Marie-Thérèse Nicolas

KCNQ1/KCNE1 potassium channels in mammalian vestibular dark cells.

The high [K(+)] in the inner ear endolymph is essential for mechanosensory transduction in hearing and balance. Several ion channels, including a slowly activating, voltage-dependent, outwardly conducting K(+) channel composed of the KCNQ1 (KvLQT1) and KCNE1 (IsK/minK) subunits, are expressed at the apical surface of vestibular dark cells. We investigated the underlying molecular mechanisms of this conductance using in situ hybridization, RT-PCR, and immunocytochemistry and by tracking the ultrastructural changes of vestibular structures in kcne1(-/-) mice. In the wild type mice, the KCNE1 and KCNQ1 proteins are expressed specifically at the apical membrane of dark cells, as early as gestational day (GD) 17 for KCNE1 while KCNQ1 mRNAs can be detected at GD 18. This is the first demonstration that the two protein components of this potassium channel co-localize in a polarized fashion at the cellular level. Although the vestibular end-organs are normal at birth in kcne1(-/-) mice, they begin to show modifications during postnatal development: we observed an increase in the height of the dark cells, in their number of mitochondria, and in basolateral membrane infoldings. Subsequently, the epithelium degenerates and the endolymphatic space collapses. Similar changes are known to occur in the cardio-auditory Jervell--Lange-Nielsen syndrome which is caused by mutations in the same channel.

Changes in D-serine levels and localization during postnatal development of the rat vestibular nuclei

The patterns of development of the vestibular nuclei (VN) and their main connections involving glutamate neurotransmission offer a good model for studying the function of the glial‐derived neuromodulator D‐serine in synaptic plasticity. In this study we show that D‐serine is present in the VN and we analyzed its distribution and the levels of expression of serine racemase and D‐amino acid oxidase (D‐AAO) at different stages of postnatal (P) development. From birth to P21, high levels of D‐serine were detected in glial cells and processes in all parts of the VN. This period corresponded to high expression of serine racemase and low expression of D‐AAO. On the other hand, in the mature VN D‐serine displayed very low levels and was mainly localized in neuronal cell bodies and dendrites. This drop of D‐serine in adult stages corresponded to an increasing expression of D‐AAO at mature stages. High levels of glial D‐serine during the first 3 weeks of postnatal development correspond to an intense period of plasticity and synaptogenesis and maturation of VN afferents, suggesting that D‐serine could be involved in these phenomena. These results demonstrate for the first time that changes in D‐serine levels and distribution occur during postnatal development in the central nervous system. The strong decrease of D‐serine levels and the glial‐to‐neuronal switch suggests that D‐serine may have distinct functional roles depending on the developmental stage of the vestibular network. J. Comp. Neurol. 497:610–621, 2006.

Localization of TREK-1, a two-pore-domain K+ channel in the peripheral vestibular system of mouse and rat.

The distribution of two-pore-domain (2P-domain) K(+) channels of the TREK subfamily was studied using immunocytochemistry in the peripheral vestibular system of mouse and rat. Using RT-PCR, the mRNA for TREK-1, but not for TREK-2 or TRAAK, were detected in mouse vestibular endorgans and ganglia. The TREK-1 channel protein was immunodetected in both nerve fibers and nerve cell bodies in the vestibular ganglion, both afferent fibers and nerve calyces innervating type I hair cells in the utricle and cristae. The post-synaptic localization in afferent calyces may suggest a neuroprotective role in glutamatergic excitotoxicity during ischemic conditions. In non-neuronal cells, TREK-1 was immunodetected in the apical membrane of dark cells and transitional cells, both of which are involved in endolymph K(+) secretion and recycling. TREK-1 may subserve some neuroprotective function in afferent nerve fibers as well as play a role in endolymph potassium homeostasis.

Cellular distribution of D-serine, serine racemase and d-amino acid oxidase in the rat vestibular sensory epithelia

Glutamate is the main neurotransmitter at the synapses between sensory cells and primary afferents in the peripheral vestibular system. Evidence has recently been obtained demonstrating that the atypical amino acid D-serine is the main endogenous co-agonist of the N-methyl-D-aspartate receptors in the CNS. We studied the distribution of D-serine and its synthesizing and degrading enzymes, serine racemase and d-amino acid oxidase in the rat vestibular sensory epithelium using immunocytochemistry. D-serine, serine racemase and D-amino acid oxidase were localized in the transitional cells, which are parasensory cells located between the sensory epithelium and the dark cells. The dark cells expressed only serine racemase. D-Serine was also detected in the supporting cells of the sensory epithelium. These cells, which are in close contact with glutamatergic synapses, express GLAST, a glial specific transporter for glutamate. They may have similar functions to glial cells in the CNS and thus expression of D-serine suggests a neuromodulator role for D-serine at the glutamatergic synapses in the peripheral vestibular system. Our data also indicate that the metabolism of D-serine is not restricted to glial cells suggesting that the amino acid may play an additional role in the peripheral nervous system.

VESTIBULAR SEMICIRCULAR CANAL EPITHELIUM OF THE RAT IN CULTURE ON FILTER SUPPORT : POLARITY AND BARRIER PROPERTIES

Abstract The inner ear of mammals contains the vestibular apparatus which is involved in the maintenance of posture and balance. The tubular structure of the apparatus is bathed by the potassium-rich endolymph and sodium-rich perilymph in the luminal and abluminal compartments, respectively. The luminal compartment is lined by a continuous epithelium with islets of receptor organs, which separates the luminal from the abluminal compartment. The present work focuses on the epithelium, without the receptor organs, and shows that it can be reconstituted in culture. The epithelium from 4-day-old Wistar rats was grown on microporous membranes. High transepithelial electrical resistances (4000–6000 Ω·cm2)were achieved after 4–8 days in culture. The epithelium was characterized by the presence of cytokeratin, ZO-1 protein, occludin, and the presence of tight junctions and kinocilia. The transepithelial resistance of the cell monolayer withstood endolymph/perilymph dual bathing when the apical pole of the cells was in contact with endolymph, but collapsed in the reverse configuration. Weak but statistically highly significant basal to apical rubidium (86Rb) transport was observed. These findings show that this epithelium maintains its in vivo polarity and could enhance the potassium composition of endolymph up to maturity. This new culture model, in which dual bathing is possible, should enable further in vitro studies of the sensory vestibular epithelia.

Prix Nobel de Chimie 2008 (Osumo Shimomura, Martin Chalfie et Roger Y. Tsien) : Osamu Shimomura

Le prix Nobel de Chimie 2008 a ete attribue a trois chercheurs, Osamu Shimomura, Roger Tsien et Martin Chalfie, pour leurs travaux sur une proteine fluorescente la GFP (Green Fluorescent Protein) extraite d’une meduse. Il n’est plus besoin de presenter cette proteine et ses utilisations. Dans cet article, nous rendons hommage au premier d’entre eux : Osamu Shimomura qui est a l’origine et represente le socle de tous ces travaux. O. Shimomura a apporte une contribution majeure dans le domaine de la biologie cellulaire avec la GFP (la vedette de ce Nobel), mais aussi dans le domaine de la chimie de la bioluminescence et surtout ne l’oublions pas dans la mesure du calcium avec l’aequorine. Il faut aussi souligner l’exemplarite de son parcours de recherche devoue exclusivement a la recherche fondamentale, a une epoque ou la rentabilite, la rapidite et la production en masse deviennent un critere « d’excellence ». Le tout allie a une rigueur implacable, une honnetete et une ethique sans faille.