Organization of the inferior colliculus of the gerbil (Meriones unguiculatus): projections from the cochlear nucleus.

Projections from the cochlear nuclear complex to the inferior colliculus in the gerbil (Meriones unguiculatus) were studied using anterograde tracing methods based on axonal transport. Methods were developed to map the results onto comparable sets of sections through the inferior colliculus so that the patterns of termination in different animals could be compared directly. Projections to the contralateral inferior colliculus are widespread and most, if not all of them, are topographically organized. Axons terminate throughout the central nucleus and also in at least three distinct regions outside the central nucleus: a caudomedial region in the dorsal cortex, the ventrolateral nucleus and the rostral pole nucleus. Projections from the dorsal and ventral cochlear nuclei appear to overlap almost completely, although those from the dorsal cochlear nucleus may be slightly more widespread at the boundaries of the central nucleus. Projections from the ipsilateral cochlear nuclei arise in both the dorsal and ventral divisions and are largely restricted to the dorsal (low-frequency) part of the inferior colliculus. In this region, the pattern of ipsilateral and contralateral projections is similar, although the terminal fields from the two sides do not appear to overlap completely. The methods developed to display the results form a framework for comparisons with the distribution of inputs from the other major sources of input to the inferior colliculus.

Improved method for the rapid determination of terpenoid aldehydes in cotton.

A simple, rapid method for the extraction of cotton terpenoid aldehydes from green tissues and seed is described. Samples were treated by ultrasonification with acidified acetonitrile/water followed by centrifugation. The resulting extract was injected directly onto a C(18) HPLC column and no sample concentration or further cleanup steps were required. The level of gossypol, the most labile of the target analytes, decreased by only 2% after 12 h of storage at room temperature, thus enabling automated analysis of individual terpenoid aldehydes by HPLC. The method gives excellent reproducibility and enables large numbers of samples to be screened quickly and accurately.

AMPA receptor binding in adult guinea pig brain stem auditory nuclei after unilateral cochlear ablation.

This study determined if an asymmetric hearing loss, due to unilateral cochlear ablation, could induce the regulation of intracellular AMPA receptors in brain stem auditory nuclei. In young adult guinea pigs, the high-affinity specific binding of [(3)H]AMPA was measured in the cochlear nucleus (CN), the superior olivary complex (SOC), and the auditory midbrain at 2-147 postlesion days. After correction for tissue shrinkage, changes in specific binding relative to that in age-matched unlesioned controls were interpreted as altered numbers and/or activity of intracellular AMPA receptors. In the CN, transient elevations and/or deficits in binding were evident in most regions, which usually recovered by 147 days. However, persistently deficient binding was evident ipsilaterally in the anterior part of the anteroventral CN (AVCNa). In the SOC, transient elevations in binding were evident at 2 days in the medial limb of the lateral superior olive (LSOmed) and the medial superior olive. Between 7 and 147 days, most SOC nuclei exhibited transient, temporally synchronized postlesion deficits in binding. However, late in the survival period, deficits persisted ipsilaterally in the LSOmed and the lateral (LSOlat) limb of the lateral superior olive. In the midbrain, transient elevations and/or deficits in binding were evident in the dorsal nucleus of the lateral lemniscus as well as in the central and dorsal nucleus of the inferior colliculus. A persistent deficit was evident in the intermediate nucleus of the lateral lemniscus. The findings implied that auditory neurons contain regulatory mechanisms that control the numbers and/or activity of intracellular AMPA receptors. Regulation was induced by cochlear nerve destruction and probably by changes in the excitation of glutamatergic neurons. Many of the regulatory changes were transient, except in the ipsilateral AVCNa and LSO, where postlesion downregulations were persistent. The downregulation in the ipsilateral AVCNa was probably induced directly by the loss of cochlear nerve endings. However, other regulatory changes may have been induced by signals carried on pathways emerging from the ipsilateral CN and on centrifugal auditory pathways.

Altered glycinergic synaptic activities in guinea pig brain stem auditory nuclei after unilateral cochlear ablation.

This paper reviews efforts to determine if a unilateral hearing loss altered inhibitory glycinergic synapses in the cochlear nucleus (CN) and the superior olive. In young adult guinea pigs, 2-147 days after unilateral cochlear ablation, we quantified the electrically evoked release and the high-affinity uptake of [(14)C]glycine as measures of transmitter release from glycinergic presynaptic endings and glycine removal from extracellular spaces. The specific binding of [(3)H]strychnine was quantified to measure synaptic glycine receptor activity and/or expression. Three types of post-lesion change were observed. First, several tissues exhibited changes consistent with a persistent deficiency in glycinergic inhibitory transmission. Deficient binding prevailed on the ablated side in the anterior and caudal anteroventral CN, the posteroventral CN and the lateral superior olive (LSO), while glycine release was near normal and uptake was elevated (except in the LSO). However, deficient release prevailed in the dorsal CN, bilaterally, and was accompanied by elevated uptake. Second, the LSO on the intact side exhibited changes consistent with strengthened glycinergic inhibition, as binding was elevated while release and uptake were near normal. Third, several tissues exhibited various transient changes in activity. These types of post-lesion change might contribute to altered auditory functions, which often accompany hearing loss.

Plastic changes in glycine and GABA release and uptake in adult brain stem auditory nuclei after unilateral middle ear ossicle removal and cochlear ablation.

[i] In young adult guinea pigs, the effects of unilateral ossicle removal and unilateral cochlear ablation were determined on [14C]glycine or [14C]GABA release and uptake measured in subdivisions of the cochlear nucleus (CN), the superior olivary complex, and the auditory midbrain, after 2 or 5, 59, and 145 postlesion days. Activities were compared to those of age-matched, unlesioned controls. [ii] [14C]Glycine release declined bilaterally in the anteroventral and dorsal CN after ossicle removal and in the dorsal CN after cochlear ablation. [iii] Transient elevations of release occurred at 59 days in the ipsilateral posteroventral CN ([14C]glycine) and bilaterally in the ventral nucleus of the lateral lemniscus ([14C]GABA) after ossicle removal, and bilaterally in the medial superior olive ([14C]glycine) after cochlear ablation. [iv] In the medial nucleus of the trapezoid body, [14C]GABA release was depressed bilaterally 5 days after ossicle removal, but was elevated at 145 days contralaterally after ossicle removal and ipsilaterally after cochlear ablation. [v] In the contralateral central nucleus of the inferior colliculus, [14C]GABA release was elevated persistently after ossicle removal. After cochlear ablation, release was elevated at 5 days, near the control at 59 days, and elevated again at 145 days. [vi] After both lesions, [14C]glycine uptake was elevated bilaterally in the CN and medial superior olive. [14C]GABA uptake became depressed by 59 or 145 days bilaterally in the auditory midbrain. [vii] These changes may stem from regulation and may contribute to mechanisms that generate symptoms such as loudness recruitment and tinnitus, which often accompany hearing loss.

Glycine receptors in adult guinea pig brain stem auditory nuclei: regulation after unilateral cochlear ablation.

In young adult guinea pigs, the effects of unilateral cochlear ablation were determined on the specific binding of [3H]strychnine measured in subdivisions of the cochlear nucleus (CN), the superior olivary complex, and the auditory midbrain, after 2, 7, 31, 60, and 147 postlesion days. Changes in binding relative to that in age-matched controls were interpreted as altered activity and/or expression of synaptic glycine receptors. Postlesion binding declined ipsilaterally in most of the ventral CN and in the lateral superior olive (LSO). Binding was modestly deficient in the ipsilateral dorsal CN and in the anterior part of the contralateral anteroventral CN. Binding was elevated in the contralateral LSO. Transient changes also occurred. Binding was elevated transiently, between 2 and 31 days, contralaterally in parts of the anteroventral CN, bilaterally in the medial superior olive (MSO), and bilaterally in most of the midbrain nuclei. Binding was deficient transiently, at 60 days, in most of the contralateral CN and bilaterally in the midbrain nuclei. The present findings, together with previously reported postlesion changes in glycine release, were consistent with persistently weakened glycinergic inhibitory transmission ipsilaterally in the ventral CN and the LSO and bilaterally in the dorsal CN. Glycinergic inhibitory transmission was strengthened in the contralateral LSO and transiently strengthened in the MSO bilaterally. A hypothetical model of the findings suggested that glycine receptor regulation may depend on excitatory and glycinergic input to auditory neurons. The present changes in glycine receptor activity may contribute to altered auditory functions, which often accompany hearing loss.

Regulation of D-aspartate release and uptake in adult brain stem auditory nuclei after unilateral middle ear ossicle removal and cochlear ablation.

In young adult guinea pigs, the effects of unilateral ossicle removal and cochlear ablation were determined on transmitter release from glutamatergic presynaptic endings and glutamate inactivation via uptake. (i) D-[3H]Aspartate release and uptake were measured in subdivisions of the cochlear nucleus (CN) and in nuclei of the superior olive (SOC) and auditory midbrain (MB) up to 145 days after placing the lesions. Activities were compared to those from age-matched unlesioned controls. Fiber degeneration was visualized histologically. (ii) In the ipsilateral CN, changes in release and uptake were governed by the type of lesion. Ossicle removal produced sparse pruning of fibers only after 112 days and decreased release and uptake at 145 days, consistent with regulatory weakening of excitatory glutamatergic transmission. Cochlear ablation deafferented the CN, producing deficient release and uptake at 2 days and abundant fiber degeneration at 7 days. Subsequently, the residual release and uptake increased in magnitude, consistent with strengthening of excitatory glutamatergic transmission. (iii) In the contralateral CN, after either lesion, changes in release and uptake usually matched those in the ipsilateral CN. Thus, the auditory pathway associated with the lesioned ear probably provided cues for the regulation of synaptic strength in the contralateral CN. (iv) Both lesions increased release in the SOC and MB, and uptake in the SOC, consistent with strengthening of excitatory glutamatergic transmission. Sparse fiber degeneration, suggesting axonal pruning, appeared in the SOC and MB after cochlear ablation. (v) The strengthening of excitatory glutamatergic transmission may facilitate and maintain symptoms such as loudness recruitment and tinnitus which often accompany hearing loss.

GABA- and glycine-immunoreactive projections from the superior olivary complex to the cochlear nucleus in guinea pig.

Retrograde transport of horseradish peroxidase was combined with immunocytochemistry to identify the origins of potential gamma-aminobutyric acid (GABA) -ergic and glycinergic inputs to different subdivisions of the cochlear nucleus. Projection neurons in the inferior colliculus, superior olivary complex, and contralateral cochlear nucleus were examined, but only those from the superior olivary complex contained significant numbers of GABA- or glycine-immunoreactive neurons. The majority of these were in periolivary nuclei ipsilaterally, with a sizeable contribution from the contralateral ventral nucleus of the trapezoid body. Overall, 80% of olivary neurons projecting to the cochlear nucleus were immunoreactive for GABA, glycine, or both. Most glycine-immunoreactive projection neurons were located ipsilaterally, in the lateral and ventral nuclei of the trapezoid body and the dorsal periolivary nucleus. This suggests that glycine is the predominant neurotransmitter used by ipsilateral olivary projections. Most GABA-immunoreactive cells were located bilaterally in the ventral nuclei of the trapezoid body. The contralateral olivary projection was primarily GABA-immunoreactive and provided almost half the GABA-immunoreactive projections to the cochlear nucleus. This suggests that GABA is the predominant neurotransmitter used by contralateral olivary projections. The present results suggest that the superior olivary complex is the most important extrinsic source of inhibitory inputs to the cochlear nucleus. Individual periolivary nuclei differ in the strength and the transmitter content of their projections to the cochlear nucleus and may perform different roles in acoustic processing in the cochlear nucleus.

Synaptophysin immunoreactivity in the cochlear nucleus after unilateral cochlear or ossicular removal.

This study determined if unilateral cochlear removal in adult guinea pigs led to synaptic loss followed by synaptogenesis in the cochlear nucleus (CN) and if unilateral middle ear ossicle removal led to synaptic loss in the CN. Synaptic endings were identified immunohistochemically, using a monoclonal antibody to synaptophysin. Immunolabeling was quantified densitometrically in the CN 4-161 days after cochlear removal and 161 days after ossicle removal. Fiber degeneration was visualized with the Nauta-Rasmussen silver method. Tissue shrinkage was measured from drawings of CN sections. Compared to the contralateral side, immunolabeling density ipsilaterally was reduced by 4 days in the anterior division of the anteroventral CN (a-AVCN) and by 7 days in the anterior part of the posteroventral CN (a-PVCN). At 7 days, preterminal fiber degeneration was abundant in both areas. These findings were consistent with the loss of cochlear nerve endings and fibers. At later times, immunolabeling density recovered. In the a-AVCN, tissue shrinkage explained approximately half the recovery of staining density; the rest was attributed to synaptogenesis. In the a-PVCN, the entire recovery was attributed to tissue shrinkage. In the polymorphic layer of the dorsal CN, immunostaining density increased transiently at 4 days, while at 7 days preterminal fiber degeneration was abundant. A net loss of synaptic endings was not detected immunohistochemically. The increased immunostaining density may reflect a transient growth of immature processes or presynaptic endings. Ossicle removal produced a deficit in immunolabeling density only in the ipsilateral a-PVCN, without fiber degeneration, suggesting a loss of presynaptic endings or of synaptophysin expression.

Uptake and release of D-aspartate, GABA, and glycine in guinea pig brainstem auditory nuclei.

This study attempts to determine if the medial (MSO) and lateral superior olive (LSO), medial nucleus of the trapezoid body (MNTB), ventral nucleus of the lateral lemniscus (VNLL), and central nucleus of the inferior colliculus (ICc) contain glutamatergic synaptic endings. Micropunch and microdissection procedures provided fresh samples of these auditory nuclei for the measurement of the high-affinity uptake and electrically evoked release of exogenous D-[3H]ASP. The study also determined if the LSO and MSO contain glycinergic synaptic endings by measuring uptake and release of [14C]Gly in these nuclei, and whether the MNTB, VNLL, and ICc contain GABAergic endings by assessing the uptake and release of [14C]GABA in these structures. Several strategies optimized the evoked Ca(2+)-dependent release of the labeled amino acids. These included the enhancement of high-affinity uptake during loading of the markers into the tissues, inhibition of uptake during the subsequent measurement of release, and use of an electrical stimulus current that evoked maximal Ca(2+)-dependent release. Each of these nuclei manifested the high-affinity uptake and the evoked Ca(2+)-dependent release of D-[3H]Asp, suggesting the presence of synaptic endings that may use Glu or Asp as a transmitter. Similar findings suggest the presence of glycinergic synaptic endings in the LSO and MSO, and of GABAergic synaptic endings in the MNTB, VNLL, and ICc.

Evidence for glutamatergic projections from the cochlear nucleus to the superior olive and the ventral nucleus of the lateral lemniscus.

This study attempts to determine if projections ascending from the guinea pig cochlear nucleus (CN) could be glutamatergic and/or aspartatergic. Multiple radio frequency lesions were made to ablate the right CN. The ablation was verified histologically. To identify the principal targets of CN efferents, silver impregnation methods were used to localize the preterminal degeneration of fibers in transverse sections of the brainstem 5 and 7 days after CN ablation. CN efferents projected heavily to the lateral superior olive (LSO) ipsilaterally, the medial superior olive (MSO) bilaterally, and contralaterally to the medial (MNTB) and ventral (VNTB) nuclei of the trapezoid body, the ventral (VNLL) and intermediate nuclei of the lateral lemniscus and the central nucleus of the inferior colliculus (ICc). There were smaller projections to the lateral nucleus of the trapezoid body ipsilaterally, the dorsal and dorsomedial periolivary nuclei bilaterally, and the dorsal nucleus of the lateral lemniscus contralaterally. There were sparse projections to the VNLL and ICc ipsilaterally and the CN contralaterally, and a very sparse projection to the contralateral LSO. To determine if CN efferents were glutamatergic and/or aspartatergic, the fresh brainstem was sectioned transversely and samples of the LSO, MSO, MNTB, VNLL, and ICc were taken to measure the electrically evoked release and the uptake of D-[3H]Asp and [14C]Gly or [14C]GABA 3-5 days after the CN ablation. The release studies suggest that only certain of the histologically identified projections ascending from the CN may be glutamatergic and/or aspartatergic. CN ablation depressed D-[3H]Asp release in the MSO bilaterally and in the contralateral MNTB and VNLL, suggesting that the CN efferents to these nuclei may use glutamate or aspartate as a transmitter. It was unclear whether a marginal depression of D-[3H]Asp release in the ipsilateral LSO reflected the presence of glutamatergic CN projections to this nucleus. D-[3H]Asp release in the ICc was unaffected, suggesting that CN efferents to this nucleus may not be glutamatergic. There were no deficits in D-[3H]Asp uptake. [14C]Gly release from the LSO and MSO was unchanged. [14C]Gly uptake was unchanged in the MSO and depressed only in the contralateral LSO, possibly reflecting subnormal uptake activity in endings contributed by contralateral MNTB cells that had lost their CN efferents.(ABSTRACT TRUNCATED AT 400 WORDS)

Evidence for a GABAergic projection from the dorsal nucleus of the lateral lemniscus to the inferior colliculus.

This study attempts to determine whether the pathways from the guinea pig dorsal nucleus of the lateral lemniscus (DNLL) to the inferior colliculus (IC) use gamma-aminobutyric acid (GABA) as a transmitter. Injections of kainic acid (KA) were used to destroy neurons in the left DNLL. Two to 4 days after the injection, Nissl-stained sections through the lesion site showed destruction of the DNLL neurons. The lesions varied in size; 12-100% of the DNLL neurons were destroyed on the injected side without damage to the ipsilateral IC. Two to 4 days after the injection, the electrically evoked, Ca(2+)-dependent release and high-affinity uptake of [3H]GABA were measured in dissected pieces of the left and right IC. These activities were compared with those in the IC taken from unlesioned controls and from sham controls, which received injections of saline instead of KA. Each IC was divided into a dorsal piece, which contained the dorsal cortex and dorsomedial nucleus, and a ventral piece, which contained the central and lateral nuclei. Lesions of the left DNLL depressed the release and uptake of [3H]GABA in the ventral pieces of the IC, but there was a greater depression in the ventral IC contralateral to the lesioned DNLL. There were good correlations between the percentage of neuronal loss in the left DNLL and deficits in [3H]GABA release and uptake activities in the ipsi- and contralateral ventral IC. By contrast, there was no depression of [3H]GABA release and uptake in the dorsal pieces of the IC. The localization of the deficits in release and uptake appears to match the distribution of the synaptic endings of the DNLL pathways in the IC. This correspondence associates GABA release and uptake activities with the DNLL projections to the IC and, therefore, suggests that GABA may be a transmitter of these pathways. The release and uptake of [14C]glycine was also measured to determine whether glycine might be a transmitter of the DNLL pathways to the IC. Lesions of the left DNLL failed to alter the Ca(2+)-dependent release or the uptake of [14C]-glycine, suggesting that DNLL neurons are unlikely to use this compound as a transmitter.

Decreased release of D-aspartate in the guinea pig spinal cord after lesions of the red nucleus.

This study attempts to determine if fibers that project from the guinea pig red nucleus to the spinal cord use L-glutamate and/or L-aspartate as transmitters. Unilateral injections of kainic acid were placed stereotaxically in the red nucleus to destroy the cells of origin of the rubrospinal tract. Six days after the injection, Nissl-stained sections through the lesion site showed that the majority of neurons in the red nucleus ipsilateral to the kainic acid injection were destroyed. In addition, the lesioned area included parts of the surrounding midbrain reticular formation. Silver-impregnated, transverse sections of the cervical spinal cord revealed the presence of degenerating fibers contralaterally in laminae IV-VII of the gray matter. Ipsilaterally, very sparse degeneration was evident in laminae VII and VIII of the gray matter. Two to six days after surgery, the electrically evoked, Ca2(+)-dependent release of both D-[3H]aspartate, a marker for glutamatergic/aspartatergic neurons, and gamma-amino[14C]-butyric acid ([14C]GABA) was measured in dissected quadrants of the spinal cervical enlargement. Lesions centered on the red nucleus depressed the release of D-[3H]aspartate by 25-45% in dorsal and ventral quadrants of the cervical enlargement contralaterally. The release of [14C]GABA was depressed by 27% in contralateral ventral quadrants. To assess the contribution of rubro- versus reticulospinal fibers to the deficits in amino acid release, unilateral injections of kainic acid were placed stereotaxically in the midbrain reticular formation lateral to the red nucleus. Nissl-stained sections through the midbrain revealed the presence of extensive neuronal loss in the midbrain and rostral pontine reticular formation, whereas neurons in the red nucleus remained undamaged. In the spinal cord, degenerating axons were present ipsilaterally in laminae VII and VIII of the gray matter. Some fiber degeneration was also evident contralaterally in laminae V and VI of the gray matter. This lesion did not affect the release of either D-[3H]aspartate or [14C]GABA in the spinal cord. The substantial decrements in D-[3H]aspartate release following red nucleus lesions suggests that the synaptic endings of rubrospinal fibers mediate the release of D-[3H]aspartate in the spinal cord. Therefore, these fibers may be glutamatergic and/or aspartatergic. Because other evidence suggests that rubrospinal neurons are probably not GABAergic, the depression of [14C]GABA release probably reflects changes in the activity of spinal interneurons following the loss of rubrospinal input.

Glycine immunoreactive projections from the dorsal to the anteroventral cochlear nucleus.

The aim of the present study was to investigate whether projections from the dorsal cochlear nucleus (DCN) to the anteroventral cochlear nucleus (AVCN) use either of two inhibitory transmitters, glycine or GABA. Retrograde HRP labeling of DCN-to-AVCN projection neurons was combined with postembedding immunocytochemistry in the DCN of guinea pigs. Following injections of HRP in the anterior or posterior divisions of AVCN, large numbers of neurons were labeled in the DCN. All of these were located in the deep layer, except for a few granule cells. Nearly all (96%) of the projection neurons were immunoreactive for glycine and most had dendritic and somatic morphologies corresponding to those of elongate neurons (so-called 'corn' cells); only a few resembled small stellate neurons. Few (3%) retrogradely labeled neurons were immunoreactive for GABA. The results suggest that projections from the deep DCN to the AVCN are formed primarily by glycinergic elongate neurons. These projections could have a substantial inhibitory influence on the output of neurons in the AVCN.

Retrograde transport of [3H]glycine from the cochlear nucleus to the superior olive in the guinea pig.

The origins of descending glycinergic projections to the guinea pig cochlear nucleus were investigated using retrograde labelling techniques. To identify the cell groups that provide descending projections to the cochlear nucleus, horseradish peroxidase, a nonspecific retrograde neuronal marker, was injected into the cochlear nucleus. After 24 or 48 hours, labelled cell bodies were evident bilaterally in all of the periolivary nuclei that surround the lateral and medial superior olive. The largest numbers of labelled neurons were located in the ventral nucleus of the trapezoid body bilaterally and in the lateral nucleus of the trapezoid body and dorsal periolivary nucleus ipsilaterally. Labelled cells were also present in the inferior colliculus bilaterally and in the contralateral cochlear nucleus. [3H]Glycine was employed as a retrograde tracer to identify the cell groups providing descending glycinergic projections to the cochlear nucleus. Three to 48 hours after injection of 19, 190, or 380 microM [3H]glycine into the cochlear nucleus, retrogradely labelled cell bodies were observed ipsilaterally in all of the periolivary nuclei. No labelled neurons were found in the inferior colliculus. After injections of the highest concentration of [3H]glycine, labelled cells were also found contralaterally in the ventral and lateral nuclei of the trapezoid body and also in the contralateral cochlear nucleus. We conclude that descending glycinergic projections to the cochlear nucleus originate mostly in ipsilateral periolivary cell groups. Minor glycinergic projections originate from the contralateral cochlear nucleus and also from the contralateral ventral and lateral nuclei of the trapezoid body.