John W. Conlee

Brainstem auditory evoked potentials correlate with morphological changes in Gunn rat pups

The relationship of brainstem structure and function in bilirubin encephalopathy is incompletely understood. The present experiments compare quantitative measures of brainstem structures with brainstem auditory evoked potentials (BAEPs) in infant jaundiced (jj) and nonjaundiced (Nj) Gunn rats. Ten jjs from 4 litters were injected with sulfadimethoxine at 11-12 days of age to raise their brain bilirubin concentration. Littermate controls were jjs given saline, and Njs given sulfadimethoxine or saline. At 15-17 days of age BAEPs were recorded, and rats were prepared for histological examination, as was reported in the previous paper (Conlee and Shapiro, 1991). Significant differences between groups were seen for BAEP wave I latency (P = 0.002). I-II interwave interval (P = 0.001), and amplitudes of waves I, II, III, and IV (each P less than 0.0005) due to increased latencies and decreased amplitudes in the jj-sulfa group. Animals with the most severe BAEP abnormalities had the most severe histological abnormalities. Cochlear nucleus volume had a positive linear correlation with the amplitude of BAEP waves I, II, and IV, and an inverse correlation with wave I latency and I-II interwave interval (P less than or equal to 0.001). The highest correlations were BAEP I-II interwave interval and amplitude of waves I and II with cochlear nucleus volume (r = -0.78, 0.71 and 0.70, respectively, P less than 0.0005).(ABSTRACT TRUNCATED AT 250 WORDS)

Morphological changes in the cochlear nucleus and nucleus of the trapezoid body in gunn rat pups

Mechanisms underlying bilirubin encephalopathy and hearing loss remain poorly understood, including the way bilirubin enters the nervous system and how bilirubin accumulates in circumscribed regions of the brain. The present experiments examined the auditory brainstem in heterozygous (Nj) and homozygous (jj) Gunn rats at an age when serum bilirubin levels were highest, and after brain bilirubin concentration was artificially raised by sulfadimethoxine administration. In four litters of 11-12 days old Gunn rats, Nj and jj littermates received a single intraperitoneal injection of sulfadimethoxine (100 mg/kg) or a comparable volume of saline. At 16-17 days of age, brainstem auditory evoked potentials were recorded to assess the severity of bilirubin toxicity in the Nj and jj animals. Following the recordings, each animal was perfusion-fixed and frozen sections of the brainstem were cut in the transverse plane from medullary through mesencephalic levels. Sections were mounted on slides, stained with thionin and coded to avoid observer bias. Quantitative analysis revealed no differences between saline and sulfa-treated Nj rats for cochlear nucleus volume, or for cell size in the cochlear nucleus or superior olive. In the sulfa-treated jj rats, cochlear nucleus volume, and cross-sectional areas of spherical cells in the anteroventral cochlear nucleus and principal cells in the nucleus of the trapezoid body, were all significantly smaller than in the combined groups of Nj animals. The affected areas in the cochlear nucleus and superior olive are innervated by large axosomatic end-bulbs of Held or calyceal endings, and were associated with bilirubin staining of glia in the most severely jaundiced jj sulfa-treated rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of aging on normal hearing loss and noise-induced threshold shift in albino and pigmented guinea pigs

In a previous investigation into noise-induced hearing loss by comparing 2-month-old albino with pigmented guinea pigs, albinos displayed significantly greater shifts in cochlear microphonic (CM) threshold and less recovery than the pigmented animals 7 days after noise exposure. The present study compared the responses of 14-month-old albino and pigmented guinea pigs to the same noise parameters used previously. Thresholds for the first detectable elicitation of CM for three pure tones were recorded prior to, at 90 min and at 7 days after a 45-min exposure to 126 dB broadband noise. Before exposure to noise, thresholds for pigmented guinea pigs were 24 dB higher than those in the albinos. Following noise exposure, the pigmented animals showed less than half the amount of threshold shift displayed by the albinos. This change ws attributed to the higher pre-exposure thresholds in the pigmented guinea pigs. Converging lines of evidence suggest that cochlear pigmentation may have both protective and toxic influences on the inner ear.

Differential Effects of Gentamicin on the Distribution of Cochlear Function in Albino and Pigmented Guinea Pigs

It has been suggested that the high affinity of melanin pigment for aminoglycoside antibiotics may cause these drugs to bind preferentially to the pigmented inner ear, producing greater ototoxicity than in the amelanotic albino cochlea. However, evidence of greater ototoxicity in albinos has led to the hypothesis that melanin inhibits the toxicity of these drugs in the pigmented inner ear. On the other hand, ototoxicity in the pigmented animals may simply be delayed relative to the albinos, only to become equal or even more severe with time. The present study was conducted to determine whether a relatively low dose of gentamicin (68.5 mg/kg) would produce differential ototoxicity between albino and pigmented guinea pigs which would persist long after drug exposure had stopped. Nine pigmented and eight albino guinea pigs were given gentamicin sulfate for 14 consecutive days, and were then allowed a two-month recovery period before cochlear analysis; 11 pairs of saline-injected or untreated albino and pigmented guinea pigs served as controls. The results showed that the gentamicin-treated albinos had significantly elevated thresholds for the compound action potential from the auditory nerve (CAP), and significantly lower endocochlear potentials (EP) and cochlear microphonic (CM) input-output voltage functions when compared to their respective controls, or to either group of pigmented guinea pigs. The CAP in drug-treated pigmented animals did not differ significantly from controls, and the differences in EP and CM were marginally significant. The results indicate that the pigmented cochlea is less susceptible to gentamicin than the albino cochlea, and support the hypothesis that melanin may inhibit aminoglycoside ototoxicity in the pigmented inner ear.

Ongoing proliferation of melanocytes in the stria vascularis of adult guinea pigs

The intermediate cells of the stria vascularis are melanocytes derived from the neural crest. These internalized pigment cells have been thought to be a static population, distinct from those found in the skin. However, this investigation demonstrates that the melanocytes of the adult stria vascularis undergo continuous replication. Cell proliferation was studied using [3H]-thymidine autoradiography and bromodeoxyuridine (BrdU) immunohistochemistry. Single or multiple injections of [3H]-thymidine within a six hour period labeled a mean of 9 intermediate cells. In pigmented guinea pigs, single daily injections of [3H]-thymidine for 2 or 4 days labeled a mean of 24 and 69 intermediate cells, respectively; Pigmented guinea pigs given BrdU once daily for 2 or 4 days labeled a mean of 38 and 75 intermediate cells. By contrast, albino littermates also given BrdU averaged only 23 and 42 labeled intermediate cells in the same 2 and 4 day experiments. The mean number of proliferating cells/mm of stria per 24 h was 1.54 in the pigmented animals and 0.88 in the albinos. Both the total number and density of labeled intermediate cells were significantly smaller in the albino than the pigmented guinea pigs. These results demonstrate that the melanocytes in the stria vascularis undergo continuous baseline mitosis, and at a rate comparable to the melanocytes of the skin. This surprising similarity promotes the speculation that the proliferative rate of the strial melanocytes may be influenced by some of the same factors known to affect replication and pigment production in the skin.

Turn-specific and pigment-dependent differences in the stria vascularis of normal and gentamicin-treated albino and pigmented guinea pigs

The aims of the present study were to determine which structures in the stria vascularis (SV) may depend upon the presence of pigmented melanocytes both for normal morphology and for the expression of gentamicin ototoxicity in the inner ear. These pigment-dependent influences were inferred through comparisons of the SV in pigmented guinea pigs and in albinos containing nonpigmented melanocytes. Results were obtained from 6 albino and 8 pigmented guinea pigs given gentamicin, and from 3 albino and 3 pigmented control animals not receiving the drug. One-month old animals received gentamicin daily (100 mg/kg) for 14 days and recovered for an additional 14 days before being prepared for electron microscopy. The SV from each of the 4 cochlear turns was analyzed using stereological point counting procedures. In control animals, differences were found in the higher cochlear turns, where volume density for the marginal cells in albinos was abnormally large (turns 3 and 4), while the volume density for intermediate cells (melanocytes) was abnormally small (turn 3). Cell volume estimates for the intermediate cells were significantly smaller in the albino than pigmented control animals in the higher cochlear turns, indicating that functional abnormalities may be found in the albino cochlea. In animals exposed to gentamicin, marginal cell volume density was reduced significantly in turn 4 of albinos, but not in any region of the pigmented inner ears. Radial area of SV and estimates of the absolute volumes for marginal cells in albinos given gentamicin also were significantly reduced in turn 1 compared to their controls; such differences were not observed in the pigmented animals. The results indicate that marginal cell size is significantly reduced in albino but not pigmented animals 14 days after gentamicin exposure, and further suggest a role of pigmented melanocytes in ameliorating gentamicin-induced cochlear damage.

Pigment-dependent differences in the stria vascularis of albino and pigmented guinea pigs and rats

Functional models of the stria vascularis (SV) have ascribed roles for the marginal and basal cells, but not for the intermediate cells, which remain poorly understood. Intermediate cells have been identified as melanocytes, which produce melanin in most pigmented animals including humans. The relationship of melanin to intermediate cell function may be addressed through comparisons with the albino inner ear. Albinos have a normal distribution of melanocytes that are unable to synthesize melanin pigment. In the present study, the SV was compared between albino and pigmented littermates in both the guinea pig and the rat. Photomicrographic montages of the SV were analyzed from each of 7 cochlear regions in the guinea pig and 5 regions in the rat. Stereological procedures were used to determine the volume density (Vv) for each of the three main cell types in the stria, the surface density (Sv) of the marginal cells, and to derive estimates of absolute cell volume and surface area. In the guinea pig, comparisons between pigment groups showed that marginal cell Vv was larger across cochlear turns in the albinos, while intermediate cell Vv was smaller. Intermediate cell cytoplasmic and total cell volumes were smaller in the albino guinea pigs; however, marginal cell Sv and absolute area were larger. In the rat, intermediate cell Vv was alos smaller across cochlear turns in the albinos. Similarly, intermediate cell cytoplasmic and total cell volumes were smaller in the albinos, while marginal cell total surface area per radial cross-section of the SV was larger. These results demonstrate that amelanotic melanocytes occupy significantly less volume than do pigmented melanocytes, and suggest that melanin may influence the structure and function of the SV.

Axon-target cell interactions in the developing auditory system.

Publisher Summary A number of significant developmental interactions between axons and their targets have been identified: for example, the role of afferent axons in shaping dendritic trees has been most clearly revealed for the Purkinje and basket/stellate neurons of the cerebellum, and the influence of target cells on neuronal survival has been studied most extensively in the chick spinal cord. Developmental regulation of the number of afferent axons converging on a target neuron has been most frequently examined in the peripheral nervous system (PNS), and the contributions of sensory experience to brain development have largely been revealed by work on the visual system of the cat. To obtain a more comprehensive view of these processes and their interrelationships, this chapter examines a variety of interactions between axons and their targets during the early life of certain auditory neurons in the avian brain stem. The striking structural and functional specializations of brain stem auditory neurons make them attractive subjects for studies of cellular interactions during the development of the central nervous system (CNS). The developing brain stem auditory nuclei of the chick embryo, in particular, are remarkable for the relative simplicity of their organization and their accessibility to the experimenter. The chapter presents the results of experimental studies involving the avian auditory system and summarizes recent work on its normal organization and development.

Differential rearing affects responsiveness of rats to depressant and convulsant drugs

The effects of postweaning housing in environmental complexity (EC) or isolation cage (IC) conditions upon the development of anesthesia following sodium pentobarbital administration and upon seizure susceptibility following metrazol administration were studied in separate groups of rats. Male and female EC rats lost the righting reflex and developed unconsciousness more rapidly following sodium pentobarbital injection (105 mg/kg, IP) than did littermate IC rats. Male EC and IC rats did not differ in the CD50 (50% convulsion dosage by log probit analysis) following injection of a range of metrazol doses (20 to 35 mg/kg, IP) when they were kept in a quiet dimly lighted room. However, the addition of stroboscopic light stimulation (1-100 Hz) to the above procedure (doses 17.5 to 30 mg/kg) reduced the CD50 for the IC rats but did not affect the CD50 for the EC rats. The results support the view that brain excitability is lower in animals reared in environments providing higher levels of sensory stimulation.

Turn-specific differences in the endocochlear potential between albino and pigmented guinea pigs

Recent findings indicate that structural differences exist in the stria vascularis (SV) between albino and pigmented guinea pigs. In the higher cochlear turns, volume density for marginal cells in the albino SV is abnormally large, while that for intermediate cells (melanocytes) is abnormally small. These anatomical variations suggest that functional differences between albino and pigmented inner ears also may be found. To examine this possibility, four strains of guinea pigs were studied, consisting of Hartley albino (N = 9) and NIH outbred pigmented (N = 15) guinea pigs, as well as albino (N = 11) and pigmented (N = 15) guinea pig siblings born to mixed litters. Tracheotomy and carotid artery cannulation were performed. Animals were mechanically ventilated, with periodic samples drawn for arterial blood gas analysis. Blood pressure, heart rate and rectal temperature were monitored. Compound action potentials were measured first to assess cochlear viability. Positive endocochlear potentials (+EP) then were recorded, beginning with the fourth turn, followed by the first, second and third turns. Results showed that the +EP in albinos remained relatively constant across cochlear turns, but decreased significantly from base to apex in the pigmented inner ears. Across all animals, mean +EPs (mV +/- S.E.M.) for turns 1-4 in albinos were: 72.5 (2.5), 68.7 (2.3), 59.2 (2.7), 68.1 (3.3); pigmented values were: 72.9 (2.9), 66.9 (2.6), 53.8 (3.0), 57.0 (2.7). One-way ANOVAs did not show a significant difference in albino +EPs between any of the cochlear turns, but did indicate a highly significant difference between turns in the pigmented inner ears (P < 0.000004). Post hoc comparisons demonstrated +EPs in turns 3 and 4 were smaller than in turn 1. Since turn 3 was recorded last in these experiments, and was reduced in value relative to turn 4 in both groups, it is likely that cochlear deterioration contributed to this result more than any other factor. These results, combined with previous anatomical data, indicate that a diminution of melanocyte cell volume in the albino SV is accompanied by an increase in marginal cell volume density and larger +EPs in the higher cochlear turns, at least at resting levels.