing properties. CAFs also exhibit spatially segregated vulnerabilities to noise. The CAF fibers contacting the modiolar side of IHCs tend to be more vulnerable. Noise vulnerability is thought to be due to the absence of neuroprotective mechanisms in the modiolar side contacting CAFs. In this study, we investigated whether the expression of neuroprotective Ca²⁺-buffering proteins is spatially segregated in CAFs. The expression patterns of calretinin, parvalbumin, and calbindin were examined in rat CAFs using immunolabeling. Calretinin-rich fibers, which made up ~50% of the neurofilament (NF)-positive fibers, took the pillar side course and contacted all IHC sides. NF-positive and calretinin-poor fibers took the modiolar side pathway and contacted the modiolar side of IHCs. Both fiber categories juxtaposed the C-terminal binding protein 2 (CtBP2) puncta and were contacted by synaptophysin puncta. These results indicated that the calretinin-poor fibers, like the calretinin-rich ones, were afferent fibers and probably formed functional efferent synapses. However, the other Ca²⁺-buffering proteins did not exhibit CAF subgroup specificity. Most CAFs near IHCs were parvalbumin-positive. Only the pillar-side half of parvalbumin-positive fibers coexpressed calretinin. Calbindin was not detected in any nerve fibers near IHCs. Taken together, of the Ca²⁺-buffering proteins examined, only calretinin exhibited spatial segregation at IHC-CAF synapses. The absence of calretinin in modiolar-side CAFs might be related to the noise vulnerability of the fibers.
Animals ; Calbindin 2* ; Calbindins ; Carrier Proteins ; Fires ; Hair Cells, Auditory, Inner ; Intermediate Filaments ; Nerve Fibers ; Noise* ; Rats ; Sensitivity and Specificity ; Synapses ; Synaptophysin

The TMEM43 has been studied in human diseases such as arrhythmogenic right ventricular cardiomyopathy type 5 (ARVC5) and auditory neuropathy spectrum disorder (ANSD). In the heart, the p.(Ser358Leu) mutation has been shown to alter intercalated disc protein function and disturb beating rhythms. In the cochlea, the p.(Arg372Ter) mutation has been shown to disrupt connexin-linked function in glia-like supporting cells (GLSs), which maintain inner ear homeostasis for hearing. The TMEM43-p.(Arg372Ter) mutant knock-in mice displayed a significantly reduced passive conductance current in the cochlear GLSs, raising a possibility that TMEM43 is essential for mediating the passive conductance current in GLSs. In the brain, the two-pore-domain potassium (K2P) channels are generally known as the “leak channels” to mediate background conductance current, raising another possibility that K2P channels might contribute to the passive conductance current in GLSs. However, the possible association between TMEM43 and K2P channels has not been investigated yet. In this study, we examined whether TMEM43 physically interacts with one of the K2P channels in the cochlea, KCNK3 (TASK-1). Utilizing co-immunoprecipitation (IP) assay and Duolink proximity ligation assay (PLA), we revealed that TMEM43 and TASK-1 proteins could directly interact. Genetic modifications further delineated that the intracellular loop domain of TMEM43 is responsible for TASK-1 binding. In the end, gene-silencing of Task-1 resulted in significantly reduced passive conductance current in GLSs. Together, our findings demonstrate that TMEM43 and TASK-1 form a protein-protein interaction in the cochlea and provide the possibility that TASK-1 is a potential contributor to the passive conductance current in GLSs.

Cochlear afferent nerve fibers (ANF) are the first neurons in the ascending auditory pathway. We investigated the low-voltage activating K + channels expressed in ANF dendrites using isolated rat cochlear segments. Whole cell patch clamp recordings were made from the dendritic terminals of ANFs. Outward currents activating at membrane potentials as low as -64 mV were observed in all dendrites studied. These currents were inhibited by 4-aminopyridine (4-AP), a blocker known to preferentially inhibit low-voltage activating K + currents (I KL ) in CNS auditory neurons and spiral ganglion neurons. When the dendritic I KL was blocked by 4-AP, the EPSP decay time was significantly prolonged, suggesting that dendritic I KL speeds up the decay of EPSPs and likely modulates action potentials of ANFs. To reveal molecular subtype of dendritic I KL , α-dendrotoxin (α-DTX), a selective inhibitor for K v 1.1, Kv 1.2, and K v 1.6 containing channels, was tested. α-DTX inhibited 23±9% of dendritic I KL . To identify the α-DTXsensitive and α-DTX-insensitive components of I KL , immunofluorescence labeling was performed. Strong K v 1.1- and K v 1.2-immunoreactivity was found at unmyelinated dendritic segments, nodes of Ranvier, and cell bodies of most ANFs. A small fraction of ANF dendrites showed K v 7.2-immunoreactivity. These data suggest that dendritic I KL is conducted through K v 1.1and K v 1.2 channels, with a minor contribution from K v 7.2 and other as yet unidentified channels.

In the auditory system, the spontaneous activity of cochlear inner hair cells (IHCs) is initiated by the release of ATP from inner supporting cells (ISCs). This ATP release sets off a cascade, activating purinergic autoreceptors, opening of Ca 2+ -activated Cl- channel TMEM16A, Cl- efflux and osmotic cell shrinkage. Then, the shrunken ISCs efficiently regain their original volume, suggesting the existence of mechanisms for refilling Cl- and K + , priming them for subsequent activity. This study explores the potential involvement of NKCCs (Na+ -K+ -Cl- cotransporters) and KCCs (K+ -Cl- cotransporters) in ISC spontaneous activity, considering their capability to transport both Cl- and K+ ions across the cell membrane. Employing a combination of immunohistochemistry, pharmacological interventions, and shRNA experiment, we unveiled the pivotal role of NKCC1 in cochlear spontaneous activity. Immunohistochemistry revealed robust NKCC1 expression in ISCs, persisting until the 2nd postnatal week. Intriguingly, we observed a developmental shift in NKCC1 expression from ISCs to synaptophysin-positive efferent terminals at postnatal day 18, hinting at its potential involvement in modulating synaptic transmission during the post-hearing period. Experiments using bumetanide, a well-known NKCC inhibitor, supported the functional significance of NKCC1 in ISC spontaneous activity. Bumetanide significantly reduced the frequency of spontaneous extracellular potentials (sEP) and spontaneous optical changes (sOCs) in ISCs. NKCC1-shRNA experiments conducted in cultured cochlear tissues further supported these findings, demonstrating a substantial decrease in event frequency and area. Taken together, we revealed the role of NKCC1 in shaping the ISC spontaneous activity that govern auditory pathway development.

No abstract available.
Humans ; Prone Position* ; Respiratory Mechanics*

No abstract available.
Humans ; Prone Position* ; Respiratory Mechanics*