Genetic structure and diversity analysis of the primary gene pool of chickpea using SSR markers.

Members of the primary gene pool of the chickpea, including 38 accessions of Cicer arietinum, six of C. reticulatum and four of C. echinospermum grown in India were investigated using 100 SSR markers to analyze their genetic structure, diversity and relationships. We found considerable diversity, with a mean of 4.8 alleles per locus (ranging from 2 to 11); polymorphic information content ranged from 0.040 to 0.803, with a mean of 0.536. Most of the diversity was confined to the wild species, which had higher values of polymorphic information content, gene diversity and heterozygosity than the cultivated species, suggesting a narrow genetic base for cultivated chickpea. An unrooted neighbor-joining tree, principal coordinate analysis and population structure analysis revealed differentiation between the cultivated accessions and the wild species; three cultivated accessions were in an intermediate position, demonstrating introgression within the cultivated group. Better understanding of the structure, diversity and relationships within and among the members of this primary gene pool will contribute to more efficient identification, conservation and utilization of chickpea germplasm for allele mining, association genetics, mapping and cloning gene(s) and applied breeding to widen the genetic base of this cultivated species, for the development of elite lines with superior yield and improved adaptation to diverse environments.

Simultaneous measurements of ossicular velocity and intracochlear pressure leading to the cochlear input impedance in gerbil.

Recent measurements of three-dimensional stapes motion in gerbil indicated that the piston component of stapes motion was the primary contributor to intracochlear pressure. In order to make a detailed correlation between stapes piston motion and intracochlear pressure behind the stapes, simultaneous pressure and motion measurements were undertaken. We found that the scala vestibuli pressure followed the piston component of the stapes velocity with high fidelity, reinforcing our previous finding that the piston motion of the stapes was the main stimulus to the cochlea. The present data allowed us to calculate cochlear input impedance and power flow into the cochlea. Both the amplitude and phase of the impedance were quite flat with frequency from 3 kHz to at least 30 kHz, with a phase that was primarily resistive. With constant stimulus pressure in the ear canal the intracochlear pressure at the stapes has been previously shown to be approximately flat with frequency through a wide range, and coupling that result with the present findings indicates that the power that flows into the cochlea is quite flat from about 3 to 30 kHz. The observed wide-band intracochlear pressure and power flow are consistent with the wide-band audiogram of the gerbil.

The vibration pattern of the hearing organ in the waltzing guinea-pig measured using laser heterodyne interferometry.

The mechanical tuning characteristics of the hearing organ were measured in response to sound stimulation using laser heterodyne interferometry in in vitro preparations of temporal bones from waltzing guinea-pigs expressing different degrees of hearing organ and sensory cell degeneration. Measurements were made at various stages of structural changes allowing us to correlate structure and mechanical function. It was found that the characteristic frequency of the response at a given location in the cochlea occurred at lower frequencies than what is normally seen and that the sharpness of the mechanical tuning was considerably reduced when sensory hair cells were absent and the hearing organ structurally altered. However, even when extensive hair cell degeneration was evident a residual mechanical tuning was present. These results further support the concept that the sensory hair cells plays a key role in determining normal auditory tuning characteristics. It is suggested that the basilar membrane mechanics gives rise to a broadly tuned mechanical response on which a sharper tuning mechanism, originating from the hair cells, is superimposed.

The tuned displacement response of the hearing organ is generated by the outer hair cells.

The motile responses of the guinea-pig hearing organ in response to a tone applied to the ear were measured by laser interferometry. Two types of responses can be recorded: (i) a vibration at the frequency of the applied tone; and (ii) a displacement response consisting of a shift in the position of the organ surface. The purpose of this study is to characterize the displacement response. The results are as follows. There is a relationship between the frequency of highest sensitivity (best-frequency) of the displacement response and the site from which it is recorded. High best-frequencies are noticed at more basal locations, low best-frequencies towards the apex. The displacement response is more frequency-selective than the vibration response. The displacement response is observed within physiological sound pressure levels. Its sharpness is dependent on the stimulus intensity, it shows biological variability and can be manipulated by drugs that are known to modify the receptor potential of the sensory cells, or to interfere with outer hair cell motility. These results suggest that the displacement response is an important step in the transduction process in the mammalian hearing organ and that it is generated by the motile action of the outer hair cells.

Mechanical demodulation of hydrodynamic stimuli performed by the lateral line organ.

Tonic displacements of the fish lateral line cupula were observed during stimulation of the organ with amplitude-modulated water motion. The modulation frequency was fixed at 2.4 Hz and the carrier frequency was varied from 25 to 500 Hz. The time waveforms of the cupular displacement at carrier frequencies below 280 Hz and above 470 Hz were essentially amplitude-modulated waves. Between 350 Hz and 410 Hz the magnitude at the modulation frequency increased sharply and the predominant shape of the displacement waveform changed to that of the modulating frequency. The mechanism for extraction of the modulation component may play a key role in the decoding of sensory information.

Shearing motion in the hearing organ measured by confocal laser heterodyne interferometry.

To investigate the presence of the postulated shearing motion in the micromechanics of the inner ear during sound stimulations we measured the vibratory response of the tectorial membrane and the reticular lamina in the third cochlear turn in an isolated temporal bone preparation using confocal laser heterodyne interferometry. The mechanical response of the tectorial membrane had the same frequency of maxima as the underlying reticular lamina, but was not as sharply tuned. When the two-dimensional motion was calculated from measurements made from several viewing angles it was found that the vibration of the reticular lamina had significant components both normal and tangential to its surface. The tectorial membrane motion, however, was primarily in a direction approximately perpendicular to the surface of the reticular lamina. The results indicate that shearing motion is produced predominantly by the radial motion of the reticular lamina.

Frequency-specific position shift in the guinea pig organ of Corti.

The organ of hearing is tuned as expressed both in the vibratory response of the cochlear partition and in the resulting receptor potentials of the sensory cells. We now demonstrate a sharply tuned response, consisting of a position shift of the surface of the organ of Corti, occurring during the presentation of a tone. The magnitude of the position shift exceeds that of the vibratory response to the stimulus. The shift is most pronounced in the region of the outer hair cells, and its affected by an inhibitor of outer hair cell motility. We conclude that the response is induced by the action of the outer hair cells.

Dose rate to the inner ear during Mössbauer experiments.

The most widely used technique for studying vibrations of the inner ear utilises the Mössbauer effect; this requires placement of a radioactive source on the basilar membrane. This source, although small in size and less than 37 MBq (1 mCi) in strength, is placed in close proximity to sensitive receptor cells. Using a series solution for the radiation field of a rectangular source the absorbed dose rate delivered to receptor cells at various depths and at points off-axis from the centre of the source is calculated. It is concluded that the dose delivered during the course of a Mössbauer experiment may well be sufficient to damage receptor cells and cause a loss of response.

Homodyne interferometer for basilar membrane measurements.

Techniques available for measuring the mechanical response of the inner ear are compared. These include capacitive probe, Mössbauer and interferometric methods. The theory of a homodyne interferometer utilized for inner ear measurements is given. Experimental apparatus built to test the interferometer performance is described. Experimental results show that the measuring system can detect vibrations as low as 3 X 10(-12) cm. Its frequency response is flat within 1 dB from 0.1 to 40 kHz. It has a linear dynamic range of over 90 dB. Immunity of the interferometer to various disturbances is demonstrated.

Non-linear response to amplitude-modulated waves in the apical turn of the guinea pig cochlea.

Mechanical vibrations of the Hensen's cells were measured in the apical turn of the cochlea in living guinea pigs, in response to amplitude-modulated (AM) sound. The FFT of the input wave consisted of spectral components at the carrier frequency C and two sidebands (C+/-M) separated from the carrier by the modulation frequency M. The FFT of the velocity response consisted of components at: (i) the modulation frequency M, and harmonics n M; (ii) Carrier frequency C and sidebands (C+/-n M); (iii) harmonics of the carrier frequency and their side bands (2C+/-n M); (3C+/-n M); (4C+/-n M); em leader n=1,2,3, em leader,10. The carrier and the first pair of side bands were broadly tuned and nearly linear. Other components were sharply tuned and highly non-linear, suggesting a different origin. Evidence is presented that these components are generated in the non-linear stereocilia dynamics. An important function of this non-linearity is to demodulate the AM wave to extract information contained in the modulation.

The response of the apical turn of cochlea modeled with a tuned amplifier with negative feedback.

In an earlier study [Hear. Res. 149 (2000) 55] velocity amplitudes of the outer Hensen's cell (HC) and basilar membrane (BM) were measured before, and at different times, after, sacrificing the animal. The velocity amplitude changed in a way that was characteristic of a negative feedback amplifier. A simple negative feedback amplifier model was proposed to explain the magnitude of the HC and BM velocity changes at CF. In the experiment tuning changed as well, both at the HC and BM. The model has now been extended to include tuning changes. The model response is compared with the experimental observations. The model is able to account quantitatively for the following experimental observations: (i) At the HC the tuning broadens and velocity decreases slowly after sacrifice. (ii) At the BM tuning sharpens and velocity increases at a faster rate. (iii) The velocity increase at BM is much larger than the decrease at HC.

Spectral characteristics of the responses of primary auditory-nerve fibers to amplitude-modulated signals.

The spectral responses of cat single primary auditory nerve fibers to sinusoidal amplitude-modulated (AM) and double-sideband (DSB) acoustic signals applied to the ear were examined. DSB is an amplitude-modulated signal with a suppressed carrier. Period histograms were compiled from the neural spike-train data, and the frequency spectrum was determined by Fourier transforming these histograms. For DSB signals, spectral components were found to be present at the frequencies of the stimulus as well as at certain combination frequencies. For AM signals, several clusters of spectral components were present. The lowest-frequency cluster consisted of components at DC, at the modulation frequency, and at its harmonics. A higher frequency cluster occurs around a component with the frequency of the carrier. The components of cluster are separated from the carrier by the modulation frequency and its harmonics. Yet higher-frequency clusters appear around multiples of the carrier frequency with components at frequencies separated from these multiples by the modulation frequency and its harmonics. The magnitudes of these spectral components were determined for carrier frequencies located below, at, and above the characteristic frequency of the units, and for different stimulus levels, modulation frequencies, and modulation depths. The low-frequency components present in the neural spike train appear to be the result of demodulation taking place in the inner ear. The demodulated components are strong and are present over a wide range of sound levels, carrier frequencies, modulation frequencies, and nerve-fiber characteristics. This demodulation may be significant for speech recognition.

Spectral characteristics of the responses of primary auditory-nerve fibers to frequency-modulated signals.

The spectral responses of cat single primary auditory nerve fibers to sinusoidal frequency-modulated acoustic signals applied to the ear are examined. Period histograms were constructed from the neural spike-train data, and the frequency spectrum was determined by Fourier transforming these histograms. Several clusters of spectral components were present. The lowest-frequency cluster consists of components at DC, at the modulation frequency, and at its harmonics. In the next cluster, components surround the carrier frequency and are separated from it by the modulation frequency and its harmonics. Higher-frequency clusters surround frequencies that are twice and three times the carrier frequency. The components in each cluster are separated from the multiples of the carrier frequency by the modulation frequency and its harmonics. The magnitudes of the spectral components were investigated for carrier frequencies located below, at, and above the unit characteristic frequency, and for different signal levels, modulation frequencies, and modulation indices. The components at the modulation frequency and its harmonics were strong and present over a wide range of signal levels, carrier frequencies, modulation frequencies, and nerve-fiber characteristics. The presence of components at the modulation frequency indicates that a demodulation process is occurring. This process may be significant for speech recognition.

Measurement of basilar membrane vibrations and evaluation of the cochlear condition.

The experimental procedure for measuring basilar membrane responses to acoustic signals is described. The surgical procedure developed for opening the cochlea with minimal trauma is presented. Each experiment included sound pressure level measurements to define the input signal, cochlear microphonic (CM) measurements to monitor the cochlear condition, interferometric measurements and histological evaluation of the cochleas. The characteristic frequency (CF) and the sensitivity at CF for the basilar membrane response is correlated with the change of CM response observed in six animals. It is demonstrated that both tuning characteristics are extremely sensitive to cochlear trauma as evidenced by changes of CM.

Relationship between basilar membrane tuning and hair cell condition.

Basilar membrane tuning characteristics were measured in 15 cats using laser interferometry. The experimental procedures introduced varying degrees of cochlear trauma. Variability from animal to animal was also observed in the characteristic frequency (CF) of tuning, the sensitivity at CF and lower frequencies, and the sharpness of tuning. The changes in the tuning characteristics of the basilar membrane are correlated with the extent of outer hair cell (OHC) damage. These observations lead to the conclusion that the tuning properties in the CF region are predominantly determined by the mechanical properties of the OHC and not the basilar membrane.

Amplification in the apical turn of the cochlea with negative feedback.

The apical turn of the anesthetized guinea pig cochlea was opened to examine the basilar membrane optically through the intact Reissner's membrane. Vibrations of the outer Hensen's cell and the basilar membrane (BM) adjacent to and about 130 microm below the level of the Hensen's cell were measured. Outer Hensen's cell vibration at the characteristic frequency was up to 900 times higher compared to the BM amplitude. After sacrifice BM vibration increased while Hensen's cell vibration decreased. The magnitude and sequence of change after sacrifice can best be explained by the presence of negative feedback between reticular lamina and BM. In other experiments using ototoxic drugs that damage outer hair cells, similar changes in Hensen's cell and BM vibration were observed. These results show that the apical turn behavior is different from that observed by other investigators in the basal turn. The potential benefits of the negative feedback are discussed. The presence of negative feedback would explain the linearity at the fundamental frequency observed in the apical turn of cochlea.

Mechanical nonlinearity in the apical turn of the guinea pig organ of Corti.

Confocal microscopy was used to view the sealed apical turn of the cochlea in a living guinea pig, and to identify the cochlear structures through the intact Reissner's membrane. X, Y and Z coordinates for each point of interest were recorded. A confocal laser heterodyne interferometer measured the cellular vibration in response to acoustical signals applied to the ear. Velocity time waveforms were recorded at 32 frequencies between 25 and 2500 Hz at each point of measurement. To characterize the vibration pattern of the organ of Corti, vibrations at multiple locations along a radial track of the reticular lamina were measured before and after sacrificing the animal. Amplitude and phase tuning curves of the fundamental and the second harmonic, velocity time waveforms, and FFTs of time waveforms are compared before and after sacrifice. The results show that a sharply tuned nonlinear part of the response disappears shortly after sacrifice.

Vibrations of the guinea pig organ of Corti in the apical turn.

Vibrations of the organ of Corti were measured in response to sound applied to the ear in the apical turn of a living guinea pig. Measurements were made at 29 points on the Reissner's membrane (RM) at 10 micro spacing along a radial track. Measurements also included 22 points on the reticular lamina (RL), Claudius' cells and osseous spiral lamina. Our goal was to characterize the vibration of the RM and the RL with high spatial resolution along a radial axis. The tuning and spatial patterns of the RM are compared in the radial direction with those for the RL at the fundamental frequency and at the second harmonic. The shape of the RM tuning curve changes with radial position, and differ significantly from those observed at the RL. These results support our earlier findings (Hao and Khanna, Hear. Res. 99 (1996) 176-189).