帕金森病MPTP食蟹猴模型嗅球细胞凋亡

目的 建立1-甲基-4-苯基-1, 2, 3, 6-四氢吡啶（MPTP）帕金森病（PD）模型,探讨模型组嗅球细胞凋亡和胶质细胞增生情况。方法 3只成年健康食蟹猴,静脉注射MPTP建立PD模型,另3只静脉注射生理盐水作为对照。取出嗅球,免疫组织化学染色检测Caspase-3、Bcl-2、离子钙接头蛋白分子1（Iba-1）和胶质纤维酸性蛋白（GFAP）在食蟹猴嗅球中的表达情况,采用Image J v1.8.0软件分析模型组和对照组之间的差异。 结果 MPTP损伤后,与对照组相比,模型组嗅球突触小球层Caspase-3阳性细胞数明显增加,而Bcl-2表达减少;与对照组相比,模型组嗅球突触小球层和外网状层的GFAP和Iba-1阳性细胞数增加。 结论 MPTP可诱导食蟹猴嗅球突触小球层细胞凋亡,并伴有星形胶质细胞增生和小胶质细胞激活,这可能与帕金森病的功能障碍有关。     

Basic fibroblast growth factor in the primary olfactory pathway: mitogenic effect on ensheathing cells.

The mitogenic effect of basic fibroblast growth factor and nerve growth factor (2.5S) on olfactory ensheathing cell culture was examined by bromodeoxyuridine uptake. It was found that, at 10 ng/ml, basic fibroblast growth factor elicited about a three-fold increase in proliferation, while the stimulatory effect of nerve growth factor was considerably less. The increased proliferation resulting from basic fibroblast growth factor could be attributed to perlecan, which was shown to be expressed by ensheathing cell in culture. Perlecan is known to induce high-affinity binding of basic fibroblast growth factor to receptors on cell membranes. Immunohistochemical staining demonstrated that basic fibroblast growth factor was abundantly expressed in select regions of the lamina propria underlying the olfactory epithelium. In these regions, contiguous patches of olfactory epithelium also showed the presence of basic fibroblast growth factor. Although basic fibroblast growth factor was present on the periphery of nerve bundles in the olfactory nerve layer of the bulb, all other laminae did not demonstrate the presence of this factor. The immunohistochemistry and cell culture results show that regions of the lamina propria and small patches of the olfactory epithelium, by their presence of basic fibroblast growth factor, are potential sites of ensheathing cell proliferation in vivo.     

Expression of growth factors in fetal human olfactory mucosa during development

The olfactory epithelium is one of the unique sites in the mammalian nervous system at which there is continual neurogenesis. Constant turnover of primary sensory neurons in the periphery results in remodeling of neuronal circuits and synapses in the olfactory bulb throughout life. Most of the specific mechanisms and factors that control this process are still unknown. Recent studies suggest that growth factors and their receptors may play a significant role in the development and continuous regeneration of olfactory neurons. In this study, standard immunohistochemical methods were used to detect the presence of epidermal growth factor-receptor, transforming growth factor-alpha and nerve growth factor-beta in the human olfactory epithelium of different developmental stages (7 to 30 weeks) in formalin-fixed tissue specimens. Immunoreactivity to epidermal growth factor-receptor was seen in basal cells, supporting cells, neurons, apical cytoplasmic region of olfactory epithelium, lamina propria and gland's cell cytoplasm. Similarly transforming growth factor-alpha immunoreactivity was recognized in basal cells, supporting cells, apical cytoplasmic region of olfactory epithelium, lamina propria and gland's cell cytoplasm but not in neurons. Nerve growth factor-beta detection was restricted on olfactory nerve cells, dentritic knob and basal cell layer. So, this study confirms the development of human olfactory mucosa and shows epidermal growth factor-receptor, transforming growth factor-alpha and nerve growth factor-beta proteins are reliable markers for developing olfactory epithelium.     

Altered glial fibrillary acidic protein immunoreactivity in rat brain following chronic hypoxia

Glial fibrillary acidic immunoreactivity in brain was examined in normal animals and in rats subjected to chronic hypoxia. Animals were exposed to a chronic normobaric adaptive hypoxia with decreasing amounts of oxygen (finally 6%) for a period of 59 and 114 days, respectively. In paraffin-embedded sections the glial fibrillary acidic protein immunoreactivity in normal and hypoxic animals was examined at three coronal levels. A mild glial fibrillary acidic protein immunoreactivity in the perivascular glial layer, external glial limitans membrane and periventricular astrocytes, as well as in some areas of hippocampus and cerebellum, was noted in normal animals. Chronic hypoxia for 114 days resulted in a marked increase of glial fibrillary acidic protein immunoreactivity in dentate gyrus of hippocampus, Bergmann glia of the cerebellum, internal capsule and pyramidal tract. On the other hand, the glial fibrillary acidic protein activity following 59 days hypoxic exposure was almost the same as controls. These results show that systemic deep chronic hypoxia (depending on the intensity and the duration) activates the endogenous expression of glial fibrillary acidic protein in astrocytes of specific brain regions. The probable significance of this finding is discussed.     

Expression of intermediate filaments and desmoplakin in vertebrate olfactory mucosa

The expression of intermediate filaments (IF) and desmoplakin was investigated in frog, bovine, and human (fetal) olfactory mucosa. IF are tissue-specific molecular cytoskeletal markers; desmoplakin is the major desmosomal protein. Positive immunoreactivity was observed in the epithelium and in the subepithelial Bowman's glands to keratin and to desmoplakin, indicating the epithelial nature of this tissue. Desmin, neurofilaments, and glial fibrillary acidic protein (GFAP) were not detected in the mucosa. The absence of neurofilaments and GFAP in the tissue containing sensory neurons and glia-like supporting cells is a unique feature and may be related to the fact that the chemosensory neurons are situated in a bonafide epithelium and are known to undergo continuous turnover. In view of the controversy regarding the expression of vimentin in the olfactory neurons, three independently derived antibodies to vimentin were used; weak or no labeling was found in the epithelium, whereas mesenchymal cells in the lamina propia were labeled with all three antibodies. Olfactory nerve fascicles in the lamina propia were heterogenously labeled: VIM 13.2 gave very weak labeling; aVimAS showed mild labeling and SBV-21 showed intensive labeling in the nerve fascicle. This heterogenous labeling pattern may suggest that olfactory vimentin is distinct in reacting only with some of the antivimentin antibodies.     

Ciliary neurotrophic factor in the olfactory bulb of rats and mice

Ciliary neurotrophic factor (CNTF) is primarily regarded as an astrocytic lesion factor, promoting neuronal survival and influencing plasticity processes in deafferented areas of the CNS. Postnatal loss of neurons in CNTF-deficient mice indicates a function of the factor also under physiological conditions. In the olfactory bulb, where neurogenesis, axo- and synaptogenesis continue throughout life, CNTF content is constitutively high. The cellular localization of CNTF in the rat olfactory bulb is not fully resolved, and species differences between mouse and rat are not yet characterized. In the present study, four different CNTF antibodies and double immunolabeling with specific markers for glial and neuronal cells were used to study the cellular localization of CNTF in rat and mouse olfactory bulb. Specificity of the detection was checked with tissue from CNTF-deficient mice, and investigations were complemented by immunolocalization of reporter protein in mice synthesizing beta-galactosidase under control of the CNTF promoter (CNTF lacZ-knock-in mice). In both species, CNTF localized to ensheathing cell nuclei, cell bodies and axon-enveloping processes. Additionally, individual axons of olfactory neurons were CNTF immunoreactive. Both CNTF protein content and immunoreaction intensity were lower in mice than in rats. Scattered lightly CNTF-reactive cells were found in the granular and external plexiform layers in rats. Some CNTF-positive cells were associated with immunoreactivity for the polysialylated form of the neural cell adhesion molecule, which is expressed by maturing interneurons derived from the rostral migratory stream. In CNTF lacZ-knock-in mice, beta-galactosidase reactivity was found in ensheathing cells of the olfactory nerve layer, and in cells of the glomerular, external plexiform and granular layers. The study proves that CNTF is localized in glial and neuronal structures in the rodent olfactory bulb. Results in mice provide a basis for investigations concerning the effects of a lack of the factor in CNTF-deficient mice.     

Migration of bipolar subependymal cells, precursors of the granule cells of the rat olfactory bulb, with reference to the arrangement of the radial glial fibers

In order to examine the relationship between radial glial fibers and the migrating bipolar subependymal cells which are considered to be post-mitotic precursors of granule cells in the rat olfactory bulb, the arrangement of radial glial fibers along the anterior lateral and olfactory ventricles was analysed by Golgi techniques, immunohistochemical demonstration of glial fibrillary acidic protein, and electron microscopy. In rats during their first 3 weeks of life, the bipolar subependymal cells migrate along the anterior lateral and olfactory ventricles into the center of the olfactory bulb, whereas the radial glial fibers radiating from the ventricular surface are arranged rather perpendicularly to the direction of migration of bipolar cells. Hence radial glial fibers in this region are not considered to act as guides for the rostralwards migration of subependymal cells.     

Immunohistochemistry of the canine vomeronasal organ

The canine's olfactory acuity is legendary, but neither its main olfactory system nor its vomeronasal system has been described in much detail. We used immunohistochemistry on paraffin-embedded sections of male and female adult dog vomeronasal organ (VNO) to characterize the expression of proteins known to be expressed in the VNO of several other mammals. Basal cell bodies were more apparent in each section than in rodent VNO and expressed immunoreactivity to anticytokeratin and antiepidermal growth factor receptor antibodies. The thin layer of neurone cell bodies in the sensory epithelium and axon fascicles in the lamina propria expressed immunoreactivity to neurone cell adhesion molecule, neurone-specific beta tubulin and protein gene product 9.5. Some neurones expressed growth-associated protein 43 (GAP43): and a number of those also expressed neurone-specific beta tubulin-immunoreactivity. Some axon fascicles were double labelled for those two proteins. The G-protein alpha subunits Gi and Go, involved in the signal transduction pathway, showed immunoreactivity in the sensory cell layer. Our results demonstrate that the canine vomeronasal organ contains a population of cells that expresses several neuronal markers. Furthermore, GAP43 immunoreactivity suggests that the sensory epithelium is neurogenic in adult dogs.     

Immunohistochemistry of the canine vomeronasal organ

The canine's olfactory acuity is legendary, but neither its main olfactory system nor its vomeronasal system has been described in much detail. We used immunohistochemistry on paraffin-embedded sections of male and female adult dog vomeronasal organ (VNO) to characterize the expression of proteins known to be expressed in the VNO of several other mammals. Basal cell bodies were more apparent in each section than in rodent VNO and expressed immunoreactivity to anticytokeratin and antiepidermal growth factor receptor antibodies. The thin layer of neurone cell bodies in the sensory epithelium and axon fascicles in the lamina propria expressed immunoreactivity to neurone cell adhesion molecule, neurone-specific beta tubulin and protein gene product 9.5. Some neurones expressed growth-associated protein 43 (GAP43): and a number of those also expressed neurone-specific beta tubulin-immunoreactivity. Some axon fascicles were double labelled for those two proteins. The G-protein alpha subunits Gi and Go, involved in the signal transduction pathway, showed immunoreactivity in the sensory cell layer. Our results demonstrate that the canine vomeronasal organ contains a population of cells that expresses several neuronal markers. Furthermore, GAP43 immunoreactivity suggests that the sensory epithelium is neurogenic in adult dogs.     

Organisation and chemical neuroanatomy of the African elephant (<Emphasis Type="Italic">Loxodonta africana</Emphasis>) olfactory bulb

The olfactory system of mammals can be divided into a main and accessory olfactory system with initial processing for each system occurring in the olfactory bulb. The main and accessory olfactory bulbs have similar structural features, even though they appear to be functionally independent. In mammals the main olfactory bulb (MOB) is also one of two established sites of lifelong generation of new cells. The present study describes the histological and immunohistochemical neuroanatomy of the olfactory bulb of the African elephant (Loxodonta africana). The morphology of MOB of the elephant does not differ significantly from that described in other mammals; however, it lacks the internal plexiform layer. In addition, the glomeruli of the glomerular layer are organised in 2–4 “honey-combed” layers, a feature not commonly observed. The cell types and structures revealed with immunohistochemical stains (parvalbumin, calbindin, calretinin, tyrosine hydroxylase, orexin-A, glial fibrillary acidic protein) were similar to other mammals. Neurogenesis was examined using the neurogenic marker doublecortin. Migration of newly generated cells was observed in most layers of the MOB. No accessory olfactory bulb (AOB) was observed. Based on the general anatomy and the immunohistochemical observations, it is evident that the morphology of the African elephant MOB is, for the most part, similar to that of all mammals, although very large in absolute size.     

Expression of connexin 57 in the olfactory epithelium and olfactory bulb

In the visual system, deletion of connexin 57 (Cx57) reduces gap junction coupling among horizontal cells and results in smaller receptive fields. To explore potential functions of Cx57 in olfaction, in situ hybridization and immunohistochemistry methods were used to investigate expression of Cx57 in the olfactory epithelium and olfactory bulb. Hybridization signal was stronger in the olfactory epithelial layer compared to the connective tissue underneath. Within the sensory epithelial layer, hybridization signal was visible in sublayers containing cell bodies of basal cells and olfactory neurons but not evident at the apical sublayer comprising cell bodies of sustentacular cells. These Cx57 positive cells were clustered into small groups to form different patterns in the olfactory epithelium. However, individual patterns did not associate with specific regions of olfactory turbinates or specific olfactory receptor zones. Patched distribution of hybridization positive cells was also observed in the olfactory bulb and accessory olfactory bulb in layers where granule cells, mitral cells, and juxtaglomerular cells reside. Immunostaining was observed in the cell types described above but the intensity was weaker than that in the retina. This study has provided anatomical basis for future studies on the function of Cx57 in the olfactory system.     

Expression of netrin-1 and netrin-1 receptor, DCC, in the rat olfactory nerve pathway during development and axonal regeneration

Netrin-1 is a bifunctional secreted protein that directs axon extension in various groups of developing axonal tracts. The transmembrane DCC (deleted in colorectal cancer) receptor is described as netrin-1 receptor and is involved in the attractive effects of netrin-1. In this study, we examined the spatio-temporal expression patterns of both netrin-1 and DCC in the rat olfactory system at different stages of development and during axonal regeneration following unilateral bulbectomy. High DCC expression was detected on the pioneer olfactory axons as they are extending toward the telencephalon. This expression was transient since from embryonic day 16 onwards, DCC was no longer detected along the olfactory nerve path. From embryonic day 14 until birth, DCC was also expressed within the mesenchyme surrounding the olfactory epithelium. During the same period, netrin-1 protein was detected along the trajectory of olfactory axons up to the olfactory bulb and its expression pattern in the nasal mesenchyme largely overlapped that of DCC. Moreover, netrin-1 continued to be present during the two first post-natal weeks, and a weak protein expression still persisted in the dorso-medial region of the olfactory epithelium in adult rats. While unilateral bulbectomy induced a transient up-regulation of netrin-1 in the lamina propria, particularly in the dorso-medial region of the neuroepithelium, no DCC expression was detected on the regenerating olfactory axons. In the developing olfactory bulb, the extension of mitral cell axons was associated with DCC presence while netrin-1 was absent along this axonal path. DCC was also highly expressed in the newly formed glomeruli after birth, and a weak DCC expression was still detected in the glomerular layer in adult rats. Taken together, these data support the notion that netrin-1, via DCC expressed on axons, may play a role in promoting outgrowth and/or guidance of pioneering olfactory axons toward the olfactory bulb primordium. Moreover, association of netrin-1 with mesenchymal DCC may provide a permissive environment to the growth of both pioneer and later-growing axons. The maintenance of netrin-1 expression in the nasal mesenchyme of adult rats as well as its regional up-regulation following unilateral bulbectomy infer that netrin-1, even in the absence of DCC, may be involved in the process of axonal growth of newly differentiated olfactory receptor neurons probably through the use of other receptors.     

Morphological characteristics of dopaminergic immunoreactive neurons in the olfactory bulb of the common marmoset monkey (Callithrix jacchus)

The present study describes the distribution of tyrosine hydroxylase (TH)-immunoreactive (IR) elements in the olfactory bulb of the common marmoset monkey (Callithrix jacchus), a primate species by immunohistochemistry. We identified six layers of the olfactory bulb of the common marmoset monkey in sections stained with cresyl violet. The majority of TH-IR cells were found in the glomerular layer. A few TH-IR cells were present in the external plexiform and granule cell layers. TH-IR fibers were identified in all layers of the olfactory bulb. The density of these nerve fibers was high in the internal plexiform and granule cell layers. The results in the olfactory bulb of the common marmoset monkey are generally similar to previous reports in some mammals. These data suggest that TH in the olfactory bulb of the common marmoset monkey may play a role in olfactory transmission via the glomeruli like in other mammals.     

Formation of an olfactory glomerulus: morphological aspects of development and organization.

We have studied the development of olfactory nerves in the rat from their first contact with the telencephalic vesicle until the formation of glomerular structures in the olfactory bulb at early postnatal period. The study is based on serial semithin and ultrathin sections of material prepared for electron microscopy and antibodies to label radial glial cells, glial fibrillary acidic protein and Rat-401. Beginning on embryonic day 12, developing olfactory axons from the olfactory placode are accompanied by migratory cells, also derived from the olfactory placode, that reach the prospective olfactory bulb by embryonic day 13. The mass of migratory cells accumulate superficial to the telencephalic vesicle. The cells increase in number by mitotic divisions. The majority of these cells represent precursor elements that will later develop into the ensheathing cells of the olfactory nerves and olfactory nerve layer of the adult. Some migratory cells penetrate into the prospective olfactory bulb early during development. The first synaptic contacts of olfactory axons with dendritic processes in the olfactory bulb were observed at embryonic day 18. Glomerular formation is initiated by penetration of cells from the migratory mass into the prospective glomerular layer by embryonic day 20 to postnatal day 0. These cells form walls surrounding zones of high synaptic density forming protoglomeruli. Postnatally, the peripheral processes of radial glial cells branch profusely delimiting glomerular formations and transform into periglomerular astrocytes. Rat-401 stains radial glial cells from embryonic day 14. Immunoreactivity becomes restricted to the olfactory glomeruli during the first postnatal weeks and it virtually disappears by the end of the first postnatal month. We conclude that the early penetration of cells from the migratory mass into the prospective olfactory bulb, observed immediately after the first synaptic contacts were established, initiates the formation of olfactory glomeruli which becomes completed by the transformation of radial glial cells into periglomerular astrocytes.     

Analysis of synaptic events in the mouse accessory olfactory bulb with current source-density techniques.

The accessory olfactory bulb of the mouse was studied by current source-density analysis of field potentials to determine the laminar and temporal distribution of synaptic currents evoked by electrical stimulation of the vomeronasal organ. The one-dimensional current source-density analysis revealed two major spatially and temporally distinct inward membrane currents (sinks): one in the glomerular layer and the other in the external plexiform layer. The glomerular layer sink preceded the external plexiform layer sink by a mean of 5.5 ms. Local infusions of the broad-spectrum excitatory amino acid antagonist, kynurenate, into the accessory olfactory bulb blocked the external plexiform layer sink without an obvious effect on the glomerular layer sink. The selective non-N-methyl-D-aspartate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione produced a dose-dependent blockade of the external plexiform layer sink, whereas the selective N-methyl-D-aspartate receptor antagonist D-2-amino-5-phosphonovalerate was without effect. These results, taken together with the cytoarchitecture of the accessory olfactory bulb, suggest that the glomerular layer sink results mainly from synaptic excitation evoked in the glomerular dendritic branches of mitral cells by the vomeronasal afferent fibres and the external plexiform layer sink mainly from non-N-methyl-D-aspartate receptor-mediated synaptic excitation in the peripheral processes of granule cells via the mitral to granule cell dendrodendritic synapse.     

成年大鼠嗅球成鞘细胞的原代培养

本实验取材于成年 Wistar大鼠嗅球的最外两层 ,进行原代培养嗅球成鞘细胞 ,用胶质纤维酸性蛋白抗体免疫组化方法确认。结果显示 :( 1)培养至第 10 d,原代培养物多见两种形态的细胞 :一种为单极、双极或多极细胞 ,带有细长的突起 ;另一种为所谓“煎蛋样”细胞 ,扁平状 ,胞质少极化 ,边缘不规则 ;( 2 )胶质纤维酸性蛋白反应显示这两种细胞均呈阳性 ,Nissl法复染后 ,可计数其纯度为 70 %～ 85 %。     

Expression of the secreted factors noggin and bone morphogenetic proteins in the subependymal layer and olfactory bulb of the adult mouse brain

The antagonism between noggin and the bone morphogenetic proteins (BMPs) plays a key role during CNS morphogenesis and differentiation. Recent studies indicate that these secreted factors are also widely expressed in the postnatal and adult mammalian brain in areas characterized by different types of neural plasticity. In particular, significant levels of noggin and BMP expression have been described in the rodent olfactory system. In the mammalian forebrain, the olfactory bulb (OB) and associated subependymal layer (SEL) are documented as sites of adult neurogenesis. Here, using multiple approaches, including the analysis of noggin-LacZ heterozygous mice, we report the expression of noggin and two members of the BMP family, BMP4 and BMP7, in these regions of the adult mammalian forebrain. We observe that along the full extent of the SEL, from the lateral ventricle to the olfactory bulb, noggin and BMP4 and 7 are mainly associated with the astrocytic glial compartment. In the OB, BMP4 and 7 proteins remain primarily associated with the SEL while strong noggin expression was also found in cells located in different OB layers (i.e. granule, external plexiform, glomerular layers). Taken together our data lead us to hypothesize that within the SEL the antagonism between noggin and BMPs, both produced by the glial tubes, act through autocrine/paracrine inductive mechanisms to maintain a neurogenetic environment all the way from the lateral ventricle to the olfactory bulb. In the OB, their expression patterns suggest multiple regulatory roles on the unusual neural plasticity exhibited by this region.     

Glial cells from adult rat olfactory bulb: immunocytochemical properties of pure cultures of ensheathing cells.

Three morphologically and immunohistochemically distinct types of cell were present in primary cultures of adult rat olfactory nerve and glomerular layers of the olfactory bulb. One cell type was multipolar and stained positively for glial fibrillary acidic protein; a second type had fried egg-like morphology and stained with antibodies to epitope ED1; the third cell type had fusiform morphology, reacted with antibodies to vimentin and laminin and was glial fibrillary acidic protein- and ED1-negative. Trypsinization of these primary cultures (3 min, 37 degrees C), detached multipolar and fusiform cells only. When detached cells were set up in secondary culture on a glass substrate, fusiform cells did not attach, resulting in a pure culture of multipolar cells. Multipolar cells were glial fibrillary acidic protein- and myelin basic protein-positive and had the properties of so-called ensheathing cells or Blanes' glia. Immunoreactivity with anti-nerve growth factor receptor and anti-fibronectin allowed us to identify four distinct populations of multipolar ensheathing cells. One population was nerve growth factor receptor-positive, fibronectin-negative. A second was nerve growth factor receptor-negative and fibronectin-positive. A third was positive for both markers and the remaining cells did not stain for either of them. The morphological and immunological characteristics of cultured cells from olfactory nerve and glomerular layers were similar to those of Schwann cells and the similarities could account for the permissivity to axonal growth of the olfactory bulb.     

Origin and migration of interneurons of the olfactory bulb in the musk shrew,Suncus murinus

The olfactory bulb of the musk shrew, Suncus murinus, is characterized by the presence of various interneurons. Our previous report (Kakuta et al., 2001) demonstrated that positive immunoreactions for calretinin were observed in periglomerular and perinidal cells in the glomerular layer, small ovoid neurons in the external plexiform layer, and granule cells in the granule cell layer of the olfactory bulb in the musk shrew aged 1 to 5 weeks, in addition to calretinin-immunoreactive bipolar cells distributed in the anterior subependymal layer and in each layer of the olfactory bulb. To examine the origin and migration of interneurons of the olfactory bulb, we labeled generated cells by injecting 28-day-old musk shrews with 5-bromo-2'-deoxyuridine (BrdU), and detected the labeled progeny cells that survived after several intervals. BrdU-labeled cells originated in the subependymal layer around the anterior horn of the lateral ventricle, and rostrally migrated in the subependymal layer from the anterior wall of the lateral ventricle into the center of the olfactory bulb, where they radially migrated into the granule cell layer, external plexiform layer, and glomerular layer. It took 2 days to migrate rostrally in the subependymal layer from the anterior lateral ventricle to the center of the olfactory bulb, and 2 to 6 days to migrate radially from the bulbar subependymal layer into the three layers mentioned. The rate of rostralward migration of the labeled cells was estimated to be 38 microm/h, while that of radial migration, 7 to 25 microm/h. The present BrdU-labeling study, together with our previous immunohistochemical study (Kakuta et al., 2001), indicates that anterior subependymal cells differentiate into granule cells in the granule cell layer, into Van Gehuchten cells in the external plexiform layer, and into periglomerular and perinidal cells in the glomerular layer of the olfactory bulb in the musk shrew.