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Localized changes in olfactory bulb morphology associated with early olfactory learning.

Young rats exposed to an odor while receiving reinforcing stimulation come to approach that odor upon subsequent presentation. In addition, such pups have increased 14C-2-deoxyglucose (2DG) uptake within focal areas of the glomerular layer in response to that odor, compared to control animals experiencing the odor for the first time. In this study, the morphology of the glomerular areas underlying these 2DG foci was examined to determine whether early olfactory learning imposed local structural changes that could produce the enhanced 2DG uptake. Alternate sections either were processed with a silver and a Nissl stain to label both cell bodies and their processes or were histochemically treated for the mitochondrial enzymes cytochrome oxidase (CO) or succinic dehydrogenase (SDH) to define the glomerular core of the bulb; 2DG autoradiographs were aligned with adjacent stained sections, and regions underlying the high 2DG uptake foci were examined. In odor-familiar animals, large glomerular clusters that protruded into the external plexiform layer or the olfactory nerve layer were associated with the focal areas of increased 2DG uptake. Morphometric analysis of these regions revealed that the glomerular layer underlying the foci of high 2DG uptake was 30% wider in odor-familiar animals than comparable areas in odor-unfamiliar animals; the cross-sectional areas of individual glomeruli were 21% larger in odor-familiar animals. The foci of enhanced 2DG uptake therefore appear to be associated with groups of enlarged glomeruli. These data demonstrate that early olfactory learning influences the morphology of the olfactory bulb.

Animals↗

A membrane-bound heparan sulfate proteoglycan that is transiently expressed on growing axons in the rat brain.

Monoclonal antibodies were raised to membrane-bound proteoglycans derived from rat brain and three monoclonal antibodies that recognized a 200-kDa heparan sulfate proteoglycan (designated H5-PG) with a core glycoprotein of 140 kDa were obtained. The expression of H5-PG was spatially and temporally regulated in the central nervous system. In the cerebellar cortex, H5-PG was associated mainly with the actively growing parallel fibers of granule cells. The expression was abruptly down-regulated in parallel with the formation of synapses on dendrites of Purkinje cells. In the cerebral cortex, the proteoglycan was widely distributed throughout the cortex. The temporal pattern of expression was similar to that in the cerebellar cortex; the peak level of expression was observed during the period from postnatal days 0 to 20 when neuritogenesis and synaptogenesis occur most extensively in the rat cerebral cortex. H5-PG in the central nervous system disappeared prior to adulthood except in the olfactory bulb. High-level expression was recognized on the olfactory nerves and glomeruli, where the renewal of both axons and synapses is occurring constantly. The data suggest that H5-PG is a glycoconjugate on axonal surface that is involved in axonal outgrowth and/or synaptogenesis.

Animals↗

Misrouting of mitral cell progenitors in the Pax6/small eye rat telencephalon.

The olfactory bulb is a protruding structure formed at the rostral end of the telencephalon. Pax6-mutant mice and rats lack the olfactory bulb and, instead, develop an olfactory bulb-like structure at the lateral part of the telencephalon. Here, we report that ectopic formation of the olfactory bulb-like structure in these mutants is caused by the abnormal migration of mitral cell progenitors, which first differentiate within the olfactory bulb. Cell-tracing experiments in whole embryos in culture indicate that, in the mutants, the mitral cell progenitors that originate from the rostral part of the telencephalon migrate caudally toward the lateral part of the telencephalon. Cell transplantation demonstrates that the abnormal cell migration is not autonomous to the mitral cell progenitors themselves. The mislocation of the olfactory bulb in the mutant is not caused by loss of olfactory nerve innervation. Furthermore, transfection of a Pax6-expression vector to the mutant telencephalon restores the normal migration of mitral cell progenitors. These results provide evidence that Pax6 is required to position the mitral cell progenitors at the rostral end of the telencephalon.

Animals↗

Localisation of 3H-GABA in the rat olfactory bulb: an in vivo and in vitro autoradiographic study.

In an attempt to further clarify the localisation of GABAergic elements in the olfactory bulb we have performed, in vivo and in vitro, autoradiographic studies with 3H-GABA (gamma-amino butyric acid) and 3H-DABA (L-2,4 diamino butyric acid). The results have shown a strong labelling with 3H-GABA of the glial cells in all the layers of the olfactory bulb. A high concentration of grains was observed in the periglomerular region. The labelling in the external plexiform layer was uniformly distributed in the neuropile with the strongest activity at the level of the dendritic processes of the granule cells, leaving the mitral cell dendrites and cell bodies almost free of grains. 3H-DABA showed a very similar pattern to 3H-GABA. When olfactory bulb slices were preincubated with beta-alanine the labelling of the glial elements almost disappeared especially at the level of the olfactory nerve layer. The labelling pattern of the other layers of the bulb remained mostly unchanged. This supports the view that a population of periglomerular and granule cells are GABAergic and that beta-alanine competes with GABA uptake sites only in glial cells.

Aminooxyacetic Acid↗

Odor psychophysics in vertebrates.

The methods used to obtain psychophysical data on the nasal chemosensory systems of all classes of vertebrates are critically reviewed and a summary of the available data on their odor detection and discrimination abilities is provided. Although there are reliable methods for training at least one member of each class to respond differentially to the presence or absence of odor stimuli, very little is known about the limits of the capacity of any of the three major nasal chemosensory systems (olfactory, vomeronasal and trigeminal) to detect pure compounds. Furthermore, studies in which rigorous procedures are followed for both the maintenance of discriminative responding and the presentation of odor stimuli often fail to determine the sensory system(s) mediating the psychophysical results. This lack of information has impeded progress on several fundamental problems in the study of nasal chemoreception.

Amphibians↗

Deafferentation up-regulates the expression of the mGlu1a metabotropic glutamate receptor protein in the olfactory bulb.

Chemical lesion of olfactory neuroepitheium induced an up-regulation of the mGlu1a metabotropic glutamate receptor protein in the olfactory bulb, as shown by Western blot analysis. At 2 days after the lesion, the increase in the receptor protein was associated with an increase in mGlu1a mRNA levels; in contrast, at longer times after the lesion (16 days), mRNA levels were reduced in spite of the high expression of the receptor protein, perhaps as a result of product-inhibition of mGlu1 gene expression. Immunohistochemical analysis showed that the increase in mGlu1a induced by olfactory denervation was confined to the glomeruli, which occupy the external portion of the olfactory bulb. Within these structures, mGlu1a receptors are mainly localized on the distal dendrites of mitral cells, which are innervated by the glutamatergic axons of the olfactory nerve. These results demonstrate that the expression of mGlu1a receptors is up-regulated in response to glutamatergic deafferentation, supporting a role for this particular receptor subtype in the physiology of synaptic transmission.

Amino Acid Sequence↗

BDNF production by olfactory ensheathing cells contributes to axonal regeneration of cultured adult CNS neurons.

Olfactory ensheathing cells (OECs) are the main glial cell type that populates mammalian olfactory nerves. These cells have a great capacity to promote the regeneration of axons when transplanted into the injured adult mammalian CNS. However, little is still known about the molecular mechanisms they employ in mediating such a task. Brain-derived neurotrophic factor (BDNF) was identified as a candidate molecule in a genomic study that compared three functionally different OEC populations: Early passage OECs (OEC Ep), Late passage OECs (OEC Lp) and the OEC cell line TEG3 [Pastrana, E., Moreno-Flores, M.T., Gurzov, E.N., Avila, J., Wandosell, F., Diaz-Nido, J., 2006. Genes associated with adult axon regeneration promoted by olfactory ensheathing cells: a new role for matrix metalloproteinase 2. J. Neurosci. 26, 5347-5359]. We have here set out to determine the role played by BDNF in the stimulation of axon outgrowth by OECs. We compared the extracellular BDNF levels in the three OEC populations and show that it is produced in significant amounts by the OECs that can stimulate axon regeneration in adult retinal neurons (OEC Ep and TEG3) but it is absent from the extracellular medium of OEC Lp cells which lack this capacity. Blocking BDNF signalling impaired axonal regeneration of adult retinal neurons co-cultured with TEG3 cells and adding BDNF increased the proportion of adult neurons that regenerate their axons on OEC Lp monolayers. Combining BDNF with other extracellular proteins such as Matrix Metalloproteinase 2 (MMP2) further augmented this effect. This study shows that BDNF production by OECs plays a direct role in the promotion of axon regeneration of adult CNS neurons.

Animals↗

Local interneurons define functionally distinct regions within lobster olfactory glomeruli

Whole-cell recording coupled with biocytin injection revealed four types of interneurons intrinsic to the olfactory lobe (OL) of the spiny lobster Panulirus argus. Each type of neuron had a distinct pattern of arborization within the three anatomically defined regions of OL glomeruli (cap, subcap and base). Type I interneurons innervated all three regions, while types II, III and IV branched only in the cap, subcap and base, respectively. Type I interneurons responded to electrical stimulation of the antennular (olfactory) nerve with a burst of 1­20 action potentials and a 1­10 s depolarization. Type II (cap) interneurons responded to the same input with a burst of 1­3 action potentials followed by a shorter hyperpolarization. Type III (subcap) interneurons responded with a burst of 1­6 action potentials followed by a delayed, 0.5­4 s depolarization. Type IV (base) interneurons responded with a brief depolarization or a burst of 1­3 action potentials followed by a 1 s hyperpolarization. The regionalized arborization and the different response properties of the type II, III and IV interneurons strongly imply that lobster olfactory glomeruli contain functionally distinct regions, a feature that should be useful in understanding the multiple synaptic pathways involved in processing olfactory input.

Journal Article↗

Glycoconjugates are stage- and position-specific cell surface molecules in the developing olfactory system, 2: Unique carbohydrate antigens are topographic markers for selective projection patterns of olfactory axons.

Three monoclonal antibodies specific for different carbohydrate antigens were used to analyze the development of the olfactory system in rats. CC2 antibodies react with a subset of main olfactory neurons, their axons, and terminals in the olfactory bulb. CC2 antigens are expressed on dorsomedial neurons in the olfactory epithelium (OE) from embryonic (E) day 15 to adults. In the olfactory bulb (OB), only dorsomedially located glomeruli express CC2 glycoconjugates from postnatal day (P) 2 to adults. Thus CC2 defines a dorsomedially organized projection that is established early in embryonic development and continues in adults. P-Path antibodies react with antigens that are expressed on the olfactory nerve in embryos, and are also detected on cell bodies in the neuroepithelium and in glomeruli of the OB at P2. At P14, P-Path staining is weaker, but remains present on many cells in the epithelium and in many glomeruli in the bulb. Postnatally, P-Path immunostaining continues to decrease in most regions of the OE and OB. At P35 and afterwards, only a few P-Path-positive neuronal cells can be detected in the OE. Furthermore, after P35 only two groups of glomeruli in the OB are P-Path immunoreactive. One is situated adjacent to the accessory olfactory bulb (AOB) at the dorsocaudal surface of the OB. The other is adjacent to the AOB at the ventrocaudal surface of the OB. Thus, in adults, P-Path glycoconjugates are expressed in neurons and axons that project only to a specific subset of caudal glomeruli of the OB. Monoclonal antibody 1B2, reacts with beta-galactose-terminating glycolipids and glycoproteins. At P2, 1B2 immunoreactivity is seen on a subset of cell bodies that are distributed throughout the OE and is expressed in most glomeruli in the OB at this age. By P35 and in adults, 1B2 continues to be expressed on a subset of neurons in the OE that project to only a small subset of glomeruli in the OB. Unlike CC2 and P-Path antigens that define specific groups of glomeruli, 1B2-immunoreactive glomeruli do not have a detectable spatial pattern. It is more likely that 1B2 antigens define a specific stage in the maturation of connections between the OE and OB.

Animals↗

Signal-to-noise ratio of chemosensory event-related potentials.

OBJECTIVE: We investigated the influence of the number of stimuli on signal-to-noise (S/N) ratio of CSERP. METHODS: CSERP from 20 normosmic subjects were obtained in response to stimulation with two olfactory (H(2)S and PEA) and a trigeminal (CO(2)) stimulant. For each of these odors, 160 stimuli were delivered into the right nostril (duration 200ms, mean ISI 30s) using a constant-flow, air-dilution olfactometer. For each EEG recording site (Fz, Cz, Pz, C3, C4), peak-to-peak amplitude N1P2 and noise amplitude levels were determined. Subsequently, S/N ratios were calculated. RESULTS: The S/N ratios for olfactory ERP generally improved for H(2)S and PEA. For responses to PEA, S/N ratios increased significantly up to 80 averages (S/N ratio=5.6). The number of stimuli for an optimal S/N ratio for trigeminal ERP was slightly lower, i.e. 60 averages (S/N ratio=7.9). CONCLUSIONS: S/N N1P2 ratios in olfactory and trigeminal ERP significantly improve with an increasing number of responses averaged under these experimental conditions. This is mainly due to a reduction of noise level. Applying more stimuli has little additional effect on S/N ratio due to a concomitant decrease in signal amplitude. SIGNIFICANCE: An optimal S/N ratio is essential when recording CSERP in neurodegenerative disorders, where responses may be of low amplitude, and for medico-legal purposes.

Adolescent↗

Nitric oxide synthase, cGMP, and NO-mediated cGMP production in the olfactory bulb of the rat.

High levels of nitric oxide synthase and cyclic 3',5'-guanosine monophosphate (cGMP) in the olfactory bulb suggest that nitric oxide, acting as a diffusible intercellular messenger molecule inducing increased synthesis of cGMP, plays an important role in olfaction. The localization of cGMP after sodium nitroprusside stimulation of in vitro slices of rat olfactory bulb was compared with the distribution of nicotinamide adenine dinucleotide phosphatediaphorase, nitric oxide synthase, and glial fibrillary acidic protein. cGMP was detected immunohistochemically in cryostat sections. In the presence of the phosphodiesterase blocker isobutyl methylxanthine, cGMP was present in neurons in the glomerular layer, axons in the external and internal plexiform layers, and in a few somata and axons of the granule cell layer. This staining was blocked by NG-nitro-L-arginine methylester hydrochloride or hemoglobin. After sodium nitroprusside stimulation, the olfactory nerve layer was intensely stained, as were the glomeruli and periglomerular cells. In the external plexiform layer, axonal staining was increased substantially, and there were occasional multipolar cGMP-positive neurons. In the internal plexiform and granule cell layers, axonal staining was greatly increased. Many granule cells were also cGMP positive after sodium nitroprusside stimulation. cGMP and nitric oxide synthase-positive neuronal elements overlapped in the glomerular and granule cell layers, but staining was not colocalized, cGMP was not found in astrocytes. The glutamatergic antagonists D-2-amino-5-phosphonovalerate and 6-cyano-7-nitroquinoxaline caused differential inhibition of cGMP accumulation in layers of the olfactory bulb. These findings support the hypothesis that nitric oxide is an intercellular messenger in the olfactory bulb (Breer and Shepherd [1993] Trends Neurosci. 16:5-9).

Animals↗

Testing a perceptual constancy model for odor strength: the effects of sniff pressure and resistance to sniffing.

The rate at which odorous vapor flows over the receptor surface is reflected in olfactory nerve activity, but the evidence for a corresponding effect on odor strength is scant and inconsistent. A perceptual constancy model for odor strength resolves this apparent contradiction: The effect of flow rate at the receptor site may be perceptually discounted when there is sensory feedback, in the form of perceived effort, from the action of sniffing responsible for changes in flow rate. Previous evidence shows that observers experience change in perceived effort when sniffing at various pressures against fixed resistance but not when sniffing with fixed pressure against varying resistance. Thus the model predicts both that odor strength varies when flow rate is manipulated by changes in olfactory resistance and that odor strength is invariant when flow rate is manipulated by changes in sniff pressure. In the present study, in which subjects made magnitude estimates of the odor strength of pyridine and n-butanol, an earlier finding for the effects of resistance change was confirmed and extended. Further, within the framework of a single experiment in which both pressure and resistance were independently manipulated, the odor strength of n-butanol conformed to the predictions of the constancy model.

1-Butanol↗

Expression pattern of alpha CaMKII in the mouse main olfactory bulb.

Multifunctional Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is highly enriched at synapses and has been implicated in regulating the formation and function of several sensory systems, including the visual and the somatosensory systems. Although there is evidence for CaMKII expression in the olfactory system, the cellular localization of CaMKII has not been well studied and its function remains unknown. In this study, we examined the normal expression patterns of the predominant alpha CaMKII in the mouse main olfactory bulb. We showed that alpha CaMKII expression levels were high in the olfactory bulb and were developmentally regulated. Immunoreactivity to alpha CaMKII was heavy in the external plexiform layer and the granule cell layer but minimal in the olfactory nerve layer and the glomerular layer. At the cellular level, alpha CaMKII was selectively expressed in the gamma-aminobutyric acid (GABA)ergic granule cells but not in the GABAergic periglomerular cells. Unexpectedly, alpha CaMKII was not detected in the glutamatergic mitral/tufted cells. At the ultrastructural level, alpha CaMKII immunoreactivity was positive in granule cell spines and dendrites, but negative in mitral/tufted cell dendrites. In contrast, in the piriform cortex, as in the majority of cortical regions, alpha CaMKII was expressed in the glutamatergic neurons but not in the GABAergic neurons. Our results set the stage for ongoing investigations of the roles of CaMKII in the formation and function of the olfactory system.

Animals↗

How chemical information processing interferes with face processing: a magnetoencephalographic study.

Magnetic field changes related to face encoding were recorded in 20 healthy young participants. Faces had to be deeply encoded under four kinds of simultaneous nasal chemical stimulation. Neutral room air, phenyl ethyl alcohol (PEA, rose flavor), carbon dioxide (CO2, pain), and hydrogen sulfide (H2S, rotten eggs flavor) were used as chemical stimuli. PEA and H2S represented odor stimuli, whereas CO2 was used for trigeminal stimulation (pain sensation). After the encoding of faces, the respective recognition performances were tested focusing on recognition effects related to specific chemical stimulation during encoding. The number of correctly recognized faces (hits) varied between chemical conditions. PEA stimulation during face encoding significantly increased the number of hits compared to the control condition. H2S also led to an increased mean number of hits, whereas simultaneous CO2 administration during face encoding resulted in a reduction. Analysis of the physiological data revealed two latency regions of interest. Compared to the control condition, both olfactory stimulus conditions resulted in reduced activity components peaking at about 260 ms after stimulus onset, whereas CO2 produced a strongly pronounced enhanced activity component peaking at about 700 ms after stimulus onset. Both olfactory conditions elicited only weak enhanced activities at about 700 ms, and CO2 did not show any difference activity at 260 ms after stimulus onset compared to the control condition. It is concluded that the early activity differences represent subconscious olfactory information processing leading to enhanced memory performances irrespective of the hedonic value, at least if they are only subconsciously processed. The later activity is suggested to reflect conscious CO2 perception negatively affecting face encoding and therefore leading to reduced subsequent face recognition. We interpret that conscious processing of nasal chemical stimulation competes with deep face encoding with respect to cortical resources, whereas subconscious processing of nasal chemical stimulation does not.

Adult↗

Delivery of insulin-like growth factor-I to the rat brain and spinal cord along olfactory and trigeminal pathways following intranasal administration.

We investigated the CNS delivery of insulin-like growth factor-I (IGF-I), a 7.65 kDa protein neurotrophic factor, following intranasal administration and the possible pathways and mechanisms underlying transport from the nasal passages to the CNS. Anesthetized adult male Sprague-Dawley rats were given [125I]-IGF-I intranasally or intravenously and then killed by perfusion-fixation within 30 min. Other animals were killed following cisternal puncture and withdrawal of cerebrospinal fluid (CSF) or intranasal administration of unlabeled IGF-I or vehicle. Both gamma counting of microdissected tissue and high resolution phosphor imaging of tissue sections showed that the tissue concentrations and distribution following intranasal administration were consistent with two routes of rapid entry into the CNS: one associated with the peripheral olfactory system connecting the nasal passages with the olfactory bulbs and rostral brain regions (e.g. anterior olfactory nucleus and frontal cortex) and the other associated with the peripheral trigeminal system connecting the nasal passages with brainstem and spinal cord regions. Intranasal administration of [125I]-IGF-I also targeted the deep cervical lymph nodes, consistent with their possible role in lymphatic drainage of both the nasal passages and the CNS. Cisternal CSF did not contain [125I]-IGF-I following intranasal administration. Intravenous [125I]-IGF-I resulted in blood and peripheral tissue exposure similar to that seen following intranasal administration but CNS concentrations were significantly lower. Finally, delivery of IGF-I into the CNS activated IGF-I signaling pathways, confirming some portion of the IGF-I that reached CNS target sites was functionally intact. The results suggest intranasally delivered IGF-I can bypass the blood-brain barrier via olfactory- and trigeminal-associated extracellular pathways to rapidly elicit biological effects at multiple sites within the brain and spinal cord.

Administration, Intranasal↗

Trigeminal and olfactory pathways mediating isolation distress and companion comfort responses in rat pups.

Isolation in a novel test cage elicits ultrasonic vocalization (USV) in 12-13-day-old rat pups, and the presence of an anesthetized littermate companion reduces USV while eliciting body contact. These responses were studied in pups with surgical lesions of trigeminal snout afferents and after olfactory denervation by nasal ZnSO4 perfusion. With lesions of only one system, body contact with the passive companion was reduced by 20%, and interpup orientation was differentially affected by the two types of lesions. However, USV rates were no different from littermate controls in the presence of the companion or when tested alone. Combined lesions of both systems reduced contact time 55% and eliminated any reduction in USV rates by the companion while leaving intact USV and locomotor responses to isolation. These results suggest that different sensory pathways may mediate isolation distress and companion comfort responses.

Animals↗

The goldfish nervus terminalis: a luteinizing hormone-releasing hormone and molluscan cardioexcitatory peptide immunoreactive olfactoretinal pathway.

Antisera to two putative neurotransmitters, luteinizing hormone-releasing hormone (LHRH) and molluscan cardioexcitatory tetrapeptide (H-Phe-Met-Arg-Phe-NH2; FMRF-amide), bind specifically to neurites in the inner nuclear and inner plexiform layers of the goldfish retina. Retrograde labeling showed that intraocular axon terminals originate from the nervus terminalis, whose cell bodies are located in the olfactory nerves. Double immunocytochemical and retrograde labeling showed that some terminalis neurons project to the retina; others may project only within the brain. All terminalis neurons having proven retinal projections were both LHRH- and FMRF-amide-immunoreactive. The activity of retinal ganglion cells was recorded with microelectrodes in isolated superfused goldfish retinas. In ON- and OFF-center double-color-opponent cells, micromolar FMRF-amide and salmon brain gonadotropin-releasing factor ( [Trp7, Leu8] LHRH) caused increased spontaneous activity in the dark, loss of light-induced inhibition, and increased incidence of light-entrained pulsatile response. The nervus terminalis is therefore a putatively peptidergic retinopetal projection. Sex-related olfactory stimuli may act through it, thereby modulating the output of ganglion cells responsive to color contrast.

Animals↗