Cytoplasmic Polyadenylation Element-Binding Protein 1 Post-transcriptionally Regulates Fragile X Mental Retardation 1 Expression Through 3' Untranslated Region in Central Nervous System Neurons.

Fragile X syndrome (FXS) is an inherited intellectual disability caused by a deficiency in Fragile X mental retardation 1 (Fmr1) gene expression. Recent studies have proposed the importance of cytoplasmic polyadenylation element-binding protein 1 (CPEB1) in FXS pathology; however, the molecular interaction between Fmr1 mRNA and CPEB1 has not been fully investigated. Here, we revealed that CPEB1 co-localized and interacted with Fmr1 mRNA in hippocampal and cerebellar neurons and culture cells. Furthermore, CPEB1 knockdown upregulated Fmr1 mRNA and protein levels and caused aberrant localization of Fragile X mental retardation protein in neurons. In an FXS cell model, CPEB1 knockdown upregulated the mRNA levels of several mitochondria-related genes and rescued the intracellular heat shock protein family A member 9 distribution. These findings suggest that CPEB1 post-transcriptionally regulated Fmr1 expression through the 3' untranslated region, and that CPEB1 knockdown might affect mitochondrial function.

Cpeb1 expression is post-transcriptionally regulated by AUF1, CPEB1, and microRNAs.

Cytoplasmic polyadenylation element binding protein 1 (CPEB1) regulates the translation of numerous mRNAs. We previously showed that AU-rich binding factor 1 (AUF1) regulates Cpeb1 expression through the 3' untranslated region (3'UTR). To investigate the molecular basis of the regulatory potential of the Cpeb1 3'UTR, here we performed reporter analyses that examined expression levels of Gfp reporter mRNA containing the Cpeb1 3'UTR. Our findings indicate that CPEB1 represses the translation of Cpeb1 mRNA and that miR-145a-5p and let-7b-5p are involved in the reduction in Cpeb1 expression in the absence of AUF1. These results suggest that Cpeb1 expression is post-transcriptionally regulated by AUF1, CPEB1, and microRNAs.

AUF1, an mRNA decay factor, has a discordant role in Cpeb1 expression.

Cytoplasmic polyadenylation element binding protein 1 (CPEB1) regulates polyadenylation and subsequent translation of CPE-containing mRNAs involved in various physiological and pathological phenomena. Although the significance of CPEB1-mediated translational regulation has recently been reported, the detailed regulatory mechanism of Cpeb1 expression remains unclear. To elucidate the post-transcriptional regulatory mechanisms of Cpeb1 expression, we constructed reporter plasmids containing various deletions or mutations in the Cpeb1 mRNA 3' untranslated region (3'UTR). We investigated their expression levels in Neuro2a neuroblastoma cells. We found that Cpeb1 expression is regulated through an AU-rich element in its 3'UTR. Furthermore, the mRNA decay factor AU-rich binding factor 1 (AUF1) regulates Cpeb1 expression, and knockdown of AUF1 upregulates Cpeb1 mRNA expression but results in a decrease in CPEB1 protein levels. These findings indicate that AUF1 has a discordant role in the expression of Cpeb1.

GATA4/6 regulate DHH transcription in rat adrenocortical autografts.

Adrenal cortex autotransplantation with ACTH stimulation may be an alternative therapy for patients with bilateral adrenalectomy to avoid adrenal crisis, but its underlying mechanism has not been elucidated. Previously, we detected Dhh upregulation in rat adrenocortical autografts after transplantation. Here, we investigated potential regulators such as Gata4, Gata6, Sry and Sox9 which affect Dhh transcription in adrenocortical autografts with or without ACTH stimulation. In ACTH-stimulated autografts, Gata4 and Gata6 were downregulated compared to control autografts. This response was linked to rDhh repression. A reporter assay using the upstream region of rDhh and a GATA binding motif revealed that rDhh promoters were significantly upregulated by co-transfection with Gata4 or Gata6 or both. Sry and Sox9 expression in autografts with or without ACTH stimulation were verified by PCR and RNAscope analyses. The ovarian differentiation factors Foxl2 and Rspo1 were also upregulated in the autografts. Gata4 and Gata6 were found to be significant factors in the regulation of rDhh expression and could be associated with adrenocortical autograft maintenance. Gonadal primordia with bipotential testicular and ovarian functions may also be present in these autografts.

Change in phospholipid species of retinal layer in traumatic optic neuropathy model.

Injured optic nerves induce death in almost all retinal ganglion cells (RGC) and cause a loss of axons. To date, we have studied injured RGC axon regeneration by using a traumatic optic nerve injury (TONI) rodent model, and we revealed that axonal regeneration is induced by the graft of an autologous peripheral nerve. The efficient approach to the regeneration of axons thus needs an environmental adjustment of RGC. However, the RGC environment induced by TONI remains unknown. Here, we analyzed female and male C57BL/6 mouse retinal tissue alterations in detail after TONI and focused on the major phospholipid species that are enriched in the whole retina. Reactive astrocyte accumulation, glia scar formation, and demyelination were observed in the injured optic nerve area, while RGC cell death, astrocyte accumulation, and Glial fibrillary acidic protein (GFAP) positive Müller cell increases were detected in the retinal layer. Furthermore, phosphatidylinositol (PI) 18:0/20:4 was localized to three nuclear layer structures: the ganglion cell layer (GCL), the inner nuclear layer (INL), and the outer nuclear layer (ONL) in control retina; however, the localization of 18:0/20:4 PI in TONI was disturbed. Meanwhile, phosphatidylserine (PS) 18:0/22:6 showed that the expression was specifically in the inner plexiform layer (IPL) with similar signal intensity in both cases. Other PS species and phosphatidylethanolamine (PE) were differentially localized in the retinal layer; however, the expressions of PE including docosahexaenoic acid (DHA) were affected by TONI. These results suggest that not only GCL but also other retinal layers were influenced by TONI.

Regulatory non-coding RNAs in nervous system development and disease.

Recent evidence demonstrates that long non-coding RNAs (lncRNAs) regulate the expression of multiple genes in an epigenetic, transcriptional, or post-transcriptional manner. They are involved in various cellular phenomena, such as the recruitment of transcription factors, epigenetic chaperoning, control of alternative splicing, mRNA stability and translational activity, as well as acting as decoys against microRNAs. In this review, we summarize the pivotal roles of lncRNAs in regulation of the gene expression involved in neural cell differentiation, synaptogenesis and synaptic plasticity in the central nervous system (CNS). We also describe the aberrant expression of multiple lncRNAs involved in the pathogenesis of neurological diseases. The abnormal expression of lncRNAs leads to altered expression levels of target genes, which contributes to neurodegenerative diseases, such as in Alzheimer's disease and Parkinson's disease, and to the formation of tumors, such as glioma. Accordingly, we discuss recent findings for the modes of action of lncRNAs in normal CNS development and for aberrant lncRNA actions in the pathogenesis of neuronal diseases.

Involvement of PLAGL1/ZAC1 in hypocretin/orexin transcription.

The hypocretin/orexin neuropeptide system coordinates the regulation of various physiological processes. Our previous study reported that a reduction in the expression of pleomorphic adenoma gene­like 1 (Plagl1), which encodes a C2H2 zinc­finger transcription factor, occurs in hypocretin neuron­ablated transgenic mice, suggesting that PLAGL1 is co­expressed in hypocretin neurons and regulates hypocretin transcription. The present study examined whether canonical prepro­hypocretin transcription is functionally modulated by PLAGL1. Double immunostaining indicated that the majority of hypocretin neurons were positive for PLAGL1 immunoreactivity in the nucleus. Notably, PLAGL1 immunoreactivity in hypocretin neurons was altered in response to several conditions affecting hypocretin function. An uneven localization of PLAGL1 was detected in the nuclei of hypocretin neurons following sleep deprivation. Chromatin immunoprecipitation revealed that endogenous PLAGL1 may bind to a putative PLAGL1­binding site in the proximal region of the hypocretin gene, in the murine hypothalamus. In addition, electroporation of the PLAGL1 expression vector into the fetal hypothalamus promoted hypothalamic hypocretin transcription. These results suggested that PLAGL1 may regulate hypothalamic hypocretin transcription.

Morphological characteristics of p75 neurotrophin receptor-positive cells define a new type of glial cell in the rat dorsal root ganglia.

In the dorsal root ganglia (DRG), two types of glial cells (Schwann cells and satellite glial cells) have been identified based on cell morphology and expression of specific markers. In the present study, we observed unknown glial cells that were positive for p75 neurotrophin receptor (p75NTR), and therefore were immunohistochemically and ultrastructurally characterized for the first time. These cells exhibited stronger immunoreactivity against an anti-p75NTR antibody than the DRG neurons (hereafter referred to as p75NTR++ cells). Moreover, these cells covered the glial cells surrounding proximal process of the large-diameter DRG neurons. The proximal process is called "dendro-axon." The p75NTR++ cells were predominantly distributed where the first myelinating Schwann cells appear. The p75NTR++ cells were also positive for the pan-glial cell markers S100, nestin, and Sox10, but negative for fibroblast and macrophage markers. Moreover, they were negative for a satellite glial cell marker, inwardly rectifying potassium channel Kir4.1, as well as a nonmyelinating Schwann cell marker, glial fibrillary acidic protein. In addition, their morphological features were distinct from those of the myelinating Schwann cells. To investigate the three-dimensional ultrastructure of the p75NTR++ cells, we used array tomography combined with correlative light and electron microscopic observation. Three-dimensional ultrastructural observation revealed that the p75NTR++ cells loosely covered glial cells around the dendro-axons with highly ramified processes. Glial cells with these morphological features have not been reported before, indicating that the p75NTR++ glial cells are a new glial cell type in the DRG. Our results will give new insights into cell-cell relationships.

Methods for array tomography with correlative light and electron microscopy.

The three-dimensional ultra-structure is the comprehensive structure that cannot be observed from a two-dimensional electron micrograph. Array tomography is one method for three-dimensional electron microscopy. In this method, to obtain consecutive cross sections of tissue, connected consecutive sections of a resin block are mounted on a flat substrate, and these are observed with scanning electron microscopy. Although array tomography requires some bothersome manual procedures to prepare specimens, a recent study has introduced some techniques to ease specimen preparation. In addition, array tomography has some advantages compared with other three-dimensional electron microscopy techniques. For example, sections on the substrate are stored semi-eternally, so they can be observed at different magnifications. Furthermore, various staining methods, including post-embedding immunocytochemistry, can be adopted. In the present review, the preparation of specimens for array tomography, including ribbon collection and the staining method, and the adaptability for correlative light and electron microscopy are discussed.

Differential expression of nuclear lamin subtypes in the neural cells of the adult rat cerebral cortex.

Lamins are type V intermediate filament proteins that are located beneath the inner nuclear membrane. In mammalian somatic cells, LMNB1 and LMNB2 encode somatic lamins B1 and B2, respectively, and the LMNA gene is alternatively spliced to generate somatic lamins A and C. Mutations in lamin genes have been linked to many human hereditary diseases, including neurodegenerative disorders. Knowledge about lamins in the nervous system has been accumulated recently, but a precise analysis of lamin subtypes in glial cells has not yet been reported. In this study we investigated the composition of lamin subtypes in neurons, astrocytes, oligodendrocyte-lineage cells, and microglia in the adult rat cerebral cortex using an immunohistochemical staining method. Lamin A was not observed in neurons and glial cells. Lamin C was observed in astrocytes, mature oligodendrocytes and neurons, but not observed in oligodendrocyte progenitor cells. Microglia also did not stain positive for lamin C which differed from macrophages, with lamin C positive. Lamin B1 and B2 were observed in all glial cells and neurons. Lamin B1 was intensely positive in oligodendrocyte progenitor cells compared with other glial cells and neurons. Lamin B2 was weakly positive in all glial cells compared to neurons. Our current study might provide useful information to reveal how the onset mechanisms of human neurodegenerative diseases are associated with mutations in genes for nuclear lamin proteins.

Involvement of DHH and GLI1 in adrenocortical autograft regeneration in rats.

Bilateral adrenalectomy forces the patient to undergo glucocorticoid replacement therapy and bear a lifetime risk of adrenal crisis. Adrenal autotransplantation is considered useful to avoid adrenal crisis and glucocorticoid replacement therapy. However, the basic process of regeneration in adrenal autografts is poorly understood. Here, we investigated the essential regeneration factors in rat adrenocortical autografts, with a focus on the factors involved in adrenal development and steroidogenesis, such as Hh signalling. A remarkable renewal in cell proliferation and increase in Cyp11b1, which encodes 11-beta-hydroxylase, occurred in adrenocortical autografts from 2-3 weeks after autotransplantation. Serum corticosterone and adrenocorticotropic hormone levels were almost recovered to sham level at 4 weeks after autotransplantation. The adrenocortical autografts showed increased Dhh expression at 3 weeks after autotransplantation, but not Shh, which is the only Hh family member to have been reported to be expressed in the adrenal gland. Increased Gli1 expression was also found in the regenerated capsule at 3 weeks after autotransplantation. Dhh and Gli1 might function in concert to regenerate adrenocortical autografts. This is the first report to clearly show Dhh expression and its elevation in the adrenal gland.

Identification of NeuN immunopositive cells in the adult mouse subventricular zone.

In the adult rodent subventricular zone (SVZ), there are neural stem cells (NSCs) and the specialized neurogenic niche is critical to maintain their stemness. To date, many cellular and noncellular factors that compose the neurogenic niche and markers to identify subpopulations of Type A cells have been confirmed. In particular, neurotransmitters regulate adult neurogenesis and mature neurons in the SVZ have been only partially analyzed. Moreover, Type A cells, descendants of NSCs, are highly heterogeneous and more molecular markers are still needed to identify them. In the present study, we systematically classified NeuN, commonly used as a marker of mature and immature post-mitotic neurons, immunopositive (+) cells within the adult mouse SVZ. These SVZ-NeuN+ cells (SVZ-Ns) were mainly classified into two types. One was mature SVZ-Ns (M-SVZ-Ns). Neurochemical properties of M-SVZ-Ns were similar to those of striatal neurons, but their birth date and morphology were different. M-SVZ-Ns were generated during embryonic and early postnatal stages with bipolar peaks and extended their processes along the wall of the lateral ventricle. The second type was small SVZ-Ns (S-SVZ-Ns) with features of Type A cells. They expressed not only markers of Type A cells, but also proliferated and migrated from the SVZ to the olfactory bulb. Furthermore, S-SVZ-Ns could be classified into two types by their spatial locations and glutamic acid decarboxylase 67 expression. Our data indicate that M-SVZ-Ns are a new component of the neurogenic niche and S-SVZ-Ns are newly identified subpopulations of Type A cells.

Origin of Oligodendrocytes in the Vertebrate Optic Nerve: A Review.

One of the unsolved problems in the research field of oligodendrocyte (OL) development has been the site(s) of origin of optic nerve OLs and its precursor cells (OPCs). It is generally accepted that OLs in the optic nerve are derived from the brain, and thus optic nerve OLs are immigrant cells. We previously demonstrated the brain origin of optic nerve OPCs in chick embryos. However, the site of optic nerve OPC origin has not been examined experimentally in developing rodents for the past two decades. We have recently reported that optic nerve OPCs in mice arise in the preoptic area by E12.5 and gradually migrate caudally and enter the optic nerve. These OPCs give rise to myelinating OLs in the optic nerve in the postnatal or adult stages. Surprisingly, there are species differences with respect to the origin of optic nerve OPCs between chicks and mice. Here, we summarize the site of OPC origin in the optic nerve based on our own previous and recent results, and discuss possible mechanisms underlying these species differences.

A Device for Ribbon Collection for Array Tomography with Scanning Electron Microscopy.

"Array tomography" is a method used to observe the fine structure of cells and tissues in a three-dimensional view. In this method, serial ultrathin sections in the ribbon state (ribbons) are mounted on a solid substrate and observed by scanning electron microscopy (SEM). The method may also be used in conjunction with post-embedding immunocytochemistry. However, it is difficult to mount many serial ribbons on a substrate manually. We developed an inexpensive laboratory-made device that mounts ribbons by pulling a nylon fishing line and lifting the substrate up from the water in a knife boat. Using this device, we succeeded in mounting several ribbons consisting a mean of 205.6 (SD: 37.7) serial ultrathin sections on 1.25 (SD: 0.06) × 1.25 (SD: 0.06)-cm silicon substrates. Furthermore, it was confirmed that our method is suitable for ribbons derived from water-soluble resin blocks. We were also able to stain the specimens by post-embedding immunocytochemistry. Thus, our method is useful in mounting numerus sections on a substrate for array tomography with SEM.

Establishment of a tumor sphere cell line from a metastatic brain neuroendocrine tumor.

Neuroendocrine tumors are rare, and little is known about the existence of cancer stem cells in this disease. Identification of the tumorigenic population will contribute to the development of effective therapies targeting neuroendocrine tumors. Surgically resected brain metastases from a primary neuroendocrine tumor of unknown origin were dissociated and cultured in serum-free neurosphere medium. Stem cell properties, including self-renewal, differentiation potential, and stem cell marker expression, were examined. Tumor formation was evaluated using intracranial xenograft models. The effect of temozolomide was measured in vitro by cell viability assays. We established the neuroendocrine tumor sphere cell line ANI-27S, which displayed stable exponential growth, virtually unlimited expansion in vitro, and expression of stem-cell markers such as CD133, nestin, Sox2, and aldehyde dehydrogenase. FBS-induced differentiation decreased Sox2 and nestin expression. On the basis of real-time PCR, ANI-27S cells expressed the neuroendocrine markers synaptophysin and chromogranin A. Intracranial xenotransplanted brain tumors recapitulated the original patient tumor and temozolomide exhibited cytotoxic effects on tumor sphere cells. For the first time, we demonstrated the presence of a sphere-forming, stem cell-like population in brain metastases from a primary neuroendocrine tumor. We also demonstrated the potential therapeutic effects of temozolomide for this disease.

Anti-Tribbles Pseudokinase 2 (TRIB2)-Immunization Modulates Hypocretin/Orexin Neuronal Functions.

Study Objectives: Recent findings showed that 16%-26% of narcolepsy patients were positive for anti-tribbles pseudokinase 2 (TRIB2) antibody, and the intracerebroventricular administration of immunoglobulin-G purified from anti-TRIB2 positive narcolepsy patients caused hypocretin/orexin neuron loss. We investigated the pathophysiological role of TRIB2 antibody using TRIB2-immunized rats and hypocretin/ataxin-3 transgenic (ataxin-3) mice. Methods: Plasma, cerebrospinal fluid (CSF), and hypothalamic tissues from TRIB2-immunized rats were collected. Anti-TRIB2 titers, hypocretin contents, mRNA expressions, the cell count of hypocretin neurons, and immunoreactivity of anti-TRIB2 antibodies on hypocretin neurons were investigated. The plasma from ataxin-3 mice was also used to determine the anti-TRIB2 antibody titer changes following the loss of hypocretin neurons. Results: TRIB2 antibody titers increased in the plasma and CSF of TRIB2-immunized rats. The hypothalamic tissue immunostained with the sera from TRIB2-immunized rats revealed positive signals in the cytoplasm of hypcretin neurons. While no changes were found regarding hypothalamic hypocretin contents or cell counts, but there were significant decreases of the hypocretin mRNA level and release into the CSF. The plasma from over 26-week-old ataxin-3 mice, at the advanced stage of hypocretin cell destruction, showed positive reactions against TRIB2 antigen, and positive plasma also reacted with murine hypothalamic hypocretin neurons. Conclusions: Our results suggest that the general activation of the immune system modulates the functions of hypocretin neurons. The absence of a change in hypocretin cell populations suggested that factors other than anti-TRIB2 antibody play a part in the loss of hypocretin neurons in narcolepsy. The increased anti-TRIB2 antibody after the destruction of hypocretin neurons suggest that anti-TRIB2 antibody in narcolepsy patients is the consequence rather than the inciting cause of hypocretin cell destruction.

Morphological and immunohistochemical comparison of intrapancreatic nerves between chronic pancreatitis and type 1 autoimmune pancreatitis.

OBJECTIVES: The abdominal pain associated with chronic pancreatitis (CP) may be related to the increased number and size of intrapancreatic nerves. On the other hand, patients with type 1 autoimmune pancreatitis (AIP) rarely suffer from the pain syndrome, and there are no previous studies concerning the histopathological findings of intrapancreatic nerves in patients with type 1 AIP. The current study is aimed at investigating the differences in the histopathological and immunohistochemical findings of intrapancreatic nerves in patients with CP and type 1 AIP. METHODS: Neuroanatomical differences between CP and type 1 AIP were assessed by immunostaining with a pan-neuronal marker, protein gene product 9.5 (PGP9.5). The number (neural density) and area (neural hypertrophy) of PGP9.5-immunopositive nerves were quantitatively analyzed. Furthermore, the expression of nerve growth factor (NGF), and a high affinity receptor for NGF, tyrosine kinase receptor A (TrkA), was assessed by immunohistochemistry. RESULTS: Both neural density and hypertrophy were significantly greater in pancreatic tissue samples from patients with CP than those with normal pancreas or type 1 AIP. NGF expression was stronger in type 1 AIP than in CP, whereas TrkA expression in type 1 AIP was poorer than in CP. CONCLUSIONS: Although CP and type 1 AIP are both characterized by the presence of sustained pancreatic inflammation, they are different in terms of the density and hypertrophy of intrapancreatic nerve fibers. It is possible that this may be related to the difference in the activity of the NGF/TrkA-pathway between the two types of pancreatitis.

Hypothalamo­hypophysial system in rats with autotransplantation of the adrenal cortex.

Patients with bilateral pheochromocytoma often require an adrenalectomy. Autotransplantation of the adrenal cortex is an alternative therapy that could potentially be performed instead of receiving glucocorticoid replacement following adrenalectomy. Adrenal cortex autotransplantation aims to avoid the side effects of long­term steroid treatment and adrenal insufficiency. Although the function of the hypothalamo­hypophysial system is critical for patients who have undergone adrenal cortex autotransplantation, the details of that system, with the exception of adrenocorticotropic hormone in the subjects with adrenal autotransplantation, have been overlooked for a long time. To clarify the precise effect of adrenal autotransplantation on the pituitary gland and hypothalamus, the current study examined the gene expression of hormones produced from the hypothalamus and pituitary gland. Bilateral adrenalectomy and adrenal autotransplantation were performed in 8 to 9­week­old male rats. The hypothalamus and pituitary tissues were collected at 4 weeks after surgery. Transcriptional regulation of hypothalamic and pituitary hormones was subsequently examined by reverse transcription­quantitative polymerase chain reaction. Proopiomelanocortin, glycoprotein hormone α polypeptide, and thyroid stimulating hormone ß were significantly elevated in the pituitary gland of autotransplanted rats when compared with sham­operated rats. In addition, there were significant differences in the levels of corticotropin releasing hormone receptor 1 (Crhr1), Crhr2, nuclear receptor subfamily 3 group C member 1 and thyrotropin releasing hormone receptor between the sham­operated rats and autotransplanted rats in the pituitary gland. In the hypothalamus, corticotropin releasing hormone and urocortin 2 mRNA was significantly upregulated in autotransplanted rats compared with sham­operated rats. The authors identified significant alterations in the function of not only the hypothalamus­pituitary­adrenal axis, but also the adenohypophysis thyrotropes in autotransplanted rats. In the future, it will be important to examine other tissues affected by glucocorticoids following adrenal cortex autotransplantation.

Sulfatide species with various fatty acid chains in oligodendrocytes at different developmental stages determined by imaging mass spectrometry.

HSO3-3-galactosylceramide (Sulfatide) species comprise the major glycosphingolipid components of oligodendrocytes and myelin and play functional roles in the regulation of oligodendrocyte maturation and myelin formation. Although various sulfatide species contain different fatty acids, it is unclear how these sulfatide species affect oligodendrogenesis and myelination. The O4 monoclonal antibody reaction with sulfatide has been widely used as a useful marker for oligodendrocytes and myelin. However, sulfatide synthesis during the pro-oligodendroblast stage, where differentiation into the oligodendrocyte lineage has already occurred, has not been examined. Notably, this stage comprises O4-positive cells. In this study, we identified a sulfatide species from the pro-oligodendroblast-to-myelination stage by imaging mass spectrometry. The results demonstrated that short-chain sulfatides with 16 carbon non-hydroxylated fatty acids (C16) and 18 carbon non-hydroxylated fatty acids (C18) or 18 carbon hydroxylated fatty acids (C18-OH) existed in restricted regions of the early embryonic spinal cord, where pro-oligodendroblasts initially appear, and co-localized with Olig2-positive pro-oligodendroblasts. C18 and C18-OH sulfatides also existed in isolated pro-oligodendroblasts. C22-OH sulfatide became predominant later in oligodendrocyte development and the longer C24 sulfatide was predominant in the adult brain. Additionally, the presence of each sulfatide species in a different area of the adult brain was demonstrated by imaging mass spectrometry at an increased lateral resolution. These findings indicated that O4 recognized sulfatides with short-chain fatty acids in pro-oligodendroblasts. Moreover, the fatty acid chain of the sulfatide became longer as the oligodendrocyte matured. Therefore, individual sulfatide species may have unique roles in oligodendrocyte maturation and myelination. Read the Editorial Highlight for this article on page 356.

Hypocretin/orexin loss changes the hypothalamic immune response.

Hypocretin, also known as orexin, maintains the vigilance state and regulates various physiological processes, such as arousal, sleep, food intake, energy expenditure, and reward. Previously, we found that when wild-type mice and hypocretin/ataxin-3 littermates (which are depleted of hypothalamic hypocretin-expressing neurons postnatally) were administered lipopolysaccharide (LPS), the two genotypes exhibited significant differences in their sleep/wake cycle, including differences in the degree of increase in sleep periods and in recovery from sickness behaviour. In the present study, we examined changes in the hypothalamic vigilance system and in the hypothalamic expression of inflammatory factors in response to LPS in hypocretin/ataxin-3 mice. Peripheral immune challenge with LPS affected the hypothalamic immune response and vigilance states. This response was altered by the loss of hypocretin. Hypocretin expression was inhibited after LPS injection in both hypocretin/ataxin-3 mice and their wild-type littermates, but expression was completely abolished only in hypocretin/ataxin-3 mice. Increases in the number of histidine decarboxylase (HDC)-positive cells and in Hdc mRNA expression were found in hypocretin/ataxin-3 mice, and this increase was suppressed by LPS. Hypocretin loss did not impact the change in expression of hypothalamic inflammatory factors in response to LPS, except for interferon gamma and colony stimulating factor 3. The number of c-Fos-positive/HDC-positive cells in hypocretin/ataxin-3 mice administered LPS injections was elevated, even during the rest period, in all areas, suggesting that there is an increase in the activity of histaminergic neurons in hypocretin/ataxin-3 mice following LPS injection. Taken together, our results suggest a novel role for hypocretin in the hypothalamic response to peripheral immune challenge. Our findings contribute to the understanding of the pathophysiology of narcolepsy.