Neuronal cell death and the loss of connectivity are two of the primary pathological mechanisms underlying Alzheimer's disease.The accumulation of amyloid-βpeptides,a key hallmark of Alzheimer's disease,is believed to induce neuritic abnormalities,including reduced growth,extension,and abnormal growth cone morphology,all of which contribute to decreased connectivity.However,the precise cellular and molecular mechanisms governing this response remain unknown.In this study,we used an innovative approach to demonstrate the effect of amyloid-βon neurite dynamics in both two-dimensional and three-dimensional cultu re systems,in order to provide more physiologically relevant culture geometry.We utilized various methodologies,including the addition of exogenous amyloid-βpeptides to the culture medium,growth substrate coating,and the utilization of human-induced pluripotent stem cell technology,to investigate the effect of endogenous amyloid-βsecretion on neurite outgrowth,thus paving the way for potential future applications in personalized medicine.Additionally,we also explore the involvement of the Nogo signaling cascade in amyloid-β-induced neurite inhibition.We demonstrate that inhibition of downstream ROCK and RhoA components of the Nogo signaling pathway,achieved through modulation with Y-27632(a ROCK inhibitor)and Ibuprofen(a Rho A inhibitor),respectively,can restore and even enhance neuronal connectivity in the presence of amyloid-β.In summary,this study not only presents a novel culture approach that offers insights into the biological process of neurite growth and inhibition,but also proposes a specific mechanism for reduced neural connectivity in the presence of amyloid-βpeptides,along with potential intervention points to restore neurite growth.Thereby,we aim to establish a culture system that has the potential to serve as an assay for measuring preclinical,predictive outcomes of drugs and their ability to promote neurite outgrowth,both generally and in a patient-specific manner.
The perception of motion is an important func-tion of vision.Neural wiring diagrams for extracting direc-tional information have been obtained by connectome recon-struction.Direction selectivity in Drosophila is thought to originate in T4/T5 neurons through integrating inputs with different temporal filtering properties.Through genetic screening based on synaptic distribution,we isolated a new type of TmY neuron,termed TmY-ds,that form recipro-cal synaptic connections with T4/T5 neurons.Its neurites responded to grating motion along the four cardinal direc-tions and showed a variety of direction selectivity.Intrigu-ingly,its direction selectivity originated from temporal filtering neurons rather than T4/T5.Genetic silencing and activation experiments showed that TmY-ds neurons are functionally upstream of T4/T5.Our results suggest that direction selectivity is generated in a tripartite circuit formed among these three neurons—temporal filtering,TmY-ds,and T4/T5 neurons,in which TmY-ds plays a role in the enhancement of direction selectivity in T4/T5 neurons.
Impaired axonal transport has been observed in patients with amyotrophic lateral sclerosis(ALS)and in animal models,suggesting that transport proteins likely play a critical role in the pathological mechanism of ALS.Dysregulation of Kinesin-family-member 5α(Kif5α),a neuron-specific isoform of heavy chain kinesin family,has been described in several neurological disorders,in humans and animal models,including ALS.In this study,we determined Kif5αexpression by gene sequencing,quantitative reverse transcription-polymerase chain reaction,and western blot assay in the cervical spinal cord of wobbler mice and immunofluorescence staining in dissociated cultures of the ventral horn.Further,we observed the distribution of Kif5αand mitochondria along motor neuronal branches by confocal imaging.Our results showed that Kif5αexpression was greatly dysregulated in wobbler mice,which resulted in altered distribution of Kif5αalong motor neuronal branches with an abnormal mitochondrial distribution.Thus,our results indicate that dysregulation of Kif5 and therefore abnormal transport in motor neuronal branches in this ALS model could be causative for several pathological findings at the cellular level,like misallocation of cytoskeletal proteins or organelles like mitochondria.
Kilian KürtenAnne-Christin GudeAimo Samuel Christian EpplenJan SteinCarsten TheissVeronika Matschke
目的:本研究旨在阐明肾细胞癌下调蛋白1(DRR1)在小鼠大脑组织中的表达谱及相关生物学功能。方法:首先,通过real time RT-PCR的方法明确在不同发育阶段的小鼠大脑组织中DRR1的表达水平;利用免疫荧光染色法检测DRR1在原代培养的神经干细胞以及神经元中的表达情况。其次,采用real time RT-PCR的方法分别检测DRR1在诱导分化前后的神经干细胞中的表达水平。最后,利用慢病毒感染的方法在原代培养的神经干细胞以及神经元中分别上调或者下调DRR1的表达,进而评价DRR1对原代培养的神经干细胞的分化以及神经元的突起形成的影响。结果:DRR1在各个发育阶段的小鼠大脑组织中均有表达,并且其表达水平随发育呈逐渐上升趋势。DRR1在分化前后的神经干细胞中均表达,但在诱导分化后的细胞中的表达水平显著高于其分化前的表达水平(P<0.01)。同时,过表达DRR1可促进神经干细胞的分化。DRR1在分化成熟的原代神经元的树突以及轴突均有表达,且DRR1的表达水平影响神经元的突起形成(P<0.01)。结论:DRR1在各发育阶段的小鼠大脑及分化前后的神经干细胞中均有表达,其作用与神经干细胞分化调控以及神经元突起形成有关。
Non-nerve cell-derived extracellular matrix(ECM)wa s coated on the aligned porous polypyrrole-poly(Llactide)(PPy-PLLA)fiber-films with the conductivity of^12 mS/m via L929 cells culture and lysing,resulting in^10%increase of PC12 cells attachment and^26μm increase of neurites length.The neurite length of^149μm in EGF/NGF group(optimal concentration radio of 12.5/50(ng/mL))on aligned and ECM-conjugated fiber-films was significantly larger than^94μm in only NGF group(50 ng/mL),confirming the synergy of EGF,NGF and aligned ECM-conjuaged PPy-PLLA fibers.When differentiated PC12 cells were exerted electrical stimulation(ES)of 100 mV/cm for 4 h/day in 2 day through ECM-PPyPLLA fiber-films,their neurite length reached to^251μm,significantly larger than^149μm of group without ES,due to the higer expression of related neural proteins in ES group.A simple mechanism was proposed to analyze synergistical effect of ECM,EGF,NGF on axons adhesion and elongation along the aligned ECM-coated fibers under ES condition.
