A transporter thought to mediate accumulation of GABA into synaptic vesicles has recently been cloned (McIntire et al., 1997). This vesicular GABA transporter (VGAT), the first vesicular amino acid transporter to be molecularly identified, differs in structure from previously cloned vesicular neurotransmitter transporters and defines a novel gene family. Here we use antibodies specific for N- and C-terminal epitopes of VGAT to localize the protein in the rat CNS. VGAT is highly concentrated in the nerve endings of GABAergic neurons in the brain and spinal cord but also in glycinergic nerve endings. In contrast, hippocampal mossy fiber boutons, which although glutamatergic are known to contain GABA, lack VGAT immunoreactivity. Post-embedding immunogold quantification shows that the protein specifically associates with synaptic vesicles. Triple labeling for VGAT, GABA, and glycine in the lateral oliva superior revealed a higher expression of VGAT in nerve endings rich in GABA, with or without glycine, than in others rich in glycine only. Although the great majority of nerve terminals containing GABA or glycine are immunopositive for VGAT, subpopulations of nerve endings rich in GABA or glycine appear to lack the protein. Additional vesicular transporters or alternative modes of release may therefore contribute to the inhibitory neurotransmission mediated by these two amino acids.

The microscopical structure of the cat cochlear nuclei was studied in Nissl preparations in order to get a suitable map for further experimental investigations. The neurons of the entire nuclear complex were classified into nine different types according to their microscopical appearance. Seven of the nine types, viz., the large spherical cells, small spherical cells, octopus cells, globular cells, pyramidal cells, giant cells, and granular cells, seem to constitute rather uniform cell groups, while the remaining types, viz., the multipolar cells and small cells, seem to be rather heteromorphic. On the basis of the distribution of these cell types the nuclear complex was divided into a corresponding number of cell areas which proved to be partly overlapping. This principle of parcellation led to a certain simplification of the previously proposed schemes of the organization of the cochlear nuclei based on cellular characteristics.

The distribution and colocalization of gamma-aminobutyric acid (GABA)- and glycine-like immunoreactivity in the cochlear nuclear complex of the guinea pig have been studied to produce a light microscopic atlas. The method used was based on post-embedding immunocytochemistry in pairs of 0.5-micron-thick plastic sections treated with polyclonal antibodies against conjugated GABA and glycine respectively. Immunoreactive cells, presumably short axon neurones, predominated in the dorsal cochlear nucleus, with mostly single-GABA-labelled cells in the superficial layer, double-labelled in the middle, and single-glycine-labelled in the deep layers. A few large single-glycine-labelled cells, interpreted as commissural neurons, occurred in the ventral nucleus. Scattered double-labelled cells, probably Golgi cells, were seen in the granule cell domain. Immunolabeled puncta of all three staining categories occurred in large numbers throughout the complex, apposed to somata and in the neuropil, showing a differential distribution onto different types of neuron. Three immunolabeled tracts were noted: the tuberculoventral tract, the commissural acoustic stria, and the trapezoidal descending fibres. Most of the fibres in these tracts were single-labelled for glycine, although in the last mentioned tract single-GABA- and double-labelled fibres were also found. Some of the immunolabeled cell types described here are proposed as the origins of the similarly labelled puncta and fibres on the basis of known intrinsic connections.

This paper defines the pattern of subdivision of the inferior colliculus in rat. It is based on serial sections of brains of albino and hooded rats cut in the frontal, sagittal and horizontal planes using Golgi, Nissl and a combined cell-myelin method. In rat, like in other mammals, the inferior colliculus consists of a central nucleus, an external cortex, and a dorsal cortex. The central nucleus is flattened in the frontal plane and confined to the caudomedial part of the inferior colliculus. It is characterized by a lamellar organization of disc-shaped neurons interspersed with multipolar cells. The cells are small to medium-sized. Although there is a dorsoventral gradient in size and packing density of cells within the nucleus, the overall size is smaller and the packing density larger than in adjacent subdivisions. The two cortices each consists of three layers. The outer-most layer is common to the two cortices, forming a fibrocellular capsule continuous along most of the circumference of the inferior colliculus. The external cortex is located lateral, rostral, ventral and ventrocaudal to the central nucleus. Its second layer, deep to the superficial capsule, is characterized by clusters of many small and a few medium-sized neurons in a myelin-dense neuropil. Layer 3, which constitutes the major portion of the subdivision, consists of relatively scattered, small, medium and large cells, the most characteristic element being large multipolar neurons with coarse Nissl granules. The dorsal cortex is located dorsocaudal and dorsomedial to the central nucleus. Its second layer is composed of small neurons, while the third, deep layer in addition contains medium-sized neurons. The cell density is intermediate to that of the central nucleus and the deep part of the external cortex. We have tried to facilitate the parcellation by reference to easily recognizable, nearby structures and to standard stereotaxic coordinates.

The cyto- and fibre-architecture of the cochlear nuclear complex of the guinea-pig has been studied in serial sections using Nissl, Golgi and combined cell-myelin staining of normal material, and a silver degeneration method after cochlear ablation. The nuclear subdivisions and major cell types can be recognised on the basis of those found in the cat, but there are some differences between the two species in the precise distribution and morphology of the neurons. The rostrodorsal part of the anteroventral cochlear nucleus (AVCN) contains predominantly spherical bushy cells, but these cannot be readily divided into large and small types as in the cat. Globular bushy cells are seen in the caudal region of the AVCN, but the majority occur in the posteroventral cochlear nucleus (PVCN), in an area extending from the nerve root right up to the boundary of the dorsal cochlear nucleus (DCN). The octopus cells constitute a distinct region in the most dorsomedial part of the PVCN underneath the DCN. Giant cells are seen scattered around the nerve root region. Multipolar and small cells are seen throughout the non-granular regions of the ventral cochlear nucleus (VCN) except for the octopus cell area, but occur mainly in the more rostral regions of the PVCN. Small cells occur in greatest abundance in the thin cap area at the dorsal edge of the VCN below a superficial granule cell layer. The latter covers the dorsolateral surface of the VCN, and a lamina of granule cells partially separates the PVCN from the DCN. The DCN can be divided into four layers. The outermost molecular layer (layer 1) is separated from the deeper regions by a prominent layer of granule cells (layer 2) which also contains the pyramidal cells. Molecular layer stellate cells are seen in layer 1 and a staggered row of cartwheel neurons is found at the boundary between layers 1 and 2. Layer 3 contains the basal dendrites of the pyramidal cells and some small (vertical) cells, and is innervated by the descending branches of the cochlear nerve. The deepest layer 4, which contains multipolar cells and giant cells, does not appear to receive this direct cochlear input.

This paper describes the fine structure of granule cells and granule-associated interneurons (termed Golgi cells) in the cochlear nuclei of cat, rat and mouse. Granule cells and Golgi cells are present in defined regions of ventral and dorsal cochlear nuclei collectively termed "cochlear granule cell domain'. The granule cells are small neurons with two or three short dendrites that give rise to a few branches with terminal expansions. These participate in glomerular synaptic arrays similar to those of the cerebellar cortex. In the glomeruli the dendrites form short type 1 synapses with a large, centrally-located mossy bouton containing round synaptic vesicles and type 2 synapses with peripherally located, smaller boutons containing pleomorphic vesicles. The granule cell axons is thin and beaded and, on its way to the molecular layer of the DCN, takes a straight course, which in ventral nucleus is parallel to the pial surface. Neurons of the second category resemble cerebellar Golgi cells and occur everywhere interspersed among the granule cells. They are usually larger than the granule cells and give rise to dendrites which may branch close to and curve around the cell body. The dendrites contain numerous mitochondria and are laden with thin appendages, giving them a hairy appearance. Both the cell body and the stem dendrites participate in glomerular synaptic arrays. Golgi cell glomeruli are distinguishable from the granule cell glomeruli by unique features of the dendritic profiles and by longer, type 1 synaptic junctions with the central mossy bouton. The Golgi cell axon forms a beaded plexus close to the parent cell body. The synaptic vesicle population of the mossy boutons suggests that they are a heterogeneous group and may have multiple origins. Apparently, each of the various classes participates in both granule and Golgi cell glomeruli. The smaller peripheral boutons with pleomorphic vesicles in the two types of glomeruli may represent Golgi cell axons which make synaptic contacts with both granule and Golgi cells. The Golgi cell axons which make synaptic contacts with both granule and Golgi cells. The Golgi cell dendrites, on the other hand, are also contacted by small boutons en passant with round synaptic vesicles, which may represent granule cell axons. A tentative scheme of the circuitry in the cochlear granule cell domain is presented. The similarity with the cerebellar granule cell layer is striking.

The cellular basis for the laminar structure of the central nucleus of the inferior colliculus has been investigated by computer-assisted 3-D reconstruction of Golgi impregnated dendritic arbors, sampled from serial sections of resin embedded material of adult rat. Two types of flattened neurone, defined here as flat (F) and less flat (LF), are described as contributing to the pattern. The dendritic arbors of F neurones had a smaller absolute thickness (mean about 50 microns) and a denser arbor. They were strikingly parallel regionally and formed laminae mostly one cell thick. The laminae appeared to be separated by interlaminar compartments populated by the LF neurones. The arbors of the latter were thicker (mean about 100 microns) and less dense than those of the F neurones. The different density of the two types may, at least in part, be responsible for the corresponding difference in density of oriented dendrites within the laminae and interlaminar compartments. The orientation planes of F and LF arbors were roughly similar, but a consistent, slight difference in orientation between F and LF arbors is not excluded. Most of the F and LF arbors were elongated in parallel with the ventrolaterally to dorsomedially oriented long axis of the laminae. A few were instead oriented rostrocaudally or in intermediate directions. The interlaminar compartments appeared less distinct in the low than in the high frequency region. The latter region also differed from the former by having F neurones with a higher number of intermediate and terminal segments and a denser arbor. It is discussed whether the observed F and LF cells constitute two distinct cell types or are varieties of one type of neurone, with the morphological differences reflecting differences in location. Further characterization of the neurones on histochemical, hodological, and other criteria is required to settle this question.

Glutamine is involved in a variety of metabolic processes, including recycling of the neurotransmitters glutamate and gamma-aminobutyric acid (GABA). The system N transporter SN1 mediates efflux as well as influx of glutamine in glial cells [Chaudhry et al. (1999), Cell, 99, 769-780]. We here report qualitative and quantitative data on SN1 protein expression in rat. The total tissue concentrations of SN1 in brain and in kidney are half and one-quarter, respectively, of that in liver, but the average concentration of SN1 could be higher in astrocytes than in hepatocytes. Light and electron microscopic immunocytochemistry shows that glutamatergic, GABAergic and, surprisingly, purely glycinergic boutons are ensheathed by astrocytic SN1 laden processes, indicating a role of glutamine in the production of all three rapid transmitters. A dedication of SN1 to neurotransmitter recycling is further supported by the lack of SN1 immunoreactivity in oligodendrocytes (cells rich in glutamine but without perisynaptic processes). All neuronal structures appear unlabelled implying that a different protein mediates glutamine uptake into nerve endings. In several regions, SN1 immunoreactivity is higher in association with GABAergic than glutamatergic synapses, in agreement with observations that exogenous glutamine increases output of transmitter glutamate but not GABA. Nerve terminals with low transmitter reuptake or high prevailing firing frequency are associated with high SN1 immunoreactivity in adjacent glia. Bergmann glia and certain other astroglia contain very low levels of SN1 immunoreactivity compared to most astroglia, including retinal Müller cells, indicating the possible existence of SN isoforms and alternative mechanisms for transmitter recycling.