Intense noise exposure alters peripheral vestibular structures and physiology.

The otolith organs play a critical role in detecting linear acceleration and gravity to control posture and balance. Some afferents that innervate these structures can be activated by sound and are at risk for noise overstimulation. A previous report demonstrated that noise exposure can abolish vestibular short-latency evoked potential (VsEP) responses and damage calyceal terminals. However, the stimuli that were used to elicit responses were weaker than those established in previous studies and may have been insufficient to elicit VsEP responses in noise-exposed animals. The goal of this study was to determine the effect of an established noise exposure paradigm on VsEP responses using large head-jerk stimuli to determine if noise induces a stimulus threshold shift and/or if large head-jerks are capable of evoking VsEP responses in noise-exposed rats. An additional goal is to relate these measurements to the number of calyceal terminals and hair cells present in noise-exposed vs. non-noise-exposed tissue. Exposure to intense continuous noise significantly reduced VsEP responses to large stimuli and abolished VsEP responses to small stimuli. This finding confirms that while measurable VsEP responses can be elicited from noise-lesioned rat sacculi, larger head-jerk stimuli are required, suggesting a shift in the minimum stimulus necessary to evoke the VsEP. Additionally, a reduction in labeled calyx-only afferent terminals was observed without a concomitant reduction in the overall number of calyces or hair cells. This finding supports a critical role of calretinin-expressing calyceal-only afferents in the generation of a VsEP response.NEW & NOTEWORTHY This study identifies a change in the minimum stimulus necessary to evoke vestibular short-latency evoked potential (VsEP) responses after noise-induced damage to the vestibular periphery and reduced numbers of calretinin-labeled calyx-only afferent terminals in the striolar region of the sacculus. These data suggest that a single intense noise exposure may impact synaptic function in calyx-only terminals in the striolar region of the sacculus. Reduced calretinin immunolabeling may provide insight into the mechanism underlying noise-induced changes in VsEP responses.

Rapamycin but not acarbose decreases age-related loss of outer hair cells in the mouse Cochlea.

Adding rapamycin or acarbose to diet at 9-10 months of age has been shown to significantly increase life span in both male and female UM-HET3 mice. The current study examined cochleae of male and female UM-HET3 mice at 22 months of age to determine if either treatment also influenced age-related loss of cochlear hair cells. A large loss of cochlear outer hair cells was observed at 22 months of age in untreated mice in both apical and basal halves of the cochlear spiral. Addition of acarbose to diet had no significant effect on the amount of outer hair cell loss at 22 months of age or in its pattern, with large loss in both apical and basal halves. The addition of rapamycin to diet, however, significantly reduced outer hair cell loss in the basal half of the cochlea at 22 months of age when compared to untreated mice. There was no significant difference between male and female mice in any of the conditions. Age-related outer hair cell loss in the apical cochlea precedes outer hair cell loss in the base in many mouse strains. The results of the present study suggest that rapamycin but not acarbose treatment can delay age-related loss of outer hair cells at doses at which each drug increases life span.

Age-related changes in auditory nerve-inner hair cell connections, hair cell numbers, auditory brain stem response and gap detection in UM-HET4 mice.

This study compared the timing of appearance of three components of age-related hearing loss that determine the pattern and severity of presbycusis: the functional and structural pathologies of sensory cells and neurons and changes in gap detection (GD), the latter as an indicator of auditory temporal processing. Using UM-HET4 mice, genetically heterogeneous mice derived from four inbred strains, we studied the integrity of inner and outer hair cells by position along the cochlear spiral, inner hair cell-auditory nerve connections, spiral ganglion neurons (SGN), and determined auditory thresholds, as well as pre-pulse and gap inhibition of the acoustic startle reflex (ASR). Comparisons were made between mice of 5-7, 22-24 and 27-29 months of age. There was individual variability among mice in the onset and extent of age-related auditory pathology. At 22-24 months of age a moderate to large loss of outer hair cells was restricted to the apical third of the cochlea and threshold shifts in the auditory brain stem response were minimal. There was also a large and significant loss of inner hair cell-auditory nerve connections and a significant reduction in GD. The expression of Ntf3 in the cochlea was significantly reduced. At 27-29 months of age there was no further change in the mean number of synaptic connections per inner hair cell or in GD, but a moderate to large loss of outer hair cells was found across all cochlear turns as well as significantly increased ABR threshold shifts at 4, 12, 24 and 48 kHz. A statistical analysis of correlations on an individual animal basis revealed that neither the hair cell loss nor the ABR threshold shifts correlated with loss of GD or with the loss of connections, consistent with independent pathological mechanisms.

Immunolocalization of vesicular glutamate transporters 1 and 2 in the rat inferior colliculus.

The inferior colliculus is a major relay nucleus in the ascending auditory pathways that receives multiple glutamatergic inputs. Vesicular glutamate transporters 1 and 2 (VGLUT1, VGLUT2) most often have complementary non-overlapping distributions and can be used to differentiate glutamatergic inputs. The present study therefore examined co-immunolabeling of VGLUT1 and VGLUT2 in three divisions of the rat inferior colliculus. Additional co-immunolabeling of microtubule-associated protein 2 and neuronal class III beta-tubulin provided visualization of neuronal soma and processes and allowed identification of axo-somatic versus axo-dendritic contacts. Results showed numerous VGLUT1 and 2 immunolabeled terminals in the central nucleus, lateral cortex and dorsal cortex. In all three divisions there was little to no co-containment of the two vesicular glutamate transporters indicating a complementary distribution. VGLUT1 made predominantly axo-dendritic connections in the neuropil, while VGLUT2 had many axo-somatic contacts in addition to axo-dendritic contacts. VGLUT2 immunolabeled terminals were numerous on the soma and proximal dendrites of many medium-to-large and large neurons in the central nucleus and medium to large neurons in the dorsal cortex. There were more VGLUT2 terminals than VGLUT1 in all divisions and more VGLUT2 terminals in dorsal and lateral cortices than in the central nucleus. This study shows that VGLUT1 and VGLUT2 differentiate complementary patterns of glutamatergic inputs into the central nucleus, lateral and dorsal cortex of the inferior colliculus with VGLUT1 endings predominantly on the dendrites and VGLUT2 on both dendrites and somas.

Deafness associated changes in two-pore domain potassium channels in the rat inferior colliculus.

Two-pore potassium channels can influence neuronal excitability by regulating background leakage of potassium ions and resting membrane potential. The present study used quantitative real time PCR and in situ hybridization to determine if the decreased activity from deafness would induce changes in two-pore potassium channel subunit expression in the rat inferior colliculus (IC). Ten subunits were assessed with quantitative real-time PCR at 3 days, 3 weeks and 3 months following bilateral cochlear ablation. TASK-1, TASK-5 and THIK-2 showed significant decreases in expression at all three times assessed. TASK-5, relatively specific to auditory neurons, had the greatest decrease. TWIK-1 was significantly decreased at 3 weeks and 3 months following deafness and TREK-2 was only significantly decreased at 3 days. TASK-3, TWIK-2, THIK-1, TRAAK and TREK-1 did not show any significant changes in gene expression. In situ hybridization was used to examine TASK-1, TASK-5, TWIK-1 and THIK-2 in the central nucleus, dorsal cortex and lateral (external) cortex of the IC in normal hearing animals and at 3 weeks following deafening. All four subunits showed expression in neurons throughout IC subdivisions in normal hearing rats, with TASK-5 having the greatest overall number of labeled neurons. There was no co-localization of subunit expression with glial fibrillary acidic protein immunostaining, indicating no expression in glia. Three weeks following deafening there was a significant decrease in the number of neurons expressing TASK-1 and THIK-2 in the IC, while TASK-5 had significant decreases in the central nucleus and dorsal cortex and TWIK-1 in the lateral and dorsal cortices.

[Characterisation of the gene expression profiles in the inner ear and the colliculus inferior of normal and deafened rats by gene-array-technology]. / Charakterisierung der Genexpression im Innenohr und der Hörbahn hörender und ertaubter Ratten mittels Gene-Arrays.

BACKGROUND: The phenotype of deafness and its mechanisms are morphologically and electrophysiologically well characterised. However, the molecular mechanisms and the consequences of deafness are poorly understood. METHODS: In this study we investigated changes in gene expression profiles in subfractions of the cochlea and the colliculus inferior, a non-cochlear tissue, of normal and deafened (10 % Neomycin) rats using the gene-array-technology. RNA was prepared from modiolus (Mo) und sensorineural epithel/lateral wall (SnE/Lw) und Colliculus inferior (IC), reverse transcribed with gene specific primers, labeled with (32)P-dATP and hybridised with its complementary sequences of 1200 rat ESTs. RESULTS: Similar gene expression profiles were detected in Mo- and SnE/Lw in normal as well in deafened rats differing significantly from those found in IC. In deafened animals differences in mRNA levels were determined in IC for 8 genes, in Mo für 17 genes and in SnE/Lw for 25 genes in comparison to those of normal rats. By using gene-arrays many genes described in the literature previously could be detected. Otherwise most of the genes found in the cochlea are unknown. CONCLUSIONS: The gene-array-technology is a valuable tool in otological research for gene expression analysis and, therefore, for comprehensive understanding of molecular processes in the inner ear. Furthermore gene screening for candidate genes could be a big step ahead in developing therapies of diseases of the inner ear.

Pathways for protection from noise induced hearing loss.

There is increasing evidence that at least one function of both the medial and the lateral olivocochlear efferent systems is to provide adjustment of the set point of activity in their postsynaptic target, the outer hair cells and afferent processes, respectively. New results, summarized in this review, suggest that both efferent systems can provide protection from noise through this mechanism. There are also intracellular pathways that can provide protection from noise-induced cellular damage in the cochlea. This review also summarizes new results on the pathways that regulate and react to levels of reactive oxygen species in the cochlea as well as the role of stress pathways for the heat shock proteins and for neurotrophic factors in protection, recovery and repair.

Expression of GABA(A) receptor subunits in the rat central nucleus of the inferior colliculus.

Expression of GABA(A) receptor (GABA(A)R) alpha(1), alpha(2), beta(2), gamma(1), gamma(2L) and gamma(2S) subunit mRNA was examined in three cell classes in the central nucleus of the rat inferior colliculus (CNIC). GABA(A)R alpha(1) and gamma(2L) subunit mRNA expression was greatest in large cells (over 25 microm long diameter), intermediate in medium sized cells (15 to 25 microm long diameter) and lowest in small cells (10 to 15 microm long diameter). GABA(A)R gamma(2S) and alpha(2) subunits had the opposite pattern, highest in the small cells, intermediate in medium cells and lowest in large cells. GABA(A)R beta(2) was significantly lower in small cells than the two other classes, while differences between large and medium cells were not significant. GABA(A)R gamma(1) subunit mRNAs expression was not above background in any of the three cell types assessed. The expression of GABA(A)R subunits suggests that cell classes in the rat CNIC may differ in their response to GABA and GABAergic drugs.

Glial cell line-derived neurotrophic factor (GDNF) and its receptor complex are expressed in the auditory nerve of the mature rat cochlea.

Glial cell line-derived neurotrophic factor (GDNF) is a survival factor for many neuronal cell types which signals through a heterodimer receptor consisting of GDNF-family receptor alpha 1 (GFRalpha-1) and Ret (rearranged during transformation). GDNF expression has previously been reported in the inner hair cells of the rat cochlea, with expression of GFRalpha-1 but not Ret in the cell bodies of the auditory nerve (spiral ganglion cells), using in situ hybridization. The present study used reverse transcription-polymerase chain reaction (RT-PCR), and immunocytochemistry to examine GDNF, GFRalpha-1 and Ret in the adult rat auditory nerve. Semi-quantitative RT-PCR showed expression of GDNF and the two receptor components, GFRalpha-1 and Ret, in the modiolar subfraction of the cochlea containing spiral ganglion cells. A shorter mRNA splice variant for GDNF was also detected. Immunocytochemistry showed immunostaining in the modiolus for GDNF, GFRalpha-1 and Ret that was confined to spiral ganglion cells. When RT-PCR expression levels were compared to the expression in the substantia nigra, GFRalpha-1 expression levels were similar, Ret mRNA was lower in the modiolus and GDNF expression was higher in the modiolus. However, when GDNF was further assessed using Western blot, while GDNF protein was found in the modiolus it was at lower levels than in substantia nigra tissue. These results demonstrate that GDNF and both of its receptor components are found in spiral ganglion cells of the adult rat cochlea. Along with the previous report of GDNF in inner hair cells, these new results provide a basis for the role of GDNF as a survival factor for the auditory nerve, as suggested by previous studies.

Intense noise induces formation of vasoactive lipid peroxidation products in the cochlea.

This study investigates the correlation between the formation of reactive oxygen species (ROS) and auditory damage in noise-induced hearing loss. The noise exposure (4-kHz octave band, 115 dB SPL, 5 h) created permanent threshold shifts at frequencies from 2 to 20 kHz. The lipid peroxidation product, 8-isoprostane, was determined biochemically and histochemically as an indicator of ROS. Noise exposure increased 8-isoprostane levels in the cochlea in a time-dependent manner. After 5 h of exposure, 8-isoprostane levels were more than 30-fold greater than baseline, and decreased rapidly after the termination of noise. The immunoreactivity to 8-isoprostane was increased in the stria vascularis, spiral ganglion cells and the organ of Corti. In the organ of Corti, immunostaining was restricted to the second turn in a region 10-12 mm from the apex. This region sustained most of the permanent hair cell damage as revealed in surface preparations. Outer hair cells were more heavily immunostained than inner hair cells while Hensen's cells showed still less immunostain. These data are consistent with the view that ROS are involved in noise-induced damage. However, the relationship between ROS formation and tissue damage appears complex. In the organ of Corti, the pattern of noise-induced lipid peroxidation correlates well with subsequent morphological damage. The stria vascularis, however, does not sustain permanent damage despite intense lipid peroxidation. Differences in endogenous antioxidant levels and commitment to different apoptotic or survival pathways may underlie such differential responses.

Diversity and plasticity in amino acid receptor subunits in the rat auditory brain stem.

Glutamate, gamma aminobutyric acid (GABA) and glycine receptors have different properties depending on the specific subunit combination utilized. The subunit composition of amino acid receptors may help to shape the responses of neurons and can provide a diversity of response properties in different neuronal types and regions. This allows a synaptic fine tuning for an optimization of processing requirements and may also allow for changes in response to changes in input. This article reviews the diversity that has been found in the subunit composition of GABA, glycine, alpha amino-3-hydroxy-5-methyl-4 isoxazole propionic acid and N-Methyl, D-aspartate (NMDA) receptors in the mammalian auditory brain stem and provides new data on how the NMDAR1 glutamate receptor subunit changes as a consequence of deafness. In the latter study, quantitative in situ hybridization was used to assess NMDAR1 mRNA expression in six cell types of the rat cochlear nucleus. A unilateral cochlear ablation was performed and expression determined in the ipsilateral and contralateral cochlear nucleus 5 and 20 days later. Significantly decreased expression, compared to normal, was found 5 days following deafness, in ipsilateral spherical bushy cells, octopus cells and shell neurons, but not in fusiform cells, corn cells or granule cells. At 20 days the expression was not significantly different from normal in any of the six cell types.

Changes in cochlear electrical stimulation induced Fos expression in the rat inferior colliculus following deafness.

Fos immunoreactive (IR) staining was used to examine changes in excitatory neuronal activity in the rat inferior colliculus (IC) between normal hearing and 21 day deaf rats evoked by basal or apical monopolar cochlear electrical stimulation. The location of evoked Fos IR neurons was consistent with expected tonotopic areas. The number of Fos IR cells increased as stimulation intensity increased in both normal and 21 day deaf animals. Stimulation at 1. 5x threshold evoked fewer Fos IR cells in 21 day deafened animals compared to normal hearing animals. At 5x and above, however, significantly increased numbers of Fos IR neurons (in a larger grouping) were evoked in 21 day deafened animals compared to normal hearing animals. Another group of animals had 7 days of deafness followed by 14 days of chronic basal cochlear electrical stimulation. In this group basal monopolar stimulation at 5x evoked not only a greater number of Fos IR neurons, compared to normal hearing animals, but the location of their grouping was slightly shifted to a more dorso-lateral region in the contralateral IC, compared to the normal hearing and 21 day deaf groups. These observations indicate that both deafness and chronic electrical stimulation may alter central auditory processing.

Differential display and gene arrays to examine auditory plasticity.

Differential gene expression forms the basis for development, differentiation, regeneration, and plasticity of tissues and organs. We describe two methods to identify differentially expressed genes. Differential display, a PCR-based approach, compares the expression of subsets of genes under two or more conditions. Gene arrays, or DNA microarrays, contain cDNAs from both known genes and novel genes spotted on a solid support (nylon membranes or glass slides). Hybridization of the arrays with RNA isolated from two different experimental conditions allows the simultaneous analysis of large numbers of genes, from hundreds to thousands to whole genomes. Using differential display to examine differential gene expression after noise trauma in the chick basilar papilla, we identified the UBE3B gene that encodes a new member of the E3 ubiquitin ligase family (UBE3B). UBE3B is highly expressed immediately after noise in the lesion, but not in the undamaged ends, of the chick basilar papilla. UBE3B is most similar to a ubiquitin ligase gene from Caenorhabditis elegans, suggesting that this gene has been conserved throughout evolution. We also describe preliminary experiments to profile gene expression in the cochlea and brain with commercially available low density gene arrays on nylon membranes and discuss potential applications of this and DNA microarray technology to the auditory system.

Upregulation of glial cell line-derived neurotrophic factor (GDNF) in the rat cochlea following noise.

There are endogenous intracellular mechanisms that provide cells with protection from stress, as well as repair from damage. These pathways often involve stress proteins and neurotrophic factors. The present study used Western blot analysis to examine changes in glial cell line-derived neurotrophic factor (GDNF) following noise overstimulation. A noise exposure was utilized which causes a temporary threshold shift and has been previously shown to upregulate heat shock protein 72 in the rat cochlea. This noise exposure also provides protection from a second noise exposure that would otherwise cause a permanent threshold shift. Experimental animals were assessed 2, 4, 8 and 12 h after cessation of noise exposure. Control animals received the same treatment except for the noise exposure and were assessed at the 8 h time point. A moderate expression of GDNF was observed in the normal cochlea. No significant change in GDNF levels was observed at 2 or 4 h following noise overstimulation. However, a significant increase was found at 8 h. At 12 h following noise overstimulation, GDNF levels were no longer significantly elevated from normal. These results suggest that GDNF is involved in the endogenous stress response in the cochlea and are consistent with the protection that exogenously applied GDNF has been shown to provide.

Differential protective effects of neurotrophins in the attenuation of noise-induced hair cell loss.

The protective efficacy of neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) at 1 or 10 microg/ml was assessed in guinea pigs exposed to 4 kHz octave band noise at 115 dB SPL for 5 h. BDNF, NT-3 or artificial perilymph was delivered to the scala tympani via a mini-osmotic pump, beginning 4 days prior to noise exposure and continuing for 1 week post-exposure. Protection was assessed physiologically by the change in auditory brainstem response (ABR) threshold, and histologically by outer hair cell (OHC) survival. There was a statistically significant increase in OHC survival and a decrease in ABR threshold shift in animals receiving NT-3 at a concentration of 10 microg/ml. In animals receiving 1 microg/ml NT-3, there was a significant increase in OHC survival in the first row of OHC, but no significant change in ABR threshold, relative to control animals. In animals treated with BDNF, no significant functional or histological protection was observed. The protection afforded by NT-3 (10 microg/ml) treatment was similar in magnitude to that reported previously with glial cell line-derived neurotrophic factor and suggests that several factors may be involved in the protective response.

Expression of glycine receptor subunit mRNAs in the rat cochlear nucleus.

The distribution of glycine receptor subunit mRNAs in the cochlear nucleus of the adult rat was examined using radioactive in situ hybridization. Expression was compared among six cell types by counting silver grains over somata. Expression of the immature alpha2 subunit was not above the threshold for detection in any neurons. Levels of expression of mature subunits varied among different cell types. Spherical bushy, small cell cap/shell neurons and fusiform cells had high expression of glycine receptor mRNA for alpha1, alpha 3 and beta subunits. Octopus cells and corn cells had high expression for alpha 3 and beta subunits, and only moderate expression for alpha1 subunit. Granule cells located between the dorsal and ventral cochlear nucleus had moderate expression of alpha 3 and beta subunits and no detectable alpha1 expression. These patterns of expression predict differences in glycinergic pharmacological properties between the cochlear nucleus neurons.

Cochleotopic fos immunoreactivity in cochlea and cochlear nuclei evoked by bipolar cochlear electrical stimulation.

Fos-like immunoreactivity evoked by basal, second or apical turn bipolar intracochlear electrical stimulation was evaluated in the spiral ganglion and cochlear nuclei. At stimulation levels of six times the electrically evoked auditory brain stem response thresholds, immunoreactive neurons were observed at appropriate discrete cochleotopic regions relative to stimulation site. The number of neurons increased with stimulus level and closely correlated to wave I amplitude. At 10 times thresholds, some spread in fos-like immunoreactivity to adjacent cochlear turns was found. However, fos-like immunoreactivity at this high level of stimulation still clearly showed a differential distribution in density of expression. These results indicated that the restricted topographic distribution of activity evoked by high levels of electrical stimulation is initiated at first order primary neurons of the system. For the profoundly deaf with cochlear implants, this indicates that place (channel) information can be maintained in the spiral ganglion and central nervous system even at very high levels of electrical stimulation. Together with our previous studies, these results indicate that fos provides a marker which can be used for evaluation of extent and pattern of cellular activation throughout the central auditory pathways, including activation of auditory nerve cells.

NMDAR1 isoforms in the rat superior olivary complex and changes after unilateral cochlear ablation.

Normal expression and deafness related changes in expression of NMDAR1 isoforms were examined in the rat superior olivary complex (SOC) using in situ hybridization with S35 labeled oligoprobes. Expression was assessed in three SOC nuclei, the lateral and medial superior olives (LSO, MSO) and the medial nucleus of the trapezoid body (MNTB). Silver grain labeling over principal cells of each region was assessed using METAMORPH image analysis system. Counts were made in ipsi- and contralateral sides after unilateral cochlear ablation and in treated and untreated animals. In the normal SOC, NMDAR1a expression was higher than 1b and 1-2 expression was followed by 1-4 and 1-1, with 1-3 below the level for detection. The levels and ratio were comparable in LSO, MSO and MNTB. Five days after cochlear ablation 1a, 1-1, 1-2 and 1-4 showed significant decreases in the ipsilateral LSO and 1-a and 1-2 showed significant decreases in the contralateral MNTB, with no significant changes in the MSO. At 20 days after deafening, no significant changes were seen for any isoform in any nucleus. The transient deafness-induced decreases in expression of NMDAR1 isoforms correlate with loss of excitation.

Glial cell line-derived neurotrophic factor has a dose dependent influence on noise-induced hearing loss in the guinea pig cochlea.

We examined the effectiveness of glial cell line-derived neurotrophic factor (GDNF) to attenuate cochlear damage from intense noise stress. Subjects were exposed to 115 dB SPL one octave band noise centered at 4 kHz for 5 h. They received artificial perilymph with or without GDNF into the left scala tympani at 0.5 microliter/h from 4 days before noise exposure through 8 days following noise exposure. Different concentrations of GDNF (1 ng/ml, 10 ng/ml, 100 ng/ml, and 1 microgram/ml) were applied chronically directly into the guinea pig cochlea via a microcannula and osmotic pump. Noise-induced hearing loss was assessed with pure tone auditory brainstem responses (at 2, 4, 8 and 20 kHz), measured prior to surgery, 1 day before noise exposure, and 7 days following noise exposure. Subjects were killed on day 8 following exposure for histological preparation and quantitative assessment of hair cell (HC) damage. A dose-dependent protective effect of GDNF on both sensory cell preservation and hearing function was found in the treated ears. At 1 ng/ml, GDNF showed no significant protection; at 10 ng/ml, GDNF showed significant HC protection; and at 100ng/ml, it was greater and bilateral. At 1 microgram/ml, GDNF appeared to have a toxic effect under noise stress in some cochleae. These findings indicate that GDNF at certain concentrations can effectively protect the inner ear from noise-induced hearing loss.

Expression of the GDNF family members and their receptors in the mature rat cochlea.

The GDNF family comprises glial cell line-derived neurotrophic factor (GDNF) and the related proteins neurturin, artemin and persephin, which form a subgroup of the TGF-beta superfamily of growth factors. All four neurotrophic factors provide neuronal cell protection and cell survival. GDNF expression was found in the cochlea, and GDNF has been shown to be effective for inner ear protection from drugs and noise-induced insults. As the other members of the GDNF family also provide protective effects on neuronal cells, they may play important roles in the inner ear. We used RT-PCR to examine the expression of GDNF, neurturin, artemin, persephin and their receptors GFRalpha-1, GFRalpha-2, GFRalpha-3 and c-ret in whole rat cochlea as well as in functionally different subfractions (modiolus and sensorineural epithelium/lateral wall) and compared the levels of neurotrophin and receptor mRNAs in the cochlea to those in substantia nigra brain region. Our results demonstrate the expression of all GDNF family members and their receptors in cochlea and substantia nigra. However, the relative levels of mRNA were different for several genes tested in subfractions of the cochlea and/or compared to expression levels in substantia nigra. The presence of mRNA for all four members of the GDNF family and their preferred receptors in the rat cochlea suggests potential functional importance of these neurotrophic factors as protection and survival factors in the inner ear.