Efferent regulation of hair cells in the turtle cochlea

Intracellular recordings were made from hair cells in the isolated cochlea of the turtle to characterize the inhibition achieved by the cochlea's efferent innervation. A short train of shocks delivered to the efferent axons produced in the hair cells slow hyperpolarizing synaptic potentials which could be reversed by shifting the membrane potential more negative than about -80 mV. Throughout the efferent hyperpolarization, there was a reduction of up to 25-fold in the amplitude of the receptor potential for tones presented at the hair cell's characteristic frequency. Efferent stimulation also was shown to degrade the cell's tuning properties. It is argued that the combined effects of the hyperpolarization and the loss in hair cell sensitivity could account for a threshold elevation of at least 70 dB in the auditory nerve fibres.     

Cell cycle regulation during mouse olfactory neurogenesis.

The development of the nervous system requires a strict control of cell cycle entry and withdrawal. The olfactory epithelium (OE) is noticeable by its ability to yield new neurons not only during development but also continuously during adulthood. The aim of our study was to investigate, by biochemical and immunohistochemical methods, which cell cycle regulators are involved in the control of neuron production during OE development and maturity. At birth, olfactory neural progenitors, the basal cells, exhibited a high mitogenic and neurogenic activity, decreasing in the following weeks together with the drop in expression of several cell cycle regulators. p27Kip1 and p18Ink4c, at birth, were expressed in the whole basal cell layer, whereas p16Ink4a, p19Ink4d, and p21Cip1 were rather located in differentiating or mature neurons. CDK inhibitors may thus act sequentially during this developmental neurogenic process. By comparison, in the adult OE, in which most neural precursors were quiescent, these cells still exhibited p18Ink4c expression but only occasionally p27Kip1 expression. It suggests that p18Ink4c may contribute to maintain basal cells in a quiescent state, whereas p27Kip1 expression in these cells may be rather linked to their neurogenic activity, which declines with age. In keeping with this hypothesis, transgenic mice that lacked p27Kip1 expression displayed a higher rate of cell proliferation versus differentiation in their OE. In these mice, a down-regulation of positive cell cycle regulators was observed that may contribute to compensate for the absence of p27Kip1. Taken together, the present data suggest distinct functions for CDK inhibitors, either in the control of cell cycle exit and differentiation during neurogenesis (respectively, p27Kip1 and p19Ink4d) or in the maintenance of a quiescent state in neural progenitors (p18Ink4c) or neurons (p21Cip1) in adults.     

Cell cycle regulation in mouse heart during embryonic and postnatal stages

The regulation of cardiomyocyte proliferation is important for heart development and function. Proliferation levels of mouse cardiomyocytes are high during early embryogenesis and start to decrease at midgestation. Many cardiomyocytes undergo mitosis without cytokinesis, resulting in binucleated cardiomyocytes during early postnatal stages, following which the cell cycle arrests irreversibly. It remains unknown how the proliferation pattern is regulated, and how the irreversible cell cycle arrest occurs. To clarify the mechanisms, fundamental information about cell cycle regulators in cardiomyocytes and cell cycle patterns during embryonic and postnatal stages is necessary. Here, we show that the expression, complex formation, and activity of main cyclins and cyclin‐dependent kinases (CDKs) changed in a synchronous manner during embryonic and postnatal stages. These levels decreased from midgestation to birth, and then showed one wave in which the peak was around postnatal day 5. Detailed analysis of the complexes suggested that CDK activities were inhibited before the protein levels decreased. Analysis of DNA content distribution patterns in mono‐ and binucleated cardiomyocytes after birth revealed changes in cell cycle distribution patterns and the transition from mono‐ to binucleated cells. These analyses indicated that the wave of cell cycle regulator expression or activities during postnatal stages mainly produced binucleated cells from mononucleated cells. The data obtained should provide a basis for the analysis of cell cycle regulation in cardiomyocytes during embryonic and postnatal stages.     

Emx2 and early hair cell development in the mouse inner ear

Emx2 is a homeodomain protein that plays a critical role in inner ear development. Homozygous null mice die at birth with a range of defects in the CNS, renal system and skeleton. The cochlea is shorter than normal with about 60% fewer auditory hair cells. It appears to lack outer hair cells and some supporting cells are either absent or fail to differentiate. Many of the hair cells differentiate in pairs and although their hair bundles develop normally their planar cell polarity is compromised. Measurements of cell polarity suggest that classic planar cell polarity molecules are not directly influenced by Emx2 and that polarity is compromised by developmental defects in the sensory precursor population or by defects in epithelial cues for cell alignment. Planar cell polarity is normal in the vestibular epithelia although polarity reversal across the striola is absent in both the utricular and saccular maculae. In contrast, cochlear hair cell polarity is disorganized. The expression domain for Bmp4 is expanded and Fgfr1 and Prox1 are expressed in fewer cells in the cochlear sensory epithelium of Emx2 null mice. We conclude that Emx2 regulates early developmental events that balance cell proliferation and differentiation in the sensory precursor population.     

Genetic mouse models to investigate cell cycle regulation

Early studies on cell cycle regulation were based on experiments in model systems (Yeast, Xenopus, Starfish, Drosophila) and have shaped the way we understand many events that control the cell cycle. Although these model systems are of great value, the last decade was highlighted by studies done in human cells and using in vivo mouse models. Mouse models are irreplaceable tools for understanding the genetics, development, and survival strategies of mammals. New developments in generating targeting vectors and mutant mice have improved our approaches to study cell cycle regulation and cancer. Here we summarize the most recent advances of mouse model approaches in dissecting the mechanisms of cell cycle regulation and the relevance to human disease. W. Li and S. Kotoshiba contributed equally.     

Cell cycle regulation

Drosophila researchers met in sunny San Diego for the 49(th) Annual Meeting of The Genetics Society of America. It was cold outside and even colder inside. Like last year, 'Mitosis, Meiosis and Cell Division' was no longer a session. Instead, we searched out and covered talks and posters in 'Cell Division and Growth Control', 'Gametogenesis', 'Cytoskeleton and Cell Biology' and 'Genome and Chromosome Structure'. We split up for maximal coverage and re-grouped later for the Workshop on Cell Cycle and Checkpoints. We apologize in advance for the brevity or omission of some reports.     

Cell cycle and cell fate interactions in neural development

Mechanisms coupling cell cycle and cell fate operate at different steps during neural development. Intrinsic factors control the cell proliferation of distinct brain regions and changes of cell fate competence, whereas components of the cell cycle machinery could play a major role in setting the appropriate timing of the generation of different cell types.     

虎皮鹦鹉耳蜗感觉细胞的发育与听觉的关系

选用18d、38d、成鸟(3月以上)三种年龄的雄性虎皮鹦鹉作为实验材料,采用测量脑干听觉诱发电位和制作内耳的石蜡切片两种方法,研究了鸟类在发育时期耳蜗感觉上皮细胞的变化以及听觉功能的发育状况。鸟的听觉能力在出生后不断得到提高,38d基本达到成鸟的水平;耳蜗毛细胞的形态、内部结构逐渐趋于成熟,其灵敏度不断提高,感受听觉的能力也在增强,38d基本与成鸟的发育程度相同。耳蜗感觉上皮细胞的发育对听觉行为产生的时间和发展具有极为重要的影响。     

Coupling the cell cycle to development and regeneration of the inner ear

Cell cycle exit and acquirement of a postmitotic state is essential for the proper development of organs. In the present review, we examine the role of the cell cycle control in the sensory epithelia of the mammalian inner ear. We describe the roles of the core cell cycle regulators in the proliferation of prosensory cells and in the initiation and maintenance of terminal mitosis of the sensory epithelia. We also discuss how other intracellular signalling may influence the cell cycle. Finally, we address the question of whether manipulations of the cell cycle may have the potential to create replacement cells for the damaged inner sensory epithelia.     

Force generation in the outer hair cell of the cochlea.

The outer hair cell of the mammalian cochlea has a unique motility directly dependent on the membrane potential. Examination of the force generated by the cell is an important step in clarifying the detailed mechanism as well as the biological importance of this motility. We performed a series of experiments to measure force in which an elastic probe was attached to the cell near the cuticular plate and the cell was driven with voltage pulses delivered from a patch pipette under whole-cell voltage clamp. The axial stiffness was also determined with the same cell by stretching it with the patch pipette. The isometric force generated by the cell is around 0.1 nN/mV, somewhat smaller than 0.15 nN/mV, predicted by an area motor model based on mechanical isotropy, but larger than in earlier reports in which the membrane potential was not controlled. The axial stiffness obtained, however, was, on average, 510 nN per unit strain, about half of the value expected from the mechanical isotropy of the membrane. We extended the area motor theory incorporating mechanical orthotropy to accommodate the axial stiffness determined. The force expected from the orthotropic model was within experimental uncertainties.     

Pocket proteins and cell cycle regulation in inner ear development

Loss of neurosensory cells of the ear, caused by genetic and non-genetic factors, is becoming an increasing problem as people age, resulting in deafness and vestibular disorders. Unveiling useful mechanisms of cell cycle regulation may offer the possibility to generate new cells out of remaining ones, thus providing the cellular basis to induce new hair cell differentiation in the mammalian ear. Here, we provide an overview of cell cycle regulating genes in general and of those studied in the ear in particular. We categorize those genes into regulators that act upstream of the pocket proteins and into those that act downstream of the pocket proteins. The three members of the pocket protein family essentially determine, through interaction with the eight members of the E2F family, whether or not the cell cycle will progress to the S-phase and thus cell division. The abundant presence of one or more members of these families in adult hair cells supports the notion that inhibition of cell cycle progression through these proteins is a lifelong process. Indeed, manipulating some of those proteins, unfortunately, leads to abortive entry into the cell cycle. Combined with recent success to induce hair cell differentiation through molecular therapy, these approaches may provide a viable strategy to restore lost hair cells in the inner ear.     

Analysis of the fifth cell cycle of mouse development

The 5th cell cycle of mouse development was analyzed to determine the lengths of each cell cycle phase. The DNA content of Feulgen-stained blastomere nuclei was measured at various times throughout the cell cycle by microdensitometry. To achieve precise timing of the start of the 5th cell cycle, experiments utilized isolated 16-cell blastomeres and cell pairs obtained by in-vitro division of isolated 8-cell blastomeres. The following estimates were made for a mixed population of polar and apolar 16-cell blastomeres: G1, less than or equal to 2 h; S, 8-9 h; G2 + M, 2 h. No significant difference was found in the timing of DNA synthesis between polar and apolar cells or between cell pairs and whole embryos.     

Cell size, cell cycle and transition probability in mouse fibroblasts

This paper describes the relationship between cell size and cell division in two situations. In the first, quiescent cells were sorted on the basis of cell size using a fluorescence-activated cell sorter and returned to culture. The results of this type of experiment are compatible with the idea that once cells have completed a size-dependent lag, the rate of entry of cells into S phase is controlled by a rate-limiting random event (or transition).The second kind of experiment follows the kinetics of complete cell cycles in rapidly proliferating cells whose mothers had been sorted on the basis of cell size. The cells born of small mother cells have longer cycle times than cells derived from large mothers. The difference in the cycle time of these two classes was due to differences in the B phase of the cell cycle [containing S, G2, M and part of G1 (G1B)], transition probability being the same in both size classes. Our results show that S, G2 and M are unaffected by size, thus confining the effect of size to G1B. It seems probable that the variability of B phase in cloned cell populations is partly due to variations of cell size at division, and correlations between the cycle times of sister cells result because sibling cells are more similar in size than unrelated cells. The major factor controlling cell division in mouse fibroblasts is shown, however, to be the transition probability; size has a more minor role.     

FGF信号通路在内耳发育调控和毛细胞再生中的作用

内耳是感受听觉和平衡觉的复杂器官。在内耳发育过程中,成纤维生长因子(fibroblast growth factor, FGF)信号通路参与了听基板的诱导、螺旋神经节(statoacoustic ganglion, SAG)的发育以及Corti器感觉上皮的分化。FGF信号开启了内耳早期发育的基因调控网络,诱导前基板区域以及听基板的形成。正常表达的FGF信号分子可促进听囊腹侧成神经细胞的特化,但成熟SAG神经元释放的过量FGF5可抑制此过程,形成负反馈环路使SAG在稳定状态下发育。FGF20在Notch信号通路的调控下参与了前感觉上皮区域向毛细胞和支持细胞的分化过程,而内毛细胞分泌的FGF8可调控局部支持细胞分化为柱细胞。人类FGF信号通路异常可导致多种耳聋相关遗传病。此外,FGF信号通路在低等脊椎动物毛细胞自发再生以及干细胞向内耳毛细胞诱导过程中都起到了关键作用。本文综述了FGF信号通路在内耳发育调控以及毛细胞再生中的作用及其相关研究进展,以期为毛细胞再生中FGF信号通路调控机制的阐明奠定理论基础。