Neuronal gap junctions in the rat main olfactory bulb,with special reference to intraglomerular gap junctions

The structural features of neuronal gap junction-forming processes in the rat olfactory bulb were analyzed electron microscopically. Gap junctions were present in glomeruli and extraglomerular regions. In extraglomerular regions, mitral/tufted cell somata, dendrites and axon hillock-initial segments made gap junctions and mixed synapses with interneuronal processes, some of which were confirmed to be GABA positive. In glomeruli gap junctions were encountered mainly between mitral/tufted cell dendrites and diverse types of processes; a small population of them were conclusively identified as periglomerular cell dendrites. Gap junction-forming processes frequently received synapses from olfactory nerve terminals, suggesting that they could be type 1 periglomerular cells. However, the majority were GABA negative or only faintly positive and none were tyrosine hydroxylase positive, indicating that they were different from previously reported type 1 periglomerular cells. Furthermore serial sectioning analyses revealed that the majority of those processes forming gap junctions with mitral/tufted dendrites were smooth cylindrical and had few presynaptic sites, indicating that they were different from previously described periglomerular cells. These findings revealed that mitral/tufted cells make gap junctions with diverse types of neurons; and some of these gap junction-forming processes originated from some types of periglomerular cells but others from hitherto uncharacterized neuron type(s).     

Neuronal gap junctions in the mouse main olfactory bulb: morphological analyses on transgenic mice

In the present study we analyzed the structural features of extraglomerular gap junction-forming processes in mouse olfactory bulb electron microscopically. This work complements a previous study in which we analyzed the structural features of neuronal gap junction-forming processes within the glomerulus itself. Furthermore we examined connexin 36 expressing cells in the mouse olfactory bulb by analyzing transgenic mice in which the connexin 36 coding sequence was replaced with histological reporters. In extraglomerular regions, the mitral/tufted cell somata, dendrites and axon hillocks made gap junctions and mixed synapses with interneuronal processes. These gap junctions and synapses were associated with various types of interneuronal processes, including a particular type of sheet-like or calyx-like process contacting the somata or large dendrites of mitral/tufted cells. In the olfactory bulbs of the transgenic mice, connexin 36 was expressed in mitral cells, tufted cells, presumed granule cells and periglomerular cells. Multiple immunofluorescent labelings further revealed that presumed interneurons expressing connexin 36 in the periglomerular region rarely expressed calbindin, calretinin or tyrosine hydroxylase and are likely to comprise a chemically uncharacterized class of neurons. Similarly, interneurons expressing connexin 36 in the granule cell layer were rarely positive for calretinin, which was expressed in numerous presumed granule cells in the mouse main olfactory bulb. In summary, these findings revealed that mitral/tufted cells make gap junctions with diverse types of neurons; in the glomeruli gap junction-forming interneuronal processes originated from some types of periglomerular cells but others from a hitherto uncharacterized neuron type(s), and in the extraglomerular region gap-junction forming processes originate mainly from a subset of cells within the granule cell layer.     

Organization of the main olfactory bulb of lesser hedgehog tenrecs

Using a confocal laser scanning microscope (CLSM) and an electron microscope, we investigated the organization of the main olfactory bulb (MOB) of tenrecs, which were previously included into insectivores but now considered to be in a new order "Afrosoricida" in the superclade 'Afrotheria'. We confirmed that the overall structural organization of the tenrec MOB was similar to that of rodents: (1) the compartmental organization of glomeruli and two types of periglomerular cells we proposed as the common organizational principles were present; (2) there were characteristic dendrodendritic and axo-dendritic synapses in the glomerulus and external plexiform layer (EPL) and gap junctions in glomeruli; and (3) no nidi, particular synaptic regions reported only in laboratory musk shrew and mole MOBs, were encountered. However, instead of nidi, we often observed a few tangled olfactory nerves (ONs) with large irregular boutons in the glomerular-external plexiform layer border zone, with which dendrites of various displaced periglomerular cells were usually found to be intermingled. Electron microscopic (EM) examinations confirmed characteristic large mossy terminal-like ON terminals making asymmetrical synapses to presumed mitral/tufted cell and displaced periglomerular cell dendrites. In addition, gap junctions were also encountered between dendritic processes in these tiny particular regions, further showing their resemblance to glomeruli.     

Immunocytochemical localization of the GABAB2 receptor subunit in the glomeruli of the mouse main olfactory bulb

The olfactory input to the brain is carried out by olfactory nerve axons that terminate in the olfactory bulb glomeruli and make synapses onto dendrites of glutamatergic projection neurons, mitral and tufted cells, and GABAergic interneurons, periglomerular cells. The dendrites are reciprocally connected through asymmetric synapses of mitral/tufted cells with periglomerular cells and symmetric synapses of the opposite direction. Transmission at the first synapse in the olfactory pathway is regulated presynaptically, and this regulation is mediated, in part, by metabotropic GABAB receptors that, when activated, inhibit transmitter release from the olfactory nerve. Functional GABAB receptors are heterodimers composed of the GABAB1 and GABAB2 subunits. Studies using double immunofluorescence have shown colocalization of both subunits in the glomerular neuropil, and ultrastructural studies have localized GABAB1 to extrasynaptic, synaptic, and perisynaptic sites on the plasma membrane of olfactory nerve terminals. We studied the subcellular localization of GABAB2 in the mouse olfactory glomeruli using a subunit-specific antibody and preembedding immunogold labeling. Immunoreactivity for GABAB2 was associated with symmetric dendrodendritic synapses of periglomerular cells with mitral/tufted cells and was localized to the extrasynaptic plasma membrane of presynaptic dendrites, and extrasynaptic, synaptic, and perisynaptic sites on the plasma membrane of postsynaptic dendrites. The results suggest that postsynaptic, and perhaps presynaptic, GABAB receptors may be expressed at GABAergic synapses between dendrites of periglomerular interneurons and projection neurons. Immunolabeling was observed at junctions of the olfactory nerve with mitral/tufted cell dendrites, providing ultrastructural evidence for the expression of the GABAB2 subunit at the primary olfactory synapse.     

Neuronal gap junctions between intraglomerular mitral/tufted cell dendrites in the mouse main olfactory bulb

In the mouse main olfactory bulb (MOB) gap junction-forming processes in glomeruli were analyzed by means of the serial electron microscopical reconstruction. Gap junctions were encountered between diverse types of dendritic processes and thus confirming our previous study on gap junctions in the rat MOB. Importantly, among more than 30 gap junctions examined in serial sections, we encountered 3 gap junctions made between mitral/tufted cell dendrites in the glomerulus. Then we must consider both direct coupling between mitral/tufted cells via gap junctions and indirect coupling between mitral/tufted cells via intervening interneuronal processes as suggested previously.     

Synaptic organization of the glomerulus in the main olfactory bulb: Compartments of the glomerulus and heterogeneity of the periglomerular cells

According to the combinatorial receptor and glomerular codes for odors, the fine tuning of the output level from each glomerulus is assumed to be important for information processing in the olfactory system, which may be regulated by numerous elements, such as olfactory nerves (ONs), periglomerular (PG) cells, centrifugal nerves and even various interneurons, such as granule cells, making synapses outside the glomeruli. Recently, structural and physiological analyses at the cellular level started to reveal that the neuronal organization of the olfactory bulb may be more complex than previously thought. In the present paper, we describe the following six points of the structural organization of the glomerulus, revealed by confocal laser scanning microscopy and electron microscopy analyses of rats, mice and other mammals: (i) the chemical heterogeneity of PG cells; (ii) compartmental organization of the glomerulus, with each glomerulus consisting of two compartments, the ON zone and the non-ON zone; (iii) the heterogeneity of PG cells in terms of their structural and synaptic features, whereby type 1 PG cells send their intraglomerular dendrites into both the ON and non-ON zones and type 2 PG cells send their intraglomerular dendrites only into the non-ON zone, thus receiving either few synapses from the ON terminals, if present, or none at all; (iv) the spatial relationship of mitral/tufted cell dendritic processes with ON terminals and PG cell dendrites; (v) complex neuronal interactions via chemical synapses and gap junctions in the glomerulus; and (vi) comparative aspects of the organization of the main olfactory bulb.     

Chemically defined neuron groups and their subpopulations in the glomerular layer of the rat main olfactory bulb--IV. Intraglomerular synapses of tyrosine hydroxylase-immunoreactive neurons

Synapses of intraglomerular processes of tyrosine hydroxylase-immunoreactive neurons in the rat main olfactory bulb were examined by electron microscopic immunocytochemistry. Prominent characteristics of intraglomerular synapses of tyrosine hydroxylase-immunoreactive elements were that the vast majority (about 80%) of their synaptic inputs were asymmetrical synapses from olfactory nerve terminals and, though far smaller in proportion, one half of the remaining were asymmetrical synapses from mitral/tufted cell dendrites and the other half were symmetrical synapses from gamma-aminobutyric acid-like immunoreactive elements. So far, we have observed no typical reciprocal synapses between tyrosine hydroxylase-immunoreactive processes and mitral/tufted dendrites; however, we have often identified serial synapses; that is, asymmetrical synapses from olfactory nerve terminals or mitral/tufted cell dendrites to tyrosine hydroxylase-immunoreactive processes, and then symmetrical synapses from the latter to different mitral/tufted cell dendrites. These synaptic connections of tyrosine hydroxylase-immunoreactive neurons were very different from those of Calbindin-D(28k)-immunoreactive neurons, which received no synaptic contact directly from olfactory nerve terminals but formed reciprocal synapses with mitral/tufted cells as we analysed previously.Thus, our present and previous electron microscopic studies combined with confocal laser scanning light microscopy clearly indicated for the first time the heterogeneity of periglomerular neurons, not only in their chemical and morphological features, but also in their synaptic organization in the olfactory glomerulus.     

Chromogranin A in the olfactory system of the rat

The olfactory bulb of the rat contains chromogranin A at a similar level as the adrenal gland or the hypophysis as revealed by immunoblots. Olfactory chromogranin A also displays the same size as chromogranin A of endocrine cells. In the hippocampus and other brain regions, we could not detect chromogranin A by immunoblotting. In contrast, chromogranin A messenger ribonucleic acid (using S1 nuclease protection assays) was observed in all brain regions examined, including the olfactory bulb. By in situ hybridization histochemistry with a complementary ribonucleic acid probe (280 nucleotides), and by immunocytochemistry, chromogranin A synthesis could be localized to cell bodies of the mitral cell layer, of the external plexiform layer and of the periglomerular region of the olfactory bulb. Immunocytochemically, chromogranin A was also detected in the central projection areas of mitral and tufted cells in the primary olfactory cortex and the anterior amygdaloid area but not in the olfactory glomeruli, where the incoming olfactory nerve fibers of the primary olfactory neurons establish synaptic contacts. Taken together the data show that chromogranin A, following biosynthesis in the perikarya of the mitral and tufted cells, is specifically transported into their axonal terminals but not into their primary dendrites. We propose that the rat olfactory system could serve as a model for the study of chromogranin A regulation and function in neurons.     

Tufted cell dendrodendritic inhibition in the olfactory bulb is dependent on NMDA receptor activity

Mitral and tufted cells constitute the primary output cells of the olfactory bulb. While tufted cells are often considered as "displaced" mitral cells, their actual role in olfactory bulb processing has been little explored. We examined dendrodendritic inhibition between tufted cells and interneurons using whole cell voltage-clamp recording. Dendrodendritic inhibitory postsynaptic currents (IPSCs) generated by depolarizing voltage steps in tufted cells were completely blocked by the N-methyl-D-aspartate (NMDA) receptor antagonist D,L-2amino-5-phosphonopentanoic acid (D,L-AP5), whereas the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist 2-3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f] quinoxaline-7-sulfonamide (NBQX) had no effect. Tufted cells in the external plexiform layer (EPL) and in the periglomerular region (PGR) showed similar behavior. These results indicate that NMDA receptor-mediated excitation of interneurons drives inhibition of tufted cells at dendrodendritic synapses as it does in mitral cells. However, the spatial extent of lateral inhibition in tufted cells was much more limited than in mitral cells. We suggest that the sphere of influence of tufted cells, while qualitatively similar to mitral cells, is centered on only one or a few glomeruli.     

Spontaneous field potentials in the glomeruli of the olfactory bulb: the leading role of juxtaglomerular cells

Field potentials recorded in the olfactory bulb glomerular layer (GL) are thought to result mainly from activation of mitral and tufted cells. The contribution of juxtaglomerular cells (JG) is unknown. We tested the hypothesis that JG are the main driving force to novel spontaneous glomerular layer field potentials (sGLFPs), which were recorded in rat olfactory bulb slices maintained in an interface chamber. We found that sGLFPs have comparable magnitudes, durations and frequencies both in standard horizontal slices, where all layers with all cell types were present, and in isolated GL slices, where only JG cells were preserved. Hence, the impact of mitral and deep/medium tufted cells to sGLFPs turned out to be minor. Therefore, we propose that the main generators of sGLFPs are JG neurons. We further explored the mechanism of generation of sGLFPs using a neuronal ensemble model comprising all types of cells associated with a single glomerulus. Random orientation and homogenous distribution of dendrites in the glomerular neuropil along with surrounding shell of cell bodies of JG neurons resulted in substantial spatial restriction of the generated field potential. The model predicts that less than 20% of sGLFP can spread from one glomerulus to an adjacent one. The contribution of JG cells to the total field in the center of the glomerulus is estimated as approximately 50% ( approximately 34% periglomerular and approximately 16% external tufted cells), whereas deep/medium tufted cells provide approximately 39% and mitral cells only approximately 10%. Occasionally, some sGLFPs recorded in adjacent or remote glomeruli were cross-correlated, suggesting involvement of interglomerular communication in information coding. These results demonstrate a leading role of JG cells in activation of the main olfactory bulb (MOB) functional modules. Finally, we hypothesize that the GL is not a set of independent modules, but it represents a subsystem in the MOB network, which can perform initial processing of odors.     

Histochemical support for a dopaminergic mechanism in the dendrites of certain periglomerular cells in the rat olfactory bulb

With the Vibratome modification of the Falck-Hillarp technique, and with the indirect immunofluorescence technique for visualizing the first three enzymes in catecholamine synthesis, evidence has been obtained that in the rat olfactory bulb several periglomerular cells are dopaminergic. Both tyrosine hydroxylase and dopadecarboxylase, converting tyrosine to DOPA and DOPA to dopamine (DA), respectively, but not dopamine-β-hydroxylase, converting DA to noradrenaline, are present in periglomerular cell bodies, as well as in their intraglomerular dendrites. These findings suggest that DA may be active at dendrodentritic synapses known to be present between periglomerular and mitral and/or tufted cells.     

Tyrosine hydroxylase-like immunoreactive neurons in the olfactory bulb of the snake,Elaphe quadrivirgata,with special reference to the colocalization of tyrosine hydroxylase- and GABA-like immunoreactivities

Summary The distribution and structural features of tyrosine hydroxylase-like immunoreactive (TH-LI) neurons were studied in the olfactory bulb of a snake, Elaphe quadrivirgata, by using pre-and post-embedding immunocytochemistry at the light microscopic level. In contrast to rodent olfactory bulbs previously reported, many TH-LI neurons were seen not only in the main olfactory bulb (MOB) but also in the accessory olfactory bulb (AOB). With regard to the TH-like immunoreactivity, there appeared no appreciable differences between MOB and AOB. As in mammalian MOB, the majority of TH-LI neurons were clustered in the periglomerular region and appeared to send their dendritic branches into glomeruli, which as a whole make an intense TH-LI band in the glomerular layer (GML). In the external plexiform/mitral cell layer (EPL/ML) of MOB and AOB as well as in the outer sublamina of the internal plexiform layer (OSL) of AOB, an appreciable number of TH-LI neurons were scattered, extending dendritic processes which appeared to make a loose meshwork. TH-LI neurons in EPL/ML (including OSL) appeared to consist of at least two morphologically different types. The first had a small perikaryon and one or two smooth dendrites which usually extended to GML and were frequently confirmed to enter into glomeruli. The second had a larger perikaryon and 2–3 dendrites which branched into several varicose processes extending in EPL/ML/OSL but appeared not to enter into glomeruli. The TH-like immunoreactivity was rarely seen in the internal plexiform layer and internal granule cell layer. The colocalization of GABA-like and TH-like immunoreactivities was further studied. Almost all TH-LI neurons in both EPL/ ML/OSL and GML contained GABA-like immunoreactivity irrespectively of the type of TH-LI cells.Abbreviations in Figures AOB accessory olfactory bulb - MOB main olfactory bulb - Hem hemisphere - ON olfactory nerve layer - VN vomeronasal nerve layer - GM glomerular layer - EP/M external plexiform layer/Mitral cell layer - IP internal plexiform layer - IG internal granular layer - OS outer sublamina of the IPL of AOB - MS middle sublamina of the IPL of AOB - IS inner sublamina of the IPL of AOB     

A Golgi study on the olfactory bulb in the lamprey, Lampetra japonica

The intrinsic organization of the olfactory bulb in the lamprey was studied using the rapid Golgi method. Although not as discrete as in many vertebrates, a laminar organization was recognized. From the periphery inward, the following layers were discernible: the layer of the olfactory fibers, the olfactory glomeruli with the mitral cells, the granule cells, and the ependymal cells. Just beneath the surface of the olfactory bulb, the olfactory fibers extended over the entire bulb forming a dense fiber plexus terminating in the olfactory glomeruli which were arranged in one to two layers internally to the layer of the olfactory fibers. The mitral cells formed no discrete layer and were located mainly around the olfactory glomeruli. The mitral cells in the lamprey were lacking in secondary dendrites, but had two or more primary dendrites which terminated in the olfactory glomeruli. The axons of the mitral cells proceeded inwardly and accumulated diffusely in the granule cell layer which occupied a wide area internally to the layer of the olfactory glomeruli with the mitral cells. The granule cell layer was composed of densely packed small spindle or fusiform axonless cells, the processes of which extended superficially to be distributed in the olfactory glomeruli. At the deepest region of the bulb was a layer of the ependymal cells lining the surface of the olfactory ventricle. The external and internal plexiform layers were not evident. Thus, while the major constituents of the olfactory bulb of the vertebrate could be identified in that of the lamprey, the general laminar organization seemed indiscrete.     

Properties of external plexiform layer interneurons in mouse olfactory bulb slices

In the external plexiform layer (EPL) of the main olfactory bulb, apical dendrites of inhibitory granule cells form large numbers of synapses with mitral and tufted (M/T) cells, which regulate the spread of activity along the M/T cell dendrites. The EPL also contains intrinsic interneurons, the functions of which are unknown. In the present study, recordings were obtained from cell bodies in the EPL of mouse olfactory bulb slices. Biocytin-filling confirmed that the recorded cells included interneurons, tufted cells, and astrocytes. The interneurons had fine, varicose dendrites, and those located superficially bridged the EPL space below several adjacent glomeruli. Interneuron activity was characterized by high frequency spontaneous excitatory postsynaptic potential/currents that were blocked by the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione and largely eliminated by the voltage-sensitive Na+ channel blocker, tetrodotoxin. Interneuron activity differed markedly from that of tufted cells, which usually exhibited spontaneous action potential bursts. The interneurons produced few action potentials spontaneously, but often produced them in response to depolarization and/or olfactory nerve (ON) stimulation. The responses to depolarization resembled responses of late- and fast-spiking interneurons found in other cortical regions. The latency and variability of the ON-evoked responses were indicative of polysynaptic input. Interneurons expressing green fluorescent protein under control of the mouse glutamic acid decarboxylase 65 promoter exhibited identical properties, providing evidence that the EPL interneurons are GABAergic. Together, these results suggest that EPL interneurons are excited by M/T cells via AMPA/kainate receptors and may in turn inhibit M/T cells within spatial domains that are topographically related to several adjacent glomeruli.     

Vasoactive intestinal polypeptide-containing elements in the olfactory bulb of the hedgehog (Erinaceus europaeus)

The distribution of vasoactive intestinal polypeptide (VIP)-immunopositive elements was analyzed in the olfactory bulb (OB) of the Western European hedgehog (Erinaceus europaeus) under light and electron microscopy. The immunoreactivity appeared in an abundant population of periglomerular cells of the glomerular layer, in interneurons of the external plexiform layer, and in a restricted group of deep short-axon cells of the internal plexiform layer, the granule cell layer and the white matter. In the glomerular layer, VIP-containing periglomerular cells constituted a population of non-GABAergic neurons and did not receive synapses from olfactory axons. In the EPL, VIP-immunoreactivity appeared in a morphologically heterogeneous population of GABAergic interneurons, most of them identified as satellite cells and Van Gehuchten cells. These interneurons exerted an abundant and selective innervation of the somata, primary and secondary dendrites of the principal mitral and tufted cells, but did not contact granule cells. Perisomatic innervation of the principal cells followed two different patterns. The first included 'normal' basket-like arrangements of VIP-containing varicosities surrounding the somata of mitral and tufted cells. In the second, a set of satellite cells gave rise to short dendritic shafts that embraced the somata of principal cells in an 'exuberant' basket-like arrangement. These two morphological patterns of perisomatic innervation of principal cells were correlated with a neurochemical specificity of the target. In this sense, the 'exuberant' basket-like structures were always found surrounding a subpopulation of principal cells that did not contain the calcium-binding protein parvalbumin (PV). By contrast, they were never found surrounding the subpopulation of PV-containing principal cells, which only showed 'normal' basket-like structures. This study provides new data on the connectivity and neurochemical features of the hedgehog olfactory bulb and suggests that the olfactory circuits in this species are more complex than those described in other mammals.     

Odor response properties of neighboring mitral/tufted cells in the rat olfactory bulb

Olfactory perception initiates in the nasal epithelium wherefrom olfactory receptor neurons--expressing the same receptor protein--project and converge in two different glomeruli within each olfactory bulb. Recent evidence suggests that glomeruli are isolated functional units, arranged in a chemotopic manner in the olfactory bulb. Exposure to odorants leads to the activation of specific populations of glomeruli. In rodents, about 25-50 mitral/tufted cells project their primary dendrites to a single glomerulus receiving similar sensory input. Yet, little is known about the properties of neighboring mitral/tufted cells connected to one or a few neighboring glomeruli. We used tetrodes to simultaneously record multiple single-unit activity in the mitral cell layer of anesthetized, freely breathing rats while exposed to mixtures of chemically related compounds. First, we characterized the odorant-induced modifications in firing rate of neighboring mitral/tufted cells and found that they do not share odorant response profiles. Individual units showed a long silent (11.01 ms) period with no oscillatory activity. Cross-correlation analysis between neighboring mitral/tufted cells revealed negligible synchronous activity among them. Finally, we show that respiratory-related temporal patterns are dissimilar among neighboring mitral/tufted cells and also that odorant stimulation results in an individual modification that is not necessarily shared by neighboring mitral/tufted cells. These results show that neighboring mitral/tufted cells frequently exhibit dissimilar response properties, which are not consistent with a precise chemotopic map at the mitral/tufted cell layer in the olfactory bulb.     

Intraglomerular inhibition: signaling mechanisms of an olfactory microcircuit

Microcircuits composed of principal neuron and interneuron dendrites have an important role in shaping the representation of sensory information in the olfactory bulb. Here we establish the physiological features governing synaptic signaling in dendrodendritic microcircuits of olfactory bulb glomeruli. We show that dendritic gamma-aminobutyric acid (GABA) release from periglomerular neurons mediates inhibition of principal tufted cells, retrograde inhibition of sensory input and lateral signaling onto neighboring periglomerular cells. We find that L-type dendritic Ca(2+) spikes in periglomerular cells underlie dendrodendritic transmission by depolarizing periglomerular dendrites and activating P/Q type channels that trigger GABA release. Ca(2+) spikes in periglomerular cells are evoked by powerful excitatory inputs from a single principal cell, and glutamate release from the dendrites of single principal neurons activates a large ensemble of periglomerular cells.     

A Golgi study on the main olfactory bulb in the snake Elaphe quadrivirgata

The intrinsic organization of the main olfactory bulb in the snake was studied using the rapid Golgi method. A distinct laminar structure was recognized. From the periphery inward, the following layers were distinguished: the layer of the olfactory fibers, the olfactory glomeruli, the mitral cells, the deep fiber plexus, the granule cells and the ependymal cells. Olfactory fibers derived from the nasal cavity reached the entire surface of the bulb, forming a dense fiber plexus, then swung deeply and terminated in the olfactory glomeruli which were arranged in 2-4 rows. The mitral cell layer occupied a wide zone and was composed of scattered mitral cells. The mitral cells had 2-9 primary dendrites proceeding externally to terminate in the olfactory glomeruli and 2-4 secondary dendrites extending tangentially in the mitral cell layer to be distributed therein. The axons of the mitral cells travelled deeply and entered the layer of the deep fiber plexus. The deep fiber plexus was the path for the bulbar efferent and afferent fibers and could be traced caudally as the main olfactory tract, up to the anterior olfactory nucleus and vicinity. The granule cell layer was composed of small cells, the granule cells, packed closely with no special arrangement. The granule cells had long processes which extended superficially to be distributed mainly in the mitral cell layer. The ependymal cells were located at the deepest layer forming the wall of the olfactory ventricle and generated a long process which extended towards the surface to terminate in the peripheral portion of the bulb. In the snake bulb, the well-documented external and internal plexiform layers were considered to be included in the wide mitral cell layer. Thus, while several specific structures were observed, the fundamental organization of the main olfactory bulb in the snake seemed to be identical to that of the main olfactory bulb in various other vertebrate species.     

Ultrastructural identification of substance P immunoreactive neurons in the main olfactory bulb of the hamster

The neurons containing substance P immunoreactivity in the main olfactory bulb of the hamster are located in the glomerular layer. Their cell bodies lie in the periglomerular region and contain spherical or ovoid nuclei which lack invaginations of the nuclear membrane and tend to be positioned eccentrically in the cell body. Dendrites of these neurons extend throughout the periglomerular region and project into the glomerular neuropil. Within the glomerular neuropil, processes with substance P immunoreactivity contain agranular, spherical synaptic vesicles. Primary olfactory axons, and processes of uncertain origin which contain pleomorphic synaptic vesicles, form synaptic contacts with substance P immunoreactive processes.These ultrastructural findings confirm that the substance P immunoreactive neurons are external tufted cells. Their likely physiological properties are considered in relation to the synaptic organization in the glomerular layer of the main olfactory bulb and to the other putative neurotransmitters or neuromodulators located in this layer.