Serum Response Factor (SRF) regulates motoneuron vulnerability and activity- dependent neuroprotection in Amyotrophic Lateral Sclerosis.

肌萎缩侧索硬化症（ALS）是一种进行性神经退行性疾病，主要影响脊髓中的运动神经元（MN），特别是下运动神经元。最近，神经元活动和放电模式显示出对鼠ALS模型如SOD 1（G93 A）ALS小鼠模型中的疾病进展具有强烈影响。在这里，MN中的化学发生学升高的放电减缓了疾病负担，而对神经元活性的干扰加速了疾病标志物的积累（例如自噬诱导）。在这项研究中，我们想分析的作用，血清反应因子（SRF），一个典型的神经元活动诱导的转录因子（TF）。SRF介导神经元活性诱导的即刻早期基因（IEG）的基因转录，所述即刻早期基因（IEG）例如c-Fos、Egr 1和Npas 4在生理和病理（例如癫痫、急性应激）诱导神经元中的神经元活性时。因此，我们假设神经元活动对延缓ALS疾病进展的有益影响通过激活MN中SRF介导的基因转录起作用。为了检验这一假设，我们已经建立了条件性Srf突变小鼠系，并记录了运动神经元中的排他性SRF耗竭（ChAT-Cre+/-; Srf fl/fl）。我们进一步将该小鼠系与已建立的小鼠ALS模型SOD 1（G93 A）交配以获得ChAT-Cre+/-;Srf fl/fl; SOD 1（G93 A）小鼠。在先前的第一项工作中，我们观察到与单独的SOD 1小鼠相比，Srf突变体/SOD 1小鼠的体重减轻和握力降低。 MN限制性SRF耗竭是否使SOD 1 ALS模型中的疾病进展恶化？为此，我们分析了几个行为参数（握力、阶梯行走、临床评分、倒置网格、开放视野），并进行了详细的组织学检查，包括自噬、MN数量、炎症和突触形成的标志物。最后，我们进行MN的激光捕获介导的分离，以通过RNA-SEQ.ii）鉴定这些小鼠中的ALS和SRF依赖性转录组。 SRF是介导MN中化学发生诱导的活性依赖性神经保护的下游靶点吗？在这里，我们使用化学遗传学在脊髓中病毒过表达激活或抑制通道（阳离子可渗透的actPSAM或阴离子可渗透的inhPSAM），分别缓解或恶化SOD 1 ALS小鼠模型中的疾病进展。与仅SOD 1小鼠相比，将在Srf突变体/SOD 1小鼠中进行这些实验。基于我们将SRF归因于在介导神经元中的神经元活动中的关键作用的假设，我们预期化学发生驱动的神经元活化不能保护免于在SRF耗尽时ALS引起的神经变性。该项目分析了神经元活动，特别是神经元活动，由SRF介导的活性介导的基因转录是ALS进展中的一个新因素，也可能作为未来潜在的药物靶点，ALS对患者的影响。

Amyotrophic Lateral Sclerosis (ALS) is a progressive neurodegenerative condition affecting primarily motoneurons (MN), in particular lower motoneurons in the spinal cord. Recently, neuronal activity and firing patterns were shown to have a strong impact on disease progression in murine ALS models such as the SOD1(G93A) ALS mouse model. Here, chemogenetically elevated firing in MNs slowed down the disease burden whereas interference with neuronal activity accelerated accumulation of disease markers (e.g. autophagy induction). In this study we want to analyze the role of SRF (serum response factor), a prototypical neuronal activity-induced transcription factor (TF). SRF mediates neuronal-activity induced gene transcription of immediate early genes (IEGs) such as c-Fos, Egr1 and Npas4 upon physiological and pathological (e.g. epilepsy, acute stress) induction of neuronal activity in neurons. Thus, we hypothesize that the beneficial impact of neuronal activity on delaying ALS disease progression works through activation of SRF-mediated gene transcription in MNs. To test this hypothesis we already established a conditional Srf mutant mouse line and documented exclusive SRF depletion in motoneurons (ChAT-Cre+/-; Srf fl/fl). We further bred this mouse line to the established mouse ALS model SOD1(G93A) to obtain ChAT-Cre+/-;Srf fl/fl;SOD1(G93A) mice. In first previous work we observed additional weight loss and grip strength reduction in Srf mutant/SOD1 mice compared to the SOD1 mice alone.With these mice at hand we follow two objectives: i) Does MN-restricted SRF depletion worsen disease progression in the SOD1 ALS model? For this, we analyze several behavioral parameters (grip strength, ladder walk, clinical score, inverted grid, open field) and perform a detailed histological inspection including markers for autophagy, MN numbers, inflammation and synapse formation. Finally, we perform a laser-capture mediated isolation of MNs to identify the ALS- and SRF-dependent transcriptome in these mice by RNA-SEQ.ii) Is SRF a downstream target in mediating chemogenetically-induced, activity-dependent neuroprotection in MNs? Here we use chemogenetics to virally overexpress activating or inhibitory channels (cation-permeable actPSAM or anion-permeable inhPSAM) in the spinal cord that alleviate or worsen disease progression in the SOD1 ALS mouse model, respectively. These experiments will be performed in the Srf mutant/SOD1 mice in comparison to SOD1 mice only. Based on our hypothesis attributing SRF a pivotal role in mediating neuronal activity in neurons we expect that chemogenetically-driven neuronal activation fails to protect from ALS inflicted neurodegeneration upon SRF depletion.In summary, this project analyzes whether neuronal activity and specifically neuronal-activity mediated gene transcription mediated by SRF is a novel factor in ALS progression that may also serve as a future potential drug target to alleviate the impact of ALS in patients.