OTOF mutations revealed by genetic analysis of hearing loss families including a potential temperature sensitive auditory neuropathy allele.

INTRODUCTION: The majority of hearing loss in children can be accounted for by genetic causes. Non-syndromic hearing loss accounts for 80% of genetic hearing loss in children, with mutations in DFNB1/GJB2 being by far the most common cause. Among the second tier genetic causes of hearing loss in children are mutations in the DFNB9/OTOF gene. METHODS: In total, 65 recessive non-syndromic hearing loss families were screened by genotyping for association with the DFNB9/OTOF gene. Families with genotypes consistent with linkage or uninformative for linkage to this gene region were further screened for mutations in the 48 known coding exons of otoferlin. RESULTS: Eight OTOF pathological variants were discovered in six families. Of these, Q829X was found in two families. We also noted 23 other coding variant, believed to have no pathology. A previously published missense allele I515T was found in the heterozygous state in an individual who was observed to be temperature sensitive for the auditory neuropathy phenotype. CONCLUSIONS: Mutations in OTOF cause both profound hearing loss and a type of hearing loss where otoacoustic emissions are spared called auditory neuropathy.

Autosomal recessive nonsyndromic neurosensory deafness at DFNB1 not associated with the compound-heterozygous GJB2 (connexin 26) genotype M34T/167delT.

Previous studies of the gap-junction beta-2 subunit gene GJB2 (connexin 26) have suggested that the 101T-->C (M34T) nucleotide substitution may be a mutant allele responsible for recessive deafness DFNB1. This hypothesis was consistent with observations of negligible intercellular coupling and gap-junction assembly of the M34T allele product expressed in Xenopus oocytes and HeLa cells. The results of our current study of a family cosegregating the 167delT allele of GJB2 and severe DFNB1 deafness demonstrate that this phenotype did not cosegregate with the compound-heterozygous genotype M34T/167delT. Since 167delT is a null allele of GJB2, this result indicates that the in vivo activity of a single M34T allele is not sufficiently reduced to cause the typical deafness phenotype associated with DFNB1. This observation raises the possibility that other GJB2 missense substitutions may not be recessive mutations that cause severe deafness and emphasizes the importance of observing cosegregation with deafness in large families to confirm that these missense alleles are mutant DFNB1 alleles.

Development of human cochlear active mechanism asymmetry: involvement of the medial olivocochlear system?

To study the functional development of the medial olivocochlear system, transient-evoked otoacoustic emission suppression experiments were conducted in 73 ears of 38 pre-term and 11 full-term neonates. The continuous contralateral stimulation was a broad band white noise, presented at 70 dB SPL. Efferent suppression was determined by subtracting the without-contralateral stimulation condition from the with-contralateral stimulation condition. Across this population, a mean suppression effect of contralateral stimulation on transient-evoked otoacoustic emissions was found, with most of the suppression effect observed after 8 ms. The amount of suppression is linearly, positively correlated with the conceptional age. In the subgroup of bilaterally tested neonates, the suppression of transient-evoked otoacoustic emissions is similar in the right ear and the left ear in subjects whose conceptional age is less than 36 weeks and significantly higher in the right ear than in the left ear in older neonates. This last observation was seen at frequencies where transient-evoked otoacoustic emission amplitudes became higher in the right ear than in the left ear as the conceptional age increased, a finding already reported in adults. This study shows that the functional adult pattern of the medial efferent system, probably involved in the detection of signals in noise such as speech sounds, seems to appear gradually in neonates and represents one of the several arguments in favor of functional auditory lateralization in humans, with a right ear advantage.

Genomics and hearing impairment.

Hearing impairment is clinically and genetically heterogeneous. There are >400 disorders in which hearing impairment is a characteristic of the syndrome, and family studies demonstrate that there are at least 30 autosomal loci for nonsyndromic hearing impairment. The genes that have been identified encode diaphanous (HDIA1), alpha-tectorin (TECTA), the transcription factor POU4F3, connexin 26 (GJB2), and two unconventional myosins (MYO7A and MYO15), and four novel proteins (PDS, COCH, DFNA5, DFNB9). The same clinical phenotype in hearing-impaired individuals, even those within the same family, can result from mutations in different genes. Conversely, mutations in the same gene can result in a variety of clinical phenotypes with different modes of inheritance. For example, mutations in the gene encoding MYO7A cause Usher syndrome type IB, autosomal-recessive nonsyndromic hearing impairment (DFNB2), and autosomal-dominant nonsyndromic hearing impairment (DFNA11). Additionally, the mouse ortholog of the MYO7A gene is the shaker-1 gene. Mouse models such as shaker-1 have facilitated the identification of genes that cause hearing impairment in humans. The availability of high-resolution maps of the human and mouse genomes and new technologies for gene identification are advancing molecular understanding of hearing impairment and the complex mechanisms of the auditory system.

Mutations in the connexin 26 gene (GJB2) among Ashkenazi Jews with nonsyndromic recessive deafness.

BACKGROUND: Mutations in the GJB2 gene cause one form of nonsyndromic recessive deafness. Among Mediterranean Europeans, more than 80 percent of cases of nonsyndromic recessive deafness result from inheritance of the 30delG mutant allele of GJB2. We assessed the contribution of mutations in GJB2 to the prevalence of the condition among Ashkenazi Jews. METHODS: We tested for mutations in GJB2 in DNA samples from three Ashkenazi Jewish families with nonsyndromic recessive deafness, from Ashkenazi Jewish persons seeking carrier testing for other conditions, and from members of other ethnic groups. The hearing of persons who were heterozygous for mutations in GJB2 was assessed by means of pure-tone audiometry, measurement of middle-ear immittance, and recording of otoacoustic emissions. RESULTS: Two frame-shift mutations in GJB2, 167delT and 30delG, were observed in the families with nonsyndromic recessive deafness. In the Ashkenazi Jewish population the prevalence of heterozygosity for 167delT, which is rare in the general population, was 4.03 percent (95 percent confidence interval, 2.5 to 6.0 percent), and for 30delG the prevalence was 0.73 percent (95 percent confidence interval, 0.2 to 1.8 percent). Genetic-linkage analysis showed conservation of the haplotype for 167delT but the existence of several haplotypes for 30delG. Audiologic examination of carriers of the mutant alleles who had normal hearing revealed subtle differences in their otoacoustic emissions, suggesting that the expression of mutations in GJB2 may be semidominant. CONCLUSIONS: The high frequency of carriers of mutations in GJB2 (4.76 percent) predicts a prevalence of 1 deaf person among 1765 people, which may account for the majority of cases of nonsyndromic recessive deafness in the Ashkenazi Jewish population. Conservation of the haplotype flanking the 167delT mutation suggests that this allele has a single origin, whereas the multiple haplotypes with the 30delG mutation suggest that this site is a hot spot for recurrent mutations.

The mouse deafness locus (dn) is associated with an inversion on chromosome 19.

Recombination data for the mouse deafness locus (dn) on chromosome 19 are consistent with the presence of an inversion for which one of the breakpoints is between D19Mit14 and D19Mit96, a distance of less than 226 kb. Fluorescence in situ hybridization studies using a bacterial artificial chromosome on interphase (G1) nuclei provide additional support for the presence of an inversion. The dn gene is probably the orthologue of the human DFNB7/DFNB11 gene on chromosome 9.

Low intensities and 1.3 ratio produce distortion product otoacoustic emissions which are larger in heterozygous (+/dn) than homozygous (+/+) mice.

The f2/f1 frequency ratio of 1.3 in combination with stimulus levels of L1/L2 = 50/60 and 50/50 dB SPL produced a higher level of distortion product otoacoustic emissions (DPOAE) in the heterozygous (+/dn) mice than in the homozygous (+/+) mice. These results suggest that the dn gene carriers have a unique cochlear trait which may be related to the dn gene locus and expressed via a frequency- and intensity-dependent DPOAE function.

Reversing click polarity may uncover auditory neuropathy in infants.

OBJECTIVE: To identify patients with primary auditory neuropathies whose cochlear potentials to a 100 microsec click persist after click cessation and simulate synchronous auditory brain stem responses (ABRs) at high intensities. DESIGN: ABRs to condensation and rarefaction clicks, as well as Maximum Length Sequence ABRs and one transtympanic electrocochleogram (ECochG), were collected from five infants with absent middle ear muscle reflexes and normal or near normal otoacoustic emissions. These infants failed ALGO screens, which used alternating polarity clicks, and/or failed full ABRs done elsewhere with alternating polarity clicks. RESULTS: When ABRs were collected in response to a single polarity pulse, they revealed robust and reproducible wave forms over 4 to 6 msec that initially were mistaken for a normal ABR by the referring agents. However, when condensation and rarefaction click data are compared, the waveforms change polarity when the stimulus is inverted. Furthermore, the waveforms fail to shift in latency as the intensity of the stimulation is reduced. Transtympanic ECochG on one of the children revealed the same polarity reversal and fixed latency functions, confirming that they were cochlear rather than neural responses. CONCLUSIONS: Comparing responses with positive versus negative polarity clicks may help separate ABRs from cochlear potentials and alert clinicians to the possibility of an auditory neuropathy. Therefore, absent or abnormal ABRs in the presence of normal otoacoustic emissions need not always implicate a purely "central disorder," but might be consistent with dysfunction between outer hair cells and primary afferent fibers.

Contralateral suppression of transient-evoked otoacoustic emissions in humans: intensity effects.

Transient evoked otoacoustic emissions (TEOAEs) were recorded to clicks presented at peak sound pressures of 50, 55, 60, 65 and 70 dB while continuous contralateral white noise was varied from 10 dB below to 10 dB above the click level. Suppression increased predictably with suppressor noise level for any given click level. However, when the suppressor noise level was held constant, suppression was greater for lower click levels. This observation is consistent with the association of suppression of otoacoustic emissions with active cochlear processes and efferent function at low intensity levels.

Phenotypic patterns of distortion product otoacoustic emission in inbred and F1 hybrid hearing mouse strains.

Distortion product otoacoustic emissions (DPOE) were obtained from five different hearing mouse groups: CBA/J, MOLF/Rk, ct (homozygous normal mice of the curly-tail stock), and the F1 hybrid offspring of the matings CBA/J x dn/dn and MOLF/Rk x dn/dn (dn/dn mice are the curly-tail stock with recessive deafness). The DPOE patterns of the CBA/J and ct strains were similar to each other and different from that of the MOLF/Rk. The two sets of F1 hybrid mice, (CBA/J x dn/dn)F1 and (MOLF/Rk x dn/dn)F1, were found to have significantly larger DPOE amplitudes than their hearing parent strains, MOLF/Rk and CBA/J, respectively. In addition, the DPOE amplitudes were greater for the offspring of the MOLF/Rk x dn/dn cross than for those of the CBA/J x dn/dn cross, even though they were lower for MOLF/Rk than for CBA/J. The distinct features of DPOE patterns among these five groups suggest that DPOE testing can be used for auditory phenotyping.

Auditory neuropathy.

Ten patients presented as children or young adults with hearing impairments that, by behavioural and physiological testing, were compatible with a disorder of the auditory portion of the VIII cranial nerve. Evidence of normal cochlear outer hair cell function was provided by preservation of otoacoustic emissions and cochlear microphonics in all of the patients. Auditory brainstem potentials showed evidence of abnormal auditory pathway function beginning with the VIII nerve: the potentials were absent in nine patients and severely distorted in one patient. Auditory brainstem reflexes (middle ear muscles; crossed suppression of otoacoustic emissions) were absent in all of the tested patients. Behavioural audiometric testing showed a mild to moderate elevation of pure tone threshold in nine patients. The extent of the hearing loss, if due to cochlear receptor damage, should not have resulted in the loss of auditory brainstem potentials. The shape of the pure tone loss varied, being predominantly low frequency in five patients, flat across all frequencies in three patients and predominantly high frequency in two patients. Speech intelligibility was tested in eight patients, and in six was affected out of proportion to what would have been expected if the pure tone loss were of cochlear origin. The patients were otherwise neurologically normal when the hearing impairment was first manifest. Subsequently, eight of these patients developed evidence for a peripheral neuropathy. The neuropathy was hereditary in three and sporadic in five. We suggest that this type of hearing impairment is due to a disorder of auditory nerve function and may have, as one of its causes, a neuropathy of the auditory nerve, occurring either in isolation or as part of a generalized neuropathic process.

Development of the VIIIth nerve compound action potential evoked by low-intensity tone pips in the Mongolian gerbil.

Maturation of the cochlea and afferent auditory units is reflected by changes in VIIIth nerve compound action potential (CAP) parameters. We studied auditory nerve CAPs evoked by low-intensity stimuli in Mongolian gerbils (Meriones unguiculatus) ranging in age from 22 to 92 days after birth. The gerbil CAP development is characterized by marked changes in latency, threshold, and amplitude during the first few weeks of life. CAP latency and CAP threshold reach adult-like values at about 1 month of age. In contrast, the CAP amplitude continues to grow in size even after 2 months. This dichotomy suggests that the development of afferent auditory nerve function in the gerbil is preceded by maturation of the mechanical processes of the middle ear and cochlea.

Auditory phenotyping of heterozygous sound-responsive (+/dn) and deafness (dn/dn) mice.

Accurate phenotyping of offspring from backcross matings between F1 heterozygous sound-responsive and deafness mice is an important step for the identification of the deafness (dn) gene (Keats et al., 1995). Here, we report the results of auditory phenotyping of backcross offspring who are either sound-responsive or deaf by recording the Preyer reflex elicited by hand clap, auditory brainstem responses (ABRs), and 2f1-f2 distortion product otoacoustic emissions (DPOEs). Our results show that the Preyer reflex observation alone is inadequate for auditory phenotyping; a more precise test such as a click-evoked ABR recording is needed for auditory phenotyping. DPOE recording results in identification of sound-responsive or deaf mice as accurately as the click-evoked ABR testing. In addition, because the DPOE amplitude function is in good agreement with the ABR threshold in frequency sensitivity and specificity for stimulus frequencies between 1 and 16 kHz, the DPOE recording can be considered as an alternate test for auditory phenotyping.

Binaural noise suppresses linear click-evoked otoacoustic emissions more than ipsilateral or contralateral noise.

We studied the efferent suppression of click-evoked otoacoustic emissions with 65 dB SPL of white noise presented to left, right, or sometimes both, ears for 408 ms. Each burst of noise preceded a series of four unipolar 80 microseconds 65 dB peak Sound Pressure clicks, presented to the left ear only. The first click of the four-click group followed the end of the noise by either 1, 2, 5, 10, 20, 50, 100 or 200 ms; each subsequent click was offset by 20 additional ms via an ILO88 system with special programming modifications. Conditions were alternated so that a 'without noise' condition preceded a 'with noise' condition for three repetitions of 600 clicks per trial. Seven subjects with normal hearing participated in the study, and three of the seven participated in a test-retest reliability study. Results showed the greatest suppression followed binaural stimulation ending within one to five ms of the first click in the pulse train. Somewhat less suppression was seen following ipsilateral stimulation. The least amount of suppression was seen following contralateral stimulation, suggesting that previous research using contralateral stimulation may underestimate efferent effects. We saw no effects when the end of the noise was 100 ms or more away from the beginning of the click train.

The deafness locus (dn) maps to mouse chromosome 19.

The deafness mouse has profound sensorineural hearing loss with degeneration of hair cells soon after birth. The mode of inheritance is recessive, and there are no associated phenotypic anomalies. Thus, this mouse provides a model for recessive, non-syndromic, prelingual deafness. We have mapped the gene causing deafness in the mouse to Chromosome (Chr) 19 by analysis of 230 intersubspecific backcross progeny. No recombinants were found with the microsatellite marker D19Mit14. The loci for two guanine nucleotide-binding proteins are tightly linked to this marker, and they are being investigated as possible candidate genes. The identification of the defective gene in the mouse will help to explain the mechanism that causes hair cell degeneration and is likely to identify a homologous gene for deafness in humans.

Cortical deafness: a longitudinal study.

We have studied a patient with MRI-confirmed bilateral absence of considerable portions of her temporal lobes resulting in cortical deafness. Although physiologic measures demonstrate normal peripheral hearing sensitivity, this patient's speech has the inflection and prosodic characteristics associated with profound peripheral hearing loss, and she is unable to understand spoken communication. Behaviorally obtained pure-tone thresholds taken over nearly 20 years range from normal to moderate hearing loss with normal middle ear muscle reflexes and normal ABRs; however, we consistently found abnormal middle latency and cortical evoked potentials. Because of her total inability to communicate auditorily, this patient was ultimately taught American Sign Language and educated at the Louisiana School for the Deaf. This rare case highlights the importance of using multiple audiologic measures sensitive to abnormalities at various levels of the auditory system.

Development of contralateral suppression of the VIIIth nerve compound action potential (CAP) in the Mongolian gerbil.

We studied whether same-frequency contralateral tones of 65 dB pSPL (peak Sound Pressure Level) suppress the VIIIth nerve compound action potential (CAP) evoked by 40-45 dB pSPL tone pips in the Mongolian gerbil from 22 to 92 days after birth (DAB). The primary stimuli were tone pips of 1, 2, 4, 8, and 10 kHz; only the 1 kHz CAP amplitude was suppressed significantly by tones of the same frequency. The suppression was seen at 22 DAB, and underwent little relative change with development.

Clinical diagnosis of the Usher syndromes. Usher Syndrome Consortium.

The Usher syndromes are genetically distinct disorders which share specific phenotypic characteristics. This paper describes a set of clinical criteria recommended for the diagnosis of Usher syndrome type I and Usher syndrome type II. These criteria have been adopted by the Usher Syndrome Consortium and are used in studies reported by members of this Consortium.

The First Jerger Lecture. Contralateral suppression of otoacoustic emissions: an index of the function of the medial olivocochlear system.

We can now distinguish, in part, between nerve deafness and hair cell deafness through the use of otoacoustic emissions. We can also assess the efferent system by carefully quantifying the effects of contralateral stimulation on these same otoacoustic emissions. The suppression of transient evoked emissions by continuous contralateral white noise is an ostensibly small effect of 2 or 3 dB when studied over a 20-msec window. However, when subjected to microstructural analysis, the effect can exceed 6 to 8 dB in the zones from 10 to 20 msec after the stimulus has subsided. Temporal and spectral analyses reveal robust effects of contralateral lateral stimulation, although in any given normal subject it may be difficult to separate middle ear effects from efferent effects. Evidence is strong that the efferent effect is mediated in part by cholinergic-primarily nicotinic-receptors in the outer hair cell. However, a unique type of patient, who shows nearly normal pure-tone audiograms and absent ABRs, shows virtually no contralateral suppression of transient evoked emissions. Some other patients, with symptoms of Charcot-Marie-Tooth disease, may paradoxically show extremely poor audiograms, but perfectly normal evoked emissions along with absent contralateral suppression. The ABR, along with middle ear muscle reflexes and masking level differences, are all absent in these patients; we therefore think they have a disorder that desynchronizes most of their primary auditory nerve fibers and thereby disconnects them from any efferent activity or masking cancellation. The existence of such an auditory disorder, characterized by severe dysfunction in speech comprehension-especially when listening in noise-suggests that what appears to be a "central auditory imperception" might stem instead from a systemic peripheral primary neuropathy.

Contralateral suppression of non-linear click-evoked otoacoustic emissions.

Click-evoked otoacoustic emissions from nominal 80 dB pSP (peak sound pressure) 80-microseconds pulses presented at 50 pulses per second were collected from the right ears of eleven normal hearing subjects using an ILO88 Otodynamic Analyzer in the non-linear mode. Clicks, pure tones, and narrow bands of noise were then presented to their left ears through insert earphones. The 80-microseconds contralateral clicks ranged in intensity from 80 dB pSP in 5 dB steps down to 60 dB pSP but data on only 10 of the subjects were collected successfully. The pure tones and narrow bands of noise centered at 250, 500, 1000, 2000, and 4000 Hz were also presented through insert phones at 20, 40, 60 and 80 dB HL (Hearing Level) to all 11 subjects. The mean overall 'echo amplitude' without contralateral stimuli was 11 dB SPL and underwent more than 3 dB of overall suppression in response to the noises which were the most effective of the contralateral suppressors. When we analyzed the echo suppression to noise in 2-ms segments, we found consistent contralateral suppression of 3-4 dB concentrated in the time zones after 8 ms. Time shifts of more than 200 microseconds between the control and experimental traces were also observed in the same zones. The clicks were the next most effective suppressors, but showed their amplitude and time effects in somewhat earlier time zones. The tones were the least effective suppressors suggesting that efferent effects we measured in the human system are not strongly tonotopic. Because 'non-linear' mode high intensity clicks were deliberately selected as stimuli to evoke the TEOAE's, the emissions and their suppression can represent neither the 'true' TEOAE nor all of the efferent system's suppression abilities.