Time-dependence and cell-type specificity of synergistic neurotrophin actions on spiral ganglion neurons.

The neurotrophins brain-derived neurotrophin (BDNF) and neurotrophin-3 (NT-3) synergistically enhance survival of spiral ganglion neurons such that simultaneous exposure to both compounds produces a larger response than would be expected from their individual effects. To elucidate the functional role of this neurotrophin interaction, we examined its temporal and cell-type specificity in vitro for both mouse and gerbil spiral ganglion neurons. Synergistic effects were transient; they were maximal within the first two postnatal days and declined during the first postnatal week. Both neurotrophins were, however, still efficacious at increasing cell survival. After postnatal day 10, the effects of coexposure to BDNF and NT-3 were additive rather than synergistic. Synergism declined more rapidly in mouse than gerbil neurons, reflecting the difference in cochlear development for each species. Only neurons without peripherin epitopes, putative type I neurons, showed synergistic survival effects; survival of peripherin-expressing neurons was purely additive. Therefore, during a restricted time period, identical neurotrophin stimuli are capable of preferentially enhancing survival of one class of neurons that compose approximately 95% of the adult spiral ganglion.

Differentiation of cyst-forming stria vascularis tissues in vitro.

The marginal cells of the stria vascularis possess distinctive morphological characteristics associated with their role in endolymph production. Interestingly, when stria-derived epithelial cells are grown in association with the underlying mesenchyme, the final differentiation of these cell types does not occur. Beyond the rudimentary polarity that is established, similar to that shown in epithelial monolayers, cells in culture bear only a slight resemblance to their marginal cell counterparts in vivo. The ultrastructural features that typify these epithelia, extensive cytoplasmic invaginations, with an abundance of mitochondria, and darkly stained cytoplasm, are not evident under standard culture conditions. In order to determine whether fluid transport, a key function of the stria vascularis, has an effect on the ultrastructural morphology, we examined de novo stria vascularis tissues that formed a fluid-filled cyst in vitro. We found that only cells associated with the luminal structure demonstrated dark cytoplasmic staining and amplification of the basolateral membrane of the marginal cells. Additionally, other epithelial features, such as mitochondria-rich and microvilli-rich cells, were observed in cyst-forming tissues. The enhancement of the marginal cell specializations was not as robust as that observed in vivo; however, they were clearly more extensive when compared to cells in the same culture that were not associated with a fluid-filled lumen. Thus it appears that fluid transport may be necessary to maximize differentiation of stria vascularis tissues in vitro.

Stria vascularis morphogenesis in vitro.

Explants of neonatal murine stria vascularis were maintained in vitro to evaluate the process of morphogenesis in cochlear tissue. Immunohistochemical and electron microscopic studies showed that the relatively undifferentiated cells in culture attained morphological features characteristic of the stria vascularis cell types in vivo (marginal, intermediate and basal cells). The three kinds of cells formed a trilaminated tissue, with the epithelial cells bordering the culture medium, basal-like cells resting on the culture substrate, and the melanocytes layered between. Furthermore, approximately 20% of these cultures displayed a unique alignment of melanocytes which formed elongated bands along the contour of the tissue edge. However, only limited cell extensions were formed between different cell types and interdigitation amongst these processes was abbreviated. Thus, cells from different embryological origins divided, migrated and reestablished appropriate cell-to-cell associations to form a layered tissue similar to the stria vascularis in vivo.