Etsuko Wada

Microglia–Müller Glia Cell Interactions Control Neurotrophic Factor Production during Light-Induced Retinal Degeneration

Activation of microglia commonly occurs in response to a wide variety of pathological stimuli including trauma, axotomy, ischemia, and degeneration in the CNS. In the retina, prolonged or high-intensity exposure to visible light leads to photoreceptor cell apoptosis. In such a light-reared retina, we found that activated microglia invade the degenerating photoreceptor layer and alter expression of neurotrophic factors such as nerve growth factor (NGF), ciliary neurotrophic factor (CNTF), and glial cell line-derived neurotrophic factor (GDNF). Because these neurotrophic factors modulate secondary trophic factor expression in Müller glial cells, microglia–Müller glia cell interaction may contribute to protection of photoreceptors or increase photoreceptor apoptosis. In the present study, we demonstrate the possibility that such functional glia–glia interactions constitute the key mechanism by which microglia-derived NGF, brain-derived neurotrophic factor (BDNF), and CNTF indirectly influence photoreceptor survival, although the receptors for these neurotrophic factors are absent from photoreceptors, by modulating basic fibroblast growth factor (bFGF) and GDNF production and release from Müller glia. These observations suggest that microglia regulate the microglia–Müller glia–photoreceptor network that serves as a trophic factor-controlling system during retinal degeneration.

Primary structure and expression of β2: A novel subunit of neuronal nicotinic acetylcholine receptors

Abstract A new subunit, β2, of the neuronal nicotinic receptor family has been identified. This subunit has the structural features of a non-agonist-binding subunit. We provide evidence that β2 can substitute for the muscle β1 subunit to form a functional nicotinic receptor in Xenopus oocytes. Expression studies performed in oocytes have demonstrated that three different neuronal nicotinic acetylcholine receptors can be formed by the pairwise injection of β2 mRNA and each of the neuronal α subunit mRNAs. The β2 gene is expressed in PC12 cells and in areas of the central nervous system where the α2, α3, and α4 genes are expressed. These results lead us to propose that the nervous system expresses diverse forms of neuronal nicotinic acetylcholine receptors by combining β2 subunits with different agonist-binding α subunits.

Mice lacking bombesin receptor subtype-3 develop metabolic defects and obesity

Mammalian bombesin-like peptides are widely distributed in the central nervous system as well as in the gastrointestinal tract, where they modulate smooth-muscle contraction, exocrine and endocrine processes, metabolism and behaviour. They bind to G-protein-coupled receptors on the cell surface to elicit their effects. Bombesin-like peptide receptors cloned so far include, gastrin-releasing peptide receptor (GRP-R), neuromedin B receptor (NMB-R), and bombesin receptor subtype-3 (BRS-3). However, despite the molecular characterization of BRS-3, determination of its function has been difficult as a result of its low affinity for bombesin and its lack of an identified natural ligand. We have generated BRS-3-deficient mice in an attempt to determine the in vivo function of the receptor. Mice lacking functional BRS-3 developed a mild obesity, associated with hypertension and impairment of glucose metabolism. They also exhibited reduced metabolic rate, increased feeding efficiency and subsequent hyperphagia. Our data suggest that BRS-3 is required for the regulation of endocrine processes and metabolism responsible for energy balance and adiposity. BRS-3-deficient mice provide a useful new model for the investigation of human obesity and associated diseases.

The distribution of mRNA encoded by a new member of the neuronal nicotinic acetylcholine receptor gene family (α5) in the rat central nervous system

The cellular localization of transcripts for a new putative agonist-binding subunit of the neuronal nicotinic acetylcholine receptor (nAChR), alpha 5, was examined using in situ hybridization in the rat central nervous system (CNS), alpha 5 subunit mRNA was localized to a small number of regions when compared with two of the other known agonist-binding subunits, alpha 3 and alpha 4, alpha 5 mRNA is expressed at relatively high levels in neurons of the subiculum (pyramidal layer), presubiculum and parasubiculum (layers IV and VI), which are components of the hippocampal formation, in the substantia nigra pars compacta and ventral tegmental area, in the interpeduncular nucleus, and in the dorsal motor nucleus of the vagus nerve. Moderate hybridization signals were detected in neurons of the isocortex (layer VIb), anterior olfactory nucleus, trigeminal ganglion, superior olivary complex, nucleus of the solitary tract, and area postrema. No hybridization above background levels was seen in the amygdala, septum, thalamus, hypothalamus, or cerebellum. These results suggest that the alpha 5 subunit differs from other known agonist-binding subunits in its distribution.

Basic Fibroblast Growth Factor (FGF) in the Central Nervous System: Identification of Specific Loci of Basic FGF Expression in the Rat Brain

The expression of basic FGF mRNA, while virtually absent in peripheral tissues, appears to be constitutively expressed in the central nervous system. As such, while it is difficult to detect any mRNA encoding basic FGF in the heart, lung, kidneys, ovaries, liver, or pituitary of rats, the levels are easily detected in brain. A regional analysis of the expression of basic FGF mRNA in brain reveals that it is widely distributed in the cortex (frontal, parietal, and occipital), the hippocampus, hypothalamus, and pons. Only a few loci of basic FGF synthesis are detected by in situ hybridization and include layers 2 and 6 of the medial (cingulate) cortex, the indusium griseum, fasciola cinereum, and field CA2 of the hippocampus. The identification of specific cell populations in the brain, and particularly in the hippocampus, that synthesize basic FGF supports the notion that this potent neurotrophic factor is involved in normal CNS function and that the presence (or absence) of its expression may be linked to the pathogenesis of the neurogenerative diseases characterizing these various loci. The significance of these findings with respect to the regulation of basic FGF expression in peripheral tissue and the central nervous system is discussed.

Clinico-pathological rescue of a model mouse of Huntington's disease by siRNA

Huntingtons disease (HD) is an autosomal dominant inheritable neurodegenerative disorder currently without effective treatment. It is caused by an expanded polyglutamine (poly Q) tract in the corresponding protein, huntingtin (htt), and therefore suppressing the huntingtin expression in brain neurons is expected to delay the onset and mitigate the severity of the disease. Here, we have used small interfering RNAs (siRNAs) directed against the huntingtin gene to repress the transgenic mutant huntingtin expression in an HD mouse model, R6/2. Results showed that intraventricular injection of siRNAs at an early postnatal period inhibited transgenic huntingtin expression in brain neurons and induced a decrease in the numbers and sizes of intranuclear inclusions in striatal neurons. Treatments using this siRNA significantly prolonged model mice longevity, improved motor function and slowed down the loss of body weight. This work suggests that siRNA-based therapy is promising as a future treatment for HD.

Maternal obesity impairs hippocampal BDNF production and spatial learning performance in young mouse offspring

Maternal obesity may affect the childs long-term development and health, increasing the risk of diabetes and metabolic syndrome. In addition to the metabolic and endocrine systems, recent reports have indicated that maternal obesity also modulates neural circuit formation in the offspring. However, this not yet been fully investigated. Here, we examined the effect of diet-induced maternal obesity on hippocampal development and function in the mouse offspring. Adult female mice were fed either a normal diet (ND, 4% fat) or a high-fat diet (HFD, 32% fat) before mating and throughout pregnancy and lactation. After weaning, all offspring were fed with a normal diet. We found that HFD offspring showed increased lipid peroxidation in the hippocampus during early postnatal development. HFD offspring had less brain-derived neurotrophic factor (BDNF) in the hippocampus than ND offspring. BDNF has been shown to play crucial roles in neuronal differentiation, plasticity and hippocampus-dependent cognitive functions such as spatial learning and memory. Using retroviral labeling, we demonstrated that dendritic arborization of new hippocampal neurons was impaired in the young HFD offspring. Finally, we evaluated cognitive function in these offspring using hippocampus-dependent behavioral tasks. The Barnes maze test demonstrated that HFD offspring showed impaired acquisition of spatial learning in the young but not adult period. This study, using a mouse model, indicates that diet-induced maternal obesity impairs hippocampal BDNF production and spatial cognitive function in young offspring, possibly due to their metabolic and oxidative changes.

Differential Gene Expression of CCKA and CCKB Receptors in the Rat Brain

Cholecystokinin (CCK) is one of the most abundant peptides in the central nervous system (CNS). Radioligand-binding studies have identified and localized both central (CCK(B)-R) and peripheral (CCK(A)-R) receptor subtypes in the CNS. However, these studies have been limited by the relative specificities of agonists and antagonists for receptor subtypes and their inability to distinguish cell bodies from dendritic projection. Recently, we isolated and cloned the cDNAs for both CCK(A) and CCK(B) receptors. In the present study, (35)S-labeled cRNA antisense probes were synthesized for both receptors and in situ hybridization studies were performed in the rat brain, allowing a direct and independent comparison of the distinct distribution of expression of CCK(A) and CCK(B) receptor mRNAs in the rat brain for the first time. mRNAs for both CCK(A) and CCK(B) receptors were expressed mainly in the cortex, olfactory regions, hippocampal formation, septum, and interpeduncular nucleus. Only CCK(A) receptor mRNAs were expressed in some of the hypothalamic nuclei (paraventricular nucleus, arcuate nucleus, and medial preoptic area). In most of amygdaloid nuclei only CCK(B) receptor mRNAs were expressed. Although the presence of receptor mRNAs does not necessarily imply the presence of functional receptor proteins in the same location, this study showed the regional distribution of mRNAs for CCK(A) and CCK(B) receptors and provides important information about the distribution of CCK receptor subtypes in the CNS which will allow better resolution of their functional roles in the brain.

Diet-induced obesity in female mice leads to peroxidized lipid accumulations and impairment of hippocampal neurogenesis during the early life of their offspring

Maternal obesity may affect the childs long‐term development and health. However, there is little information about the involvement of maternal obesity in the brain development of offspring. Here, we investigated the effects of maternal obesity on the hippocampal formation of offspring. Adult female mice were fed either a normal diet (ND, 4% fat) or a high‐fat diet (HFD, 32% fat) 6 wk before mating and throughout pregnancy and the majority of lactation. We found that infants from HFD‐fed dams (HFD offspring) showed obesity and hyperlipidemia during suckling. In HFD offspring, lipid peroxidation was promoted in serum and the hippocampal dentate gyrus, where neurogenesis takes place throughout postnatal life. Using a BrdU‐pulse labeling study, we showed that malondialdehyde, a product of peroxidized lipids, reduced the proliferation of hippocampal progenitor cells in vitro and that neurogenesis in HFD offspring during postnatal development was similarly lowered relative to the ND animals. These results indicated that maternal obesity impairs hippocampal progenitor cell division and neuronal production in young offspring possibly due to metabolic and oxidative changes.—Tozuka, Y.,Wada, E., Wada, K. Diet‐induced obesity in female mice leads to peroxidized lipid accumulations and impairment of hippocampal neurogenesis during the early life of their offspring. FASEB J. 23, 1920–1934 (2009)

Neurotensin increases mortality and mast cells reduce neurotensin levels in a mouse model of sepsis.

Sepsis is a complex, incompletely understood and often fatal disorder, typically accompanied by hypotension, that is considered to represent a dysregulated host response to infection. Neurotensin (NT) is a 13-amino-acid peptide that, among its multiple effects, induces hypotension. We find that intraperitoneal and plasma concentrations of NT are increased in mice after severe cecal ligation and puncture (CLP), a model of sepsis, and that mice treated with a pharmacological antagonist of NT, or NT-deficient mice, show reduced mortality during severe CLP. In mice, mast cells can degrade NT and reduce NT-induced hypotension and CLP-associated mortality, and optimal expression of these effects requires mast cell expression of neurotensin receptor 1 and neurolysin. These findings show that NT contributes to sepsis-related mortality in mice during severe CLP and that mast cells can lower NT concentrations, and suggest that mast cell–dependent reduction in NT levels contributes to the ability of mast cells to enhance survival after CLP.