Histological and cytological study of the bed nuclei of the stria terminalis in adult rat. II. Oval nucleus: extrinsic inputs, cell types, neuropil, and neuronal modules

The oval nucleus (Ov) of the bed nuclei of the stria terminalis was studied in adult rats. The Ov is composed of 11 neuron types distributed into a shell and a core domain. The stria terminalis, internal capsule, ventral amygdaloid pathway, and medial forebrain bundle are the main sources of afferents to the neuropil of the Ov. The nucleus shell contains abundant intrinsic neurons possibly connected among themselves and with the core by centripetal axon collaterals. Series of intrinsic neurons in the shell, linked with both short-axon and projecting neurons in the core, suggest a centripetal control of projecting neurons. In situ hybidization for vesicular glutamate transporter (VGlu) and glutamic acid decarboxylase (GAD) show numerous GAD-synthesizing neurons and an absence of VGlu-synthesizing neurons. In the electron microscope, the neuropil of the Ov contains axospinous, axoshaft, axosomatic, mixed (i.e., chemical-electrical), and axoaxonic synapses, in order of frequency. Synaptic boutons in apposition with the initial segment, represent an additional axoaxonic interaction. Further neural synchronization of the Ov occurs via gap junctions between somata, soma-dendrite, and possibly by apposition between axon terminals. The putative inputs from the major tracts of the forebrain coupled with the cytological organization of the Ov make it one of the most complex structures of the mammalian brain.     

Juxtacapsular nucleus of the stria terminalis of the adult rat: extrinsic inputs, cell types, and neuronal modules: a combined Golgi and electron microscopic study

This study unravels the microscopic organization of the juxtacapsular nucleus of the bed nuclei of the stria terminalis (Ju) by using silver impregnation and electron microscopic techniques. Examination of Golgi-impregnated specimens demonstrates that the Ju has precise boundaries primarily determined by a conical condensation of fibers of the stria terminalis (StT) around the nucleus. The internal capsule, ansa peduncularis, and medial forebrain bundle together with the StT provide extrinsic afferents to the neuropil of the Ju. Two main neuron types are found in the Ju: interneurons (including basket and neurogliaform cells) and projection neurons (bipolar and small pyramidal cells). The bipolar cell type accounts for about 80% of the sampled neurons. Short-axon neurons located within the dorsal part of the Ju send descending fibers that appear to terminate on the bipolar neurons, suggesting the existence of vertically oriented functional units within the nucleus. With the electron microscope, Ju neurons are seen in clusters of two or three neurons coupled by gap junctions. The neuropil contains numerous dendrites, axons, myelinated axons, and several types of synaptic interactions, including axospinous, axoshaft, and axosomatic. Within the neuropil, Ju neurons appear to be presynaptically modulated by axoaxonal interactions. The present findings suggest a model wherein bipolar neurons represent the output system of the Ju controlled by the interneurons, which would, in turn, be modulated by collaterals arising from the tributary fiber tracts. Additional neural interaction between Ju neurons utilizes gap junction-mediated electrotonic coupling.     

Modulation of dendrodendritic interactions and mitral cell excitability in the mouse accessory olfactory bulb by vaginocervical stimulation

When female mice are mated, they form a memory to the pheromonal signal of their male partner. The neural changes underlying this memory occur in the accessory olfactory bulb, depend upon vaginocervical stimulation at mating and involve changes at the reciprocal synapses between mitral and granule cells. However, the action of vaginocervical stimulation on the reciprocal interactions between mitral and granule cells remains to be elucidated. We have examined the effects of vaginocervical stimulation on paired-pulse depression of amygdala-evoked field potentials recorded in the external plexiform layer of the accessory olfactory bulb (AOB) and the single-unit activity of mitral cells antidromically stimulated from the amygdala in urethane-anaesthetized female mice. Artificial vaginocervical stimulation reduced paired-pulse depression (considered to be due to feedback inhibition of the mitral cell dendrites from the granule cells via reciprocal dendrodendritic synapses) recorded in the AOB external plexiform layer. As would be expected from this result, vaginocervical stimulation also enhanced the spontaneous activity of a proportion of the mitral cells tested. These results suggest that vaginocervical stimulation reduces dendrodendritic feedback inhibition to mitral cells and enhances their activity.     

Inverse expression of olfactory cell adhesion molecule in a subset of olfactory axons and a subset of mitral/tufted cells in the developing rat main olfactory bulb

The projection of olfactory sensory neuron (OSN) axons from the olfactory epithelium (OE) to the olfactory bulb (OB) is highly organized but topographically complex. Evidence suggests that odorant receptor expression zones in the OE map to the OB about orthogonal axes. One candidate molecule for the formation of zone-specific targeting of OSN axon synapses onto the OB is the olfactory cell adhesion molecule (OCAM). OCAM(+) OSNs are restricted to three of the four zones in the OE and project their axons to the ventral OB where they form synapses with mitral/tufted (M/T) cells. To determine when this zonal connection is established, we have examined OCAM expression in rat olfactory system, during seminal periods of glomerular formation. OCAM(+) axons sort out in the ventral olfactory nerve layer of the OB before glomerular formation. Surprisingly, OCAM was also expressed transiently by subsets of M/T cell dendrites located in the dorsal OB. The expression of OCAM by OSN axons and M/T dendrites was asymmetrical; in the dorsal OB, OCAM(-) OSN axons synapsed on OCAM(+) M/T dendrites, whereas in the ventral OB, OCAM(+) OSN axons synapsed on OCAM(-) M/T dendrites. The restricted spatial map of OCAM(+) M/T cells appeared earlier in development than the zonal segregation of OCAM(+) OSN axons. Thus, OCAM on M/T cell dendrites may act in a spatiotemporal window to specify regions of the developing rat OB, thereby establishing a foundation for mapping of the OE zonal organization onto the OB.     

Sexually dimorphic activation of the accessory, but not the main, olfactory bulb in mice by urinary volatiles

Previous research suggests that volatile body odourants detected by the main olfactory epithelium (MOE) are processed mainly by the main olfactory bulb (MOB) whereas nonvolatile body odourants detected by the vomeronasal organ (VNO) are processed via the accessory olfactory bulb (AOB). We asked whether urinary volatiles from males and females differentially activate the AOB in addition to the MOB in gonadectomized mice of either sex. Exposure to urinary volatiles from opposite-sex but not same-sex conspecifics augmented the number of Fos-immunoreactive mitral and granule cells in the AOB. Volatile urinary odours from male as well as female mice also stimulated Fos expression in distinct clusters of MOB glomeruli in both sexes. Intranasal administration of ZnSO(4), intended to disrupt MOE function, eliminated the ability of volatile urinary odours to stimulate Fos in both the MOB and AOB. In ovariectomized, ZnSO(4)-treated females a significant, though attenuated, AOB Fos response occurred after direct nasal exposure to male urine plus soiled bedding, suggesting that VNO signaling remained partially functional in these mice. Future studies will determine whether MOE or VNO signaling, or both types of input, drive the sexually dimorphic response of the AOB to volatile opposite-sex odours and whether this AOB response contributes to reproductive success.     

Increased expression of neuronal nitric oxide synthase mRNA in the accessory olfactory bulb during the formation of olfactory recognition memory in mice

The mouse accessory olfactory bulb contains a high density of nitric oxide synthase and, in females, is involved in the formation of a mating and pheromone-specific recognition memory. The exact role of nitric oxide in this memory model is not yet clearly understood. In this study, in situ hybridization was used to assess neuronal nitric oxide synthase mRNA expression during the critical interval associated with synaptic plasticity in the accessory olfactory bulb of female mice leading to the formation of a recognition memory for the stud male pheromones present following mating. Nitric oxide synthase mRNA was significantly increased following mating and 120-min stud male exposure compared with oestrus mice. The mRNA expression was more predominant in the anterior than the posterior regions of the bulb. These observations indicate a stimulus-specific activation of nitric oxide gene expression in the female mouse accessory olfactory bulb and support the hypothesis that nitric oxide may modulate intermediate synaptic pathways during the formation of a pheromone-dependent olfactory recognition for stud males.     

Vomeronasal and olfactory nerves of adult and larval bullfrogs: II. Axon terminations and synaptic contacts in the accessory olfactory bulb

The ultrastructure of the accessory olfactory bulb (AOB) of the bullfrog tadpole and adult was examined, and the main difference between tadpole and adult is that the latter is more compact and shows more synapses. Except for vomeronasal (VMN) glomeruli, the AOB is not highly organized, with mitral cell neurons scattered throughout the neuropil. VMN axon terminals form asymmetric synapses with mitral cell dendrites in glomeruli; in VMN axon terminals, dense-cored vesicles are seen along with the more abundant lucent vesicles 40-50 nm in diameter. Counts indicated that more than 90% of the dendro-dendritic synapses between mitral cells and presumed granule cells are of the asymmetrical type, and reciprocal asymmetrical-symmetrical synapses are not common. Lucent vesicles with round or slightly ellipsoidal profiles and less abundant dense-cored vesicles 60-90 nm in diameter are found in pre- and postsynaptic dendrites; sometimes the dense-cored vesicles lie against or near the presynaptic membrane. Microtubules were often seen to be closely associated with pre- and postsynaptic elements of dendro-dendritic synapses. The most characteristic feature of mitral cell bodies, apart from their large size, is an extensive Golgi system that may extend well into their major dendritic extensions. Dense-cored vesicles are associated with Golgi membranes, from which they probably originate. Centrioles are associated with the Golgi system, and some become basal bodies and give rise to cilia in some mitral cells.     

Errors in lamina growth of primary olfactory axons in the rat and mouse olfactory bulb.

In the adult olfactory nerve pathway of rodents, each primary olfactory axon forms a terminal arbor in a single glomerulus in the olfactory bulb. During development, axons are believed to project directly to and terminate precisely within a glomerulus without any exuberant growth or mistargeting. To gain insight into mechanisms underlying this process, the trajectories of primary olfactory axons during glomerular formation were studied in the neonatal period. Histochemical staining of mouse olfactory bulb sections with the lectin Dolichos biflorus-agglutinin revealed that many olfactory axons overshoot the glomerular layer and course into the deeper laminae of the bulb in the early postnatal period. Single primary olfactory axons were anterogradely labelled either with the lipophilic carbocyanine dye, 1,1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI), or with horse-radish peroxidase (HRP) by localized microinjections into the nerve fiber layer of the rat olfactory bulb. Five distinct trajectories of primary olfactory axons were observed in DiI-labelled preparations at postnatal day 1.5 (P1.5). Axons either coursed directly to and terminated specifically within a glomerulus, branched before terminating in a glomerulus, bypassed glomeruli and entered the underlying external plexiform layer, passed through the glomerular layer with side branches into glomeruli, or branched into more than one glomerulus. HRP-labelled axon arbors from eight postnatal ages were reconstructed by camera lucida and were used to determine arbor length, arbor area, and arbor branch number. Whereas primary olfactory axons display errors in laminar targeting in the mammalian olfactory bulb, axon arbors typically achieve their adult morphology without exuberant growth. Many olfactory axons appear not to recognize appropriate cues to terminate within the glomerular layer during the early postnatal period. However, primary olfactory axons exhibit precise targeting in the glomerular layer after P5.5, indicating temporal differences in either the presence of guidance cues or the ability of axons to respond to these cues.     

Distribution of astroglia in glomeruli of the rat main olfactory bulb: Exclusion from the sensory subcompartment of neuropil

During an entire lifetime, sensory axons of regenerating olfactory receptor neurons can enter glomeruli in the olfactory bulb and establish synaptic junctions with central neurons. The role played by astrocytes in this unique permissiveness is still unclear. Glomerular astrocytes have been identified by immunocytochemistry for glial fibrillary acidic protein and S100 proteins at the light and electron microscopic levels. The latter labeling included submicroscopic lamellar and filopodial extensions of astroglial processes. Cell bodies and processes accumulate along the border between juxtaglomerular walls and glomerular neuropil. Within glomeruli, a network of astroglial processes encloses mesh-like neuropil zones devoid of astroglia. Electron microscopy confirmed the division into subcompartments of glomerular neuropil: 1) The “sensory-synaptic subcompartment” includes all sensory axon terminals and terminal dendritic branches receiving sensory input, whereas astroglia are excluded; 2) in the “central-synaptic subcompartment,” astroglial processes are intermingled with other neuropil components: dendrites of relay cells and interneurons, dendrodendritic synapses, centrifugal (cholinergic and serotonergic) axons, their axodendritic synapses, and blood vessels. Unevenly distributed astroglial processes in this subcompartment are attached to vascular basal laminae, stem dendrites, and subpopulations of dendrodendritic synapses, especially those colocalized with centrifugal projections (“triadic synapses”). Astroglia-free parts of the “central” subcompartment contain segments of dendrites and subpopulations of dendrodendritic synapses. Because of the subdivision of the glomerular neuropil into portions with and without glial components, glia do not completely demarcate the border between the “sensory” and the “central” subcompartments. Interdigitation between the subcompartments varies among glomeruli and even within a single glomerulus. The mesh width of astroglial networks covaries with numerical relations between sensory and dendrodendritic synapses. This distribution pattern of astrocytes suggests that these glial cells monitor brain-derived effects on olfactory glomerular neuropil rather than olfactory input and that astroglial processes are (re-)arranged accordingly. J. Comp. Neurol. 388:191–210, 1997. © 1997 Wiley-Liss, Inc.     

The afferent connections of the main and the accessory olfactory bulb formations in the rat: an experimental HRP-study

The afferent connections of the main and accessory olfactory bulbs in the rat were examined by injecting horseradish peroxidase (HRP) into one or the other of these structures either by microelectrophoresis or by hydraulic pressure. Alternate sections were stained with newly developed HRP-procedures using either benzidine dihydrochloride (de Olmos and Heimer, '77) or tetramethyl-benzidine. Eighteen to twenty-four hours after unilateral HRP injections confined to the main olfactory bulb, a large number of HRP-labeled perikaria appeared in the following telencephalic structures on the ipsilateral side: All portions of the anterior olfactory nucleus (AON) except its external part, the lateral transitional field (LT) between AON and the paleocortex, the whole extent of the primary olfactory cortex (POC); the medial forebrain bundle area deep to the olfactory tubercle, the nucleus of the horizontal limb of the diagonal band (NHDB) and the nucleus of the lateral olfactory tract (NLOT). A moderate to small number of labeled cells, furthermore, were seen in the dorsal (DT) and medial (MT) transition fields, the ventral praecommissural hippocampus (tt2), the ventral superficial part of the nucleus of the vertical limb of the diagonal band (NVDB), the sublenticular part of the substantia innominata (SI), the anterior amygdaloid area, the posterolateral cortical amygdaloid nucleus (C2) and the transition region (28 L') between the olfactory cortex and the lateral entorhinal area proper. On the contralateral side a large number of labeled cells were found in all parts of the AON, with especially heavy labeling in its external part. A moderate number of labeled cells could also be detected in the lateral transition field (LT) and the NLOT. In the diencephalon and the brain stem a moderate number of HRP-labeled perikaria were observed in the dorsal, perifornical, and lateral hypothalamus, as well as in locus coeruleus and the dorsal and medial raphae nuclei. Following large HRP injections in the main olfactory bulb a moderate to small number of labeled cells were seen also in the posterior and premammillary hypothalamus and in field CA1 of the retrocommissural hippocampus on the ipsilateral side, as well as in POC on the contralateral side. It is possible, however, that the uptake of label took place in an undetected pool of HRP in the very rostal part of AON rather than in the olfactory bulb. HRP injections in the accessory olfactory bulb resulted in labeled neurons in the posterior ventro-lateral part of the bed nucleus of the stria terminalis, the nucleus of the accessory olfactory tract, the rostrodorsal portions of the medial amygdaloid nucleus, and the whole extent of the posteromedial cortical amygdaloid nucleus (C3) on the ipsilateral side. A few lightly labeled cells were seen also in the contralateral C3.     

Regionalization of Fos immunostaining in rat accessory olfactory bulb when the vomeronasal organ was exposed to urine.

The distribution of Fos-immunoreactive (Fos-ir) cells in the accessory olfactory bulb (AOB) of rats following vomeronasal organ exposure to urine was studied. Following exposure to male and female Wistar rat urine, Fos-ir cells were found in the mitral/tufted cell layer, granule cell layer and periglomerular cell layer of the AOB of female Wistar rat, with the highest number in the granule cell layer. Exposure to water or removal of the vomeronasal organ suppressed the expression of Fos-ir cells. These results suggest that female Wistar rats specifically detect urinary substances derived from male or female Wistar rats via the vomeronasal organ. Exposure of the vomeronasal organ of female Wistar rats to male Wistar urine induced the appearance of many more Fos-ir cells in all layers of the AOB than exposure to female Wistar urine. As for the mitral/tufted cell layer, the density of Fos-ir cells in the rostral portion (Gi2alpha-positive) of all regions of the AOB was about twice as high as that in the caudal portion when male urine was given. The distribution pattern of Fos-ir cells in response to female urine was not identical to that in response to male urine. That is, the density of Fos-ir cells in the caudal portion was slightly larger than that in the rostral portion in the lateral region, while in other regions the density in the rostral portion was higher than that in the caudal portion. It is likely that information from different pheromones is transmitted to the higher brain regions through the different regions of the AOB.     

Differential projections of the main and accessory olfactory bulb in the frog.

The central projections of the main olfactory bulb and the accessory olfactory bulb of the adult leopard frog (Rana pipiens) were reexamined, by using a horseradish peroxidase anterograde tracing method that fills axons with a continuous deposit of reaction product. The fine morphology preserved by this method allowed the terminal fields of the projection tracts to be delineated reliably, and for the first time. Herrick's amygdala has been newly subdivided into cortical and medial nuclei on the basis of cytoarchitecture, dendritic morphology, and the differential projections of the main and accessory olfactory tracts. The main olfactory bulb projects through the medial and lateral olfactory tracts to the postolfactory eminence, the rostral end of the medial cortex, the rostral end of the medial septal nucleus, the cortical amygdaloid nucleus, the nucleus of the hemispheric sulcus, and both the dorsal and ventral divisions of the lateral cortex, including its retrobulbar fringe. The lateral olfactory tract overlaps the dorsal edge of the striatal plate along the ventral border of the lateral cortex, but it is not certain whether any striatal cells are postsynaptic to the tract fibers. The lateral cortex is the largest of these territories, and receives the terminals of the main olfactory projection throughout its extent. It extends from the olfactory bulb to the posterior pole, and from the striatum to the summit of the hemisphere, where it borders the dorsal cortex. The medial and lateral olfactory tracts combine in the region of the amygdala to form a part of the stria medullaris thalami. These fibers cross in the habenular commissure and terminate in the contralateral cortical amygdaloid nucleus and periamygdaloid part of the lateral cortex. Cells projecting to the main olfactory bulb are found in the diagonal band and adjacent cell groups, but there is no evidence of an interbulbar projection arising from either the olfactory bulb proper or a putative anterior olfactory nucleus. The accessory olfactory bulb projects through the accessory olfactory tract to the medial and cortical amygdaloid nuclei. A fascicle of the tract crosses in the anterior commissure to terminate in the contralateral amygdala. While the main and accessory olfactory projections may converge in the cortical amygdaloid nucleus, the medial amygdaloid nucleus is connected exclusively with the accessory olfactory bulb.     

Light microscopic Golgi study of mitral/tufted cells in the accessory olfactory bulb of the adult rat.

Mitral/tufted cells (MTCs) of the accessory olfactory bulb (AOB) of adult rats were investigated light microscopically with the rapid Golgi method. The somata of the MTCs, appearing ovoid or triangular in shape, are distributed throughout the external plexiform layer. The soma size varies from small to large (12-26 microns). Apical dendrites originating from the soma enter the glomerular layer to provide branches that form the glomerular arbors. After making a glomerular arbor, some dendrites develop a second arbor (en passant and terminal arbors, respectively). The MTCs have a very diverse dendritic branching pattern and most have a variable number of glomerular arbors per cell (up to 6); we have tentatively classified the MTCs into simple, intermediate, and complex. Of the glomerular arbors, 80% have a diameter of less than 50 microns. The glomerular arbors have been classified as baskets (small spherical or ovoid) with short loopy processes; balls of yarn (large and nearly spherical) with loosely intermingled thick loops; and bushes (small to large and rather polymorphic) with irregular processes. The MTCs send dendritic arbors to terminate in one or more glomeruli where they are arranged in several different types of endings. Since it is generally believed that the dendrites of mitral and tufted cells of the main olfactory bulb terminate in only one glomerulus, the difference in the termination of the dendrites of the MTCs may represent a morphological characteristic that is relevant to the coding and/or integration of sensory information.     

Characterization of somatostatin- and cholecystokinin-immunoreactive periglomerular cells in the rat olfactory bulb

Periglomerular cells (PG) are interneurons of the olfactory bulb (OB) that modulate the first synaptic relay of the olfactory information from the olfactory nerve to the dendrites of the bulbar principal cells. Previous investigations have pointed to the heterogeneity of these interneurons and have demonstrated the presence of two different types of PG. In the rat OB, type 1 PG receive synaptic contacts from the olfactory axons and are gamma-aminobutyric acid (GABA)-ergic, whereas type 2 PG do not receive synaptic contacts from the olfactory axons and are GABA immunonegative. In this study, we analyze and characterize neurochemically a group of PG that has not been previously classified either as type 1 or type 2. These PG are immunoreactive for the neuropeptides somatostatin (SOM) or cholecystokinin (CCK). By using double immunocytochemistry, we demonstrate that neither the SOM- nor the CCK-immunoreactive PG contain GABA immunoreactivity, which is a neurochemical feature of type 1 PG. Moreover, they do not contain the calcium-binding proteins calbindin D-28k and calretinin, which are neurochemical markers of the type 2 PG. Electron microscopy demonstrates that the dendrites of the SOM- and CCK-containing PG are distributed in the synaptic and sensory subcompartments of the glomerular neuropil and receive synaptic contacts from the olfactory axons. Therefore, they should be included in the type 1 group rather than in the type 2. Altogether, these data indicate that the SOM- and the CCK-containing PG may constitute a group of GABA-immunonegative type 1 PG that has not been previously described. These results further extend the high degree of complexity of the glomerular circuitry.     

Clonal mixing, clonal restriction, and specification of cell types in the developing rat olfactory bulb

To understand the clonal relationship of various olfactory bulb (OB) cell types, OB progenitor cells were infected at embryonic day (E) 14, E15, and E17 with retroviral libraries encoding alkaline phosphatase or beta-galactosidase. After survival to postnatal day 10-15, sibling relationships were identified by polymerase chain reaction DNA amplification of distinct sequences in the retroviral constructs. Within the OB, clonal progeny dispersed widely in all directions. In sharp contrast, however, clonal dispersion between the OB and neocortex was not observed, although occasional clonal dispersion between the OB and pyriform and hippocampal regions could not be excluded. Most clones (84%) contained a single cell type, especially after E17 injections, suggesting the existence of either restricted precursors, or multipotential progenitors instructed by a restricted cellular environment. Mixed OB clones (16%) contained multiple cell types in the OB, or occasionally glial or neuronal cells outside the OB, demonstrating the existence of multipotential OB progenitors, likely at a stage before formation of the olfactory rostral migratory stream. Surprisingly, OB glial cells were not labeled, suggesting distinct lineages or perhaps distinct migratory paths for glia and neurons into the OB. A hierarchical cell lineage is proposed that involves a multipotential progenitor that gives rise to potentially more limited progenitors.     

Different granule cell populations innervate superficial and deep regions of the external plexiform layer in rat olfactory bulb

Studies on the morphological organization of the main olfactory bulb have indicated that there are subpopulations of granule cells with different dendritic patterns in the external plexiform layer (EPL). Small, extracellular injections of horseradish peroxidase (HRP) were made iontophoretically into superficial and deep parts of the EPL and the granule cell layer (GCL) in adult rats. Superficial EPL injections principally labeled superficial granule cell somata, whereas deep EPL injections labeled both superficial and deep granule cell somata. Injections in the superficial GCL labeled granule cell dendritic processes extending across the entire EPL. However, deep GCL injections labeled few granule cell dendrites in the superficial EPL, but labeled many such processes in the deep EPL. These results were the same in material processed with the Hanker-Yates procedure, where the morphology of individual neurons could be studied, and in the more sensitive tetramethyl benzidineprocedure. Serial reconstructions of individual granule cells were made from both HRP and Golgi-Kopsch material. The distal dendrites of deep granule cells reached only as far as the deep EPL, where they branched extensively and had many dendritic spines. The dendrites of superficial granule cells, however, reached the most superficial part of the EPL where they ramified most extensively. The superficial granule cells typically had a higher spine density in the superficial part of the EPL than in the deep part. On the basis of these results, we conclude that the superficial granule cells predominantly innervate the superficial EPL and that the deep granule cells exclusively innervate the deep EPL. Granule cells are believed to exert inhibitory influences on the bulbar output neurons, the mitral and tufted cells, through reciprocal dendrodendritic synapses. Since the secondary dendrites of the tufted cells ramify in the superficial EPL and the dendrites of most mitral cells ramify in deep EPL, the superficial and deep granule cells may preferentially modulate the responses of tufted and mitral cells, respectively.     

Immunoreactive GAP-43 in the neuropil of adult rat neostriatum: localization in unmyelinated fibers, axon terminals, and dendritic spines

GAP-43 is a neuron-specific phosphoprotein that has been implicated in neuronal development, axonal regeneration, and synaptic plasticity. Although in mammals the caudate-putamen is among those brain areas that retain a high content of GAP-43 throughout life, the role of the phosphoprotein in the neostriatum is unknown. In order to understand better the possible function(s) of GAP-43 in the adult striatum, its cellular localization was examined with immunohistochemistry at the light and electron microscopic levels by using a sheep polyclonal antibody. At the light microscopic level immunoreactive GAP-43 was abundant throughout the neostriatal neuropil but was absent from neuronal somata. At the ultrastructural level, labeling was most prevalent in small unmyelinated axons (0.12-0.15 microns diameter). Reaction product was distributed along fibers in discrete patches about 1 micron apart and in preterminal sites from which vesicle-filled boutons arose. Staining was also present in small (0.35 microns) axon terminals that contained round vesicles and formed asymmetric synapses, mostly with thin spines. Following unilateral cortical lesions, some degenerating cortical axons in the neostriatum exhibited GAP-43 labeling. Unexpectedly, in normal striatum, GAP-43 was also occasionally found in the heads of dendritic protrusions and in thin spines that received asymmetric contacts. We speculate that in the adult neostriatum, the protein may be important in the remodeling of synapses onto medium spiny neurons that involve, in part, the corticostriatal pathway.     

Dendritic and axonal organization of mitral and tufted cells in the rat olfactory bulb

The output cells of the main olfactory bulb, the mitral and tufted cells, can be categorized into subclasses on the basis of their intrabulbar dendritic and axonal characteristics. Their form was studied in rats following labeling by iontophoretic injection of horseradish peroxidase into the external plexiform layer (EPL). The fact that these extracellular injections labeled small numbers of neurons permitted reconstruction of individual cells. The injection depth within the EPL determined the type of cells labeled. Secondary dendrites of each cell type lay in one of three partially overlapping zones in the EPL. The deepest zone contained the secondary dendrites of one group of mitral cells (Type I), which had the deepest and longest dendrites of the output cells. An intermediate zone of the EPL contained the secondary dendrites of middle tufted and a second class of mitral cells (Type II). The superficial zone, adjacent to the glomerular layer, contained the relatively short, asymmetric dendritic fields of external tufted cells. The few labeled internal tufted cells had secondary dendrites in either the intermediate or deep zones. Every cell type, except the Type I mitral cells, had axon collaterals in the internal plexiform and upper granule cell layers. No cell types had axons re-entering the EPL. These results for output cells combined with our previous observations on granule cells point to a functional sublaminar organization of the EPL that has not previously been proposed.     

Segregated pathways to the vomeronasal amygdala: differential projections from the anterior and posterior divisions of the accessory olfactory bulb

Apically and basally located receptor neurons in the vomeronasal sensory epithelium express G(i2 alpha)- and G(o alpha)-proteins, V1R and V2R vomeronasal receptors, project to the anterior and posterior accessory olfactory bulb and respond to different stimuli, respectively. The extent to which secondary projections from the two portions of the accessory olfactory bulb are convergent in the vomeronasal amygdala is controversial. This issue is addressed by using anterograde and retrograde tract-tracing methods in rats including electron microscopy. Injections of dextran-amines, Fluoro Gold, cholera toxin-B subunit and Fast Blue were delivered to the anterior and posterior accessory olfactory bulb, bed nucleus of the stria terminalis, dorsal anterior amygdala and bed nucleus of the accessory olfactory tract/anteroventral medial amygdaloid nucleus. We have demonstrated that, apart from common vomeronasal-recipient areas, only the anterior accessory olfactory bulb projects to the bed nucleus of the stria terminalis, medial division, posteromedial part, and only the posterior accessory olfactory bulb projects to the dorsal anterior amygdala and deep cell layers of the bed nucleus of the accessory olfactory tract and the anteroventral medial amygdaloid nucleus. These results provide evidence that, excluding areas of convergence, the V1R and V2R vomeronasal pathways project to specific areas of the amygdala. These two vomeronasal subsystems are therefore anatomically and functionally separated in the telencephalon.     

Spatial dimension in olfactory coding: a representation of the 2-deoxyglucose patterns of glomerular labeling in the olfactory bulb

A method has been developed for visualizing the patterns of uptake of radioactive 2-deoxyglucose (2-DG) induced in the glomerular layer of rat olfactory bulbs by various olfactory stimuli. Some characteristics of these patterns such as shape, contrast, symmetry, specificity and variability, are discussed. This method is thought to be useful for understanding olfactory coding at the glomerular level.