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     

Ketamine and MK801 attenuate paired pulse inhibition in the olfactory bulb of the rat

Summary We have investigated the effects of the phencyclidine like-compounds ketamine and MK801 on the evoked field potentials of rat olfactory bulb. Low doses of ketamine (3–6 mg/kg) blocked the inhibition of mitral cells by granule cells evoked by stimulation of lateral olfactory tract fibres or by stimulation of olfactory nerve. This blockade was not accompanied by a decrease in granule cell excitation as revealed by field potential recording. MK801 had a similar effect on the inhibition of mitral cells evoked by stimulation of the lateral olfactory tract. As ketamine does not influence the inhibitory action of GABA (Anis et al. 1983) these results suggest that both ketamine and MK801 block inhibition by an action on intrinsic excitatory feed-back circuits in the olfactory bulb.     

Selective neuroinhibitory effects of taurine in slices of rat main olfactory bulb

Taurine is abundant in the main olfactory bulb, exceeding glutamate and GABA in concentration. In whole-cell patch-clamp recordings in rat olfactory bulb slices, taurine inhibited principal neurons, mitral and tufted cells. In these cells, taurine decreased the input resistance and caused a shift of the membrane potential toward the chloride equilibrium potential. The taurine actions were sustained under the blockade of transmitter release and were reversible and dose-dependent. At a concentration of 5 mM, typically used in this study, taurine showed 90% of its maximal effect. GABA(A) antagonists, bicuculline and picrotoxin, blocked the taurine actions, whereas the glycine receptor antagonist strychnine and GABA(B) antagonists, CGP 55845A and CGP 35348, were ineffective. These findings are consistent with taurine directly activating GABA(A) receptors and inducing chloride conductance. Taurine had no effect on periglomerular and granule interneurons. The subunit composition of GABA(A) receptors in these cells, differing from those in mitral and tufted cells, may account for taurine insensitivity of the interneurons. Taurine suppressed olfactory nerve-evoked monosynaptic responses of mitral and tufted cells while chloride conductance was blocked. This action was mimicked by the GABA(B) agonist baclofen and abolished by CGP 55845A; CGP 35348, which primarily blocks postsynaptic GABA(B) receptors, was ineffective. The taurine effect most likely was due to GABA(B) receptor-mediated inhibition of presynaptic glutamate release. Neither taurine nor baclofen affected responses of periglomerular cells. The lack of a baclofen effect implies that functional GABA(B) receptors are absent from olfactory nerve terminals that contact periglomerular cells. These results indicate that taurine decreases the excitability of mitral and tufted cells and their responses to olfactory nerve stimulation without influencing periglomerular and granule cells. Selective effects of taurine in the olfactory bulb may represent a physiologic mechanism that is involved in the inhibitory shaping of the activation pattern of principal neurons.     

Sex steroids modulate the signals from volatile female odors in the accessory olfactory bulb of male mice

We previously reported that male mice detect volatile female odors via the accessory olfactory system, and that these odors activate granule cells in the accessory olfactory bulb (AOB) with a characteristic pattern. We also reported that sex steroids modulate the attraction of male mice to volatile female odors. The present study investigated hormonal modulation of signals from volatile female odors in the AOB with c-Fos immunostaining. After intact male mice were exposed to volatile female odors, there were more c-Fos positive cells in the caudal granule cell layer (GCL) than in the rostral GCL of the AOB. This effect was observed 3 days but not 7 days after castration, suggesting that hormonal deficiency causes the reorganization of the AOB after 3 days. There was no difference in the number of c-Fos positive cells between the rostral and caudal GCL of castrated male mice treated with 17 beta-estradiol (E). In contrast, there were more c-Fos positive cells in the caudal GCL than in the rostral GCL of castrated male mice treated with dihydrotestosterone (DHT). In both DHT- and E-treated castrated male mice, there was no difference in the number of c-Fos positive cells between the rostral GCL and caudal GCL. This finding suggests that E disrupts the effect of DHT, and that androgen is required for maintaining the intact neuronal network of the AOB. The present study suggests that sex steroids modulate the signals from volatile female odors in the AOB of male mice.     

Mapping of benzodiazepine-like immunoreactivity in the rat brain as revealed by a monoclonal antibody to benzodiazepines.

A monoclonal antibody against benzodiazepines (21-7F9) was used to study the distribution of benzodiazepine-like immunoreactivity in the rat brain. Immunodensitometry in combination with image analysis were used for quantification. The results showed a ubiquitous distribution of benzodiazepine-like immunoreactivity throughout the brain. Very high levels of benzodiazepine-like immunoreactivity were found in the Purkinje cell layer of the cerebellum, in the primary olfactory cortex, in the stratum pyramidale of the hippocampus and in the mitral cell layer of the olfactory bulb. High densities of benzodiazepine-like immunoreactivity were found in the granule cell layer of the cerebellum, the pyramidal cell layer of the olfactory tubercle, the granule layer of the dentate gyrus, the arcuate nucleus of the hypothalamus, the mammillary bodies, the interstitial nucleus of Cajal and superficial grey layer of superior colliculus. The substantia nigra pars compacta, the islands of Calleja and layers II, III, V and VI of the cerebral cortex had moderate levels of benzodiazepine-like immunoreactivity. Lower densities were found in the internal granular layer and the external plexiform layer of the olfactory bulb, in the molecular layer of the dentate gyrus, in layers I and IV of the cerebral cortex, in the nucleus caudate-putamen and most of the thalamic nuclei. The lowest density of immunoreactivity was found in the globus pallidus, and the strata radiatum, oriens and lacunosum-moleculare of the hippocampus. The distribution of endogenous benzodiazepine-like immunoreactivity was compared with the distribution of the GABA/benzodiazepine receptor by using both immunocytochemistry and receptor autoradiography. Our studies have shown a clear mismatch between the localization of the benzodiazepine-like immunoreactivity and the GABA/benzodiazepine receptors.     

Complementary postsynaptic activity patterns elicited in olfactory bulb by stimulation of mitral/tufted and centrifugal fiber inputs to granule cells

Main olfactory bulb (MOB) granule cells receive spatially segregated glutamatergic synaptic inputs from the dendrites of mitral/tufted cells as well as from the axons of centrifugal fibers (CFFs) originating in olfactory cortical areas. Dendrodendritic synapses from mitral/tufted cells occur on granule cell distal dendrites in the external plexiform layer (EPL), whereas CFFs preferentially target the somata/proximal dendrites of granule cells in the granule cell layer (GCL). In the present study, tract tracing, and recordings of field potentials and voltage-sensitive dye optical signals were used to map activity patterns elicited by activation of these two inputs to granule cells in mouse olfactory bulb slices. Stimulation of the lateral olfactory tract (LOT) produced a negative field potential in the EPL and a positivity in the GCL. CFF stimulation produced field potentials of opposite polarity in the EPL and GCL to those elicited by LOT. LOT-evoked optical signals appeared in the EPL and spread subsequently to deeper layers, whereas CFF-evoked responses appeared in the GCL and then spread superficially. Evoked responses were reduced by N-methyl-d-aspartate (NMDA) receptor antagonists and completely suppressed by AMPA receptor antagonists. Reduction of extracellular Mg(2+) enhanced the strength and spatiotemporal extent of the evoked responses. These and additional findings indicate that LOT- and CFF-evoked field potentials and optical signals reflect postsynaptic activity in granule cells, with moderate NMDA and dominant AMPA receptor components. Taken together, these results demonstrate that LOT and CFF stimulation in MOB slices selectively activate glutamatergic inputs to the distal dendrites versus somata/proximal dendrites of granule cells.     

Deeply located granule cells and mitral cells undergo apoptosis after transection of the central connections of the main olfactory bulb in the adult rat

The main olfactory bulb (MOB) is the first relay station of the olfactory system: it receives afferents from sensory neurons and sends efferents to the primary olfactory cortex. The MOB also receives many centrifugal afferents from various regions. Transection of peripheral afferents to the MOB has been reported to induce cell death in granule cells. However, little is known about the effect of transection of these central connections of the MOB in adult rats. Here, we used a unilateral olfactory peduncle transection model in the adult rat to examine neuronal degeneration in the MOB. In the MOB ipsilateral to the surgery, the granule cell layer (GCL) was smaller, and the number of mitral cells was decreased compared with the contralateral MOB at 7 days after surgery. Many degenerating cells were present in both the mitral cell layer (MCL) and GCL in the ipsilateral MOB at 3 days after surgery, although there were no obvious changes in the gross morphology. We also found terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick end labeling (TUNEL)-positive cells in the MCL and GCL in the ipsilateral MOB at 3 days after surgery. The majority of the degenerating and TUNEL-positive cells were located in the deep, rather than the superficial, GCL. Immunohistochemistry for activated caspase-9 further supported the occurrence of apoptotic cell death in the mitral and deeply located granule cells. These results indicate that not only axotomized mitral cells, but also deeply located granule cells that were not directly injured, underwent apoptosis after transection of the central connections, and suggest that sensitivities to transection of the central connections differ among granule cells according to their depth in the GCL.     

Morphological characteristics of dopaminergic immunoreactive neurons in the olfactory bulb of the common marmoset monkey (Callithrix jacchus)

The present study describes the distribution of tyrosine hydroxylase (TH)-immunoreactive (IR) elements in the olfactory bulb of the common marmoset monkey (Callithrix jacchus), a primate species by immunohistochemistry. We identified six layers of the olfactory bulb of the common marmoset monkey in sections stained with cresyl violet. The majority of TH-IR cells were found in the glomerular layer. A few TH-IR cells were present in the external plexiform and granule cell layers. TH-IR fibers were identified in all layers of the olfactory bulb. The density of these nerve fibers was high in the internal plexiform and granule cell layers. The results in the olfactory bulb of the common marmoset monkey are generally similar to previous reports in some mammals. These data suggest that TH in the olfactory bulb of the common marmoset monkey may play a role in olfactory transmission via the glomeruli like in other mammals.     

Vasoactive intestinal polypeptide-immunoreactive neurons in the main olfactory bulb of the hedgehog (Erinaceus europaeus)

Vasoactive intestinal polypeptide (VIP) immunoreactivity was localized by the indirect antibody enzyme method (PAP technique) in the main olfactory bulb of the hedgehog. Most VIP-immunoreactive cells were located in the glomerular layer and throughout the external plexiform layer. Fewer cells were observed in the granule cell layer. At the morphological level they exhibit the characteristics of periglomerular, external tufted, superficial short axon, horizontal and Van Gehuchten cells. It should be mentioned that another specific neuronal type was found in the inner third of the external plexiform layer, which is not described in other animals. These results revealed that a high number of intrinsic neuronal types of the olfactory bulb of the hedgehog display a strong VIP immunoreactivity.     

GABAergic and glutamatergic synaptic input to identified granule cells in salamander olfactory bulb.

1. Whole-cell patch clamp recording techniques were applied to granule cells in an in vitro salamander olfactory bulb preparation to study their morphology, membrane properties and pharmacology of postsynaptic responses to electrical stimulation of either the olfactory nerve (ON) or medial olfactory tract (MOT). Optical recordings of the same preparations stained with the voltage-sensitive dye RH414 were also made. 2. Anatomical reconstructions of biocytin-filled granule cells showed that they extend widespread spine-bearing dendrites and an axon-like process that branched within the external plexiform layer. 3. ON or MOT stimulation evoked a long-lasting depolarization, usually generating only a single action potential, in granule cells studied under standard recording conditions. Bath application of bicuculline methiodide (BMI, a GABAA receptor antagonist, 20 or 25 microM) enhanced the spontaneous and electrically evoked excitatory drive to granule cells. 4. The electrically evoked synaptic responses consisted of both excitatory and inhibitory synaptic inputs. Using symmetrical Cl- conditions inside and outside the cell to enhance Cl- currents, spontaneous and electrically driven BMI-sensitive inhibitory postsynaptic currents (IPSCs) were revealed, indicating that granule cells receive GABAergic synaptic input. 5. Bath application of GABA (250 microM to 1 mM) shunted and hyperpolarized granule cells as observed directly from whole-cell recordings and indirectly from cell-attached patch single channel recordings. 6. Bath application of the glutamate receptor antagonists 6-cyano-2,3-dihydroxy-7-nitroquinoxaline (CNQX, 10 microM) and/or DL-2-amino-5-phosphonopentanoic acid (DL-AP5, 100 microM) showed that granule cell dendrodendritic EPSPs are shaped by both non-NMDA and NMDA receptors. 7. The time course and pharmacological sensitivity of both single granule cell responses and ensemble responses recorded optically in the deeper layers of the bulb correlated well. 8. It is concluded that salamander granule cells integrate several types of synaptic input, may have both dendritic and axonal output, and play a major role in generating voltage-sensitive dye signals in the olfactory bulb.     

Distribution of enkephalin-like immunoreactivity in the rat main olfactory bulb

Enkephalin-like immunoreactivity was localized within the main olfactory bulb of the rat using immunohistochemical techniques. These studies utilized well characterized antisera directed to either leu⁵- or met⁵-enkephalin. Specificity was established by absorption of the antisera with either 10 μM synthetic leu⁵- or met⁵-enkephalin.Specific enkephalin-like immunoreactivity was observed within several different cell populations including (1) periglomerular cells, (2) granule cells and their processes within the external plexiform layer and (3) occasional short-axon (horizontal) cells within the granule and external plaxiform layers. The granule cell layer contained the greatest number of immunoreactive cells. Only a limited number of immunoreactive cells were found in both the periglomerular and granule cell layers, suggesting the enkephalin-containing neurons represent a sub-population within each layer.The absence of immunoreactive processes in the periventribular white matter, as well as the morphologies of immunoreactive bulbar neurons, indicates that enkephalin is found exclusively within intrinsic olfactory bulb neurons.     

Immunocytochemical localization of GABA neurons and dopamine neurons in the rat main and accessory olfactory bulbs

Immunocytochemical localization of GABA neurons and dopamine neurons in the rat olfactory bulb was obtained with sheep antiserum to glutamate decarboxylase (GAD) and rabbit antiserum to tyrosine hydroxylase (TH). GAD-positive neurons include periglomerular cells, granule cells, superficial and deep short axon cells. TH-positive neurons represent periglomerular cells. Two-color immunocytochemistry shows that GABA and dopamine periglomerular cells are separate populations. The accessory olfactory bulb has rare dopamine cells and few superficial short axon cells. Radial gradients of GAD-immunostaining are evident in the main but not in the accessory olfactory bulb.     

多巴胺受体在大鼠嗅球的表达及其在帕金森病大鼠嗅觉障碍中的作用

目的 观察多巴胺受体在大鼠嗅球（OB）的表达与分布,探讨左旋多巴（L-DOPA）治疗对帕金森病（PD）大鼠嗅觉的影响。 方法 采用免疫印迹、免疫荧光等方法观察多巴胺受体在大鼠OB中的表达;6-羟多巴胺（6-OHDA）双侧注射建立PD大鼠模型,检测L-DOPA治疗对PD大鼠嗅觉功能及谷氨酸脱羧酶（GAD）和脑源性神经营养因子（BNDF）表达的影响。 结果 嗅球内D1和D2两种多巴胺受体亚型表达含量高。D1和D2在颗粒细胞层（GCL）内GAD阳性的γ-氨基丁酸（GABA）能神经元上大量表达,被酪氨酸羟化酶（TH）阳性神经纤维终末包绕。PD大鼠OB内GCL层TH蛋白表达明显下降（0.05±0.01 vs 0.01±0.00,P<0.001）。L-DOPA治疗后,PD大鼠找寻食物小球时间显著降低［（624.4±113.4）s vs(312.4±79.35)s,P<0.05］,OB内BDNF表达显著升高（0.02±0.01 vs 0.07±0.01,P<0.01）。 结论 D1和D2在GCL层GABA能神经元大量表达。L-DOPA治疗可缓解PD大鼠嗅觉障碍,可能与激活OB内GABA能神经元上的D1和D2复合体,进而改善BDNF表达有关。     

Expression of connexin 57 in the olfactory epithelium and olfactory bulb

In the visual system, deletion of connexin 57 (Cx57) reduces gap junction coupling among horizontal cells and results in smaller receptive fields. To explore potential functions of Cx57 in olfaction, in situ hybridization and immunohistochemistry methods were used to investigate expression of Cx57 in the olfactory epithelium and olfactory bulb. Hybridization signal was stronger in the olfactory epithelial layer compared to the connective tissue underneath. Within the sensory epithelial layer, hybridization signal was visible in sublayers containing cell bodies of basal cells and olfactory neurons but not evident at the apical sublayer comprising cell bodies of sustentacular cells. These Cx57 positive cells were clustered into small groups to form different patterns in the olfactory epithelium. However, individual patterns did not associate with specific regions of olfactory turbinates or specific olfactory receptor zones. Patched distribution of hybridization positive cells was also observed in the olfactory bulb and accessory olfactory bulb in layers where granule cells, mitral cells, and juxtaglomerular cells reside. Immunostaining was observed in the cell types described above but the intensity was weaker than that in the retina. This study has provided anatomical basis for future studies on the function of Cx57 in the olfactory system.     

Olfactory system modulation of hippocampal cell death

The hippocampal dentate gyrus is a major recipient of olfactory input in rodents, via connections from the olfactory (piriform) cortex and the olfactory bulb to the entorhinal cortex. Given this connectivity and the known role of activity in dentate gyrus granule cell survival, the present experiment examined the immediate effects of loss of olfactory input to the hippocampus on apoptosis. Adults rats underwent unilateral or bilateral olfactory bulb ablations (OBX), and allowed to recover 24–72 h before the piriform cortex and hippocampal dentate gyrus were processed for terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling [TUNEL] of apoptotic cells. OBX transiently increased TUNEL-positive cells in the ipsilateral piriform cortex and dentate gyrus. Increased TUNEL-labeling was apparent within 24 h in both structures, but was more extensive and prolonged in piriform cortex. The results suggest a trans-synaptic regulation of cell survival through at least two synapses.     

Microglia in the olfactory bulb of rats during postnatal development and olfactory nerve injury with zinc sulfate: a lectin labeling and ultrastrucutural study

Using isolectin (GSA I-B4) as a marker, this study examined the possible alterations of lectin-labeled membranous glycoproteins in microglial cells in the olfactory bulb of normal development and under experimentally induced degeneration. In light microscopy, several morphological types of microglial cells representing different degrees of cell differentiation were distributed in the bulb laminae. A gradient of microglial differentiation extending from the intermediate to superficial and intermediate to deep occurs in the bulb layers. The differentiation gradient and lectin labeling pattern of microglial cells in the developing bulb resembled those in other areas of the brain tissues. Differentiating microglia showed a gradual diminution of lectin staining when the nascent round cells transformed into the mature ramified cells. Microglia in the external plexiform layer of the olfactory bulb were the first to mature and the cells expressed very weak lectin reactivity. In mature or adult rats, some microglial cells showing intense lectin labeling were observed in the olfactory nerve layer, granule cell layer and subependymal layer. Ultrastructurally, lectin labeling was localized at the trans saccules of the Golgi apparatus. Microglial cells in other bulb laminae, however, exhibited a negative reaction for the isolectin at the Golgi apparatus. Following intranasal irrigation of zinc sulfate, some microglial cells in the olfactory nerve layer and glomerular layer were activated to become phagocytic cells with increased lectin labeling at their ramified processes. GSA I-B4 staining was also localized at their trans saccules of the Golgi apparatus. The lectin labeling pattern of these phagocytic cells resembled that of differentiating microglia in postnatal bulbs, suggesting that bulb microglia in the lesioned sites were activated through cell dedifferentiation into macrophages.     

The bilateral bulbar projections of the primary olfactory neurons in the frog

Summary Whether or not the frog olfactory neuroreceptor cells project bilaterally to the olfactory bulb is still a debated question. We therefore decided to ascertain whether bilateral projections of the primary olfactory input exist and if so to investigate their extent. Reproducible extracellular bilateral bulbar potentials were recorded in the frog following electrical stimulation of dorsal or ventral olfactory nerve bundles. The general features of the contralateral evoked responses were very similar to those of the ipsilateral response. The contralateral response disappeared after transection of the rostral part of the olfactory interbulbar adhesion but not following transection of the habenular or anterior commissures. Horseradish peroxidase labelling showed that the fiber terminations of the olfactory nerve bundle was not restricted to the ipsilateral olfactory bulb but included the medial aspects of the contralateral bulb. The intertelencephalic sections increased the magnitude of the ipsilateral evoked responses. Olfactory bulb isopotential maps revealed a rough topographical correspondence between the olfactory neuroepithelium and bulb along the medio-lateral axis as well as along the dorso-ventral axis. In addition, a projection of the medial and central part of the olfactory sac to the medial part of the contralateral olfactory bulb through the interbulbar adhesion was confirmed. These findings suggest first, that the fibers from the neuro-receptors located in either the ventral or the dorsal olfactory mucosae project to both olfactory bulbs, and second, that the left and right bulbs exert a constant inhibition on each other via the habenular commissure.Abbreviations AON anterior olfactory nucleus - ax olfactory neuroreceptor axon - BA bulbar adhesion - D_I latero-dorsal olfactory nerve bundle - D_II centro-dorsal olfactory nerve bundle - D_III mediodorsal olfactory nerve bundle - EPL external plexiform layer - GL glomerular layer - gl glomerulus - GRL granular cell layer - MOB main olfactory bulb - m mitral cell - MBL mitral cell body layer - ON olfactory nerve - V lateral ventricule - V_I latero-ventral ol-factory nerve bundle - V_II centro-ventral olfactory nerve bundle - V_III medio-ventral olfactory nerve bundle - VN vomero-nasal nerve     

Tyrosine hydroxylase immunoreactive structures in the aged human olfactory bulb and olfactory peduncle

We studied the anatomical distribution of dopaminergic structures in the normal, aged, human olfactory bulb and olfactory peduncle with a monoclonal antibody against tyrosine hydroxylase. Three different tyrosine hydroxylase containing cell groups are present in the olfactory bulbs: (1) a group of round, medium-sized cells within and around the glomeruli; (2) cells in the external plexiform layer; and (3) cells that are scattered in the stratum album. Occasionally, a few labeled neurons can be observed in the granule cell layer. In the olfactory peduncle a few labeled cells are present in the superficial layers just underneath the pia. Tyrosine hydroxylase containing terminal-like structures are present in the glomerular layer and the external plexiform layer. In a few cases dense terminal labeling is also observed in the cell groups that constitute the anterior olfactory nucleus. In the olfactory peduncle scattered labeled fibers are present. In addition, the present study makes clear that quantitative differences exist between the individual cases for which no explanation could be found.     

Olfactory bulb interneurons releasing NO exhibit the Reelin receptor ApoEr2 and part of those targeted by NO express Reelin

Nitric oxide (NO) and Reelin both modulate neuronal plasticity in developing and mature synaptic networks. We recently showed a loss of neuronal nitric oxide synthase (nNOS) protein in the olfactory bulb of reeler mutants and advanced the hypothesis that the Reelin and NO signalling pathways may influence each other. We now studied the distribution of NO sensitive guanylyl cyclase (NOsGC), Reelin and its receptor Apolipoprotein E2 (ApoEr2) in the olfactory bulb by multiple fluorescence labelling and tested whether nNOS and ApoEr2 colocalize in this area. We also essayed the protein content of NOsGC in the reeler olfactory bulb and tested whether there are any changes in nNOS and NOsGC protein in other reeler brain areas. Olfactory bulb interneurons expressing ApoEr2 and nNOS are only few in the glomerular layer but represent the large majority of granule cell layer interneurons. Conversely, NOsGC interneurons are rare in the granule cell layer and abundant as periglomerular cells. Reelin containing periglomerular cells almost entirely belong to the NOsGC subset. These data further support the hypothesis of a reciprocal signalling between Reelin/NOsGC and ApoEr2/nNOS expressing neurons to affect olfactory bulb activity. We also show that a significant rise in NOsGC content accompanies the decrease of nNOS protein in the reeler olfactory bulb. The same reciprocal changes present in the cortex/striatum and the hippocampus of reeler mice. Thus, the influence that the deficit of extracellular Reelin seems to exert on nNOS and its receptor is not limited to the olfactory bulb but is a general feature of the reeler brain.     

GABAergic basal forebrain afferents innervate selectively GABAergic targets in the main olfactory bulb

In this work we have analyzed the targets of the GABAergic afferents to the main olfactory bulb originating in the basal forebrain of the rat. We combined anterograde tracing of 10 kD biotinylated dextran amine (BDA) injected in the region of the horizontal limb of the diagonal band of Broca that projects to the main olfactory bulb, with immunocytochemical detection of GABA under electron microscopy or vesicular GABA transporter (vGABAt) under confocal fluorescent microscopy. GABAergic afferents were identified as double labeled BDA-GABA boutons. Their targets were identified by their ultrastructure and GABA content. We found that GABAergic afferents from the basal forebrain were distributed all over the bulbar lamination, but were more abundant in the glomerular and inframitral layers (i.e. internal plexiform layer and granule cell layer). The fibers had thick varicosities with abundant mitochondria and large perforated synaptic specializations. They contacted exclusively GABAergic cells, corresponding to type 1 periglomerular cells in the glomerular layer, and to granule cells in inframitral layers. This innervation will synchronize the bulbar inhibition and consequently the response of the principal cells to the olfactory input. The effect of the activation of this pathway will produce a disinhibition of the bulbar principal cells. This facilitation might occur at two separate levels: first in the terminal tufts of mitral and tufted cells via inhibition of type 1 periglomerular cells; second at the level of the firing of the principal cells via inhibition of granule cells. The GABAergic projection from the basal forebrain ends selectively on interneurons, specifically on type 1 periglomerular cells and granule cells, and is likely to control the activity of the olfactory bulb via disinhibition of principal cells. Possible similarities of this pathway with the septo-hippocampal loop are discussed.