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.

Stellate neurons in the outer two layers of the rat dorsal cochlear nucleus (DCN) were studied by the Golgi-EM method. Stellate cell bodies are usually spherical or ovoidal and range from 9 microns to 14 microns in mean diameter. The smallest cells are situated underneath the ependymal layer and the largest cells in layer 2. Primary dendrites are short, thin and smooth and arise abruptly from the perikaryon, without a tapering main stem. Meandering secondary and tertiary dendrites extend in all directions, carry few pleomorphic spines lacking a spine apparatus and often show artifactual beading. The axons are impregnated only for a short distance (10-45 microns). The nucleus is indented, the nucleolus varies in position, and the chromatin, evenly dispersed in the centre, forms small clumps along the nuclear envelope. The cytoplasm is rich in free polyribosomes and contains scattered cisterns of granular endoplasmic reticulum. Varicosities of thin fibres, containing round synaptic vesicles, form asymmetric synapses on perikarya, dendritic shafts and spines of stellate cells. Such fibres run parallel to the long axis of the DCN or are oriented radially and are interpreted as axons of cochlear granule cells. Two kinds of bouton containing pleomorphic vesicles, one kind electron lucent and the other electron dense, form symmetric synapses on perikarya and dendritic shafts of stellate cells. The lucent boutons occur more frequently than the dense boutons, especially on the distal dendritic branches. The boutons with pleomorphic vesicles presumably represent terminals of local circuit neurons, probably the stellate and cartwheel cells. In addition, stellate cells show numerous dendro-somatic and dendro-dendritic appositions characterized by gap junctions and puncta adhaerentia. Most of the dendrites involved in these appositions resemble stellate cell dendrites and it is concluded that DCN stellate cells are coupled electrotonically with one another. The axons of stellate cells acquire a thin myelin sheath. Since the Golgi impregnation did not stain axons of stellate cells past this point, we were unable to demonstrate the synaptic targets of stellate cells.

Activity of glutamic acid decarboxylase GluDCase, the biosynthetic enzyme of y-aminobutyric acid (GABA), was y-Aminobutyric acid (GABA) has been implicated in modulating secretion of pituitary hormones (1-5) and hypothalamichypophysial hormones (6, 7). GABAergic neuroendocrine modulation may occur in the hypothalamus, for example, in the median eminence, where GABAergic terminals are present (8), or directly at the pituitary level (5, 9, 10).Measurable amounts of endogenous GABA have been reported in the anterior pituitary lobe, but much higher amounts are present in the posterior lobe (5, 11). GABA is synthesized by the hypophysis in vitro (12, 13). Furthermore, glutamic acid decarboxylase (GluDCase, the GABA biosynthetic enzyme) activity has been found in the neurointermediate lobe (14) but not in the anterior lobe (5). GABA receptors appear to be present in the anterior lobe (15) and on axons of hypothalamicneurohypophysial neurons (16). However, the organization of GABAergic structures in the hypophysis has not been examined.In the present study, we have measured GluDCase activity in the anterior and neurointermediate pituitary lobes, with and without stalk transection, and used a specific antiserum to rat brain GluDCase (17) to localize GluDCase in the hypophysis. MATERIALS AND METHODSSprague-Dawley male rats (body weight 150-200 g) were purchased from Zivic Miller (Allison Park, PA). The rats were subdivided into three groups: unoperated, pituitary-stalk transected, and sham operated. Six days after surgery, rats from the three groups (see Table 1) were decapitated, the brains were removed, and the pituitary glands were dissected out. The neurointermediate lobe was separated from the anterior lobe. The tissues were homogenized with a glass/glass homogenizer on ice, in 150 ,ul of a buffer solution containing 20 mM potassium phosphate buffer (pH 7.2), 1 mM 2-aminoethylisothiouronium bromide hydrobromide, 0.2 mM. pyridoxal phosphate, 0.1 mM EDTA, and 0.25% Triton X-100. Aliquots (30-40 p1) of the whole homogenate or the supernatant fluid, obtained after centrifugation for 30 min at 25,000 X g at 4°C, were assayed for GluDCase (18) and protein (19).Four unoperated, two sham-operated, and four stalk-transected rats were used for light microscopy, and four unoperated rats were used for electron microscopy. The animals were fixed by perfusion through the ascending aorta with 200 ml of a solution containing 1% formaldehyde and 0.5% glutaraldehyde in 0.065 M sodium phosphate buffer (pH 7.3 at 37°C, delivered at a pressure of 80 cm ofwater) followed by 300 ml of 4% formaldehyde in the same buffer at 20°C. The dissected pituitary glands were sectioned on a Vibratome, and the sections, after treatment with 3% H202 in 10% methanol to eliminate endogenous peroxidase-like activity, were processed with the unlabeled antibody-enzyme method of Sternberger (20) for light and electron microscopy. This procedure, described in detail elsewhere (21), uses our GluDCase antiserum S3 (second bleed) (17) at dilutions of 1...

Experimental light and electron microscopic studies were carried out to determine the length of parallel fibers in the cat cerebellar cortex. Using a fine surgical knife, vermal and hemispheral folia were cut perpendicular to their long axis. The animals were sacrificed 1-10 days after the operation. Sections of the transected folia were then stained with a Fink-Heimer procedure. The resulting degeneration appeared as fine dots that extended lateral to the lesion, as predictable from the course of the parallel fibers. Densitometer readings indicate that the density of degeneration declines gradually lateral to the lesion. The specificity of the silver impregnation was checked by processing silver stained sections for electron microscopy. This confirmed the location of the silver precipitate on degenerating parallel fibers. The pattern of parallel fiber degeneration in the molecular layer has a trapezoidal configuration centered on the lesion. The shorter parallel fibers are located at the base of the molecular layer and extend for 5 mm. The parallel fibers become progressively longer as they approach the pial surface where they attain a maximum length of 7 mm. Our studies suggest that in folia longer than 7 mm parallel fibers are 6 mm long on the average. In addition, it was determined on Golgi sections that the average center-to-center distance between en passant boutons of individual parallel fibers is 5.2 mum. The data indicate that an average parallel fiber, 6 mm long, forming approximately 1100 boutons, may synapse with each Purkinje dendritic tree it traverses.

Two procedures are described for vascular perfusion of the nervous system with a zinc-aldehyde fixative. The procedures, simple and economical, combine the advantages of perfusion fixation with an aldehyde solution and matrix stabilization by a mordating agent, and improve the sensitivity of the peroxidase-antiperoxidase (PAP) method for the immunocytochemical localization of several antigens. Procedure A is intended for the light-microscopic immunostaining of cellular elements containing high concentrations of antigen. Penetration of the immunoreagents is adequate without the use of detergents. Procedure B is particularly advantageous for the light-microscopic immunostaining of cellular elements that contain low concentrations of antigen, and for high-resolution microphotography. With procedure B, the tissue penetration of immunoreagents is more limited than with procedure A; however, neuronal cell bodies and dendrites are more easily penetrated by the immunoreagents than are axons. Neuronal cell bodies and dendrites thus become clearly detectable in the light-microscope, even when they are surrounded by numerous immunoreactive axon terminals, and especially after the blockage of axoplasmic transport by the topical injection of colchicine.