Olfactory deficit is associated with mitral cell dysfunction in the olfactory bulb of P301S tau transgenic mice.

Neurofibrillary tangles consisting of hyperphosphorylated tau (P-tau) are the neuropathological hallmark of Alzheimer's disease (AD), and olfaction disorder is an early symptom of AD. However, the link between P-tau aggregation and olfaction disorder remains unclear. In this study, the expression of P-tau in the olfactory bulb (OB), particularly in the mitral cell layer (MCL), external plexiform layer (EPL), and granule cell layer (GCL), of AD patients was found to be significantly higher than that in the OB of normal aging subjects, which suggested that these layers in the OB were susceptible to P-tau. The P301S tau transgenic mice (P301S mice) exhibit AD-like features, which can be characterized by olfactory dysfunction that precedes cognitive disorder. Importantly, the excessive P-tau expression in the OB of P301S mice, particularly in MCs, was associated with MC loss at 9 months of age, and decreased MC firing activities started to be observed at 2 months of age. Our results revealed that MCs might contribute to olfactory dysfunction in P301S mice. Furthermore, we described an aberrant dendro-dendritic synaptic structure between granule cells (GCs) and MCs and abnormal gamma oscillations in the EPL of the OB, and these findings indicated that P-tau might disrupt the regulation of MCs by GCs in P301S mice starting at 5 months of age. These data showed that the reduction in the MC firing frequency at 2 months of age might not be caused by GC suppression. Based on these findings, we speculated that MCs are a putative target for the treatment of P-tau-induced early olfactory dysfunction, and thus, an exploration of the specific causes and mechanisms of MC functional changes in P301S mice is crucial.

Impairment of Dendrodendritic Inhibition in the Olfactory Bulb of APP/PS1 Mice.

Olfactory dysfunction is an early event in Alzheimer's disease (AD). However, the mechanism underlying the AD-related changes in the olfactory bulb (OB) remains unknown. Granule cells (GCs) in the OB regulate the activity of mitral cells (MCs) through reciprocal dendrodendritic synapses, which is crucial for olfactory signal processing and odor discrimination. Nevertheless, the relationships between the morphological and functional changes of dendrodendritic synapses, particularly the local field potentials (LFPs) as a consequence of olfactory disorders in patients with AD have not been investigated. Here, we studied the morphological and functional changes induced by dendrodendritic inhibition in GCs onto MCs in the OB of amyloid precursor protein (APP)/PS1 mice and age-matched control mice during aging, particular, we focused on the effects of olfactory disorder in the dendrodendritic synaptic structures and the LFPs. We found that olfactory disorder was associated with increased amyloid-ß (Aß) deposits in the OB of APP/PS1 mice, and those mice also exhibited abnormal changes in the morphology of GCs and MCs, a decreased density of GC dendritic spines and impairments in the synaptic interface of dendrodendritic synapses between GCs and MCs. In addition, the aberrant enhancements in the γ oscillations and firing rates of MCs in the OB of APP/PS1 mice were recorded by multi-electrode arrays (MEAs). The local application of a GABAAR agonist nearly abolished the aberrant increase in γ oscillations in the external plexiform layer (EPL) at advanced stages of AD, whereas a GABAAR antagonist aggravated the γ oscillations. Based on our findings, we concluded that the altered morphologies of the synaptic structures of GCs, the dysfunction of reciprocal dendrodendritic synapses between MCs and GCs, and the abnormal γ oscillations in the EPL might contribute to olfactory dysfunction in AD.

MiR-124 Promotes Newborn Olfactory Bulb Neuron Dendritic Morphogenesis and Spine Density.

Using microarray analysis, we detected microRNA-124 (miR-124) to be abundantly expressed in the olfactory bulb (OB). miR-124 regulates adult neurogenesis in the subventricular zone (SVZ). However, much less is known about its role in newborn OB neurons. Here, using both gain-of-function and loss-of-function approaches, we demonstrate that brain-specific miR-124 affects dendritic morphogenesis and spine density in newborn OB neurons. Functional Annotation Clustering of miR-124 targets was enriched in "cell morphogenesis involved in neuron differentiation."

Exposure to Sevoflurane Affects the Development of Parvalbumin Interneurons in the Main Olfactory Bulb in Mice.

Sevoflurane is widely used in adult and pediatric patients during clinical surgeries. Although studies have shown that exposure to sevoflurane impairs solfactory memory after an operation, the neuropathological changes underlying this effect are not clear. This study detected the effect of sevoflurane exposure on the development of calcium-binding proteins-expressing interneurons in the main olfactory bulb (MOB). We exposed neonatal mice to 2% sevoflurane at two different developmental time points and found that exposing mice to sevoflurane at postnatal day (PD) 7 significantly decreased the expression of GAD67 and parvalbumin (PV) in the olfactory bulb (OB) but did not alter the expression of calretinin (CR) or calbindin D28k (CB). The number and dendritic morphology of PV-expressing interneurons in the MOB were impaired by exposure to sevoflurane at PD7. However, exposure to sevoflurane at PD10 had no effect on calcium-binding protein expression or the number and dendritic morphology of PV-expressing interneurons in the MOB. These results suggest that exposing neonatal mice to sevoflurane during a critical period of olfactory development affects the development of PV-expressing interneurons in the MOB.

Altered Morphologies and Functions of the Olfactory Bulb and Hippocampus Induced by miR-30c.

Adult neurogenesis is considered to contribute to a certain degree of plasticity for the brain. However, the effects of adult-born neurons on the brain are still largely unknown. Here, we specifically altered the expression of miR-30c in the subventricular zone (SVZ) and dentate gyrus (DG) by stereotaxic injection with their respective up- and down-regulated lentiviruses. Results showed an increased level of miR-30c enhanced adult neurogenesis by prompting cell-cycles of stem cells, whereas down-regulated miR-30c led to the opposite results. When these effects of miR-30c lasted for 3 months, we detected significant morphological changes in the olfactory bulb (OB) and lineage alteration in the hippocampus. Tests of olfactory sensitivity and associative and spatial memory showed that a certain amount of adult-born neurons are essential for the normal functions of the OB and hippocampus, but there also exist redundant newborn neurons that do not further improve the functioning of these areas. Our study revealed the interactions between miRNA, adult neurogenesis, brain morphology and function, and this provides a novel insight into understanding the role of newborn neurons in the adult brain.

microRNA Profiling of Amniotic Fluid: Evidence of Synergy of microRNAs in Fetal Development.

Amniotic fluid (AF) continuously exchanges molecules with the fetus, playing critical roles in fetal development especially via its complex components. Among these components, microRNAs are thought to be transferred between cells loaded in microvesicles. However, the functions of AF microRNAs remain unknown. To date, few studies have examined microRNAs in amniotic fluid. In this study, we employed miRCURY Locked Nucleotide Acid arrays to profile the dynamic expression of microRNAs in AF from mice on embryonic days E13, E15, and E17. At these times, 233 microRNAs were differentially expressed (p< 0.01), accounting for 23% of the total Mus musculus microRNAs. These differentially-expressed microRNAs were divided into two distinct groups based on their expression patterns. Gene ontology analysis showed that the intersectional target genes of these differentially-expressed microRNAs were mainly distributed in synapse, synaptosome, cell projection, and cytoskeleton. Pathway analysis revealed that the target genes of the two groups of microRNAs were synergistically enriched in axon guidance, focal adhesion, and MAPK signaling pathways. MicroRNA-mRNA network analysis and gene- mapping showed that these microRNAs synergistically regulated cell motility, cell proliferation and differentiation, and especially the axon guidance process. Cancer pathways associated with growth and proliferation were also enriched in AF. Taken together, the results of this study are the first to show the functions of microRNAs in AF during fetal development, providing novel insights into interpreting the roles of AF microRNAs in fetal development.

miR-30c and semaphorin 3A determine adult neurogenesis by regulating proliferation and differentiation of stem cells in the subventricular zones of mouse.

OBJECTIVES: Mechanisms that regulate proliferation of adult neural stem cells are largely unknown. Here, we have investigated the role of microR-30c (miR-30c) and its target, semaphoring 3A (sema3A), in regulating adult neurogenesis and mechanisms underlying this process. MATERIALS AND METHODS: In situ hybridization, immunofluorescence and quantitative real-time PCR were used to assess complementary expression patterns of miR-30c and sema3A in mice. Effects of miR-30c in the subventricular zone (SVZ) were examined by stereotaxic injection of up- and down-regulating lentiviruses. 5'-bromo-2'-deoxyuridine labelling was performed to investigate effects of miR-30c and sema3A on adult neurogenesis. Real-time cell assays, morphological analysis and cell cycle measurements were used to reveal the mechanisms by which miR-30c and sema3A regulate adult neurogenesis. RESULTS: Expression of miR-30c negatively correlated with that of sema3A in neurons, and levels of miR-30c and sema3A correlated positively with numbers of newborn cells in the SVZ and rostral migration stream. miR-30c and sema3A affected adult neurogenesis by regulating proliferation and differentiation, as well as cycles of stem cells in the SVZ. CONCLUSIONS: miR-30c and sema3A regulate adult neurogenesis by controlling proliferation and differentiation of stem cells in the SVZ. This finding reveals a novel regulatory mechanism of adult neurogenesis.

Formaldehyde exposure alters miRNA expression profiles in the olfactory bulb.

It has been reported that inhaling formaldehyde (FA) causes damage to the central nervous system. However, it is unclear whether FA can disturb the function of the olfactory bulb. Using a microarray, we found that FA inhalation altered the miRNA expression profile. Functional enrichment analysis of the predicted targets of the changed miRNA showed that the enrichment canonical pathways and networks associated with cancer and transcriptional regulation. FA exposure disrupts miRNA expression profiles within the olfactory bulb.

Neuroprotective effects of DAHP and Triptolide in focal cerebral ischemia via apoptosis inhibition and PI3K/Akt/mTOR pathway activation.

Triptolide (TP), one of the major active components of the traditional Chinese herb Tripterygium wilfordii Hook F, and 2, 4-diamino-6-hydroxypyrimidine (DAHP), an inhibitor of tetrahydrobiopterin (BH4) synthesis, have been reported to have potent anti-inflammatory and immunosuppressive properties. However, the protective effects of TP and DAHP on cerebral ischemia have not been reported yet. In this study, we investigated the neuroprotective effects of TP and DAHP in a middle cerebral artery occlusion (MCAO) rat model. Furthermore, we examined whether the neuroprotective effects of TP and DAHP were associated with the inhibition of apoptosis through suppressing BH4 and inducible NOS (iNOS) synthesis or the activation of the phosphoinositide-3-kinase/serine-threonine kinase Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway. Our results showed that pretreatments with TP (0.2 mg/kg) and DAHP (0.5 g/kg) significantly reduced ischemic lesion volume, water content, and neuronal cell death compared with the vehicle MCAO rats. In addition, compared with the MCAO group, TP, and DAHP pretreatment groups significantly reduced astrocyte numbers, caspase-3, cleaved caspase-3, and NF-κB up-regulation, while increased Bcl-2 expression. Moreover, protein expressions of PI3K, Akt, and mTOR increased, while extracellular signal-regulated protein kinases 1 and 2 (ERK1 and ERK2) phosphorylation decreased in both the TP-treated rats and DAHP-treated rats. These results demonstrate that TP and DAHP can decrease cell apoptosis in focal cerebral ischemia rat brains and that the mechanism may be related to the activation of the PI3K/Akt/mTOR pathway and inactivation of the ERK1/2 pathway. Thus our hypothesis was reached PI3K/Akt/mTOR and ERK1/2 pathways may provide distinct cellular targets for a new generation of therapeutic agents for the treatment of stroke, and TP and DAHP may be potential neuroprotective agents for cerebral ischemia/reperfusion injury.

Autophagy upregulation and apoptosis downregulation in DAHP and triptolide treated cerebral ischemia.

It has previously been demonstrated that ischemic stroke activates autophagy pathways; however, the mechanism remains unclear. The aim of this study is to further investigate the role that autophagy plays in cerebral ischemia. 2, 4-diamino-6-hydroxy-pyrimidine (DAHP), for its nitric oxide synthase (NOS) inhibiting neuroprotective effect, and triptolide (TP), for its anti-inflammatory property, were selected to administer pre middle cerebral artery occlusion (MCAO). The drugs were administered 12 hours prior to MCAO. Both magnetic resonance imaging (MRI) and 2, 3, 5-triphenyltetrazolium chloride (TTC) staining showed that the drugs reduce the area of infarction. Immunoblotting analysis revealed increases in Beclin-1 and myeloid cell leukelia-1(Mcl-1) in treated rats. This could be a contributing factor to the reduction in autophagy induced damage. Immunochemistry and western blot showed that mTOR expression in treated rats was marginally different 24 h after injury, and this could also be significant in the mechanism. Furthermore, terminal deoxynucleotidyl transferase- (TdT-) mediated dUTP nick end labeling (TUNEL) staining proved that the drugs are effective in reducing apoptosis. The upregulation of Beclin-1 and Mcl-1 and downregulation of Bcl-2, caspase-3, and the Bcl-2/Beclin-1 ratio infer that the neuroprotective effect of DAHP and TP act via the mediation of autophagy and apoptosis pathways.

Identification of a microRNA regulator for axon guidance in the olfactory bulb of adult mice.

Semaphorin3A (sema3a), mainly localized in the olfactory neuron layer and periglomerular layer, is essential for the normal arrangement of axons in the olfactory bulb both in embryonic and adult mice functioning through its dynamic spatiotemporal expression. The regulators that can modulate the expression of sema3a by direct interaction, however, are unknown. In order to find the regulators of sema3a in the olfactory bulb, we focused on microRNAs, well-known post-transcriptional regulators. We found that axon guidance is the main molecular and biological process ongoing in the steady-state olfactory bulb of the adult mouse by screening the abundant microRNAs and exploring their functions in the olfactory bulb via our customized microRNA arrays, Gene Ontology and Kyoto Encyclopedia of Genes annotation. Furthermore, we traced the expression of three candidate regulators (miR-30c, miR-200b, and miR-429) and sema3a by the quantitative real-time polymerase chain reaction and immunohistochemistry. The results showed that only miR-30c expression corresponded inversely with sema3a. Finally, miR-30c was verified to be a specific regulator of sema3a by dual luciferase reporter assay in vitro. Taken together, our results suggested that miR-30c is a potential regulator in axon-guidance by suppressing the expression of sema3a, which will give new insights in elucidating the mechanism of architectonic and functional maintenance of the olfactory bulb.

Systematic hypothesis for post-stroke depression caused inflammation and neurotransmission and resultant on possible treatments.

Post-stroke depression (PSD) is a prevalent complex psychiatric disorder that causes delay to functional recovery from rehabilitation and also increases cognitive impairment. The etiology of PSD remains controversial and appears to be physical and psycho-social in origin, alone or in combination. The causes of PSD as well as the mechanisms conferring beneficial antidepressant effects in the context of ischemic brain injury are still unknown. In addition, appropriate treatment strategies for therapy to prevent stroke-induced depression-like behavior remain to be developed. This paper, therefore, proposes two hypotheses for post-stroke depression: The inflammatory hypothesis, which is the increased production of proinflammatory cytokines resulting from brain ischemia in cerebral areas causing the pathogenesis of post-stroke depression and the glutamate hypothesis, where the excess glucocorticoids released from stress-induced over-activation of hypothalamus-pituitary-adrenal (HPA) lead to dysfunction of glutamatergic transmission. Neurotrophins, especially brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) both play various roles in the central nervous system (CNS), attenuate apoptosis in cultured neurons, stimulate neurogenesis and increase survival and protect neuronal tissues from cell death induced by ischemia or depression. We also touch upon recent treatment strategies including inhibition of pro-inflammatory cytokines, SSRI, neurotrophins and cell-based therapies. In the present review, we provide an overview of recent evidence concerning the mechanisms of post-stroke depression and propose four prospective treatment strategies so as to provide references for clinical evidence-based medications.

17ß-estradiol protects human eyelid-derived adipose stem cells against cytotoxicity and increases transplanted cell survival in spinal cord injury.

Stem cell transplantation represents a promising strategy for the repair of spinal cord injury (SCI). However, the low survival rate of the grafted cells is a major obstacle hindering clinical success because of ongoing secondary injury processes, which includes excitotoxicity, inflammation and oxidative stress. Previous studies have shown that 17b-estradiol (E2) protects several cell types against cytotoxicity. Thus, we examined the effects of E2 on the viability of human eyelid adipose-derived stem cells (hEASCs) in vitro with hydrogen peroxide (H2O2)-induced cell model and in vivo within a rat SCI model. Our results showed that E2 protected hEASCs against H2O2-induced cell death in vitro, and enhanced the survival of grafted hEASCs in vivo by reducing apoptosis. Additionally, E2 also enhanced the secretion of growth factors by hEASCs, thereby making the local microenvironment more conducive for tissue regeneration. Overall, E2 administration enhanced the therapeutic efficacy of hEASCs transplantation and facilitated motor function recovery after SCI. Hence, E2 administration may be an intervention of choice for enhancing survival of transplanted hEASCs after SCI.

[Research progress of external tufted cells in olfactory glomerulus].

External tufted (ET) cells are the major excitatory elements coordinating the activities of glomerulars and mediating the input from the olfactory neurons to mitral cells. The ET cells participate in inter-and intra-glomerular microcircuits in the olfactory bulb, link the isofunctional odor columns within the same olfactory bulb, and play an important role in olfactory information processing. This paper reviews the research progress of the anatomy and physiological properties and electrophysiological modeling of ET cells, elaborate the problems and defects in the field. And then it further gives some proposals for the future research of electrophysiological properties, development of olfactory information coding and performance of modeling of ET cells.

Endoplasmic reticulum stress-mediated hippocampal neuron apoptosis involved in diabetic cognitive impairment.

Poor management of DM causes cognitive impairment while the mechanism is still unconfirmed. The aim of the present study was to investigate the activation of C/EBP Homology Protein (CHOP), the prominent mediator of the endoplasmic reticulum (ER) stress-induced apoptosis under hyperglycemia. We employed streptozotocin- (STZ-) induced diabetic rats to explore the ability of learning and memory by the Morris water maze test. The ultrastructure of hippocampus in diabetic rats and cultured neurons in high glucose medium were observed by transmission electron microscopy and scanning electron microscopy. TUNEL staining was also performed to assess apoptotic cells while the expression of CHOP was assayed by immunohistochemistry and Western blot assay in these hippocampal neurons. Six weeks after diabetes induction, the escape latency increased and the average frequency in finding the platform decreased in diabetic rats (P < 0.05). The morphology of neuron and synaptic structure was impaired; the number of TUNEL-positive cells and the expression of CHOP in hippocampus of diabetic rats and high glucose medium cultured neurons were markedly altered (P < 0.05). The present results suggested that the CHOP-dependent endoplasmic reticulum (ER) stress-mediated apoptosis may be involved in hyperglycemia-induced hippocampal synapses and neurons impairment and promote the diabetic cognitive impairment.

C16 peptide shown to prevent leukocyte infiltration and alleviate detrimental inflammation in acute allergic encephalomyelitis model.

Integrins are important adhesion receptors for leukocytes binding to endothelial cellular adhesion molecules. Previous studies have suggested that blocking relevant integrins might prevent leukocyte infiltration and suppress clinical and pathological features of neuroinflammatory disease. Experimental autoimmune encephalomyelitis (EAE), a rodent model of Multiple sclerosis (MS), is characterized by chronic inflammatory disorder of the central nervous system in which circulating leukocytes enter the brain and spinal cord leading to inflammation, myelin damage and subsequent paralysis. To prove this hypothesis and explore a promising application for MS treatment, the effects of C16, an ανß3 integrin-binding peptide, were tested in vitro and in vivo by transendothelial assay, electron microscopy observation, multiple histological and immunohistochemical staining. The results showed C16 inhibited transendothelial migration of the C8166-CD4 lymphoblast cells, and alleviated extensive spinal cord and brain infiltration of leukocytes and macrophages in the EAE model. Furthermore, a significant amelioration of astrogliosis and a dramatic decrease in demyelination and axonal loss were observed in C16 treated animals. The attenuating inflammatory progression may improve the regional environment and trigger further neuroprotective effects on myelin and axons, all this suggests that C16 peptide may be a promising therapeutic agent for multiple sclerosis.

Contribution of Rag1 to spatial memory ability in rats.

Rag1 plays a critical role in the development and maturation of lymphocytes, and is related to immune memory functions. Deletion of Rag1 results in a lack of mature functional B and T lymphocytes. Rag1 transcription is most apparent in regions of the postnatal brain with high neuronal cell density--the cerebellum and the hippocampal formation, both of which are relevant to learning and memory function. In this research, three pairs of shRNA targeting Rag1 and a pair of scrambled sequences were constructed, packaged within a lentiviral vector system, and transferred into the cultured rat hippocampal neuron cells in vitro. Meanwhile, the lentivirus was injected stereotaxically into CA3 of the rat hippocampus, where the positive immunofluorescence for GFP expression was located. Rag1 mRNA expression was detected by RT-PCR 7, 14 and 28 days after stereotaxic injection. Assessment in the Morris water maze test 28 days post stereotaxic injection showed a loss of spatial learning and memory in the experimental rats. In long-term potentiation research, the experimental group did not display remarkable disparity in comparison with the control group. These findings indicate that the knockdown of Rag1 expression in the hippocampus may impair spatial learning and memory ability in rats.

Progress in defining heterogeneity and modeling periglomerular cells in the olfactory bulb.

In recent years the evolution of olfactory bulb periglomerular cells, as well as the function of periglomerular cells in olfactory encoding, has attracted increasing attention. Studies of neural information encoding based on the analysis of simulation and modeling have given rise to electrophysiological models of periglomerular cells, which have an important role in the understanding of the biology of these cells. In this review we provide a brief introduction to the anatomy of the olfactory system and the cell types in the olfactory bulb. We elaborate on the latest progress in the study of the heterogeneity of periglomerular cells based on different classification criteria, such as molecular markers, structure, ion channels and action potentials. Then, we discuss the several existing electrophysiological models of periglomerular cells, and we highlight the problems and defects of these models. Finally, considering our present work, we propose a future direction for electrophysiological investigations of periglomerular cells and for the modeling of periglomerular cells and olfactory information encoding.

The alteration of 5-HT(2A) and 5-HT(2C) receptors is involved in neuronal apoptosis of goldfish cerebellum following traumatic experience.

5-HT receptor changes remain controversial in posttraumatic stress disorder (PTSD) models. This study looks at the relationship between traumatic injuries and the alterations in 5-HT(2A) and 5-HT(2C) receptors in the goldfish brain. The effect of treatment with doxepin and fluoxetine, known to be selective serotonin reuptake inhibitor (SSRI) antidepressants, on 5-HT receptor expression in goldfish with fin ablation was also investigated. We demonstrated that fin ablation induced anxiety-like behavioural alterations and significant up-regulation of c-fos expression in goldfish cerebellum. The behavioural alterations correlated well with an increased expression of 5-HT(2A) receptors in the cerebellum of the fish with traumatic injury. An increase in the number of apoptotic cells and a higher caspase-8 protein level was present in the brains of goldfish with fin ablation compared to the control. Our findings suggest that neuronal apoptosis occured in the cerebellum as a result of fin ablation and may be related to the alterations of 5-HT(2A) and 5-HT(2C) levels and that the beneficial clinical effects of doxepin/fluoxetine treatment are due to the down-regulation of 5-HT(2A) and up-regulation of 5-HT(2C) receptors in the brain.

[GAD67-GFP expression and co-localization with bNOS in main olfactory bulb of GAD67-GFP knock-in mouse].

OBJECTIVE: To investigate the distribution of GAD67 and the co-localization with bNOS in the main olfactory bulb of GAD67-GFP knock-in mouse. METHODS: Polymerase chain reaction was applied to identify the genotype of GAD67-GFP knock-in mouse, the animals were sacrificed and frozen sections of olfactory bulb were prepared. The Nissl-staining was performed to show an framework of the neuron in the olfactory bulb. The distribution of GAD67 and co-localization with bNOS were detected by immunofluorescence technique. RESULTS: The proportion of GAD67-positive cells among DAPI-positive cells were (42.98 ± 0.92)% in glomerular layer, (23.64 ± 0.84)% in mitral cell layer and (77.75 ± 0.84)% in granule cell layer; the bNOS-positive cells mainly existed in glomerular layer and mitral cell layer, very few in granule cell layer. No co-localization of GAD67 and bNOS in granule cell layer and mitral cell layer was found, but there was dispersed distribution in glomerular layer. CONCLUSION: GAD67-positive neurons mainly appear in glomerular layer and granule cell layer, and the bNOS is mostly expressed in glomerular layer and mitral cell layer; while the co-localization of GAD67 and bNOS only occurs in glomerular layer of olfactory bulb.