Neurotrophins are not required for normal embryonic development of olfactory neurons.
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Neurons of the vertebrate olfactory epithelium (OE) regenerate continuously throughout life. The capacity of these neurons to regenerate and make new and precise synaptic connections in the olfactory bulb provides a useful model to study factors that may control or mediate neuronal regeneration. Expression and in vitro studies have suggested potential roles for the neurotrophins in the olfactory system. To directly examine whether neurotrophins are required for olfactory neuron development, we characterized in vivo the role of the neurotrophins in the primary olfactory system. For this, we generated mutant mice for TrkA, TrkB, TrkC, and also for BDNF and NT3 together with P2-IRES-tau-LacZ trangenic mice. Histochemical staining for beta-galactosidase at birth allowed in vivo analysis of the P2 subpopulation of olfactory neurons as well as their projections to the olfactory bulb. Our data indicate that Trk signaling is not required for normal embryonic development of the olfactory system. (+info)
Multiple sites of L-histidine decarboxylase expression in mouse suggest novel developmental functions for histamine.
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Histamine mediates many types of physiologic signals in multicellular organisms. To clarify the developmental role of histamine, we have examined the developmental expression of L-histidine decarboxylase (HDC) mRNA and the production of histamine during mouse development. The predominant expression of HDC in mouse development was seen in mast cells. The HDC expression was evident from embryonal day 13 (Ed13) until birth, and the mast cells were seen in most peripheral tissues. Several novel sites with a prominent HDC mRNA expression were revealed. In the brain, the choroid plexus showed HDC expression at Ed14 and the raphe neurons at Ed15. Close to the parturition, at Ed19, the neurons in the tuberomammillary (TM) area and the ventricular neuroepithelia also displayed a clear HDC mRNA expression and histamine immunoreactivity (HA-ir). From Ed14 until birth, the olfactory and nasopharyngeal epithelia showed an intense HDC mRNA expression and HA-ir. In the olfactory epithelia, the olfactory receptor neurons (ORN) were shown to have very prominent histamine immunoreactivity. The bipolar nerve cells in the epithelium extended both to the epithelial surface and into the subepithelial layers to be collected into thick nerve bundles extending caudally toward the olfactory bulbs. Also, in the nasopharynx, an extensive subepithelial network of histamine-immunoreactive nerve fibers were seen. Furthermore, in the peripheral tissues, the degenerating mesonephros (Ed14) and the convoluted tubules in the developing kidneys (Ed15) showed HDC expression, as did the prostate gland (Ed15). In adult mouse brain, the HDC expression resembled the neuronal pattern observed in rat brain. The expression was restricted to the TM area in the ventral hypothalamus, with the main expression in the five TM subgroups called E1-E5. A distinct mouse HDC mRNA expression was also seen in the ependymal wall of the third ventricle, which has not been reported in the rat. The tissue- and cell-specific expression patterns of HDC and histamine presented in this work indicate that histamine could have cell guidance or regulatory roles in development. (+info)
Rhinotopy is disrupted during the re-innervation of the olfactory bulb that follows transection of the olfactory nerve.
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Re-innervation of the olfactory bulb was investigated after transection of the olfactory nerve using monoclonal antibody RB-8 to assess whether rhinotopy of the primary olfactory projection is restored. In normal animals RB-8 heavily stains the axons, and their terminals, that project from the ventrolateral olfactory epithelium onto glomeruli of the ventrolateral bulb (termed RB-8(+)). In contrast, axons from dorsomedial epithelium are unlabeled (RB-8(-)) and normally terminate in the dorsomedial bulb. Sprague-Dawley rats underwent unilateral olfactory nerve transection and survived for 6 weeks prior to perfusion, sectioning and immunostaining with RB-8. Nerve lesion does not shift the position of the boundary between RB-8(+) and RB-8(-) regions of the epithelium. However, following transection and bulb re-innervation, the distribution of RB-8(+) and RB-8(-) axons is markedly abnormal. First, in all 10 experimental animals RB-8(-) axons displace RB-8(+) axons from anterior glomeruli. Furthermore, the usual target of the RB-8(-) fibers, i.e. the dorsomedial bulb at more posterior levels of the bulb, remains denervated, judging by the lack of staining with antibodies that label axons derived from all epithelial zones. Finally, RB-8(+) fibers invade foreign territory in the dorsolateral bulb on the lesioned side in some cases. The shifts in terminal territory in the bulb after transection contrast with the restoration of the normal zonal patterning of the projection after recovery from methyl bromide lesion, but is consistent with reports of mistargeting by a receptor-defined subset of neurons after transection. (+info)
The development of the olfactory mucosa in the mouse: electron microscopy.
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The development of the olfactory epithelium from the 10th day of gestation of postnatal life has been examined electron miscroscopically in the mouse. At 10 days' gestation the epithelium is already differentiated into dark and pale cells, the former representing embryonic stem cells and the latter the developing receptors. Axons are also visible at this stage. At 11 days the first signs of dendrite formation appear, and at 12 days spheroidal terminal swellings containing numerous microtubules are present at the apices of receptor dendrites. Centriole clusters also appear in the receptor cell bodies and dendrites. From the 12th to the 16th day of gestation a few cilia are formed on the receptor endings. Final steps in the maturation of differentiating receptors begin on the 17th day of gestation, when membranous organelles and lysosomes increase greatly in numbers. However, immature receptors can still be found in the base of the epithelium in postnatal life. Supporting cells are first recognizable on the 17th day of gestation, derived apparently from the remaining stem cells. At the same time differentiated basal cells and glands of Bowman begin to appear. In the early develoment of the olfactory nerve bundles the axons have large and varying diameters, but later on axonal sizes are progressively reduced and the adult size range is achieved at about 18 days of gestation. The significance of these findings is discussed. (+info)
Pituitary adenylyl cyclase-activating peptides and alpha-amidation in olfactory neurogenesis and neuronal survival in vitro.
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We investigated the role of amidated neuropeptides, and specifically pituitary adenylyl cyclase-activating polypeptide (PACAP), in olfactory neurogenesis and olfactory receptor neuronal survival. Using both immunohistochemistry and in situ hybridization, we find that both peptidylglycine alpha-amidating monooxygenase (PAM), the enzyme responsible for amidation and therefore activation of all amidated neuropeptides, and amidated PACAP are expressed in developing and adult olfactory epithelium. Amidated PACAP is highly expressed in proliferative basal cells and in immature olfactory neurons. The PACAP-specific receptor PAC(1) receptor is also expressed in this population, establishing that these cells can be PACAP responsive. Experiments were conducted to determine whether amidated neuropeptides, such as PACAP38, might function in olfactory neurogenesis and neuronal survival. Addition of PACAP38 to olfactory cultures increased the number of neurons to >250% of control and stimulated neuronal proliferation and survival. In primary olfactory cultures, pharmacologically decreased PAM activity, as well as neutralization of PACAP38, caused neuron-specific loss that was reversed by PACAP38. Mottled (Brindled) mice, which lack a functional ATP7A copper transporter and serve as a model for Menkes disease, provided an in vivo partial loss-of-function PAM knock-out. These mice had decreased amidated PACAP production and concomitant decreased numbers of olfactory receptor neurons. These data establish amidated peptides and specifically PACAP as having important roles in proliferation in the olfactory system and suggest that a similar function exists in vivo. (+info)
The OMP-lacZ transgene mimics the unusual expression pattern of OR-Z6, a new odorant receptor gene on mouse chromosome 6: implication for locus-dependent gene expression.
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Reporter gene expression in the olfactory epithelium of H-lacZ6 transgenic mice mimics the cell-selective expression pattern known for some odorant receptor genes. The transgene construct in these mice consists of the lacZ coding region, driven by the proximal olfactory marker protein (OMP) gene promoter, and shows expression in a zonally confined subpopulation of olfactory neurons. To address mechanisms underlying the odorant receptor-like expression pattern of the lacZ construct, we analyzed the transgene-flanking region and identified OR-Z6, the first cloned odorant receptor gene that maps to mouse chromosome 6. OR-Z6 bears the highest sequence similarity (85%) to a human odorant receptor gene at the syntenic location on human chromosome 7. We analyzed the expression pattern of OR-Z6 in olfactory tissues of H-lacZ6 mice and show that it bears strong similarities to that mapped for beta-galactosidase. Expression of both genes in olfactory neurons is primarily restricted to the same medial subregion of the olfactory epithelium. Axons from both neuronal subpopulations project to the same ventromedial aspect of the anterior olfactory bulbs. Furthermore, colocalization analyses in H-lacZ6 mice demonstrate that OR-Z6-reactive glomeruli receive axonal input from lacZ-positive neurons as well. These results suggest that the expression of both genes is coordinated and that transgene expression in H-lacZ6 mice is regulated by locus-dependent mechanisms. (+info)
Differential and overlapping expression patterns of X-dll3 and Pax-6 genes suggest distinct roles in olfactory system development of the African clawed frog Xenopus laevis.
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In Xenopus laevis, the formation of the adult olfactory epithelium involves embryonic, larval and metamorphic phases. The olfactory epithelium in the principal cavity (PC) develops during embryogenesis from the olfactory placode and is thought to respond to water-borne odorants throughout larval life. During metamorphosis, the PC undergoes major transformations and is exposed to air-borne odorants. Also during metamorphosis, the middle cavity (MC) develops de novo. The olfactory epithelium in the MC has the same characteristics as that in the larval PC and is thought to respond to water-borne odorants. Using in situ hybridization, we analyzed the expression pattern of the homeobox genes X-dll3 and Pax-6 within the developing olfactory system. Early in development, X-dll3 is expressed in both the neuronal and non-neuronal ectoderm of the sense plate and in all cell layers of the olfactory placode and larval PC. Expression becomes restricted to the neurons and basal cells of the PC by mid-metamorphosis. During metamorphosis, X-dll3 is also expressed throughout the developing MC epithelium and becomes restricted to neurons and basal cells at metamorphic climax. This expression pattern suggests that X-dll3 is first involved in the patterning and genesis of all cells forming the olfactory tissue and is then involved in neurogenesis or neuronal maturation in putative water- and air-sensing epithelia. In contrast, Pax-6 expression is restricted to the olfactory placode, larval PC and metamorphic MC, suggesting that Pax-6 is specifically involved in the formation of water-sensing epithelium. The expression patterns suggest that X-dll3 and Pax-6 are both involved in establishing the olfactory placode during embryonic development, but subtle differences in cellular and temporal expression patterns suggest that these genes have distinct functions. (+info)
Immunohistochemical and ultrastructural studies of 2,6-dimethylaniline-induced nasal proliferative lesions in a rat two-stage nasal carcinogenesis model initiated with N-bis(2-hydroxypropyl)nitrosamine.
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Proliferative lesions induced by 2,6-dimethylaniline (DMA) in a two-stage rat nasal carcinogenesis model were immunohistochemically and ultrastructurally investigated. Male F344 rats received diet containing 3,000 ppm DMA for 52 weeks after initiation with a single subcutaneous injection of 2400 mg/kg of N-bis(2-hydroxypropyl)nitrosamine (DHPN). Histopathologically, proliferation of Bowman's glands, glandular hyperplasias, dysplastic foci, adenomas, and carcinomas were observed in treated rats. These nasal lesions mostly arose in the olfactory mucosa of the nasal cavity. Immunohistochemically, they were positive for cytokeratin and/or collagen type IV antibodies. Ultrastructurally, intracytoplasmic dense secretory granules (200-850 nm in diameter), identical to those in normal Bowman's glands, were observed in all the lesions, providing further support from an origin from these glands. Based on their cellular characterization, growth pattern and/or proliferative activity, two morphological continua were evident, one from dysplastic foci to carcinomas and the other from proliferation of Bowman's glands to glandular hyperplasias and adenomas. These results suggest that dysplastic foci arise from Bowman's glands and progress to carcinomas, while proliferation of Bowman's glands result in glandular hyperplasias and adenomas. (+info)