Temporal relationship between the auditory brainstem response and focal responses of auditory nerve root and cochlear nucleus during development in the tammar wallaby (Macropus eugenii)

Thirty-two pouch-young tammar wallabies were used to discover the generators of the auditory brainstem response (ABR) during development by the use of simultaneous ABR and focal brainstem recordings. A click response from the auditory nerve root (ANR) in the wallaby was recorded from postnatal day (PND) 101, when no central auditory station was functional, and coincided with the ABR, a simple positive wave. The response of the cochlear nucleus (CN) was detected from PND 110, when the ABR had developed 1 positive and 1 negative peak. The dominant component of the focal ANR response, the N-1 wave, coincided with the first half of the ABR P wave, and that of the focal CN response, the N-1 wave, coincided with the later two thirds. In older animals, the ANR response coincided with the ABR's N-1, wave, while the CN response coincided with the ABR's P-2, N-2 and P-3 waves, with its contribution to the ABR P-2 dominant. The protracted development of the marsupial auditory system which facilitated these correlations makes the tammar wallaby a particularly suitable model. Copyright (C) 2001 S. Karger AG, Basel.

Functional development of the inferior colliculus (IQ and its relationship with the auditory brainstem response (ABR) in the tammar wallaby (Macropus eugenii)

To discover the developmental relationship between the auditory brainstem response (ABR) and the focal inferior colliculus (IC) response, 32 young tammar wallabies were used, by the application of simultaneous ABR and focal brainstem recordings, in response to acoustic clicks and tone bursts of seven frequencies. The ic or the tammar wallaby undergoes a rapid functional development from postnatal day (PND) 114 to 160. The earliest (PND 114) auditory evoked response was recorded from the rostral IC. With development, more caudal parts of the IC became functional until age about PND 127, when all parts of the IC were responsive to sound. Along a dorsoventral direction, the duration of the IC response decreased, the peak latency shortened, while the amplitude increased, reaching a maximum value at the central IC, then decreased. After PND 160, the best frequency (BF) of the ventral IC was the highest, with values between 12.5 and 16 kHz, the BF of the dorsal IC was the lowest, varying between 3.2 and 6.4 kHz, while the BF of the central IC was between 6.4 and 12.5 kHz. Between PND 114 and 125, the IC response did not have temporal correlation with the ABR. Between PND 140 and 160, only the early components of the responses from the ventral and central IC correlated with the P4 waves of the ABR. After PND 160, responses recorded from different depths of the IC had a temporal correlation with the ABR. (C) 2001 Published by Elsevier Science B.V.

Automated Analysis of the Auditory Brainstem Response Using Derivative Estimation Wavelets

In this paper, we describe an algorithm that automatically detects and labels peaks I - VII of the normal, suprathreshold auditory brainstem response (ABR). The algorithm proceeds in three stages, with the option of a fourth: ( 1) all candidate peaks and troughs in the ABR waveform are identified using zero crossings of the first derivative, ( 2) peaks I - VII are identified from these candidate peaks based on their latency and morphology, ( 3) if required, peaks II and IV are identified as points of inflection using zero crossings of the second derivative and ( 4) interpeak troughs are identified before peak latencies and amplitudes are measured. The performance of the algorithm was estimated on a set of 240 normal ABR waveforms recorded using a stimulus intensity of 90 dBnHL. When compared to an expert audiologist, the algorithm correctly identified the major ABR peaks ( I, III and V) in 96 - 98% of the waveforms and the minor ABR peaks ( II, IV, VI and VII) in 45 - 83% of waveforms. Whilst peak II was correctly identified in only 83% and peak IV in 77% of waveforms, it was shown that 5% of the peak II identifications and 31% of the peak IV identifications came as a direct result of allowing these peaks to be found as points of inflection. Copyright (C) 2005 S. Karger AG, Basel.

On the dual structure of the auditory brainstem response in dogs

Objective: To use the over-complete discrete wavelet transform (OCDWT) to further examine the dual structure of auditory brainstem response (ABR) in the dog. Methods: ABR waveforms recorded from 20 adult dogs at supra-threshold (90 and 70 dBnHL) and threshold (0-15 dBSL) levels were decomposed using a six level OCDWT and reconstructed at individual scales (frequency ranges) A6 (0-391 Hz), D6 (391-781 Hz), and D5 (781-1563 Hz). Results: At supra-threshold stimulus levels, the A6 scale (0-391 Hz) showed a large amplitude waveform with its prominent wave corresponding in latency with ABR waves II/III; the D6 scale (391-781 Hz) showed a small amplitude waveform with its first four waves corresponding in latency to ABR waves I, II/III, V, and VI; and the D5 scale (781-1563 Hz) showed a large amplitude, multiple peaked waveform with its first six waves corresponding in latency to ABR waves I, II, III, IV, V, and VI. At threshold stimulus levels (0-15 dBSL), the A6 scale (0-391 Hz) continued to show a relatively large amplitude waveform, but both the D6 and D5 scales (391781 and 781-1563 Hz, respectively) now showed relatively small amplitude waveforms. Conclusions: A dual structure exists within the ABR of the dog, but its relative structure changes with stimulus level. Significance: The ABR in the dog differs from that in the human both in the relative contributions made by its different frequency components, and the way these components change with stimulus level. (c) 2006 International Federation of Clinical Neurophysiology. Published by Elsevier Ireland Ltd. All rights reserved.

The effect of high-pass filtering on TEOAE in 2-month-old infants

The aim of the present study was to investigate the effect of high-pass filtering on TEOAE obtained from 2-month-old infants as a function of filter cut-off frequency, activity states and pass/fail status of infants. Two experiments were performed. In Experiment 1, 100 2-month-old infants (200 ears) in five activity states (asleep, awake but peaceful, sucking a pacifier, feeding, restless) were tested by use of TEOAE technology. Five different filter conditions were applied to the TEOAE responses post hoc. The filter conditions were set at 781 Hz (default setting), 1.0, 1.2, 1.4 and 1.6 kHz. Results from this experiment showed that TEOAE parameters, such as whole-wave reproducibility (WR) and signal-to-noise ratio (SNR) at 0.8 kHz and 1.6 kHz, changed as a function of the cut-off frequency. The findings suggest that the 1.6 kHz and 1.2 kHz filter conditions are optimal for WR and SNR pass/fail criteria, respectively. Although all infant recordings appeared to benefit from the filtering, infants in the noisy states seemed to benefit the most. In Experiment 2, the high-pass filtering technique was applied to 23 infants (35 ears) who apparently failed the TEOAE tests on initial screening but were subsequently awarded a pass status based on the results from a follow-up auditory brainstem response (ABR) assessment. The findings showed a significant decrease in noise contamination of the TEOAE with a corresponding significant increase in WR. With high-pass filtering at 1.6 kHz, 21/35 ears could be reclassified into the pass category.

Conductive hearing loss in preterm infants with bronchopulmonary dysplasia

Objective: To determine the risk of conductive hearing loss in preterm infants with bronchopulmonary dysplasia (BPD) and preterm controls. Methodology: The study population consisted of 78 infants with BPD of 26-33 weeks gestation and 78 controls of similar gestational age matched for broad-based birthweight categories. An auditory brainstem response (ABR) audiology was performed shortly before hospital discharge. Visual reinforcement orientation audiometry (VROA) and impedance audiometry were performed at 8-12 months corrected for prematurity. Infants with persistent audiological abnormalities were referred for evaluation to paediatric ENT surgeons. Results: Infants with BPD had a significantly higher rate of ABR abnormalities (BPD: 22%, controls: 9%; P = 0.028). On VROA and impedance audiometry, the infants with BPD also had a higher rate of persistent abnormalities. Following ENT assessment, 22.1% of infants with BPD and 7.7% of controls had persistent conductive dysfunction requiring myringotomy and grommet tube insertion (P = 0.03). Most of these infants had normal ABR audiometry at hospital discharge. Conclusions: Preterm infants with BPD are at high risk of persistent conductive hearing loss late in the first year of life compared to controls. An ABR audiology conducted at the time of hospital discharge does not predict accurately later conductive hearing problems. Infants with BPD should have routine audiological evaluation toward the end of the first year of life.

Characterization of calcium-activated chloride channels in patches excised from the dendritic knob of mammalian olfactory receptor neurons

We investigated the properties of calcium-activated chloride channels in inside-out membrane patches from the dendritic knobs of acutely dissociated rat olfactory receptor neurons. Patches typically contained large calcium-activated currents, with total conductances in the range 30-75 nS. The dose response curve for calcium exhibited an EC50 of about 26 mu M. In symmetrical NaCl solutions, the current-voltage relationship reversed at 0 mV and was linear between -80 and +70 mV. When the intracellular NaCl concentration was progressively reduced from 150 to 25 mM, the reversal potential changed in a manner consistent with a chloride-selective conductance. Indeed, modeling these data with the Goldman-Hodgkin-Katz equation revealed a P-Na/P-Cl of 0.034. The halide permeability sequence was P-Cl > P-F > P-I > P-Br indicating that permeation through the channel was dominated by ion binding sites with a high field strength. The channels were also permeable to the large organic anions, SCN-, acetate(-), and gluconate(-), with the permeability sequence P-Cl > P-SCN > gluconaie. Significant permeation to gluconate ions suggested that the channel pore had a minimum diameter of at least 5.8 Angstrom.

Autoinhibition by an internal nuclear localization signal revealed by the crystal structure of mammalian importin alpha

Importin alpha is the nuclear import receptor that recognizes classical monopartite and bipartite nuclear localization signals (NLSs). The structure of mouse importin alpha has been determined at 2.5 Angstrom resolution. The structure shows a large C-terminal domain containing armadillo repeats, and a less structured N-terminal importin beta-binding domain containing an internal NLS bound to the NLS-binding site. The structure explains the regulatory switch between the cytoplasmic, high-affinity form, and the nuclear, low-affinity form for NLS binding of the nuclear import receptor predicted by the current models of nuclear import. Importin beta conceivably converts the low- to high-affinity form by binding to a site overlapping the autoinhibitory sequence. The structure also has implications for understanding NLS recognition, and the structures of armadillo and HEAT repeats.

Crystal structure of mammalian purple acid phosphatase

Background: Mammalian purple acid phosphatases are highly conserved binuclear metal-containing enzymes produced by osteoclasts, the cells that resorb bone. The enzyme is a target for drug design because there is strong evidence that it is involved in bone resorption. Results: The 1.55 Angstrom resolution structure of pig purple acid phosphatase has been solved by multiple isomorphous replacement. The enzyme comprises two sandwiched beta sheets flanked by or-helical segments. The molecule shows internal symmetry, with the metal ions bound at the interface between the two halves. Conclusions: Despite less than 15% sequence identity, the protein fold resembles that of the catalytic domain of plant purple acid phosphatase and some serine/threonine protein phosphatases. The active-site regions of the mammalian and plant purple acid phosphatases differ significantly, however. The internal symmetry suggests that the binuclear centre evolved as a result of the combination of mononuclear ancestors. The structure of the mammalian enzyme provides a basis for antiosteoporotic drug design.

Crystallization and preliminary X-ray diffraction studies of mammalian purple acid phosphatase

The oxidized form of purple acid phosphatase from pig allantoic fluid has been crystallized in the presence of phosphate using the hanging-drop technique. The crystals belong to the space group P2(1)2(1)2(1) and have unit-cell parameters a = 66.8, b = 70.3, c = 78.7 Angstrom. Diffraction data collected from a cryocooled crystal using a conventional X-ray source extend to 1.55 Angstrom resolution. A knowledge of the three-dimensional structure of mammalian purple acid phosphatase will aid in understanding the substrate specificity of the enzyme and will be important in the rational design of inhibitors, with potential in the treatment of bone diseases.

A rapid selection/amplification procedure for high-level expression of recombinant protein in a metal-amplifiable mammalian expression system

We describe a strategy for the selection and amplification of foreign gene expression in Chinese hamster ovary (CHO) cells employing a metallothionein gene-containing expression vector. This report describes an amplification procedure that results in an enrichment of clones exhibiting high levels of recombinant protein production and reduces the labour required for screening recombinant cell lines.

Single amino acid substitutions in alpha-conotoxin PnIA shift selectivity for subtypes of the mammalian neuronal nicotinic acetylcholine receptor

The alpha-conotoxins, a class of nicotinic acetylcholine receptor (nAChR) antagonists, are emerging as important probes of the role played by different nAChR subtypes in cell function and communication, In this study, the native alpha-conotoxins PnIA and PnIB were found to cause concentration-dependent inhibition of the ACh-induced current in all rat parasympathetic neurons examined, with IC50 values of 14 and 33 nM, and a maximal reduction in current amplitude of 87% and 71%, respectively. The modified alpha-conotoxin [N11S]PnIA reduced the ACh-induced current with an IC50 value of 375 nM and a maximally effective concentration caused 91% block, [A10L]PnIA was the most potent inhibitor, reducing the ACh-induced current in similar to 80% of neurons, with an IC50 value of 1.4 nM and 46% maximal block of the total current, The residual current was not inhibited further by alpha-bungarotoxin, but was further reduced by the cu-conotoxins PnIA or PnIB, and by mecamylamine. H-1 NMR studies indicate that PnIA, PnIB, and the analogues, [A10L]PnIA and [N11S]PnIA, have identical backbone structures. We propose that positions 10 and II of PnIA and PnIB influence potency and determine selectivity among alpha 7 and other nAChR subtypes, including alpha 3 beta 2 and alpha 3 beta 4, Four distinct components of the nicotinic ACh-induced current in mammalian parasympathetic neurons have been dissected with these conopeptides.

Structural basis of recognition of monopartite and bipartite nuclear localization sequences by mammalian importin-alpha

Importin-alpha is the nuclear import receptor that recognizes cargo proteins which contain classical monopartite and bipartite nuclear localization sequences (NLSs), and facilitates their transport into the nucleus. To determine the structural basis of the recognition of the two classes of NLSs by mammalian importin-alpha, we co-crystallized an N-terminally truncated mouse receptor protein with peptides corresponding to the monopartite NLS from the simian virus 40 (SV40) large T-antigen, and the bipartite NLS from nucleoplasmin. We show that the monopartite SV40 large T-antigen NLS binds to two binding sites on the receptor, similar to what was observed in yeast importin-alpha. The nucleoplasmin NLS-importin-alpha complex shows, for the first time, the mode of binding of bipartite NLSs to the receptor. The two basic clusters in the NLS occupy the two binding sites used by the monopartite NLS, while the sequence linking the two basic clusters is poorly ordered, consistent with its tolerance to mutations. The structures explain the structural basis for binding of diverse NLSs to the sole receptor protein. (C) 2000 Academic Press.