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168 117 P ARF BRAG/IQSEC Brefeldin Aresistant ArfGEF 2 BRAG2/IQSEC1 ADP 6 Arf6 GEF BRAG2 AMPA BRAG2 BRAG2 BRAG2pan C BRAG2LC BRAG2 BRAG2LC PSD AMPA BRAG2pan PSD BRAG2 PSD BRAG2Arf6 AMPA P Arf EFAA GTP ADP 6 ARF6 ARF6 GEF EFA6A EFA6A EFA6A PSD95 EFA6A EFA6A PSD PSD EFA6A PSD PSD ARF6 P Roles of BMP signaling in synapse development Continual formation and elimination of synapses is required for refinement of the neuronal circuit. Synapse remodeling should be regulated by specific signaling mechanisms, but few molecules involved in synapse elimination have been identified, despite a large body of work on molecular cascades that initiate or enhance synapse formation. Here, we report that BMP4 negatively regulates hippocampal synapse formation in vitro. First, we confirmed that hippocampal neurons respond to BMP4 by measuring accumulation of activated Smad by immunocytochemistry. Next, we constructed BMP4EGFP and visualized its localization in living hippocampal neurons. Timelapse fluorescence imaging revealed that BMP4EGFP clusters showed bidirectional movements in the somatodendritic compartment. Finally, overexpression of BMP4 in neurons induced reduction of both density and immunoreactivity of VGluT1positive presynaptic puncta associated with BMP4expressing dendrites. These results indicate the presence of transsynaptic retrograde signaling in BMP4dependent suppression of synapse formation. P Roles of ACF, a large linker protein interacting with both microtubules and Factin, in the postsynaptic functions Two major components of the cytoskeletal system, Factin and microtubules MTs, have been shown to be involved in the postsynaptic functions. Actin network regulates the spine shape and motility. MTs invade into spines intermittently and help spine head expansion through the remodeling of actin network. Although these observations suggest importance of coordination between two cytoskeletal systems, there have been few reports on molecular mechanisms that regulate their interaction. ACF7/MACF1 is a member of spectraplakin family protein and links between Factin and MTs. By manipulating ACF7 content in cultured hippocampal neurons, we assessed the impact of ACF7dependent cytoskeletal reorganization on spine morphology. First, overexpression of ACF7GFP revealed accumulation of ACF7 in a subset of dendritic spines. This selective targeting may reflect specific functional states of spines. Second, downregulation of ACF7 by shRNA induced elongation of spines, which was rescued by expression of shRNA registant ACF7 cDNA. This phenotype may indicate that reduction of ACF7 blocks MTdependent spine head expansion and reversibly enhances excess growth of thin spines. P Imaging dynamics of axonal mitochondria The complex geometry of neuron requires specialized mechanism to allocate sufficient number of organelle to neurites and synapses. Proper distribution of mitochondria is critical for multiple neuronal functions including energy production, calcium homeostasis, neuronal apoptosis, synaptic transmission and plasticity. However, mitochondrial dynamics underlying the maintenance of their proper distribution is largely unknown. To study mitochondrial dynamics in neurons, we established a live imaging system to visualize neuronal mitochondria in hippocampal dissociated culture. We are now investigating neuronal mitochondria distribution and trafficking by expressing mitochondrial outer membrane protein Omp25C tagged with fluorescent proteins and performing time lapse confocal imaging. The size, distribution and motility of mitochondria in the axon and synaptic contact sites are analyzed quantitatively and these data will be integrated to estimate the rate and extent of mitochondria replacement in specific neuronal compartments. P ERRγ estrogenrelated receptor: ERR estrogen receptor: ERαßγ 3 ERR DNA ER ERR ER ER ERR 3 ERRγR 10 ERRγR ER ERRγR ER ERRγR ER
117 169 P Agerelated changes in estrogen receptorβ mRNA expression in male rat brain Estrogen plays important roles not only in reproductive function but also in other functions such as cognition and emotion. Estrogen acts mainly via its receptor ER, ERα and β, in target tissues. During aging, estrogenic actions are altered in both females and males, raising the possibility that expression level of ER may be altered with age. Agerelated changes in ER expression in female rat brain have been well demonstrated with regard to reproductive aging, while very little is known about the effects of age on the expression of ERs, especially ERβ, in males. In this study, to elucidate the effects of aging on ERβ expression in the male brain at the transcriptional level, we performed in situ hybridization using young, middleaged and old male rats. We revealed the number of ERβ mRNA positive cells was decreased with age in the cerebral cortex, hippocampus, dorsal endopiriform nucleus, medial septal nucleus, amygdala, anteroventral periventricular nucleus, substantia nigra, raphe and locus coeruleus. These results suggest that ERβ expression in male rat brain decreases with age at the transcriptional level and that these aging effects are regionspecific. P PRLR 1,2 2 2 1 2 1 2 ARC PRLR PRLR TH PRLR PRLR ARC TH PRLR TH PRLR PRLR ARC PRL ARC PRLR PRL PRLR P RTPCR 2 Wistar OVX Sham 1 2 17β 1 2 17α/C1720 1 OVX 2 OVX Sham 1 2 P Sporadically lurking HAPimmunoreactive cells in the hippocampus and their morphological relation with steroid receptors Md. Nabiul Islam, Ryutaro Fujinaga, Akie Yanai, Mir Rubayet Jahan, Yukio Takeshita, Keiji Kokubu, Koh Shinoda Division of Neuroanatomy, Yamaguchi University Graduate School of Medicine Huntingtinassociated protein 1 HAP1 is a neural huntingtin interactor that is widely expressed in the limbic and hypothalamic regions. Although HAP1 has been reported to be associated with steroid receptors, HAP1immunoreactive HAP1ir cells remain to be identified in the hippocampus. We determined the distribution of hippocampal HAP1ir cells in light and fluorescence microscopy and characterized their morphological relationships with steroid receptors, markers of adult neurogenesis and the GABAergic system in adult Wistar rats of both sexes. HAP1ir cells that were sporadically distributed in the subgranular zone SGZ of the dentate gyrus and in the interface between the stratum lacunosummoleculare and stratum radiatum of Ammon’s horn, were identified as the “sporadically lurking HAP1-ir (SLH)” cells. The SLH cells showed no clear association with the markers of adult neurogenesis in the SGZ, while all the SLH cells expressed neuN. More than 90% of the SLH cells expressed nuclear ERα, while more than 65% of them exhibited GABA immunoreactivity. We conclude that SLH cells might be involved in estrogendependent hippocampal functions through ERα and GABAergic regulation. P GR GR GR GR vimentin GR GR GR coactivator p300 SRC1 ADX GR coactivator vimentin GR vimentin p300 SRC1 ADX vimentin GR p300SRC1 GR GR P 1 1 2 1 1 2 24 cfos, Per1 cfos, Per1 gate 10 1 3 2 30 cfos Per1 in situ hybridization
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