Konstantina M. Stankovic

Survival of adult spiral ganglion neurons requires erbB receptor signaling in the inner ear

Degeneration of cochlear sensory neurons is an important cause of hearing loss, but the mechanisms that maintain the survival of adult cochlear sensory neurons are not clearly defined. We now provide evidence implicating the neuregulin (NRG)-erbB receptor signaling pathway in this process. We found that NRG1 is expressed by spiral ganglion neurons (SGNs), whereas erbB2 and erbB3 are expressed by supporting cells of the organ of Corti, suggesting that these molecules mediate interactions between these cells. Transgenic mice in which erbB signaling in adult supporting cells is disrupted by expression of a dominant-negative erbB receptor show severe hearing loss and 80% postnatal loss of type-I SGNs without concomitant loss of the sensory cells that they contact. Quantitative RT-PCR analysis of neurotrophic factor expression shows a specific downregulation in expression of neurotrophin-3 (NT3) in the transgenic cochleas before the onset of neuronal death. Because NT3 is critical for survival of type I SGNs during development, these results suggest that it plays similar roles in the adult. Together, the data indicate that adult cochlear supporting cells provide critical trophic support to the neurons, that survival of postnatal cochlear sensory neurons depends on reciprocal interactions between neurons and supporting cells, and that these interactions are mediated by NRG and neurotrophins.

energy extraction from the biologic battery in the inner ear

Endocochlear potential (EP) is a battery-like electrochemical gradient found in and actively maintained by the inner ear. Here we demonstrate that the mammalian EP can be used as a power source for electronic devices. We achieved this by designing an anatomically sized, ultra-low quiescent-power energy harvester chip integrated with a wireless sensor capable of monitoring the EP itself. Although other forms of in vivo energy harvesting have been described in lower organisms, and thermoelectric, piezoelectric and biofuel devices are promising for mammalian applications, there have been few, if any, in vivo demonstrations in the vicinity of the ear, eye and brain. In this work, the chip extracted a minimum of 1.12 nW from the EP of a guinea pig for up to 5 h, enabling a 2.4 GHz radio to transmit measurement of the EP every 40–360 s. With future optimization of electrode design, we envision using the biologic battery in the inner ear to power chemical and molecular sensors, or drug-delivery actuators for diagnosis and therapy of hearing loss and other disorders.

Localization of pH regulating proteins H+ATPase and Cl-/HCO3- exchanger in the guinea pig inner ear.

Mechanisms that regulate endolymphatic pH are unknown. It has long been recognized that, because of the large positive endolymphatic potential in the cochlea, a passive movement of protons would be directed out of endolymph leading to endolymphatic alkalization. However, endolymphatic pH is close to that of blood, suggesting that H+ is being secreted into endolymph. Since the kidney and the inner ear are both actively engaged in fluid and electrolyte regulation, we attempted to determine whether proteins responsible for acid secretion in the kidney also exist in the guinea pig inner ear. To that end, a monoclonal antibody against a 31 kDa subunit of a vacuolar vH+ATPase and a polyclonal, affinity purified antibody against the AE2 Cl-/HCO3- exchanger (which can also recognize AE1 under some conditions) were used. In the cochlea, the strongest immunoreactivity for the vH+ATPase was found in apical plasma membranes and apical cytoplasm of strial marginal cells. These cells were negative for the Cl-/HCO3- exchanger. Certain cells of the inner ear demonstrated both apical staining for vH+ATPase and basolateral staining for the Cl-/HCO3- exchanger; these included interdental cells and epithelial cells of the endolymphatic sac. Cochlear cell types with diffuse cytoplasmic staining for vH+ATPase and a basolaterally localized Cl-/HCO3- exchanger included inner hair cells, root cells and a subset of supporting cells in the organ of Corti. Hair cells of the utricle, saccule and cristae ampullaris also expressed both vH+ATPase and the Cl-/HCO3- exchanger, but immunostaining for the vH+ATPase was less intense and less polarized than in the cochlea. These immunocytochemical results support a role for the vH+ATPase and Cl-/HCO3- exchanger in the regulation of endolymphatic pH and suggest that certain cells (including strial marginal cells and epithelial cells of the endolymphatic sac) may be specialized for this regulation.

Medial efferent inhibition produces the largest equivalent attenuations at moderate to high sound levels in cat auditory‐nerve fibers

Previous work has shown that medial efferents can inhibit responses of auditory-nerve fibers to high-level sounds and that fibers with low spontaneous rates (SRs) are inhibited most. However, quantitative interpretation of these data is made difficult by effects of adaptation. To minimize systematic differences in adaptation, efferent inhibition was measured with a randomized presentation of both sound level and efferent stimulation. In anesthetized cats, efferents were stimulated with 200/s shocks and auditory-nerve-fiber responses were recorded for tone bursts (0-100 dB SPL, 5-dB steps) at their characteristic frequencies. Below 50 dB SPL, efferent inhibition (measured as equivalent attenuation) was similar for all fibers with similar CFs in the same cat. At 45-75 dB SPL, low-SR and medium-SR fibers often showed much larger inhibition, and substantial inhibition even at 100 dB SPL. Expressed as a fractional decrease in rate, at 90-100 dB SPL the inhibition was 0%, 6%, and 13% for high-, medium-, and low-SR fibers (differences statistically significant). Finding the largest equivalent attenuations at 45-75 dB SPL does not fit with the hypothesis that medial-efferent inhibition is due solely to a reduction of basilar-membrane motion. The large attenuations, some over 50 dB, indicate that medial efferent inhibition is more potent than previously reported.

GENE EXPRESSION PROFILING OF NASAL POLYPS ASSOCIATED WITH CHRONIC SINUSITIS AND ASPIRIN-SENSITIVE ASTHMA

Objective: To identify genes whose expression is most characteristic of chronic rhinosinusitis and aspirin‐sensitive asthma through genome‐wide transcriptional profiling of nasal polyp tissue.

Dynamic patterns of neurotrophin 3 expression in the postnatal mouse inner ear

Recent studies indicate that neurotrophin 3 (NT3) may be important for the maintenance and function of the adult inner ear, but the pattern of postnatal NT3 expression in this organ has not been characterized. We used a reporter mouse in which cells expressing NT3 also express β‐galactosidase, allowing for their histochemical visualization, to determine the pattern of NT3 expression in cochlear and vestibular organs. We analyzed animals from birth (P0) to adult (P135). At P0, NT3 was strongly expressed in supporting cells and hair cells of all vestibular and cochlear sense organs, Reissners membrane, saccular membrane, and the dark cells adjacent to canal organs. With increasing age, staining disappeared in most cell types but remained relatively high in inner hair cells (IHCs) and to a lesser extent in IHC supporting cells. In the cochlea, by P0 there is a longitudinal gradient (apex > base) that persists into adulthood. In vestibular maculae, staining gradients are: striolar > extrastriolar regions and supporting cells > hair cells. By P135, cochlear staining is restricted to IHCs and their supporting cells, with stronger expression in the apex than the base. By the same age, in the vestibular organs, NT3 expression is weak and restricted to saccular and utricular supporting cells. These results suggest that NT3 might play a long‐term role in the maintenance and functioning of the adult auditory and vestibular systems and that supporting cells are the main source of this factor in the adult. J. Comp. Neurol. 501:30–37, 2007.

23.2 A 1.1nW energy harvesting system with 544pW quiescent power for next-generation implants

A wireless sensor that is powered from the endocochlear potential (EP), a 70-to-100mV bio-potential inside the mammalian ear, has been demonstrated in [1]. Due to the anatomical size and physiological constraints inside the ear, a maximum of 1.1 to 6.25nW can be extracted from the EP. The nanowatt power budget of the sensor gives rise to unique challenges with power conversion efficiency and quiescent current reduction in the power management unit (PMU). While [1] presents the system aspects of the biomedical harvesting including the biologic interface and system measurements, this work presents the details of the nanowatt PMU required to power the electronics. More specifically, it focuses on the low-power circuit design techniques needed to realize a nW power converter that is applicable to a broad spectrum of emerging biomedical applications with ultra-low energy-harvesting sources.

Real-time quantitative RT-PCR for low-abundance transcripts in the inner ear: analysis of neurotrophic factor expression

Real-time quantitative reverse transcription-PCR is a highly sensitive technology that allows high throughput quantification of gene expression. Application of this technique to the inner ear is potentially very important, but is not straightforward because tissue harvesting can be challenging, RNA yield from individual inner ears is low, and cDNA synthesis from scant RNA can be inefficient. To overcome these challenges, we tested many parameters and reagents, and developed an approach to reliably quantitate small changes in low-abundance transcripts. Using this technique we demonstrate the presence and quantify amounts of the neurotrophic factors neurotrophin 3 (NT-3), brain-derived neurotrophic factor (BDNF) and glial cell-line-derived neurotrophic factor (GDNF), in the cochlea and vestibular end organs of postnatal murine inner ear (P26). We show that out of the factors tested, BDNF is the only one differentially expressed between the cochlea and vestibular end organs, being 23.4+/-0.3 times more abundant in the vestibular end organs. Within the cochlea, GDNF gene expression is 4.9+/-0.2 times greater than NT-3 expression. Within the combined vestibular end organs, BDNF expression is 43.0+/-1.5 times greater than NT-3 expression. Our results suggest that neurotrophic factors continue to play a role in the postnatal inner ear, in addition to their previously shown essential role during development.

Nonneuronal cells regulate synapse formation in the vestibular sensory epithelium via erbB-dependent BDNF expression

Recent studies indicate that molecules released by glia can induce synapse formation. However, what induces glia to produce such signals, their identity, and their in vivo relevance remain poorly understood. Here we demonstrate that supporting cells of the vestibular organ—cells that have many characteristics of glia—promote synapse formation only when induced by neuron-derived signals. Furthermore, we identify BDNF as the synaptogenic signal produced by these nonneuronal cells. Mice in which erbB signaling has been eliminated in supporting cells have vestibular dysfunction caused by failure of synapse formation between hair cells and sensory neurons. This phenotype correlates with reduced BDNF expression in supporting cells and is rescued by reexpression of BDNF in these cells. Furthermore, knockdown of BDNF expression in supporting cells postnatally phenocopies the loss of erbB signaling. These results indicate that vestibular supporting cells contribute in vivo to vestibular synapse formation and that this is mediated by reciprocal signals between sensory neurons and supporting cells involving erbB receptors and BDNF.

Medial efferent effects on auditory-nerve responses to tail-frequency tones. I. Rate reduction

One way medial efferents are thought to inhibit responses of auditory-nerve fibers (ANFs) is by reducing the gain of the cochlear amplifier thereby reducing motion of the basilar membrane. If this is the only mechanism of medial efferent inhibition, then medial efferents would not be expected to inhibit responses where the cochlear amplifier has little effect, i.e., at sound frequencies in the tails of tuning curves. Inhibition at tail frequencies was tested for by obtaining randomized rate-level functions from cat ANFs with high characteristic frequencies (CF > or = 5 kHz), stimulated with tones two or more octaves below CF. It was found that electrical stimulation of medial efferents can indeed inhibit ANF responses to tail-frequency tones. The amplitude of efferent inhibition depended on both sound level (largest near to threshold) and frequency (largest two to three octaves below CF). On average, inhibition of high-CF ANFs responding to 1 kHz tones was around 5 dB. Although an efferent reduction of basilar-membrane motion cannot be ruled out as the mechanism producing the inhibition of ANF responses to tail frequency tones, it seems more likely that efferents produce this effect by changing the micromechanics of the cochlear partition.