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Soluble factors from the olfactory bulb attract olfactory Schwann cells.

Olfactory Schwann cells (OSCs) extend processes that ensheathe bundles of olfactory axons as they course from the olfactory epithelium to the olfactory bulb (OB). Results of morphological and immunohistochemical studies have led to speculation that OSCs may be involved in guiding the olfactory axons to their target tissue. In this study we have explored this possibility by investigating the relationship between OSCs and the OB. Olfactory Schwann cells labelled with 1,1'-dioctadecyl 3,3,3',3'-tetramethylindocarbocyanine perchlorate (Dil) were injected into the nasal region of E14 rat embryos and entire embryos were cultured for 24 hr. It was found in some embryos, that the OSCs had migrated toward the presumptive OB. Cocultures of neonatal OB explants on OSC monolayers showed that the OSCs were attracted to the OB and formed a ring-like aggregate around the explant after 48 hr culture. This attraction was absent when a piece of cerebrum was used in place of the OB. When medium conditioned by neonatal OBs was placed in the lower compartment of the chemotaxis chamber, OSCs seeded in the upper compartment migrated through the pores of the nucleopore filter to reach the underside which was in contact with the conditioned medium. After 6 hr of incubation, scanning electron microscopy was performed on the underside of the nucleopore filters. Cell counts of OSCs showed that the cell density was significantly higher when medium conditioned by OBs was used instead of unconditioned medium or medium conditioned by cerebrum. The results of these experiments show that the OSCs migrate toward the OB under the influence of soluble factor(s) secreted by the target tissue.

Animals↗

Differentiation of chemically defined neuronal populations in the transplanted olfactory bulb without olfactory receptor innervation.

Olfactory bulbs (OBs) from embryonic day 15 and 17 and postnatal day 1 mice were transplanted into the lateral ventricle of juvenile host mice without bulbectomy, and fine structural and chemical features of neurons and glia in the OB transplants were investigated immunocytochemically and electron microscopically. In the OB transplants there were neither clearly defined glomeruli nor layers, nor olfactory marker protein immunoreactive elements. However, chemically defined neuronal populations resembling those in the normal OBs such as those immunoreactive for gamma-aminobutyric acid (GABA), tyrosine hydroxylase and Ca(2+)-binding proteins (calbindin-D28K, calretinin, parvalbumin) were observed. Electron microscopically, dendrodendritic and somatodendritic reciprocal synapses, that is, synapses characteristic of the OB, were occasionally observed in the OB transplants. These results indicated that at least some embryonic or newborn mouse OB neurons and/or precursor cells could exhibit chemical properties and form typical synaptic contacts observed in normal OB, even when they received no inputs from olfactory receptor cells.

Animals↗

A circadian clock in the olfactory bulb controls olfactory responsivity.

Recently, it has been shown that multiple mammalian cell types express daily rhythms in vitro. Although the suprachiasmatic nucleus (SCN) of the hypothalamus is known to regulate a wide range of circadian behaviors, the role for intrinsic rhythmicity in other tissues is unknown. We tested whether the main olfactory bulb (OB) of mice mediates daily changes in olfaction. We found circadian rhythms in cedar oil-induced c-Fos, a protein marker of cellular excitation, in the mitral and granular layers of the OB and in the piriform cortex (PC). These oscillations persisted in constant darkness with a fourfold change in amplitude and a peak approximately 4 h after the onset of daily locomotor activity. Electrolytic lesions of the SCN abolished circadian locomotor rhythms, but not odor-induced c-Fos rhythms in the OB or PC. Furthermore, removal of the OB abolished spontaneous circadian cycling of c-Fos in the PC, shortened the free-running period of locomotor rhythms, and accelerated re-entrainment after a 6 h advance and slowed re-entrainment after a 6 h delay in the light schedule. OB ablation or odorant altered the amplitude of c-Fos rhythms in the SCN and ablation of one OB abolished c-Fos rhythms in the ipsilateral PC, but not in the contralateral OB and PC. We conclude that the OB comprises a master circadian pacemaker, which enhances olfactory responsivity each night, drives rhythms in the PC, and interacts with the SCN to coordinate other daily behaviors.

Analysis of Variance↗

Inputs from the olfactory bulb and olfactory cortex to the entorhinal cortex in the cat. II. Physiological studies.

Field potentials and unit activity elicited by electrical stimulation of the olfactory bulb (OB) and anterior and posterior prepiriform cortex (PPCa and PPCp) were measured extracellularly in the entorhinal cortex (EC) of the cat. Different topographic distributions of the amplitude and peak latency of average evoked potentials (AEPs) were obtained depending on the stimulated area. The maximal evoked activity in the EC showed a gradient in a latero-medial direction with the extremes corresponding to the stimulation of OB and PPCp respectively. Analysis of firing patterns of units in the EC in response to stimulation of the OB, PPCa and PPCp showed that an appreciable number of units responded to stimulation of different areas, mainly PPCa and PPCp. It was found that the pathways being stimulated differed in conduction velocities with the PPCp-EC being the slowest. Most responding units lay in layer I and II of the EC. The AEPs to PPCa- and PPCp-stimulation presented different types of depth profiles. Stimulation of the PPCa evoked an initial surface-negative depth-positive potential whereas the PPCp evoked a different type of AEP with an initial positive component at the surface and negative in depth. It is assumed that the stimulated fibres have their active synapses at different levels within the superficial layers of the EC. The possibility of direct influence of olfactory inputs on the hippocampus mediated by one synapse in the EC is discussed.

Animals↗

Olfactory bulb transplantation into the olfactory bulb of neonatal rats.

Embryonic olfactory bulbs (E14-E18) were homotopically transplanted into neonatal rats (P1-P5) after partial or total bulb removal. After different survival times, the animals were sacrificed and their brains processed for histological and immunohistochemical observations. The growing sensory axons randomly formed glomeruli within the transplanted tissue which often acquired a typical laminar organization. Myelinated fibers connecting the transplant with the telencephalon were clearly identified in one animal. Thus, the present study has shown that olfactory bulb transplants were able to differentiate, mature and establish relationship with both the periphery and the brain.

Animals↗

Kindling and electrode effects on the benzodiazepine receptors density of olfactory bulb and hippocampus after olfactory bulb kindling.

The olfactory bulb (OB) kindling is a model of limbic secondary generalized epilepsy. Ten days after the completion of OB kindling, we have studied the long term effects of both electrode insertion and kindling on the binding of [3H]diazepam to crude mitochondrial fractions. On the one hand, we have shown that electrode implantation in sham-operated controls induced an obvious increase in benzodiazepine (BZD) receptor density (Bmax) only at the site of the electrode in comparison to sham-unoperated rats. These results might indicate an additional mechanism extending earlier observations reported by others, who have shown that prolonged electrode implantation induced changes in sham-operated and kindled rats. On the other hand, the long lasting effect of OB kindling on the binding parameters of [3H]diazepam was examined in the focus and in the hippocampus. The results indicate a bilateral increase of BZD receptors in the OB and an ipsilateral increase in the hippocampus. These changes might be a regulation phenomenon in response to a hyperexcitability state and to focal stimulations.

Animals↗

Olfactory bulb transplantation into the olfactory bulb of neonatal rats: a WGA-HRP study.

After unilateral bulbectomy in neonatal (P1-P5) rats, autoradiographically prelabeled presumptive olfactory bulbs from E15 and E17 embryos were transplanted in place of the removed tissue. After 2-7 months, the animals received injections of wheat germ agglutinin-horseradish peroxidase (WGA-HRP) into the piriform cortex. Nine of the twenty animals revealed WGA-HRP-positive neurons among neurons autoradiographically labeled, providing thus evidence that the axons of the output neurons from the homotopically transplanted olfactory bulb reconnect with the host piriform cortex.

Animals↗

Olfactory bulb transplantation into the olfactory bulb of neonatal rats: an autoradiographic study.

Tritiated thymidine prelabeled presumptive olfactory bulbs (E15-E17, and E19) were homotopically transplanted in unilaterally partially or totally bulbectomized neonatal rats (P1-P5). [3H]thymidine was injected to pregnant rats at the time when the large neurons of the bulb were undergoing cellular division. After postoperative survival times from 20 days to 7 months, the animals were sacrificed and processed for histological, immunohistochemical and autoradiographic observations. The nuclear autoradiographic label allowed easy recognition of the transplanted tissue in totally bulbectomized animals after short survival and in partially bulbectomized animals after long survival. The autoradiographic label was strictly confined to the transplanted tissue and intermingling of host and donor neurons was never observed. The reliability of the autoradiographic technique in our study will enable us to mark those neurons whose axons can be demonstrated, by retrograde tracing methods, to establish connections with the host brain.

Animals↗

Volumetric and horseradish peroxidase tracing analysis of rat olfactory bulb following reversible olfactory nerve lesions.

Olfactory receptor neurons can regenerate from basal stem cells. Receptor neuron lesion causes degenerative changes in the olfactory bulb followed by regeneration as new olfactory receptor axons innervate the olfactory bulb. To our knowledge, parametric analyses of morphometric changes in the olfactory bulb during degeneration and regeneration do not exist except in abstract form. To better characterize olfactory bulb response, we performed morphometric analysis in rats following reversible olfactory nerve lesion with diethyldithiocarbamate. We also performed anterograde tracing of the olfactory nerve with wheatgerm agglutinin linked to horseradish peroxidase. Results of morphometry and tracing were complementary. The glomerular layer and external plexiform layer showed shrinkage of 45 and 26%, respectively, at 9 days. No significant shrinkage occurred in any other layer. Individual glomeruli shrank by 40-50% at 3 and 9 days following lesion. These data show that degenerative changes occur both in the glomeruli and transneuronally in the external plexiform layer. Olfactory nerve regeneration (identified by WGA-HRP transport) paralleled volumetric recovery. Recovery occurred first in ventral and lateral glomeruli between 9 and 16 days followed by recovery in medial and dorsal glomeruli. These data indicate substantial transynaptic degeneration in the olfactory bulb and a heretofore unrecognized gradient in olfactory nerve regeneration that can be used to systematically study recovery of a cortical structure.

Animals↗

Levels of amino acid neurotransmitters during mouse olfactory bulb neurogenesis and in histotypic olfactory bulb cultures.

The developmental changes in the levels of amino acid neurotransmitters were analyzed by high pressure liquid chromatography during mouse olfactory bulb neurogenesis, from embryonic day (E)13 until the young adult age, between postnatal days (P)30 and P40. During the embryonic period, high levels of glutamate, aspartate and GABA were observed, with the values of GABA about 2-fold higher than those of glutamate and aspartate. At P0, the production of these neurotransmitters experienced birth stress as shown by a significant 2-fold reduction in their levels. During the first two postnatal weeks, a progressive increase in the glutamate content was detected diminishing slightly in the adult stage. The aspartate concentrations showed a maximal value at P3 and then decreased gradually until the second postnatal week; in the young adult age, its concentration was comparable with that of glutamate. The postnatal GABA contents increased progressively from birth to maturity, showing maximal levels at P3, P11 and in the adult. Throughout the studied developmental period, the concentration of glycine remained relatively low. With regard to taurine, very low concentrations were detected during the prenatal period but after birth, the taurine content gradually increased with age, and in the adult animal, its concentration was comparable with those of GABA and glutamate. Our data demonstrate the predominance of GABA and glutamate during olfactory bulb synaptogenesis, however, in the adult animal, both glutamate and aspartate exert the same influence in the excitatory synaptic transmission; in the adult inhibitory synaptic transmission, taurine appears to play an important neuromodulatory or neurotransmitter role as that of GABA. To determine the intrinsic neurotransmitter production, primary histotypic olfactory bulb cultures were prepared from mice at P10. The comparative analysis of in vitro neurotransmitter contents with those in in situ adult animal showed higher levels of endogenously produced glutamate, glycine and GABA in the olfactory bulb than the extrinsic ones coming from olfactory nerve axons and higher olfactory brain centers. On the other hand, most of aspartate and taurine neurotransmitters apparently come from extrinsically located neurons.

Age Factors↗

Localization of corticotropin-releasing factor-like immunoreactivity in monkey olfactory bulb and secondary olfactory areas.

Electrophysiological and anatomical observations suggest that terminals of olfactory bulb mitral cells ending in rat primary olfactory cortex exert certain postsynaptic effects via an excitatory amino acid neurotransmitter. Recent anatomical studies have shown that several peptides, most notably corticotropin-releasing factor (CRF) (Imaki et al., '89) Brain Res., 496: 35-44), are also localized within rat olfactory bulb projection neurons, thus raising the possibility that there is a peptide cotransmitter in this system. In contrast to the availability of data for rodents, very little is known about the distribution of peptides and other putative transmitters in the olfactory systems of primate species. In the present study, sections through the olfactory bulb and its target areas were obtained from two monkey species (Saimiri sciureus and Macaca fascicularis) and processed for immunohistochemistry with a well-characterized polyclonal antiserum directed against the human form of CRF. Virtually identical results were obtained in the two species. Within the olfactory bulb, nearly all mitral and many tufted cells contained CRF-like immunoreactivity. CRF-positive fibers were seen within the olfactory tract and olfactory stria, which contain the axons of mitral and tufted cells. Within the anterior olfactory nucleus and layer Ia of the olfactory tubercle and piriform cortex, immunoreactivity was seen within fine processes, as well as in coarse, varicose fibers and isolated puncta. CRF-positive cells were seen within layer III of the olfactory tubercle and piriform cortex. Immunoreactive fibers and varicosities were also seen within olfactory-recipient regions of the amygdala and entorhinal cortex. These observations suggest that CRF may act as a transmitter and/or neuromodulator in primate olfactory system.

Amygdala↗