A unique cell population in the mouse olfactory bulb displays nuclear beta-catenin signaling during development and olfactory sensory neuron regeneration

Olfactory sensory neurons (OSNs) in the nose form precise connections with neurons in the brain. However, mechanisms that account for the formation of such precise neuronal connections are incompletely understood. Recent studies implicate the function of Wnt growth factors in the formation of neuronal connections. To assess the role of Wnt signaling in the olfactory system, we examined the expression of beta-galactosidase (beta-gal) in the TOPGAL mouse, a transgenic strain in which beta-gal expression reports the activation of the canonical Wnt signaling pathway. In the olfactory epithelium, no beta-gal expression was observed at any developmental stages. In the olfactory bulb (OB), beta-gal expression was observed in a population of cells located at the interface of the olfactory nerve layer and the glomerular layer. The beta-gal expression was developmentally regulated with the peak expression occurring at late embryonic and early postnatal stages and a greatly reduced expression in adulthood. Further, forced OSN regeneration and subsequent reinnervation of the OB led to a reactivation of beta-gal expression in mature animals. The temporal coincidence between the peak of beta-gal expression and formation of OSN connections, together with the spatial localization of these cells, suggests a potential role of these cells and canonical Wnt signaling in the formation of OSN connections in the OB during development and regeneration.     

Flexible categorization in the mouse olfactory bulb

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Characterization of the synaptic properties of olfactory bulb projections

The olfactory bulb directly projects to several diverse telencephalic structures, but, to date, few studies have investigated the physiological characteristics of most of these areas. As an initial step towards understanding the odor processing functions of these secondary olfactory structures, we recorded evoked field potentials in response to lateral olfactory tract stimulation in vivo in urethane-anesthetized Sprague-Dawley rats in the following brain structures: anterior olfactory nucleus, ventral and dorsal tenia tecta, olfactory tubercle, anterior and posterior piriform cortex, the anterior cortical nucleus of the amygdala, and lateral entorhinal cortex. Using paired-pulse stimulation with interpulse intervals of 25-1000 ms, we observed facilitation of the response to the second pulse in every structure examined, although the degree of facilitation varied among the target structures. Additionally, pulse train stimulation at three different frequencies (40, 10 and 2 Hz) produced facilitation of evoked field potentials that also varied among target structures. We discuss the potential utility of such short-term facilitation in olfactory processing.     

Electrophysiological demonstration of independent olfactory receptor types and associated neuronal responses in the trout olfactory bulb

The present study attempts to highlight the principles by which peripheral olfactory information of across- and within-class odorant signals is transformed into bulbar neuron responses. For this purpose, we performed electro-olfactogram cross-adaptation and mixture experiments as well as single unit recording of olfactory bulb neurons using amino acid, bile acid and F-prostaglandin stimulants in brown and rainbow trout. The results show that amino acids, a bile acid and a F-prostaglandin activate independent receptor types. However, within the class of amino acids, different receptor types are only partially independent. Neurons responsive to bile acid and amino acids were segregated to the mid-dorsal and latero-posterior olfactory bulb, respectively. Of the 43 responsive olfactory bulb neurons studied in brown trout, 41 showed specificity for one odorant class. Olfactory bulb neurons gained responsiveness to new amino acids with increasing stimulant concentration. We conclude that different odorant classes activate specific neurons located in different regions of the trout olfactory bulb, and that information distinguishing related amino acids can be represented in a limited number of bulbar neurons with distinct response profiles under the conditions investigated.     

Axon mis-targeting in the olfactory bulb during regeneration of olfactory neuroepithelium

During development, primary olfactory axons typically grow to their topographically correct target zone without extensive remodelling. Similarly, in adults, new axons arising from the normal turnover of sensory neurons essentially project to their target without error. In the present study we have examined axon targeting in the olfactory pathway following extensive chemical ablation of the olfactory neuroepithelium in the P2-tau:LacZ line of mice. These mice express LacZ in the P2 subpopulation of primary olfactory neurons whose axons target topographically fixed glomeruli on the medial and lateral surfaces of the olfactory bulb. Intraperitoneal injections of dichlobenil selectively destroyed the sensory neuroepithelium of the nasal cavity without direct physical insult to the olfactory neuron pathway. Primary olfactory neurons regenerated and LacZ staining revealed the trajectory of the P2 axons. Rather than project solely to their topographically appropriate glomeruli, the regenerating P2 axons now terminated in numerous inappropriate glomeruli which were widely dispersed over the olfactory bulb. While these errors in targeting were refined over time, there was still considerable mis-targeting after four months of regeneration.     

Transregulation of erbB expression in the mouse olfactory bulb.

Previously, we have shown that erbB-3 expression is restricted to the ensheathing cells of the olfactory nerve layer, while erbB-4 is found in the periglomerular and mitral/tufted cells of the olfactory bulb and in cells coming out from the rostral migratory stream of the subependymal layer. In the present work, we have treated adult mice with zinc sulfate intranasal irrigation and analyzed erbB-3 and erbB-4 expression in the deafferented olfactory bulb. Following treatment, olfactory axons undergo degeneration, as indicated by the loss of OMP expression in the deafferented olfactory bulb. The thickness of the olfactory nerve layer is reduced, but the specific intensity of erbB-3 labeling in the remaining olfactory nerve layer is increased with respect to control. Interestingly, following deafferentation, erbB-4 immunoreactivity decreases specifically in cell types that normally make synaptic contacts with primary olfactory neurons in the glomeruli, i.e. periglomerular and mitral/tufted cells. Partial lesion of the olfactory epithelium allows regenerative axon growth of olfactory neurons to the olfactory bulb. Following olfactory axon regeneration, erbB-3 and erbB-4 immunoreactivity in the olfactory bulb is similar to control. Thus, like tyrosine hydroxylase, the down regulation of erbB-4 expression in the periglomerular cells is reversible.     

Activity-dependent changes in cholinergic innervation of the mouse olfactory bulb

The interplay between olfactory activity and cholinergic modulation remains to be fully understood. This report examines the pattern of cholinergic innervation throughout the murine main olfactory bulb across different developmental stages and in naris-occluded animals. To visualize the pattern of cholinergic innervation, we used a transgenic mouse model, which expresses a fusion of the microtubule-associated protein, tau, with green fluorescence protein (GFP) under the control of the choline acetyltransferase (ChAT) promoter. This tau-GFP fusion product allows for a remarkably vivid and clear visualization of cholinergic innervation in the main olfactory bulb (MOB). Interestingly, we find an uneven distribution of GFP label in the adult glomerular layer (GL), where anterior, medial, and lateral glomerular regions of the bulb receive relatively heavier cholinergic innervation than other regions. In contrast to previous reports, we find a marked change in the pattern of cholinergic innervation to the GL following unilateral naris occlusion between the ipsilateral and contralateral bulbs in adult animals.     

Cholinergic modulation of dopaminergic neurons in the mouse olfactory bulb

Considerable evidence exists for an extrinsic cholinergic influence in the maturation and function of the main olfactory bulb. In this study, we addressed the muscarinic modulation of dopaminergic neurons in this structure. We used different patch-clamp techniques to characterize the diverse roles of muscarinic agonists on identified dopaminergic neurons in a transgenic animal model expressing a reporter protein (green fluorescent protein) under the tyrosine hydroxylase promoter. Bath application of acetylcholine (1 mM) in slices and in enzymatically dissociated cells reduced the spontaneous firing of dopaminergic neurons recorded in cell-attached mode. In whole-cell configuration no effect of the agonist was observed, unless using the perforated patch technique, thus suggesting the involvement of a diffusible second messenger. The effect was mediated by metabotropic receptors as it was blocked by atropine and mimicked by the m2 agonist oxotremorine (10 muM). The reduction of periglomerular cell firing by muscarinic activation results from a membrane-potential hyperpolarization caused by activation of a potassium conductance. This modulation of dopaminergic interneurons may be important in the processing of sensory information and may be relevant to understand the mechanisms underlying the olfactory dysfunctions occurring in neurodegenerative diseases affecting the dopaminergic and/or cholinergic systems.     

Slice culture of the olfactory bulb of Xenopus laevis tadpoles

We report on the development of a slice culture of amphibian brain tissue. In particular, we cultured slices from Xenopus laevis tadpoles that contain the olfactory mucosae, the olfactory nerves, the olfactory bulb and the telencephalon. During 6 days in roller tubes the slices flattened, starting from 250 microm and decreasing to approximately 40 microm, corresponding to about three cell layers. Dendritic processes could be followed over distances as long as 200 microm. Neurons in the cultured slice could be recorded using the patch clamp technique and simultaneously imaged using an inverted laser scanning microscope. We characterized the main neuron types of the olfactory bulb, i.e. mitral cells and granule cells, by correlating their typical morphological features in the acute slice with the electrophysiological properties in both the acute slice and slice culture. This correlation allowed unambiguous identification of mitral cells and granule cells in the slice culture.     

Acid-sensing ion channels in mouse olfactory bulb M/T neurons

The olfactory bulb contains the first synaptic relay in the olfactory pathway, the sensory system in which odorants are detected enabling these chemical stimuli to be transformed into electrical signals and, ultimately, the perception of odor. Acid-sensing ion channels (ASICs), a family of proton-gated cation channels, are widely expressed in neurons of the central nervous system. However, no direct electrophysiological and pharmacological characterizations of ASICs in olfactory bulb neurons have been described. Using a combination of whole-cell patch-clamp recordings and biochemical and molecular biological analyses, we demonstrated that functional ASICs exist in mouse olfactory bulb mitral/tufted (M/T) neurons and mainly consist of homomeric ASIC1a and heteromeric ASIC1a/2a channels. ASIC activation depolarized cultured M/T neurons and increased their intracellular calcium concentration. Thus, ASIC activation may play an important role in normal olfactory function.     

New automated fluorometric peptide microassay for carnosine in mouse olfactory bulb

A procedure for assaying peptides at the picomole level in tissue extracts has been developed and used to measure the dipeptide carnosine in mouse olfactory bulb. In this procedure the tissue extract is reacted with 2-methoxy-2,4-diphenyl-3(2H)-furanone (MDPF), and the resultant fluorophors are resolved on a high performance reverse-phase column. Quantitation is performed in a filter fluorometer equipped with a flow cell. Carnosine was found to be present at a level of 1.93 ± 0.44 nmol/mg of tissue (mean + SD of 11 samples), in agreement with previous findings by other methods.