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Analysis of gamma-Protocadherin conditional targeted mice
S98
Abstracts
noids suppress this GABA release. In the present study, we investigated involvement of the endocannabinoid metabolic pathway and the cannabinoid CB2 receptor in PC dendrogenesis. Using several endocannabinoid metabolic enzyme inhibitors, we show that 2-arachidonoylglycerol (2-AG), one of the endocannabinoids, controls PC dendrogenesis; an inhibitor for the 2-AG synthesizing enzyme promotes PC dendritic development, whereas that for the 2-AG degrading enzyme prevents dendrite elongation of PCs. Next, using CB2-selective and non-selective ligands, we found a possibility that the CB2 receptor may contribute to development of PC dendrites. In addition, we also report results that receptors for glycine, another inhibitory neurotransmitter, may play a role in PC dendrogenesis.
tumor-suppressor in various tissues. In the present study, therefore, the functions of OBCAM were investigated in cultured glial cells such as astrocytes and C6 glioma. Immunocytochemistry and western blotting revealed using type 1 astrocytes and C6 glioma cells that OBCAM expression was high at growing culture stages but low at stational culture stages. High magnification view of confocal images further showed that OBCAM was localized at pericellular filopodia. The inhibition of OBCAM function with the specific monoclonal antibody promoted the proliferation of astrocyte growth. These results indicate that OBCAM expression is regulated with growing conditions and participates in controlling the growth of glial cells. doi:10.1016/j.neures.2009.09.422
doi:10.1016/j.neures.2009.09.419
P1-f01 Roles of the chondroitin sulfate chain on the core protein of neuroglycan C, a brain-specific transmembrane proteoglycan Sachiko Aono, Fumiko Matsui, Atsushi Suzuki, Yoshihito Tokita Inst. Develop. Res., Aichi Human Service Ctr., Kasugai, Japan Neuroglycan C (NGC), a transmembrane chondroitin sulfate proteoglycan that is predominantly expressed in the brain, is a typical part-time proteoglycan that changes its structure from a proteoglycan form to a non-proteoglycan form without chondroitin sulfate during the development of the cerebellum and retina. Modification of the NGC core protein with chondroitin sulfate is also differently regulated depending on cell types. In the present study, the mechanism of chondroitin sulfate glycosylation was investigated using co-transfectants of GRP78 (one of chaperon) and NGC cDNAs. Further, the roles of the chondroitin sulfate chain were investigated using mice that express NGC in non-proteoglycan form due to a mutation (S123A) at the chondroitin sulfate chain binding site. Uncited reference Aono and Oohira (2007).
P1-f04 Spatial and temporal distribution patterns of PSA-NCAM positive pericytes of the embryonic brain Momoko Miyakawa 1 , Tatsunori Seki 2 , Yasuo Uchiyama 1 1 Department of Cell Biol. and Neurosci., Juntendo University Sch. Med. Tokyo, Japan; 2 Div. of Dev. Neurosci., Tohoku University Grad. Sch. of Med., Sendai, Japan
Highly polysialylated NCAM (PSA-NCAM) that was expressed in pericyte-like cells was found around capillaries in the chick embryonic brains. The present study examined spatiotemporal correlation of PSA-NCAM expression and angiogenesis in chick embryos from embryonic day 4(E4) to E10, using immunohistochemistry. PSA-NCM expression was observed around capillaries in the caudal telencephalon, diencephalon and rostral mesencephalon during the earliest stage of the angiogeneses, from E4 to E7. The PSA-NCAM-positive capilaries appeared immature since basement membrane of the capillaries was not continuous and blood cells in the lumen were sparse. In the ventricular zone, cellular processes doubly immunostained for PSA-NCAM and Alpha-SMA were found at the tip of the capillaries. PSA-NCAM was expressed around capillaries on the brain surface from the caudal telencephalon to the rostral mesencephalon. Our present data showed that PSA-NCAM-positive pericytes appeared restricted spatiotemporally in early developmental brains. doi:10.1016/j.neures.2009.09.423
Reference Aono, S., Oohira, A., 2007. Neuroglycan C, a part-time proteoglycan, in the central nervous system. In: Maeda, N. (Ed.), Neural Proteoglycans. Research Signpost, Kerala, pp. 37–50. doi:10.1016/j.neures.2009.09.420
P1-f02 Activity-dependent changes in expression of chondroitin sulfate proteoglycan Shoko Morita 1 , Atsuhiko Oohira 2 , Seiji Miyata 1 1 2
Department of Applied Biology, Kyoto Institute of Technology, Kyoto, Japan; Aichi Medical School of Medicine, Japan
The hypothalamo-neurohypophysial system, synthesizing arginine vasopressin (AVP) and oxytocin (OXT), is well known to show structural plasticity during chronic physiological stimulation. The immunoreactivity of phosphacan was primarily observed to surround AVP-positive somata and that of neurocan was seen chiefly around OXT-positive somata in the supraoptic nucleus (SON), although the immunoreactivity of neuroglycan C was seen around both AVP- and OXT-positive somata. In the neurohypophysis (NH), the immunoreactivity of phosphacan and neurocan was observed at AVP-positive magnocellular terminals but not OXT-positive ones. Salt-loading, known as the chronic stimulation to cause the structural plasticity, decreased significantly phosphacan levels in the SON and those of phosphacan and neurocan immunoreactivity in the NH. Interestingly, salt-loading increased neuroglycan C levels in the SON. RT-PCR revealed that the chronic stimulation did not change mRNA levels of phosphacan in the SON and NH but increased those of neurocan in the SON. doi:10.1016/j.neures.2009.09.421
P1-f03 Expression of immunoglobulin superfamily cell adhesion molecule OBCAM in glial cells Chiaki Sugimoto 1 , Shohei Maekawa 2 , Seiji Miyata 1 1 2
Department of Applied Biology, Kyoto Institute of Technology, Kyoto, Japan; Kobe University, Japan
Opioid-binding cell adhesion molecule (OBCAM) is the member of the IgLON family, a subgroup of the immunoglobulin superfamily. OBCAM has been shown to be expressed in neurons of brains, but recent studies indicate that OBCAM is a possible
P1-f05 Analysis of gamma-Protocadherin conditional targeted mice Yasuto Itoga 1 , Kayoko Makino 2 , Shun Hamada 3 , Takahiro 1 1 Hirabayashi , Takeshi Yagi 1
Grad. Sch. of Frontier Biosci., Osaka University, Osaka, Japan; 2 TAKARA BIO INC., Japan; 3 Dev. Nutrition and health Sci, Fukuoka Women’s University, Fukuoka, Japan The gamma-Protocadherin (Pcdh␥) gene clsuster can generate 22 distinct isoforms by cis-splicing of the mRNA and Pcdh␥ proteins have been proposed to be involved in neural diversity. According to the previous reports, gene targeted mice lacking the Pcdh␥ gene cluster showed apoptotic neural loss in the spinal cord and died shortly after birth. To study the function of Pcdh␥ more detail, we generated conditional alleles in which Pcdh␥ can be disrupted from specific cell types by Cre-mediated recombination. We crossed the conditional Pcdh␥ mice with ␣ calmodulin kinase 2 Cre (CaMK2-Cre) transgenic mice to selectively inactivate Pcdh␥ in excitatory forebrain neurons. In the mutant, we detected apoptotic cells in their hippocampus, amygdala, and piriform cortex. Interestingly, many apoptotic cells were not excitatory neurons in which CaMK2 promoter can drive, but inhibitory neurons. This result indicates that Pcdh␥ regulate cell survival non cell-autonomously. doi:10.1016/j.neures.2009.09.424
P1-f06 Expression and function of protocadherin9 in the developing cortex Asuka Matsui 1 , Ryosuke Kaneko 2 , Takeshi Yagi 2 , Nobuhiko Yamamoto 1 1
Dept. Frontier Bio., Univ. of Osaka, Osaka, Japan; Univ. of Osaka, Osaka, Japan
2
Dept. Frontier Bio.,
Laminar specificity is one of the most striking aspects of the neocortical structure. To explore the molecular basis, we searched for layer-specific genes by constructing a subtraction cDNA library which is enriched for the genes expressed in layer 4. As a result, protocadherin9 (pcdh9) was found to be specifically expressed in layer 6 as well as layer 4. In the developing cortex, pcdh9 expression was already observed at E18, gradually increased until P7 and then decreased. In terms of area specificity, pcdh9 was strongly expressed in part of the sensory cortices. To reveal the function, we generated pcdh9 knockout mice. Although the mutant cortex was properly laminated, cortical thickness in the sensory area was slightly thinner in the mutant than wild-type mice. In particular, layer 6, in which pcdh9 is strongly expressed, was
Abstracts
noids suppress this GABA release. In the present study, we investigated involvement of the endocannabinoid metabolic pathway and the cannabinoid CB2 receptor in PC dendrogenesis. Using several endocannabinoid metabolic enzyme inhibitors, we show that 2-arachidonoylglycerol (2-AG), one of the endocannabinoids, controls PC dendrogenesis; an inhibitor for the 2-AG synthesizing enzyme promotes PC dendritic development, whereas that for the 2-AG degrading enzyme prevents dendrite elongation of PCs. Next, using CB2-selective and non-selective ligands, we found a possibility that the CB2 receptor may contribute to development of PC dendrites. In addition, we also report results that receptors for glycine, another inhibitory neurotransmitter, may play a role in PC dendrogenesis.
tumor-suppressor in various tissues. In the present study, therefore, the functions of OBCAM were investigated in cultured glial cells such as astrocytes and C6 glioma. Immunocytochemistry and western blotting revealed using type 1 astrocytes and C6 glioma cells that OBCAM expression was high at growing culture stages but low at stational culture stages. High magnification view of confocal images further showed that OBCAM was localized at pericellular filopodia. The inhibition of OBCAM function with the specific monoclonal antibody promoted the proliferation of astrocyte growth. These results indicate that OBCAM expression is regulated with growing conditions and participates in controlling the growth of glial cells. doi:10.1016/j.neures.2009.09.422
doi:10.1016/j.neures.2009.09.419
P1-f01 Roles of the chondroitin sulfate chain on the core protein of neuroglycan C, a brain-specific transmembrane proteoglycan Sachiko Aono, Fumiko Matsui, Atsushi Suzuki, Yoshihito Tokita Inst. Develop. Res., Aichi Human Service Ctr., Kasugai, Japan Neuroglycan C (NGC), a transmembrane chondroitin sulfate proteoglycan that is predominantly expressed in the brain, is a typical part-time proteoglycan that changes its structure from a proteoglycan form to a non-proteoglycan form without chondroitin sulfate during the development of the cerebellum and retina. Modification of the NGC core protein with chondroitin sulfate is also differently regulated depending on cell types. In the present study, the mechanism of chondroitin sulfate glycosylation was investigated using co-transfectants of GRP78 (one of chaperon) and NGC cDNAs. Further, the roles of the chondroitin sulfate chain were investigated using mice that express NGC in non-proteoglycan form due to a mutation (S123A) at the chondroitin sulfate chain binding site. Uncited reference Aono and Oohira (2007).
P1-f04 Spatial and temporal distribution patterns of PSA-NCAM positive pericytes of the embryonic brain Momoko Miyakawa 1 , Tatsunori Seki 2 , Yasuo Uchiyama 1 1 Department of Cell Biol. and Neurosci., Juntendo University Sch. Med. Tokyo, Japan; 2 Div. of Dev. Neurosci., Tohoku University Grad. Sch. of Med., Sendai, Japan
Highly polysialylated NCAM (PSA-NCAM) that was expressed in pericyte-like cells was found around capillaries in the chick embryonic brains. The present study examined spatiotemporal correlation of PSA-NCAM expression and angiogenesis in chick embryos from embryonic day 4(E4) to E10, using immunohistochemistry. PSA-NCM expression was observed around capillaries in the caudal telencephalon, diencephalon and rostral mesencephalon during the earliest stage of the angiogeneses, from E4 to E7. The PSA-NCAM-positive capilaries appeared immature since basement membrane of the capillaries was not continuous and blood cells in the lumen were sparse. In the ventricular zone, cellular processes doubly immunostained for PSA-NCAM and Alpha-SMA were found at the tip of the capillaries. PSA-NCAM was expressed around capillaries on the brain surface from the caudal telencephalon to the rostral mesencephalon. Our present data showed that PSA-NCAM-positive pericytes appeared restricted spatiotemporally in early developmental brains. doi:10.1016/j.neures.2009.09.423
Reference Aono, S., Oohira, A., 2007. Neuroglycan C, a part-time proteoglycan, in the central nervous system. In: Maeda, N. (Ed.), Neural Proteoglycans. Research Signpost, Kerala, pp. 37–50. doi:10.1016/j.neures.2009.09.420
P1-f02 Activity-dependent changes in expression of chondroitin sulfate proteoglycan Shoko Morita 1 , Atsuhiko Oohira 2 , Seiji Miyata 1 1 2
Department of Applied Biology, Kyoto Institute of Technology, Kyoto, Japan; Aichi Medical School of Medicine, Japan
The hypothalamo-neurohypophysial system, synthesizing arginine vasopressin (AVP) and oxytocin (OXT), is well known to show structural plasticity during chronic physiological stimulation. The immunoreactivity of phosphacan was primarily observed to surround AVP-positive somata and that of neurocan was seen chiefly around OXT-positive somata in the supraoptic nucleus (SON), although the immunoreactivity of neuroglycan C was seen around both AVP- and OXT-positive somata. In the neurohypophysis (NH), the immunoreactivity of phosphacan and neurocan was observed at AVP-positive magnocellular terminals but not OXT-positive ones. Salt-loading, known as the chronic stimulation to cause the structural plasticity, decreased significantly phosphacan levels in the SON and those of phosphacan and neurocan immunoreactivity in the NH. Interestingly, salt-loading increased neuroglycan C levels in the SON. RT-PCR revealed that the chronic stimulation did not change mRNA levels of phosphacan in the SON and NH but increased those of neurocan in the SON. doi:10.1016/j.neures.2009.09.421
P1-f03 Expression of immunoglobulin superfamily cell adhesion molecule OBCAM in glial cells Chiaki Sugimoto 1 , Shohei Maekawa 2 , Seiji Miyata 1 1 2
Department of Applied Biology, Kyoto Institute of Technology, Kyoto, Japan; Kobe University, Japan
Opioid-binding cell adhesion molecule (OBCAM) is the member of the IgLON family, a subgroup of the immunoglobulin superfamily. OBCAM has been shown to be expressed in neurons of brains, but recent studies indicate that OBCAM is a possible
P1-f05 Analysis of gamma-Protocadherin conditional targeted mice Yasuto Itoga 1 , Kayoko Makino 2 , Shun Hamada 3 , Takahiro 1 1 Hirabayashi , Takeshi Yagi 1
Grad. Sch. of Frontier Biosci., Osaka University, Osaka, Japan; 2 TAKARA BIO INC., Japan; 3 Dev. Nutrition and health Sci, Fukuoka Women’s University, Fukuoka, Japan The gamma-Protocadherin (Pcdh␥) gene clsuster can generate 22 distinct isoforms by cis-splicing of the mRNA and Pcdh␥ proteins have been proposed to be involved in neural diversity. According to the previous reports, gene targeted mice lacking the Pcdh␥ gene cluster showed apoptotic neural loss in the spinal cord and died shortly after birth. To study the function of Pcdh␥ more detail, we generated conditional alleles in which Pcdh␥ can be disrupted from specific cell types by Cre-mediated recombination. We crossed the conditional Pcdh␥ mice with ␣ calmodulin kinase 2 Cre (CaMK2-Cre) transgenic mice to selectively inactivate Pcdh␥ in excitatory forebrain neurons. In the mutant, we detected apoptotic cells in their hippocampus, amygdala, and piriform cortex. Interestingly, many apoptotic cells were not excitatory neurons in which CaMK2 promoter can drive, but inhibitory neurons. This result indicates that Pcdh␥ regulate cell survival non cell-autonomously. doi:10.1016/j.neures.2009.09.424
P1-f06 Expression and function of protocadherin9 in the developing cortex Asuka Matsui 1 , Ryosuke Kaneko 2 , Takeshi Yagi 2 , Nobuhiko Yamamoto 1 1
Dept. Frontier Bio., Univ. of Osaka, Osaka, Japan; Univ. of Osaka, Osaka, Japan
2
Dept. Frontier Bio.,
Laminar specificity is one of the most striking aspects of the neocortical structure. To explore the molecular basis, we searched for layer-specific genes by constructing a subtraction cDNA library which is enriched for the genes expressed in layer 4. As a result, protocadherin9 (pcdh9) was found to be specifically expressed in layer 6 as well as layer 4. In the developing cortex, pcdh9 expression was already observed at E18, gradually increased until P7 and then decreased. In terms of area specificity, pcdh9 was strongly expressed in part of the sensory cortices. To reveal the function, we generated pcdh9 knockout mice. Although the mutant cortex was properly laminated, cortical thickness in the sensory area was slightly thinner in the mutant than wild-type mice. In particular, layer 6, in which pcdh9 is strongly expressed, was