Centre-surround inhibition among olfactory bulb glomeruli.

Centre-surround inhibition--the suppression of activity of neighbouring cells by a central group of neurons--is a fundamental mechanism that increases contrast in patterned sensory processing. The initial stage of neural processing in olfaction occurs in olfactory bulb glomeruli, but evidence for functional interactions between glomeruli is fragmentary. Here we show that the so-called 'short axon' cells, contrary to their name, send interglomerular axons over long distances to form excitatory synapses with inhibitory periglomerular neurons up to 20-30 glomeruli away. Interglomerular excitation of these periglomerular cells potently inhibits mitral cells and forms an on-centre, off-surround circuit. This interglomerular centre-surround inhibitory network, along with the well-established mitral-granule-mitral inhibitory circuit, forms a serial, two-stage inhibitory circuit that could enhance spatiotemporal responses to odours.

Developmental regulation of metabotropic glutamate receptor 1 splice variants in olfactory bulb mitral cells.

Alternative splicing of the metabotropic glutamate receptor 1 (mGluR1) receptor gene generates two major receptor isoforms, mGluR1a and mGluR1b, differing in intracellular function and distribution. However, little is known on the expression profiles of these variants during development. We examined the mRNA expression profile of mGluR1a/b in microdissected layers and acutely isolated mitral cells in the developing mouse olfactory bulb. This analysis showed that the two mGluR1 variants are differentially regulated within each bulb layer. During the first postnatal week, the mGluR1a isoform replaces GluR1b in the microdissected mitral cell layer (MCL) and in isolated identified mitral cells, coinciding with a developmental epoch of mitral cell dendritic reorganization. Although mGluR1a mRNA is expressed at high levels in both the adult external plexiform layer (EPL) and MCL, Western blotting analysis reveals a marked reduction of the mGluR1a protein in the MCL, where mitral cell bodies are located, and strong labeling in the EPL, which contains mitral cell dendrites. This suggests that there is increased dendritic trafficking efficiency of the receptor in adult. The temporal and spatial shift in mGluR1b/a expression suggests distinct roles of the mGluR1 isoforms, with mGluR1b potentially involved in the early mitral cell maturation and mGluR1a in dendritic and synapse function.

Quantitative analysis of neuronal diversity in the mouse olfactory bulb.

Olfactory sensory information is processed and integrated by circuits within the olfactory bulb. Golgi morphology suggests the olfactory bulb contains several major neuronal classes. However, an increasingly diverse collection of neurochemical markers have been localized in subpopulations of olfactory bulb neurons. While the mouse is becoming the animal model of choice for olfactory research, little is known about the proportions of neurons expressing and coexpressing different neurochemical markers in this species. Here we characterize neuronal populations in the mouse main olfactory bulb, focusing on glomerular populations. Immunofluorescent labeling for: 1) calretinin, 2) calbindin D-28K (CB), 3) parvalbumin, 4) neurocalcin, 5) tyrosine hydroxylase (TH), 6) the 67-kDa isoform of GAD (GAD67), and 7) the neuronal marker NeuN was performed in mice expressing green fluorescent protein under the control of the glutamic acid decarboxylase 65kDa (GAD65) promoter. Using unbiased stereological cell counts we estimated the total numbers of cells and neurons in the bulb and the number and percentage of neurons expressing and coexpressing different neurochemical populations in each layer of the olfactory bulb. Use of a genetic label for GAD65 and immunohistochemistry for GAD67 identified a much larger percentage of GABAergic neurons in the glomerular layer (55% of all neurons) than previously recognized. Additionally, while many glomerular neurons expressing TH or CB coexpress GAD, the majority of these neurons preferentially express the GAD67 isoform. These data suggest that the chemospecific populations of neurons in glomeruli form distinct subpopulations and that GAD isoforms are preferentially regulated in different neurochemical cell types.

Reduced levels of Cacna1c attenuate mesolimbic dopamine system function.

Genetic variation in CACNA1C, which codes for the L-type calcium channel (LTCC) Cav 1.2, is associated with clinical diagnoses of bipolar disorder, depression and schizophrenia. Dysregulation of the mesolimbic-dopamine (ML-DA) system is linked to these syndromes and LTCCs are required for normal DAergic neurotransmission between the ventral tegmental area (VTA) and nucleus accumbens (NAc). It is unclear, however, how variations in CACNA1C genotype, and potential subsequent changes in expression levels in these regions, modify risk. Using constitutive and conditional knockout mice, and treatment with the LTCC antagonist nimodipine, we examined the role of Cacna1c in DA-mediated behaviors elicited by psychomotor stimulants. Using fast-scan cyclic voltammetry, DA release and reuptake in the NAc were measured. We find that subsecond DA release in Cacna1c haploinsufficient mice lacks normal sensitivity to inhibition of the DA transporter (DAT). Constitutive haploinsufficiency of Cacna1c led to attenuation of hyperlocomotion following acute administration of stimulants specific to DAT, and locomotor sensitization of these mice to the DAT antagonist GBR12909 did not reach the same level as wild-type mice. The maintenance of sensitization to GBR12909 was attenuated by administration of nimodipine. Sensitization to GBR12909 was attenuated in mice with reduced Cacna1c selectively in the VTA but not in the NAc. Our findings show that Cacna1c is crucial for normal behavioral responses to DA stimulants and that its activity in the VTA is required for behavioral sensitization. Cacna1c likely exerts these effects through modifications to presynaptic ML-DA system function.

Sex-dependent mitophagy and neuronal death following rat neonatal hypoxia-ischemia.

Males are more susceptible than females to long-term cognitive deficits following neonatal hypoxic-ischemic encephalopathy (HIE). Mitochondrial dysfunction is implicated in the pathophysiology of cerebral hypoxia-ischemia (HI), but the influence of sex on mitochondrial quality control (MQC) after HI is unknown. Therefore, we tested the hypothesis that mitophagy is sexually dimorphic and neuroprotective 20-24h following the Rice-Vannucci model of rat neonatal HI at postnatal day 7 (PN7). Mitochondrial and lysosomal morphology and degree of co-localization were determined by immunofluorescence in the cerebral cortex. No difference in mitochondrial abundance was detected in the cortex after HI. However, net mitochondrial fission increased in both hemispheres of female brain, but was most extensive in the ipsilateral hemisphere of male brain following HI. Basal autophagy, assessed by immunoblot for the autophagosome marker LC3BI/II, was greater in males suggesting less intrinsic reserve capacity for autophagy following HI. Autophagosome formation, lysosome size, and TOM20/LAMP2 co-localization were increased in the contralateral hemisphere following HI in female, but not male brain. An accumulation of ubiquitinated mitochondrial protein was observed in male, but not female brain following HI. Moreover, neuronal cell death with NeuN/TUNEL co-staining occurred in both hemispheres of male brain, but only in the ipsilateral hemisphere of female brain after HI. In summary, mitophagy induction and neuronal cell death are sex dependent following HI. The deficit in elimination of damaged/dysfunctional mitochondria in the male brain following HI may contribute to male vulnerability to neuronal death and long-term neurobehavioral deficits following HIE.

Expression of galectin-1 in the mouse olfactory system.

Primary sensory olfactory axons arise from the olfactory neuroepithelium that lines the nasal cavity and then project via the olfactory nerve into the olfactory bulb. The beta-galactoside binding lectin, galectin-1, and its laminin ligand have been implicated in the growth of these axons along this pathway. In galectin-1 null mutant mice, a subpopulation of primary sensory olfactory axons fails to reach its targets in the olfactory bulb. In the present study we examined the spatiotemporal expression pattern of galectin-1 in normal mice in order to understand its role in the development of the olfactory nerve pathway. At E15.5, when olfactory axons have already contacted the olfactory bulb, galectin-1 was expressed in the cartilage and mesenchyme surrounding the nasal cavity but was absent from the olfactory neuroepithelium, nerve and bulb. Between E16.5 and birth galectin-1 began to be expressed by olfactory nerve ensheathing cells in the lamina propria of the neuroepithelium and nerve fibre layer. Galectin-1 was neither expressed by primary sensory neurons in the olfactory neuroepithelium nor by their axons in the olfactory nerve. Laminin, a galectin-1 ligand, also exhibited a similar expression pattern in the embryonic olfactory nerve pathway. Our results reveal that galectin-1 is dynamically expressed by glial elements within the nerve fibre layer during a discrete period in the developing olfactory nerve pathway. Previous studies have reported galectin-1 acts as a substrate adhesion molecule by cross-linking primary sensory olfactory neurons to laminin. Thus, the coordinate expression of galectin-1 and laminin in the embryonic nerve fibre layer suggests that these molecules support the adhesion and fasciculation of axons en route to their glomerular targets.

N-acetyl-lactosamine in the rat olfactory system: expression and potential role in neurite growth.

Primary sensory olfactory neurons exhibit a mosaic topographical projection from the olfactory neuroepithelium in the nasal cavity to the olfactory bulb formation of the telencephalon. Axons from primary neurons that are widely scattered in the epithelium terminate in discrete regions of the olfactory bulb. It has been hypothesised that carbohydrates present on the surface of primary olfactory axons mediate selective fasciculation and the formation of the topographical pathway. We examined the expression of the disaccharide N-acetyl-lactosamine in both the developing and the adult rat olfactory system. N-acetyl-lactosamine was expressed by all primary sensory olfactory neurons and by their terminations in the olfactory bulb throughout embryonic development and early postnatal life. In adults, N-acetyl-lactosamine was restricted to a subpopulation of primary sensory olfactory neurons that were dispersed throughout the neuroepithelium but that projected predominantly to the ventrolateral and ventromedial surfaces of the olfactory bulb. The axons of these neurons sort out in the outer layer of the bulb and preferentially self-fasciculate to form distinct axon bundles that terminate within select glomeruli. The role of N-acetyl-lactosamine in axon growth was tested by culturing primary sensory olfactory neurons on substrate-bound carbohydrates. Olfactory neuroepithelium cultures from both embryonic and postnatal rats revealed that substrate-bound N-acetyl-lactosamine was a strong and specific neurite growth-promoting agent. These data suggest that, during development of the olfactory projection, N-acetyl-lactosamine, which is present on all olfactory axons, acts as a nonselective permissive substrate for axon growth. In adults, however, the restricted distribution of N-acetyl-lactosamine on a subpopulation of axons may facilitate sorting out and self-fasciculation, which is necessary for preserving the mosaic nature of the olfactory pathway in this highly plastic region of the nervous system. These results support the hypothesis that cell surface carbohydrates are involved in axon growth in the olfactory system.

Identification of cells expressing galectin-1, a galactose-binding receptor, in the rat olfactory system.

Interactions between carbohydrate ligands and their receptors play an important role in cell adhesion and migration in many tissues. Cell-surface carbohydrates that contain terminal galactose have previously been implicated in primary sensory axon growth in the rodent olfactory system. The aim of the present study was to determine whether galectin-1, a galactose-binding receptor, was expressed within the rat primary olfactory pathway. Immunohistochemical and in situ hybridisation analyses revealed expression of galectin-1 by primary sensory olfactory neurons during the major embryonic period of axonogenesis as well as in maturity. In the adult olfactory bulb, galectin-1 was expressed by both second-order projection neurons and interneurons and was selectively localised to the synaptic neuropil layers. Mitral cells, the principal postsynaptic target of primary olfactory axons, began expressing this lectin soon after genesis and maintained high levels into adulthood. The expression of galectin-1 in the primary olfactory pathway and olfactory bulb neuropil suggests a role for this lectin both in the initial formation and in the subsequent maintenance of neuronal connections between the peripheral and the central olfactory neurons as well as between neurons within the bulb.

Radial glia development in the mouse olfactory bulb.

Radial glia are critical for cell migration and lamination of the cortex. In most developing cortical structures, radial glia, as their name suggests, extend processes from the ventricle to the pia in regular parallel arrangements. However, immunohistochemical labeling from several laboratories suggests that radial glia have a more branched morphology in the olfactory bulb. To investigate the morphology of radial glia in the mouse olfactory bulb we (1) labeled radial glia and olfactory receptor neuron axons at 24-hour intervals by immunohistochemistry; and (2) developed a novel method of generating and applying "nanocrystals" of 1,1'-dioctadecyl-3,3,3',3'- tetramethylindocarbocyanine perchlorate (DiI) to the ventricle surface such that the processes of single olfactory bulb radial glia are labeled in the embryonic olfactory bulb. We examined the structure and interactions of radial glia with ingrowing olfactory receptor neuron (ORN) axons in late embryonic olfactory bulb development. These results showed that olfactory bulb radial glia do not form straight parallel structures as do radial glia in the neocortex but rather have a convoluted trajectory from the ventricle to the bulb surface. Moreover, olfactory bulb radial glia consistently extend tangential branches at the level of the internal plexiform layer. Beginning at embryonic day 17.5, two types of radial glia can be distinguished: type I radial glia have a process that extends from the ventricle into the glomerular layer. These apical processes form highly restricted tufts, or "glial glomeruli" at the same time that ORN axons are forming "axonal glomeruli." In type II radial glia the apical process does not enter the glomerular layer but instead ramifies within the external plexiform layer. The tight spatiotemporal relationship between the glomerulization of radial glia processes and ORN axons during development suggest that radial glia processes could play a role in the formation and/or stabilization of mammalian glomeruli.

Development of the olfactory bulb: evidence for glia-neuron interactions in glomerular formation.

Olfactory bulb (OB) glomeruli have long been considered functional units in the processing of odor information. Recently, it has been shown that axons from olfactory receptor neurons (ORNs) expressing the same odorant receptor gene converge onto two or a few topographically fixed glomeruli in the OB. The interactions between ORN axons, mitral/tufted cell dendrites, juxtaglomerular (JG) cells, and glial cells during the development of glomeruli is of great importance in light of this receptor gene glomerular topography in the primary olfactory projection. To explore the development of mammalian olfactory glomeruli, we investigated the relationships among radial glia (RG), astrocytes, ORNs, JG cells, mitral/tufted cell dendrites, and olfactory Schwann cells throughout embryonic and early postnatal development. Our results indicate that glomeruli are formed through an invariant sequence of cellular events: (1) pioneering ORN axons contact the rostral telencephalon at approximately E11-14, which coincides with the onset of morphologic changes in telencephalic RG; (2) at E15-16, RG branch and begin to form two plexuses, one located in the subventricular layer and the other superficial to the presumptive mitral cell layer; (3) at E17-18, ORN axons accumulate in a dense band superficial to the outer radial glia plexus; (4) at E19-20, processes from RG and astrocytes begin to ramify to form glial tufts, or glial glomeruli. Coincident with the formation of these glial glomeruli, ORN axons intermingle with the glial processes and form proto-glomeruli; (5) at E21 to P0, JG cells begin to migrate into position surrounding glomeruli, (6) and at P4, the apical tuft of mitral cells becomes restricted to a single glomerulus. Interestingly, glomerular development also occurs in a distinct rostral to caudal gradient. That is, glomeruli in the rostral OB develop earlier than those in the caudal OB, but the sequence of cellular events at any point in the bulb is invariant. These results demonstrate that glomeruli are formed in a specific spatiotemporal sequence beginning with ORN axon-glia contacts, then JG cell arrival, and finally mitral cell apical dendrite restriction.

Expression of semaphorins in developing and regenerating olfactory epithelium.

Semaphorins provide signals that guide growing axons to their appropriate destinations. The secreted semaphorin, Sema3A, mediates repulsive effects on axons from various neuronal populations in embryonic rats. The authors localized Sema3A mRNA expression in the primary olfactory pathway during development, in adult rats, and in adult rats that were subjected to a unilateral olfactory bulbectomy. Developing rats at ages from embryonic day 14 (E14) to E19 expressed Sema3A in the olfactory receptor neurons (ORNs) of the olfactory epithelium and in chondrogenic structures surrounding the nasal cavity. In vitro, ORN axons at E14 avoided substrate-bound Sema3A. Low levels of Sema3A expression persisted in the normal adult epithelium both in ORNs scattered throughout the epithelium and in small clusters. Three days after a unilateral olfactory bulbectomy, Sema3A transcript levels increased in regenerating neurons. High levels of Sema3A transcript were found at 1 week postbulbectomy, persisted for 2 weeks, and diminished by 3 weeks. Several other murine semaphorins (Sema4A, Sema4B, and Sema4C) were expressed differentially in the primary olfactory pathway both during development and regeneration. These findings suggest that Sema3A and perhaps other semaphorins play a role in directing ORNs out of the epithelium and to the olfactory bulb, their target structure, during both development and regeneration.

Olfactory receptor neurons express D2 dopamine receptors.

The role of the dopamine (DA) in the olfactory bulb (OB) was explored by determining which of the potential target cells express dopamine receptors (DARs). Previously, it was reported that D2-like DAR (D2, D3, and D4 subtypes) radioligand binding is restricted to the outer layers of the OB. The neuronal elements present only in these layers are the axons of the olfactory receptor neurons (ORNs) and the juxtaglomerular (JG) neurons of the glomerular layer. Based on this pattern of D2-like ligand binding, it was suggested that D2-like receptors might be located presynaptically on ORN terminals. The present study was undertaken to investigate this hypothesis. In the outer bulb layers of rats in which the ORNs were destroyed by nasal lavage with ZnSO(4), D2-like radioligand binding was reduced severely. The receptor subtype D2 mRNA, but not D3 mRNA, was detected in adult rat olfactory epithelial tissue. By using in situ hybridization, this D2 mRNA was located preferentially in epithelial layers that contain ORN perikarya. D2 mRNA was eliminated after bulbectomy, a manipulation known to cause retrograde degeneration of the mature ORNs. Taken together, the surgical manipulations indicate that mature ORNs express D2 DARs and are consistent with the hypothesis that functional receptors are translocated to their axons and terminals in the bulb. This suggests that dopamine released from JG interneurons could be capable of presynaptically influencing neurotransmission from the olfactory nerve terminals to OB target cells through the D2 receptor.

Substrate-bound carbohydrates stimulate signal transduction and neurite outgrowth in an olfactory neuron cell line.

Subpopulations of olfactory receptor neurons, which are dispersed throughout the olfactory neuroepithelium, express specific cell surface carbohydrates and project to discrete regions of the olfactory bulb. Cell surface carbohydrates such as N-acetyl-lactosamine have been postulated to mediate sorting and selective fasciculation of discrete axon subpopulations during development of the olfactory pathway. Substrate-bound N-acetyl-lactosamine promotes neurite outgrowth by both clonal olfactory receptor neuron cell lines and olfactory receptor neurons in vitro, indicating that cell surface carbohydrates may be ligands for receptor-mediated stimulation of axon growth in vivo. In the present study, the role of transmembrane signaling in N-acetyl-lactosamine-stimulated neurite outgrowth was examined in the clonal olfactory neuron cell line 4.4.2. Substrate-bound N-acetyl-lactosamine stimulated neurite outgrowth which was specifically inhibited by antagonists to N- and L-type calcium channels and to tyrosine kinase phosphorylation. These results indicate that N-acetyl-lactosamine can evoke transmembrane receptor-mediated responses capable of influencing neurite outgrowth.

Sensory experience selectively regulates transmitter synthesis enzymes in interglomerular circuits.

Sensory experience influences brain organization and function. A particularly striking example is in the olfactory bulb where reduction of odorant sensory signals profoundly down-regulates dopamine in glomerular neurons. There are two large populations of glomerular inhibitory interneurons: (1) GABAergic periglomerular (PG) cells, whose processes are limited to a single glomerulus, regulate intraglomerular processing and (2) DAergic-GABAergic short axon (SA) cells, whose processes contact multiple glomeruli, regulate interglomerular processing. The inhibitory neurotransmitter GABA is synthesized from L-glutamic acid by the enzyme glutamic acid decarboxylase (GAD) of which there are two major isoforms: GAD65 and GAD67. GAD65 is expressed in uniglomerular PG cells. GAD67 is expressed by SA cells, which also co-express the rate-limiting enzyme for dopamine synthesis, tyrosine hydroxylase (TH). Deafferentation or sensory deprivation decreases TH expression but it is not known if sensory input alters GAD isoforms. Here we report that either deafferentation or reduction of sensory input by nares occlusion significantly reduced GAD67 protein and the number of SA cells expressing GAD67. However, neither manipulation altered GAD65 protein or the number of GAD65 PG cells. These findings show that sensory experience strongly impacts transmitter regulation in the circuit that controls neural processing across glomeruli but not in the circuit that regulates intraglomerular processing.

Control of RSV-induced lung injury by alternatively activated macrophages is IL-4R alpha-, TLR4-, and IFN-beta-dependent.

Severe respiratory syncytial virus (RSV)-induced bronchiolitis has been associated with a mixed "Th1" and "Th2" cytokine storm. We hypothesized that differentiation of "alternatively activated" macrophages (AA-M phi) would mediate the resolution of RSV-induced lung injury. RSV induced interleukin (IL)-4 and IL-13 by murine lung and peritoneal macrophages, IL-4R alpha/STAT6-dependent AA-M phi differentiation, and significantly enhanced inflammation in the lungs of IL-4R alpha(-/-) mice. Adoptive transfer of wildtype macrophages to IL-4R alpha(-/-) mice restored RSV-inducible AA-M phi phenotype and diminished lung pathology. RSV-infected Toll-like receptor (TLR)4(-/-) and interferon (IFN)-beta(-/-) macrophages and mice also failed to express AA-M phi markers, but exhibited sustained proinflammatory cytokine production (e.g., IL-12) in vitro and in vivo and epithelial damage in vivo. TLR4 signaling is required for peroxisome proliferator-activated receptor gamma expression, a DNA-binding protein that induces AA-M phi genes, whereas IFN-beta regulates IL-4, IL-13, IL-4R alpha, and IL-10 expression in response to RSV. RSV-infected cotton rats treated with a cyclooxygenase-2 inhibitor increased expression of lung AA-M phi. These data suggest new treatment strategies for RSV that promote AA-M phi differentiation.

Two GABAergic intraglomerular circuits differentially regulate tonic and phasic presynaptic inhibition of olfactory nerve terminals.

Olfactory nerve axons terminate in olfactory bulb glomeruli forming excitatory synapses onto the dendrites of mitral/tufted (M/T) and juxtaglomerular cells, including external tufted (ET) and periglomerular (PG) cells. PG cells are heterogeneous in neurochemical expression and synaptic organization. We used a line of mice expressing green fluorescent protein under the control of the glutamic acid decarboxylase 65-kDa gene (GAD65+) promoter to characterize a neurochemically identified subpopulation of PG cells by whole cell recording and subsequent morphological reconstruction. GAD65+ GABAergic PG cells form two functionally distinct populations: 33% are driven by monosynaptic olfactory nerve (ON) input (ON-driven PG cells), the remaining 67% receive their strongest drive from an ON-->ET-->PG circuit with no or weak monosynaptic ON input (ET-driven PG cells). In response to ON stimulation, ON-driven PG cells exhibit paired-pulse depression (PPD), which is partially reversed by GABA(B) receptor antagonists. The ON-->ET-->PG circuit exhibits phasic GABA(B)-R-independent PPD. ON input to both circuits is under tonic GABA(B)-R-dependent inhibition. We hypothesize that this tonic GABA(B)R-dependent presynaptic inhibition of olfactory nerve terminals is due to autonomous bursting of ET cells in the ON-->ET-->PG circuit, which drives tonic spontaneous GABA release from ET-driven PG cells. Both circuits likely produce tonic and phasic postsynaptic inhibition of other intraglomerular targets. Thus olfactory bulb glomeruli contain at least two functionally distinct GABAergic circuits that may play different roles in olfactory coding.

Odor-induced, activity-dependent transneuronal gene induction in vitro: mediation by NMDA receptors.

Expression of tyrosine hydroxylase (TH) by juxtaglomerular (JG) neurons of the olfactory bulb (OB) requires innervation of the bulb by olfactory receptor neurons (ORNs). ORN lesion selectively downregulates TH in JG neurons. In reversible odor deprivation, TH expression is downregulated as the naris is closed and then upregulated upon naris reopening. The mechanism or mechanisms regulating this dependence are unknown. TH expression could be regulated by trophic factor release and/or synaptic activity from ORN terminals. We investigated TH expression in cocultures of dissociated postnatal rat OB cells and embryonic olfactory neuroepithelium (OE) slice explants. TH-positive neurons in control dissociated OB cell cultures alone comprise only a small fraction of the total population of cells present in the culture. However, when OE slice explants are cocultured with dispersed OB cells, there is a mean 2.4-fold increase in the number of TH-positive neurons. ORNs in vivo use glutamate as a neurotransmitter. Broad spectrum excitatory amino acid antagonists (kyurenic acid) or selective antagonists of the NMDA receptor (APV) both prevent induction of TH expression in OE-OB cocultures. Furthermore, pulse application of NMDA stimulates TH expression in OB neurons in the absence of OE. In vitro, OB TH neurons express NMDA receptors, suggesting that NMDA stimulation is acting directly on TH neurons. Exposure of OE explants to natural odorants results in upregulation of TH, presumably through increased ORN activity, which could be blocked by APV. These findings indicate that odorant-stimulated glutamate release by ORN terminals regulates TH expression via NMDA receptors on JG dopaminergic neurons.

Role of galectin-1 in the developing mouse olfactory system.

Primary sensory olfactory neurons reside in a neuroepithelium lining the nasal cavity and project topographically onto the surface of the olfactory bulb, a rostral extension of the telencephalon. Galectin-1, a bivalent galactose-binding vertebrate lectin, is expressed in the developing rodent olfactory system. In the present study, the mouse olfactory neuron cell line 4.4.2 was used to examine the role of galectin-l in neurite outgrowth in vitro. Recombinant galectin-l has neurite outgrowth-promoting activity when used as a substrate for 4.4.2 cells. When either galectin-1 or lactose was added to the culture media, the neurite outgrowth-promoting activity was abolished. These results demonstrate that galectin-1 can modulate neurite growth in vitro. The in vivo role of galectin-1 was investigated by examining the topographical organization of the olfactory pathway in mice carrying a null mutation for galectin-1. Using Dolichos biflorus agglutinin as a convenient histochemical marker of a subpopulation of primary sensory olfactory neurons which project topographically to the dorsomedial olfactory bulb, we show an aberrant topography of olfactory axons in the null mutants. A subset of primary sensory olfactory axons failed to project to their correct target sites in the caudal olfactory bulb. These data indicate that galectin-1 is involved in the growth and/or guidance of primary sensory olfactory axons between the nasal cavity and the olfactory bulb. This is the first demonstration that a lectin has neurite outgrowth-promoting activity and plays a role in neuronal pathfinding in the mammalian nervous system.

GABAergic phenotypic differentiation of a subpopulation of subventricular derived migrating progenitors.

Olfactory bulb interneurons are continuously generated throughout development and in adulthood. These neurons are born in the subventricular zone (SVZ) and migrate along the rostral migratory stream into the olfactory bulb where they differentiate into local interneurons. To investigate the differentiation of GABAergic interneurons of the olfactory bulb we used a transgenic mouse which expresses green fluorescent protein (GFP) under the control of the glutamic acid decarboxylase 65 kDa (GAD65) promoter. During development and in adulthood GFP was expressed by cells in the SVZ and along the entire length of its rostral extension including the distal portion within the olfactory bulb. The occurrence of GAD65 mRNA in these zones was confirmed by PCR analysis on microdissected regions along the pathway. Polysialic acid neural cell adhesion molecule, a marker of migrating neuroblasts in adults, was coexpressed by the majority of the GFP-positive SVZ-derived progenitor cells. Cell tracer injections into the SVZ indicated that approximately 26% of migrating progenitor cells expressed GFP. These data show the early differentiation of migrating SVZ-derived progenitors into a GAD65-GFP-positive phenotype. These cells could represent a restricted lineage giving rise to GAD65-positive GABAergic olfactory bulb interneurons.

Ultrasensitive pheromone detection by mammalian vomeronasal neurons.

The vomeronasal organ (VNO) is a chemoreceptive organ that is thought to transduce pheromones into electrical responses that regulate sexual, hormonal and reproductive function in mammals. The characteristics of pheromone signal detection by vomeronasal neurons remain unclear. Here we use a mouse VNO slice preparation to show that six putative pheromones evoke excitatory responses in single vomeronasal neurons, leading to action potential generation and elevated calcium entry. The detection threshold for some of these chemicals is remarkably low, near 10(-11) M, placing these neurons among the most sensitive chemodetectors in mammals. Using confocal calcium imaging, we map the epithelial representation of the pheromones to show that each of the ligands activates a unique, nonoverlapping subset of vomeronasal neurons located in apical zones of the epithelium. These neurons show highly selective tuning properties and their tuning curves do not broaden with increasing concentrations of ligand, unlike those of receptor neurons in the main olfactory epithelium. These findings provide a basis for understanding chemical signals that regulate mammalian communication and sexual behaviour.