Directional selectivity and frequency tuning of midbrain cells in the oyster toadfish, Opsanus tau.

Single-unit recordings were made from areas in the midbrain (torus semicircularis) of the oyster toadfish. We evaluated frequency tuning and directional responses using whole-body oscillation to simulate auditory stimulation by particle motion along axes in the horizontal and mid-sagittal planes. We also tested for bimodality in responses to auditory and hydrodynamic stimuli. One recording location in each animal was marked by a neurobiotin injection to confirm the recording site. Recordings were made in nucleus centralis, nucleus ventrolateralis, and the deep cell layer. Most units were frequency-selective with best frequencies between 50 and 141 Hz. Suppression of activity was apparent in 10% of the cells. Bimodality was common, including inhibition and suppression of background activity by auditory or hydrodynamic stimulation. The majority of the cells were directionally selective with directional response patterns that were sharpened compared with those of primary saccular afferents. The best directional axes were arrayed widely in spherical space, covering most azimuths and elevations. This representation is adequate for the computation of the motional axis of an auditory stimulus for sound source localization.

Directional encoding by fish auditory systems.

This paper reviews and discusses several investigations of the peripheral neural code for the directional axis of acoustical particle motion in the saccule of two fishes: goldfish (Carassius auratus) and toadfish (Opsanus tau). Most saccular afferents are directional in the manner of hair cells, having a cosine-shaped directional response pattern. The saccular sensory epithelia are orientated almost vertically in a parasagittal plane. In the horizontal plane, these epithelia are orientated obliquely with respect to the midline. Hair-cell stereocilia project perpendicularly. Thus, directional response patterns of saccular afferents tend to be orientated in azimuth parallel to the orientation of the epithelia in the head. The oblique angle of the toadfish saccule is greater than that of the goldfish, and the range of best directions in the horizontal plane for each species reflects those differing orientations. The azimuth of acoustical particle motion could be computed by comparing the relative activation of the two saccules, as is the case for the ears of most terrestrial vertebrates. The spatial patterns of saccular hair-cell orientation of most fishes thus appear to have little function in azimuthal source location, but for toadfish are probably most important for determining the elevation of monopole sources.

Dendritic arbors on the saccule and lagena in the ear of the goldfish, Carassius auratus.

The ear of the goldfish (Carassius auratus) contains three otolithic endorgans: the saccule, lagena, and utricle. The saccule has an auditory function in most teleost fishes for whom data are available, and there is evidence that the lagena is also an auditory endorgan in the goldfish. This study was conducted to compare the innervation of the saccule and the lagena to one another and to previously published data from goldfish and other species. We placed cobaltous-lysine in saccular and lagenar nerves in vivo and permitted uptake over 18-24 h. A total of 59 saccular and 59 lagenar dendritic arbors were labeled in 10 fishes. Our data indicate that arbors on the saccule and lagena have similar morphologies, but differ in relative size. Saccular arbors tend to be smaller than lagenar arbors, with median arbor widths of 50 micrometer on the saccule and 74 micrometer on the lagena. Fiber diameters on the two endorgans are similar. A regional analysis of the saccule indicated that a wide range of arbor sizes are found along the rostral-caudal axis, with larger arbors more common caudally. Our data do not support the presence of two distinct categories of saccular afferents with non-overlapping distributions. Moderate arbor widths (50-99 micrometer) were most common in all regions of the lagena. Maximum arbor width and hair cell density do not appear to be correlated with one another on either the saccule or the lagena. Comparisons with published data from goldfish and oscar revealed similarities and differences that may be attributable to variations in label uptake or transport as well as potential species differences.

Dendritic arbors and central projections of physiologically characterized auditory fibers from the saccule of the toadfish, Opsanus tau.

Neurobiotin was injected iontophoretically into saccular afferents of toadfish (Opsanus tau) after intracellular recording to examine dendritic arbors and central projections with respect to the physiological and directional response properties of the cells. Dendritic arbors of 36 afferents were examined in detail. Maximum diameter of the arbor and the number of terminal points were positively correlated with each other, but neither was predictive of spontaneous activity or sensitivity. Best azimuths were centered around 30 degrees -40 degrees, which corresponds to the angle of the saccule with respect to the fish's midline. In general, best elevations for afferents corresponded to hair cell orientations in the region innervated; unexpectedly low elevations obtained from afferents innervating the middle saccule may reflect curvature of the sensory epithelium against the otolith. Three efferent cells were filled partially. The location and large size of the efferent projections indicate that activity along the saccule could be modulated by a single efferent. All afferents projected to the dorsal zone of the descending octaval nucleus (dDON); many afferents bifurcated to terminate in the anterior octaval nucleus, and a few of those also had terminal fields in the medial zone of DON. All afferent projections into the dDON consisted of multiple axon collaterals projecting to numerous sites along the rostral-caudal extent of the nucleus. Variation in terminal field sites also was noted in the medial to lateral axis of the dDON; however, there were no consistent correlations between terminal field locations, physiology, and best directions of the saccular afferents.

Anatomical evidence for binaural processing in the descending octaval nucleus of the toadfish (Opsanus tau).

The connections of a potential auditory circuit were determined in the medulla of the toadfish (Opsanus tau). Fluorescent dextran amines placed in the medial torus semicircularis (mTS) retrogradely filled cells primarily in the dorsal region of the descending octaval nuclei (DON) with contralateral predominance. Fluorescent dextran amines placed in the DON revealed commissural fibers that cross the midline with the internal arcuate tract. The interconnections are consistent with a dorsal-ventral organization of the DON: reciprocal innervation is present for the left and right dorsal zones of the DON and for the left and right ventral zones of the DON. Based on projections to the medial (auditory) TS and the reciprocal connections, the dorsal region of the DON appears to be the major auditory processing site in the medulla and also may be a site for directional, binaural comparisons. The ventral region of the DON may be a site for bilateral vestibular processing. Double-labelling experiments revealed that some of the descending octaval cells projecting to the contralateral DON also project to the mTS. Based on the auditory pathway indicated by this study, future neurophysiological investigations of sensitivity to directional sound stimuli should begin in the dorsal DON of the toadfish.

Projections of primary afferents from regions of the saccule in toadfish (Opsanus tau).

The hair cell orientation patterns present on the saccules of fishes may be important for encoding the direction of a sound source. This study was conducted to determine whether primary afferent projections to the medulla are organized by the best directions for the hair cells they innervate. The toadfish saccule has hair cells oriented primarily in the vertical plane: both the rostral and the caudal saccule have hair cell orientations sweeping from 0 degrees to 45 degrees, and the middle saccule has hair cells oriented at 90 degrees. Fluorescent dextran amines were used singly and in combination to label regions of the saccular nerve innervating rostral, middle, and caudal saccule. The projections of those afferents were examined in detail in the anterior and descending octaval nuclei, which are auditory nuclei in this species. There was no evidence of topographic projections based on location along the length of the saccule or based on hair cell orientation. There was some evidence that parallel inputs are present from each region of the saccule examined, which may be based on the 180 degrees opposition of hair cells found throughout the saccule; however, afferents from the rostral, middle, and caudal saccule appear to have overlapping projections to the anterior and descending octaval nuclei. These data suggest that in toadfish, calculations of the direction of the sound source may begin in either of these primary auditory nuclei by comparing afferent input from along the saccule.

Diversity in frequency response properties of saccular afferents of the toadfish, Opsanus tau.

The frequency response of primary saccular afferents of toadfish (Opsanus tau) was studied in the time and frequency domains using the reverse correlation (revcor) method. Stimuli were noise bands with flat acceleration spectra delivered as whole-body motion. The recorded acceleration waveform was averaged over epochs preceding and following each spike. This average, termed the revcor, is an estimate of the response of an equivalent linear filter intervening between body motion and spike initiation. The spectrum of the revcor estimates the shape of the equivalent linear filter. Revcor responses were brief, damped oscillations indicative of relatively broadly tuned filters. Filter shapes were generally band-pass and differed in bandwidth, band edge slope, and characteristic frequency (74 Hz to 140 Hz). Filter shapes tend to be independent of stimulus level. Afferents can be placed into two groups with respect to characteristic frequency (74-88 Hz and 140 Hz). Some high-frequency afferents share a secondary peak at the characteristic frequency of low-frequency afferents, suggesting that an afferent may receive differently tuned peripheral inputs. For some afferents having similar filter shapes, revcor responses often differ only in polarity, probably reflecting inputs from hair cells oriented in opposite directions. The origin of frequency selectivity and its diversity among saccular afferents may arise from a combination of hair cell resonance and micromechanical processes. The resulting frequency analysis is the simplest yet observed among vertebrate animals. During courtship, male toadfish produce the 'boatwhistle' call, a periodic vocalization having several harmonics of a 130 Hz fundamental frequency. The saccule encodes the waveform of acoustic particle acceleration between < 50 and about 250 Hz. Thus, the fundamental frequency component of the boatwhistle is well encoded, but the successive higher harmonics are filtered out. The boatwhistle is thus encoded as a time-domain representation of its fundamental frequency or pulse repetition rate.

Directional response properties of saccular afferents of the toadfish, Opsanus tau.

The displacement sensitivity, frequency response, and directional response properties of primary saccular afferents of toadfish (Opsanus tau) were studied in response to a simulation of acoustic particle motion for which displacement magnitudes and directions were manipulated in azimuth and elevation. Stimuli were 50, 100, and 200 Hz sinusoidal, translatory oscillations of the animal at various axes in the horizontal and midsagittal planes. Thresholds in these planes defined a cell's characteristic axis (the axis having the lowest threshold) in spherical coordinates. Recordings were made from afferents in rostral, middle, and caudal bundles of the saccular nerve. The most sensitive saccular afferents responded with a phase-locked response to displacements as small as 0.1 nm. This sensitivity rivals that of the mammalian cochlea and is probably common to the sacculi and other otolith organs of most fishes. Most afferents showed lower thresholds at 100 Hz than at 50 or 200 Hz. Eighty percent of afferents have three-dimensional directional properties that would be expected if they innervated a group of hair cells having the same directional orientation on the saccular epithelium. Of the afferents that are not perfectly directional, most appear to innervate just two groups of hair cells having different orientations. The directional characteristics of afferents are qualitatively correlated with anatomically defined patterns of hair cell orientation on the saccule. In general, azimuths of best sensitivity tend to lie parallel to the plane of the otolith and sensory epithelium. Elevations of best sensitivity correspond well with hair cell orientation patterns in different regions of the saccular epithelium. Directional hearing in the horizontal plane probably depends upon the processing of interaural differences in overall response magnitude. These response differences arise from the gross orientations of the sacculi and are represented, in part, as time differences among nonspontaneous afferents that show level-dependent phase angles of synchronization. Directional hearing in the vertical plane may be derived from the processing of across-afferent profiles of activity within each saccule. Fishes were probably the first vertebrates to solve problems in sound source localization, and we suggest that their solutions formed a model for those of their terrestrial inheritors.

Hair cell orientation patterns on the saccules of juvenile and adult toadfish, Opsanus tau.

Saccules from 10 adult (five female, five male) toadfish (Opsanus tau) 13.5-26 cm standard length, were examined for individual/sexual variation in the hair cell orientation pattern. In addition, saccules from two juveniles (5 and 6 cm standard length) were compared with those of the adults to determine whether maturational differences exist in the hair cell orientations. The hair cell orientation pattern is unlike any reported previously for this species or its congener, O. beta. There are no major differences between the hair cell orientations of males and females, nor between the juveniles and the adults. A slight individual variation is present in the proportion of hair cells oriented in a particular direction in a specific area of the sensory epithelium. Potential ramifications of this hair cell orientation pattern are discussed with regard to development and auditory processing.

Structural diversity in the inner ear of teleost fishes: implications for connections to the Mauthner cell.

A body of literature suggests that the Mauthner cell startle response can be elicited by stimulation of the ear. While we know that there are projections to the M-cell from the ear, the specific endorgan(s) of the ear projecting to the M-cell are not known. Moreover, there are many reasons to question whether there is one pattern of inner ear to M-cell connection or whether the endorgan(s) projection to the M-cell varies in species that have different hearing capabilities of hearing structures. In this paper, we briefly review the structure of fish ears, with an emphasis on structural regionalization within the ear. We also review the central projections of the ear, along with a discussion of the limited data on projections to the M-cell.

The effects of ICI 182,780, a pure anti-oestrogen, on the hypothalamic-pituitary-gonadal axis and on endometrial proliferation in pre-menopausal women.

ICI 182,780 has shown pure oestrogen antagonism in vitro and in vivo in animals. A total of 17 women with normal menstrual cycles were administered ICI 182,780, 12 mg daily for 7 days in the follicular phase prior to hysterectomy; 11 normal women were used as controls. Of the 17 patients, three (18%) experienced a luteinizing hormone (LH) surge in the treatment group compared with five (45%) in the controls (P = 0.24), and these patients were only included up to the surge. There were no differences in the daily mean plasma LH and follicle stimulating hormone concentrations between the treatment (n = 17) and control (n = 10) groups. The mean plasma oestradiol was higher in the treatment group than controls (P < 0.05) on days 5, 6 and 7. However, there was no increase in endometrial thickness in the treatment group throughout the study. In the control group, endometrial thickness increased during the study and was significantly higher (P < 0.05) on day 7. There was no ultrasonic evidence of ovarian hyperstimulation and no serious adverse events reported. This study shows that treatment for 7 days with ICI 182,780 does not cause ovarian hyperstimulation and has a potent anti-oestrogenic action on the endometrium. We conclude that ICI 182,780 may be a useful compound in the treatment of oestrogen-dependent gynaecological disease.

Alterations in the distribution and activity of transglutaminase during tumour growth and metastasis.

Transglutaminase activity and subcellular distribution have been examined in both normal and tumour tissue. Subcellular fractionation of rat liver demonstrated a bimodial distribution for transglutaminase between the particulate (approximately 40%) and cytosol (approximately 60%) fractions. Isolation of enriched plasma membrane fractions indicated the presence of membrane associated transglutaminase activity which co-distributed with that of 5'-nucleotidase and Na+/K+-ATPase. Induction of hepatocellular carcinomas in rats by treatment with either diethylnitrosamine or 6-p-dimethylaminophenylazobenzothiazole resulted in a reduction in transglutaminase activity which was accompanied by redistribution of the enzyme to the particulate fraction of the cell. The tumour bearing liver appeared to represent an intermediate stage between the hepatocellular carcinoma and control liver when assayed for content and distribution of transglutaminase activity. The transglutaminase activity of four transplantable rat sarcomas (P7, P8, MC3 and CC5) was found to be greatly reduced when compared with the normal tissues of rat liver, lung and spleen. A further reduction in this activity occurred in the primary growths of the sarcomas P7 and P8 following detection of metastases. Our data suggest that such changes in the distribution and content of transglutaminase may be a feature of tumour tissue and may be of value in both monitoring and investigating the carcinogenic process.

Effects of in vitro incorporation of cholesterol and cholesterol analogues into rat platelets.

Cholesterol and analogues of cholesterol bearing shorter side chains were incorporated into rat platelet membranes by incubation with sterol-rich liposomes in vitro. Cholesterol-enriched platelets showed increased aggregability to collagen compared with controls. Platelets containing the cholesterol analogues pregn-5-en-3 beta-ol and chol-5-en-3 beta-ol were even more sensitive to aggregation and could aggregate spontaneously on stirring. The size of the platelets containing pregn-5-en-3 beta-ol was markedly reduced when compared with controls in the scanning electron microscope. The results suggest that the sterol content and structure of the platelet membrane can have a critical role in maintaining the normal function of the cell.