Developmental expression of GLUT3 glucose transporter in the rat brain

AbstractThe ontogeny of the GLUT3 glucose transporter gene and protein expression was studied in rat brain. Northern blot analysis using total RNA from rat brains at different developmental stages revealed that the levels of GLUT3 mRNA were very low during the embryonic stage and increased towards the postnatal stage. Immunohistochemistry using a specific antibody showed that the expression of GLUT3 protein was barely detectable in the embryonic stage, but was clearly detected on the plasma membrane of neuronal cells from 10 days after birth to the adult. Expression of GLUT3 mRNA and protein in the cerebral neuronal cell cultures was also examined during the maturation of neurons. GLUT3 glucose transporter of primary neuronal cultured cerebral cortical neurons was only detected in mature neurons after they were cultured for 14 days. These results indicate that GLUT3 plays an important role in glucose homeostasis postnatally in neurons of the rat brain

Glinide, but Not Sulfonylurea, Can Evoke Insulin Exocytosis by Repetitive Stimulation: Imaging Analysis of Insulin Exocytosis by Secretagogue-Induced Repetitive Stimulations

To investigate the different effects between sulfonylurea (SU) and glinide drugs in insulin secretion, pancreatic β-cells were repeatedly stimulated with SU (glimepiride) or glinide (mitiglinide). Total internal reflection fluorescent (TIRF) microscopy revealed that secondary stimulation with glimepiride, but not glucose and mitiglinide, failed to evoke fusions of insulin granules although primary stimulation with glucose, glimepiride, and mitiglinide induced equivalent numbers of exocytotic responses. Glimepiride, but not glucose and mitiglinide, induced abnormally sustained [Ca2+]i elevations and reductions of docked insulin granules on the plasma membrane. Our data suggest that the effect of glinide on insulin secretory mechanisms is similar to that of glucose

Imaging analysis reveals mechanistic differences between first- and second-phase insulin exocytosis

The mechanism of glucose-induced biphasic insulin release is unknown. We used total internal reflection fluorescence (TIRF) imaging analysis to reveal the process of first- and second-phase insulin exocytosis in pancreatic β cells. This analysis showed that previously docked insulin granules fused at the site of syntaxin (Synt)1A clusters during the first phase; however, the newcomers fused during the second phase external to the Synt1A clusters. To reveal the function of Synt1A in phasic insulin exocytosis, we generated Synt1A-knockout (Synt1A−/−) mice. Synt1A−/− β cells showed fewer previously docked granules with no fusion during the first phase; second-phase fusion from newcomers was preserved. Rescue experiments restoring Synt1A expression demonstrated restoration of granule docking status and fusion events. Inhibition of other syntaxins, Synt3 and Synt4, did not affect second-phase insulin exocytosis. We conclude that the first phase is Synt1A dependent but the second phase is not. This indicates that the two phases of insulin exocytosis differ spatially and mechanistically

骨芽細胞系列細胞のビタミンD受容体は、in vivo において1α,25(OH)2D3の骨吸収促進活性に必須である

要旨我々は以前、活性型ビタミンD3 [1α,25(OH)2D3]製剤であるエルデカルシトールの薬用量の投与は、骨吸収を抑制することで、骨量を増加させることを報告した。この骨吸収抑制効果は、骨芽細胞系列細胞のビタミンD受容体（VDR）を介することを明らかにした。本研究において我々は、骨芽細胞系列細胞特異的VDR欠損（Ob-VDR-cKO）マウスを用いて、1α,25(OH)2D3の大量投与によって誘導される骨吸収促進が骨芽細胞系列細胞のVDRを介するか否かを調べた。4日間の野生型マウスへの1α,25(OH)2D3（5 µg/kg体重/日）投与は、骨における破骨細胞数を増加させ、骨吸収マーカーであるI型コラーゲンC末端テロペプチド（C-terminal crosslinked telopeptide of type I collagen, CTX-I）の血清濃度を上昇させた。骨吸収促進は、血清カルシウム（Ca）値、線維芽細胞増殖因子23（Fibroblast Growth Factor, FGF-23）値の上昇と、体重の減少を伴っていた。このことは、中毒量の1α,25(OH)2D3は、骨吸収促進と高Ca血症を誘導することを示している。対照的に、野生型マウスへの抗Receptor Activator of NF-κB Ligand（RANKL）中和抗体前投与は、1α,25(OH)2D3が誘導する血清CTX-I, CaおよびFGF23値の上昇を抑制した。また、抗RANKL中和抗体前投与は、1α,25(OH)2D3が誘導する体重減少を抑制した。抗RANKL中和抗体前投与マウスの所見と一致して、1α,25(OH)2D3をOb-VDR-cKOマウスに大量投与しても、破骨細胞数、血清CTX-I値、血清Ca値、血清FGF-23値は、有意に上昇せず、また体重も減少しなかった。以上、本研究において、1α,25(OH)2D3の大量投与による骨吸収促進、血清Ca値上昇および毒性作用は、骨芽細胞系列細胞のVDRを介して発揮されることを明らかにした。2020博士（歯学）松本歯科大

Deletion of CDKAL1 Affects Mitochondrial ATP Generation and First-Phase Insulin Exocytosis

A variant of the CDKAL1 gene was reported to be associated with type 2 diabetes and reduced insulin release in humans; however, the role of CDKAL1 in β cells is largely unknown. Therefore, to determine the role of CDKAL1 in insulin release from β cells, we studied insulin release profiles in CDKAL1 gene knockout (CDKAL1 KO) mice.Total internal reflection fluorescence imaging of CDKAL1 KO β cells showed that the number of fusion events during first-phase insulin release was reduced. However, there was no significant difference in the number of fusion events during second-phase release or high K(+)-induced release between WT and KO cells. CDKAL1 deletion resulted in a delayed and slow increase in cytosolic free Ca(2+) concentration during high glucose stimulation. Patch-clamp experiments revealed that the responsiveness of ATP-sensitive K(+) (K(ATP)) channels to glucose was blunted in KO cells. In addition, glucose-induced ATP generation was impaired. Although CDKAL1 is homologous to cyclin-dependent kinase 5 (CDK5) regulatory subunit-associated protein 1, there was no difference in the kinase activity of CDK5 between WT and CDKAL1 KO islets.We provide the first report describing the function of CDKAL1 in β cells. Our results indicate that CDKAL1 controls first-phase insulin exocytosis in β cells by facilitating ATP generation, K(ATP) channel responsiveness and the subsequent activity of Ca(2+) channels through pathways other than CDK5-mediated regulation

The impact of CALL instruction on classroom computer use: A foundation for rethinking technology in teacher education

This purpose of this study is to examine how language teachers apply practical experiences from computer-assisted language learning (CALL) coursework to their teaching. It also examines ways in which teachers continue their CALL professional development. Participants in the study were 20 English as a second language and foreign language teachers who had, within the last 4 years, completed the same graduate-level CALL course and who are currently teaching. Surveys and follow-up interviews explored how participants learn about CALL activities; how what they learned in the course interacts with their current teaching contexts; the factors that influence whether or not they use technology in their classrooms; and how they continue to acquire and master new ideas in CALL. The findings support previous research on technology teacher education as it suggests that teachers who use CALL activities are often those teachers who had experience with CALL prior to taking the course; that lack of time, support, and resources prohibits the use of CALL activities in some classrooms; and that colleagues are the most common resource of new CALL activity ideas outside of formal coursework. Implications for teacher education are that teachers learn better in situated contexts, and technology courses should be designed accordingly