Lac z Histochemistry and Immunohistochemistry Reveal Ephrin-B Ligand Expression in the Inner Ear

Bianchi, Lynne M.; Dinsio, Kyl J.; Davoli, Katherine; Gale, Nicholas W.

doi:10.1177/002215540205001208

Cited by 10 publications

(9 citation statements)

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“…Cochleae were decalcified in 5% EDTA for three days at 4°C, washed in rinse buffer (0.01% sodium deoxycholate, 0.02% Nonidet P-40, 2 mM magnesium chloride in PBS) for 1 hour and stained in 1 mg/ml X-gal/5 mM potassium ferrocyanide/5 mM potassium ferricyanide in rinse buffer overnight at 37°C (Bianchi, et al, 2002; Corradi, et al, 2003). Cochleae were dissected for whole-mount preparation or processed further for sections; Cochleae were cryoprotected in 20% sucrose overnight at 4°C before embedding in Tissue-Tek O.C.T.…”

Section: Methodsmentioning

confidence: 99%

Localization of kainate receptors in inner and outer hair cell synapses

et al. 2014

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Glutamate plays a role in hair cell afferent transmission, but the receptors that mediate neurotransmission between outer hair cells (OHCs) and type II ganglion neurons are not well defined. A previous study using in situ hybridization showed that several kainate-type glutamate receptor (KAR) subunits are expressed in cochlear ganglion neurons. To determine whether KARs are expressed in hair cell synapses, we performed X-gal staining on mice expressing lacZ driven by the GluK5 promoter, and immunolabeling of glutamate receptors in whole-mount mammalian cochleae. X-gal staining revealed GluK5 expression in both type I and type II ganglion neurons and OHCs in adults. OHCs showed X-gal reactivity throughout maturation from postnatal day 4 (P4) to 1.5 months. Immunoreactivity for GluK5 in IHC afferent synapses appeared to be postsynaptic, similar to GluA2 (GluR2; AMPA-type glutamate receptor (AMPAR) subunit), while GluK2 may be on both sides of the synapses. In OHC afferent synapses, immunoreactivity for GluK2 and GluK5 was found, although GluK2 was only in those synapses bearing ribbons. GluA2 was not detected in adult OHC afferent synapses. Interestingly, GluK1, GluK2 and GluK5 were also detected in OHC efferent synapses, forming several active zones in each synaptic area. At P8, GluA2 and all KAR subunits except GluK4 were detected in OHC afferent synapses in the apical turn, and GluA2, GluK1, GluK3 decreased dramatically in the basal turn. These results indicate that AMPARs and KARs (GluK2/GluK5) are localized to IHC afferent synapses, while only KARs (GluK2/GluK5) are localized to OHC afferent synapses in adults. Glutamate spillover near OHCs may act on KARs in OHC efferent terminals to modulate transmission of acoustic information and OHC electromotility.

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Section: Methodsmentioning

confidence: 99%

Localization of kainate receptors in inner and outer hair cell synapses

et al. 2014

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“…Whole-mount staining for ␤-Gal in LYVE-1 ϩ/Ϫ and LYVE-1 Ϫ/Ϫ mice and tumor cryostat sections was performed as described previously (2). For whole-mount preparations of dermal lymphatics, ear tissues were fixed overnight in 4% paraformaldehyde in phosphate-buffered saline (PBS), and the dorsal segment was blocked for 3 h in 30 mg/ml milk powder-10% fetal bovine serum (FBS) in PBS-0.1% (wt/vol) Triton X-100 prior to double staining overnight with rat anti-mouse CD31 (MEC 13.3)/ Alexa594-conjugated goat anti-rat and hamster anti-mouse podoplanin (8.1.1)/ Alexa488-conjugated goat anti-hamster antibodies.…”

Section: Methodsmentioning

confidence: 99%

Normal Lymphatic Development and Function in Mice Deficient for the Lymphatic Hyaluronan Receptor LYVE-1

Gale

Prevo

Espinosa

et al. 2007

Molecular and Cellular Biology

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187

143

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The hyaluronan receptor LYVE-1 is expressed abundantly on the surfaces of lymphatic vessels and lymph node sinus endothelial cells from early development, where it has been suggested to function both in cell adhesion/transmigration and as a scavenger for hyaluronan turnover. To investigate the physiological role(s) of LYVE-1, we generated mice in which the gene for the receptor was inactivated by replacement with a ␤-galactosidase reporter. LYVE-1 ؊/؊ mice displayed an apparently normal phenotype, with no obvious alteration in lymphatic vessel ultrastructure or function and no apparent change in secondary lymphoid tissue structure or cellularity. In addition, the levels of hyaluronan in tissue and blood were unchanged. LYVE-1 ؊/؊ mice also displayed normal trafficking of cutaneous CD11c؉ dendritic cells to draining lymph nodes via afferent lymphatics and normal resolution of oxazolone-induced skin inflammation. Finally, LYVE-1 ؊/؊ mice supported normal growth of transplanted B16F10 melanomas and Lewis lung carcinomas. These results indicate that LYVE-1 is not obligatory for normal lymphatic development and function and suggest either the existence of compensatory receptors or a role more specific than that previously envisaged.

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“…Eph proteins, including the Eph receptors and their ligands, the ephrins, are broadly expressed in the developing auditory system (for reviews see Bianchi et al, 2002, Pickles, 2003, Cramer, 2005, Gabriele et al, 2011). They provide multiple mechanisms for signaling and for generating precise connectivity.…”

Section: Eph Receptors and Ephrinsmentioning

confidence: 99%

“…Several studies have demonstrated that Eph receptors and ephrins are expressed in the developing inner ear in mice (Henkemeyer et al, 1994, Bianchi and Gale, 1998, Bianchi et al, 2002, Pickles et al, 2002, Zhou et al, 2011, Coate et al, 2012, Defourny et al, 2013). SGNs express ephrin-B1 and ephrin-B2, and growth of SGN processes is inhibited in vitro by EphB2 through reverse signaling (Bianchi and Gray, 2002, Zhou et al, 2011).…”

Section: Peripheral Auditory Pathwaysmentioning

confidence: 99%

Axon guidance in the auditory system: Multiple functions of Eph receptors

Cramer

Gabriele

2014

Neuroscience

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The neural pathways of the auditory system underlie our ability to detect sounds and to transform amplitude and frequency information into rich and meaningful perception. While it shares some organizational features with other sensory systems, the auditory system has some unique functions that impose special demands on precision in circuit assembly. In particular, the cochlear epithelium creates a frequency map rather than a space map, and specialized pathways extract information on interaural time and intensity differences to permit sound source localization. The assembly of auditory circuitry requires the coordinated function of multiple molecular cues. Eph receptors and their ephrin ligands constitute a large family of axon guidance molecules with developmentally regulated expression throughout the auditory system. Functional studies of Eph/ephrin signaling have revealed important roles at multiple levels of the auditory pathway, from the cochlea to the auditory cortex. These proteins provide graded cues used in establishing tonotopically ordered connections between auditory areas, as well as discrete cues that enable axons to form connections with appropriate postsynaptic partners within a target area. Throughout the auditory system, Eph proteins help to establish patterning in neural pathways during early development. This early targeting, which is further refined with neuronal activity, establishes the precision needed for auditory perception.

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Lac z Histochemistry and Immunohistochemistry Reveal Ephrin-B Ligand Expression in the Inner Ear

Cited by 10 publications

References 10 publications

Localization of kainate receptors in inner and outer hair cell synapses

Localization of kainate receptors in inner and outer hair cell synapses

Normal Lymphatic Development and Function in Mice Deficient for the Lymphatic Hyaluronan Receptor LYVE-1

Axon guidance in the auditory system: Multiple functions of Eph receptors

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