ラット ソクザカク ノ IPSP ニ タイスル アデノシン ト カンナビノイド ニヨル ダツブンキョク ユウドウ ヨクセイ ノ ユウドウ

A short period of depolarization evokes transient suppression of neurotransmitter release (Depolarization-induced Suppression of IPSP: DSI). Recent studies suggest that cannabinoids are candidates for the retrograde messenger required to induce DSI in the hippocampal CA1 area. A G protein-coupled process and inhibition of N-type calcium channels at the presynaptic terminals are believed to be involved in the induction mechanism. We examined DSI induction in the nucleus accumbens in which inputs from multiple sites in the telencephalon, including the hippocampus and basolateral amygdala, converge. Using whole-cell recording from a horizontal slice preparation of nucleus accumbens from the rat brain, we monitored the effect of endogenous cannabinoids on DSI induction and found that 300 nM AM281, a selective cannabinoid CB1 receptor antagonist, resulted in blockade of DSI. Adenosine is reported to suppress neurotransmitter release by inhibiting N-type calcium channels, which is a similar mechanism to that reported for cannabinoid-induced DSI. We therefore examined the effect of endogenous adenosine on DSI induction using DPCPX, a selective A1 receptor antagonist, and found that DSI induction was blocked in the presence of 100 nM DPCPX. These results suggest that cooperative activation of CB1 and A1 receptors contributes to DSI induction in the rat nucleus accumbens.　Key words: Nucleus accumbens, DSI, Cannabinoid, Adenosine, Whole-cell recording短時間の脱分極は一過性に伝達物質の放出を抑制する（脱分極誘導 IPSP 抑制：DSI）。最近の研究によればカンナビノイドは海馬 CA1 領域における DSI 誘導に関わる逆行性伝達物質であることが示されている。誘導のメカニズムとしてはＧ蛋白に結合した過程やＮ型のカルシウムチャンネルが関与していると考えられている。我々は、海馬・基底外側扁桃体などを含んだ終脳における多くの部位からの入力を統合している場所である側座核において、DSI の誘導を検討した。側座核の水平断薄切標本を用いて whole cell recording を行い、DSI誘導に関する内因性カンナビノイドの影響を調べたところ、選択的 CB1 カンナビノイド受容体阻害薬である 300nM の AM281 が DSI の誘導を阻害した。アデノシンはＮ型カルシウムチャンネルを阻害することによって伝達物質の放出を抑制することが報告されているが、この効果はカンナビノイドの効果と類似している。それ故我々は選択的な A1 アデノシン受容体の阻害薬である DPCPX を用いて、DSI 誘導に関する内因性アデノシンの効果を調べてみた。その結果、100nM の DPCPX が DSI の誘導を阻害することがわかった。これらの結果から CB1 と A1 受容体活性が側座核における DSI の誘導に関わっていることが示された。　キーワード：側座核、DSI 、カンナビノイド、アデノシン、whole-cell recordin

Facilitation of NMDAR-Independent LTP and Spatial Learning in Mutant Mice Lacking Ryanodine Receptor Type 3

AbstractTo evaluate the role in synaptic plasticity of ryanodine receptor type 3 (RyR3), which is normally enriched in hippocampal area CA1, we generated RyR3-deficient mice. Mutant mice exhibited facilitated CA1 long-term potentiation (LTP) induced by short tetanus (100 Hz, 100 ms) stimulation. Unlike LTP in wild-type mice, this LTP was not blocked by the NMDA receptor antagonist D-AP5 but was partially dependent on L-type voltage-dependent Ca2+ channels (VDCCs) and metabotropic glutamate receptors (mGluRs). Long-term depression (LTD) was not induced in RyR3-deficient mice. RyR3-deficient mice also exhibited improved spatial learning on a Morris water maze task. These results suggest that in wild-type mice, in contrast to the excitatory role of Ca2+ influx, RyR3-mediated intracellular Ca2+ ([Ca2+]i) release from endoplasmic reticulum (ER) may inhibit hippocampal LTP and spatial learning

Tissue-specific splicing regulator Fox-1 induces exon skipping by interfering E complex formation on the downstream intron of human F1γ gene

Fox-1 is a regulator of tissue-specific splicing, via binding to the element (U)GCAUG in mRNA precursors, in muscles and neuronal cells. Fox-1 can regulate splicing positively or negatively, most likely depending on where it binds relative to the regulated exon. In cases where the (U)GCAUG element lies in an intron upstream of the alternative exon, Fox-1 protein functions as a splicing repressor to induce exon skipping. Here we report the mechanism of exon skipping regulated by Fox-1, using the hF1γ gene as a model system. We found that Fox-1 induces exon 9 skipping by repressing splicing of the downstream intron 9 via binding to the GCAUG repressor elements located in the upstream intron 8. In vitro splicing analyses showed that Fox-1 prevents formation of the pre-spliceosomal early (E) complex on intron 9. In addition, we located a region of the Fox-1 protein that is required for inducing exon skipping. Taken together, our data show a novel mechanism of how RNA-binding proteins regulate alternative splicing

Viral RNA recognition by LGP2 and MDA5, and activation of signaling through step-by-step conformational changes

細胞内のウイルスを認識する蛋白質の仕組みを解明 --ウイルスから我々の体を守る影のヒーロー--. 京都大学プレスリリース. 2020-12-04.Cytoplasmic RIG-I-like receptor (RLR) proteins in mammalian cells recognize viral RNA and initiate an antiviral response that results in IFN-β induction. Melanoma differentiation-associated protein 5 (MDA5) forms fibers along viral dsRNA and propagates an antiviral response via a signaling domain, the tandem CARD. The most enigmatic RLR, laboratory of genetics and physiology (LGP2), lacks the signaling domain but functions in viral sensing through cooperation with MDA5. However, it remains unclear how LGP2 coordinates fiber formation and subsequent MDA5 activation. We utilized biochemical and biophysical approaches to observe fiber formation and the conformation of MDA5. LGP2 facilitated MDA5 fiber assembly. LGP2 was incorporated into the fibers with an average inter-molecular distance of 32 nm, suggesting the formation of hetero-oligomers with MDA5. Furthermore, limited protease digestion revealed that LGP2 induces significant conformational changes on MDA5, promoting exposure of its CARDs. Although the fibers were efficiently dissociated by ATP hydrolysis, MDA5 maintained its active conformation to participate in downstream signaling. Our study demonstrated the coordinated actions of LGP2 and MDA5, where LGP2 acts as an MDA5 nucleator and requisite partner in the conversion of MDA5 to an active conformation. We revealed a mechanistic basis for LGP2-mediated regulation of MDA5 antiviral innate immune responses