Control of Cortical Axon Elongation by a GABA-Driven Ca<sup style="margin: 0px; padding: 0px; border: 0px; outline-style: none; font-weight: inherit; font-style: inherit; font-size: 0.85em; font-family: inherit; line-height: 0; text-align: inherit; vertical-align: super;">2+/Calmodulin-Dependent Protein Kinase Cascade</sup>

Ca(2+) signaling plays important roles during both axonal and dendritic growth. Yet, whether and how Ca(2+) rises may trigger and contribute to the development of long range cortical connections remains largely unknown. Here we demonstrate that two separate limbs of CaMK kinase (CaMKK) - CaMKI cascades, CaMKK-CaMKIα and CaMKK-CaMKIγ, critically coordinate axonal and dendritic morphogenesis of cortical neurons, respectively. The axon-specific morphological phenotype required a diffuse cytoplasmic localization and a strikingly α-isoform-specific kinase activity of CaMKI. Unexpectedly, treatment with muscimol, a GABA(A) receptor agonist, selectively stimulated elongation of axons but not of dendrites, and the CaMKK-CaMKIα cascade critically mediated this axonogenic effect. Consistent with these findings, during early brain development, in vivo knockdown of CaMKIα significantly impaired the terminal axonal extension, and thereby perturbed the refinement of the interhemispheric callosal projections into the contralateral cortices. Our findings thus indicate a novel role for the GABA-driven CaMKK-CaMKIα cascade as a mechanism critical for accurate cortical axon pathfinding, an essential process which may contribute to fine-tuning the formation of interhemispheric connectivity during the perinatal development of the central nervous system

シンキ マク アンカー タンパクシツ CLICK-3 CaMKIガンマ ノ クローニング ナラビニ ブンシ セイギョ キコウ ノ カイセキ

京都大学0048新制・課程博士博士(医学)甲第10449号医博第2648号新制||医||844(附属図書館)UT51-2003-T275京都大学大学院医学研究科脳統御医科学系専攻(主査)教授 月田 承一郎, 教授 中西 重忠, 教授 武藤 誠, 教授 成宮 周学位規則第4条第1項該当Doctor of Medical ScienceKyoto UniversityDA

Nonlinear Decoding and Asymmetric Representation of Neuronal Input Information by CaMKIIα and Calcineurin

How information encoded in glutamate release rates at individual synapses is converted into biochemical activation patterns of postsynaptic enzymes remains unexplored. To address this, we developed a dual fluorescence resonance energy transfer (FRET) imaging platform and recorded CaMKIIα and calcineurin activities in hippocampal neurons while varying glutamate uncaging frequencies. With little spine morphological change, 5 Hz spine glutamate uncaging strongly stimulated calcineurin, but not CaMKIIα. In contrast, 20 Hz spine glutamate uncaging, which induced spine growth, activated both CaMKIIα and calcineurin with distinct spatiotemporal kinetics. Higher temporal resolution recording in the soma revealed that CaMKIIα activity summed supralinearly and sensed both higher frequency and input number, thus acting as an input frequency/number decoder. In contrast, calcineurin activity summated sublinearly with increasing input number and showed little frequency dependence, thus functioning as an input number counter. These results provide evidence that CaMKIIα and calcineurin are fine-tuned to unique bandwidths and compute input variables in an asymmetric manner

A Flp-dependent G-CaMP9a transgenic mouse for neuronal imaging in vivo

脳の神経活動を可視化する新規マウス系統を開発 --高感度・高速カルシウムセンサーによる神経活動の計測に成功--. 京都大学プレスリリース. 2022-02-15.Genetically encoded calcium indicators (GECIs) are widely used to measure calcium transients in neuronal somata and processes, and their use enables the determination of action potential temporal series in a large population of neurons. Here, we generate a transgenic mouse line expressing a highly sensitive green GECI, G-CaMP9a, in a Flp-dependent manner in excitatory and inhibitory neuronal subpopulations downstream of a strong CAG promoter. Combining this reporter mouse with viral or mouse genetic Flp delivery methods produces a robust and stable G-CaMP9a expression in defined neuronal populations without detectable detrimental effects. In vivo two-photon imaging reveals spontaneous and sensory-evoked calcium transients in excitatory and inhibitory ensembles with cellular resolution. Our results show that this reporter line allows long-term, cell-type-specific investigation of neuronal activity with enhanced resolution in defined populations and facilitates dissecting complex dynamics of neural networks in vivo

Additional file 3: Figure S1. of Facilitation of axon outgrowth via a Wnt5a-CaMKK-CaMKIα pathway during neuronal polarization

Activation of Wnt5a-Ca2+ signaling in immature cultured cortical neurons. Addition of Wnt5a (100 ng/ml, final concentration) to the culture medium evoked a sustained Ca2+ oscillatory response in immature cultured cortical neurons (Cell #1 and Cell #2). Vertical lines indicate the time points of Wnt5a application and of the respective image frames. Scale bar: 5 μm. (PDF 74 kb

Additional file 1: Movie 1. of Facilitation of axon outgrowth via a Wnt5a-CaMKK-CaMKIÎą pathway during neuronal polarization

Fluo-4 Ca2+ imaging of immature migrating cortical neurons during spontaneous Ca2+ activity. Movie 1 (related to Fig. 1c) shows transient Fluo-4 fluorescence signals induced by spontaneous Ca2+ rises, in migrating neurons in CP and IZ as identified by TagRFP expression. Migrating neurons in CP and IZ are magnified from boxed areas in the left panel of the movie. (AVI 3964 kb