TRPV4 channels mediate the infrared laser-evoked response in sensory neurons.

Infrared laser irradiation has been established as an appropriate stimulus for primary sensory neurons under conditions where sensory receptor cells are impaired or lost. Yet, development of clinical applications has been impeded by lack of information about the molecular mechanisms underlying the laser-induced neural response. Here, we directly address this question through pharmacological characterization of the biological response evoked by midinfrared irradiation of isolated retinal and vestibular ganglion cells from rodents. Whole cell patch-clamp recordings reveal that both voltage-gated calcium and sodium channels contribute to the laser-evoked neuronal voltage variations (LEVV). In addition, selective blockade of the LEVV by micromolar concentrations of ruthenium red and RN 1734 identifies thermosensitive transient receptor potential vanilloid channels as the primary effectors of the chain reaction triggered by midinfrared laser irradiation. These results have the potential to facilitate greatly the design of future prosthetic devices aimed at restoring neurosensory capacities in disabled patients.

Three-dimensional culture of newborn rat utricle using an extracellular matrix promotes formation of a cyst.

The vestibule is the end organ devoted to sensing of head movements in space. To function properly, its mechano-receptors require the presence of a unique apical extracellular medium, the endolymph. Numerous studies have elucidated the mechanisms involved in the production and homeostasis of this unique medium and the responses of sensory cells to stimulation. However, anatomical constraints have prevented direct and simultaneous studies of their relationships. The aim of this study was the development of an in vitro model that would allow concomitant investigations on maturation and physiological properties of both the hair cells and their endolymphatic compartment. A three-dimensional (3D) culture of newborn rat utricles using an extracellular matrix sustaining 3D cellular growth was developed during 3, 6, or 10 days in vitro (DIV). Using morphological and electrophysiological techniques, we describe the de novo formation of a cyst. It was composed of the sensory epithelium and non-sensory cells-canalar, dark and intermediate cells-that polarized so that their apical surface faced its lumen. During the time of culture, the utricular potential (UP) was steady (-1.1+/-5.0 mV) in oxygenated condition, while in anoxia, the UP significantly decreased to -8.4+/-1.0 mV at 8 DIV. Over the same period, the K+ concentration in the cyst increased up to 86.1+/-33.9 mM (versus 5.6+/-1.5 mM in the bath). These observations indicated that the mechanisms generating the UP and the K-secretory activity were functional at this stage. Concomitantly, the hair cells acquired mature and functional properties: the type 1 and type 2 phenotypes, a mean resting membrane potential of -68.1+/-4.6 mV and typical electrophysiological responses. This preparation provides a powerful means to simultaneous access the hair cells and their endolymphatic compartment, with the possibility to use multi-technical approaches to investigate their interdependent relationships.

Differential expression of Bcl-2-related proteins in differentiating NT2 cells.

Although the role of Bcl-2-related proteins as regulators of the apoptotic process has been well documented, recent studies suggest that they might also be implicated in neuronal differentiation. We have studied by immunocytochemistry, Western blotting and RT-PCR the expression pattern of Bcl-xL, Bcl-2 and BAX in the in vitro model of neuronal differentiation constituted by retinoic acid (RA)-treated NTera-2/D1 (NT2/D1) cells. Whereas BAX level did not change significantly during the RA treatment, Bcl-xL level increased markedly during the first week, before returning to basal level during the second week. Bcl-2 expression, undetectable in undifferentiated cells, increased progressively from the first week. From our results, we suggest that, at least in our model, Bcl-2-related proteins might be involved in neuronal differentiation.