Microbes modulate sympathetic neurons via a gut-brain circuit

Gut-brain connections monitor the intestinal tissue and its microbial and dietary content1, regulating both intestinal physiological functions such as nutrient absorption and motility2,3, and brain–wired feeding behaviour2. It is therefore plausible that circuits exist to detect gut microbes and relay this information to central nervous system (CNS) areas that, in turn, regulate gut physiology4. We characterized the influence of the microbiota on enteric–associated neurons (EAN) by combining gnotobiotic mouse models with transcriptomics, circuit–tracing methods, and functional manipulation. We found that the gut microbiome modulates gut-extrinsic sympathetic neurons; while microbiota depletion led to increased cFos expression, colonization of germ-free mice with short-chain fatty acid-producing bacteria suppressed cFos expression in the gut sympathetic ganglia. Chemogenetic manipulations, translational profiling, and anterograde tracing identified a subset of distal intestine-projecting vagal neurons positioned to play an afferent role in microbiota-mediated modulation of gut sympathetic neurons. Retrograde polysynaptic neuronal tracing from the intestinal wall identified brainstem sensory nuclei activated during microbial depletion, as well as efferent sympathetic premotor glutamatergic neurons that regulate gastrointestinal transit. These results reveal microbiota-dependent control of gut extrinsic sympathetic activation through a gut-brain circuit. Actively translated mRNA profiles from immunoprecipitated ribosome-bound mRNA from the nodose ganglion, dorsal root ganglion, and celiac superior mesentetic of Snap25:RiboTag specific pathogen free or germ free mice were prepared by deep sequencing on an Illumina NextSeq. Actively translated mRNA profiles from immunoprecipitated ribosome-bound mRNA from the nodose ganglion of SNS:RiboTag, Nav1.8:RiboTag, and Advillin:RiboTag mice were prepared by deep sequencing on an Illumina NextSeq 16S RNA profiles, from cecal samples of C57BL6/J mice given streptomycin or PBS, were prepared for deep sequencing on an Illumina MiSeq.肠-脑环路可监测肠道组织及其微生物与膳食组成1，同时调控肠道生理功能（如营养吸收与肠道运动2,3）以及脑调控的摄食行为2。因此，存在能够感知肠道微生物并将信号传递至中枢神经系统（CNS）区域、进而调控肠道生理功能的神经环路，该推测具有合理性4。本研究通过将悉生小鼠模型与转录组学、环路示踪技术及功能操控手段相结合，解析了微生物群对肠相关神经元（enteric-associated neurons, EAN）的调控作用。研究发现肠道微生物组可调控肠外交感神经元：微生物群耗竭会导致cFos表达上调，而给无菌小鼠定植产短链脂肪酸（short-chain fatty acid）细菌，则可抑制肠道交感神经节内的cFos表达。化学遗传学操控、翻译谱分析及顺行示踪技术鉴定出一类投射至远端肠道的迷走神经元亚群，该亚群可在微生物群介导的肠交感神经元调控中发挥传入作用。通过对肠壁进行多突触逆行神经元示踪，本研究鉴定出微生物群耗竭过程中激活的脑干感觉核团，以及调控胃肠道转运的传出交感前运动谷氨酸能神经元。上述结果揭示了通过肠-脑环路实现的、依赖于微生物群的肠外交感神经激活调控机制。本研究对Snap25:RiboTag无特定病原体（specific pathogen free, SPF）或无菌小鼠的结状神经节、背根神经节及腹腔肠系膜上神经节的免疫沉淀核糖体结合mRNA进行活跃翻译mRNA谱分析，并通过Illumina NextSeq平台完成深度测序。本研究对SNS:RiboTag、Nav1.8:RiboTag及Advillin:RiboTag小鼠的结状神经节的免疫沉淀核糖体结合mRNA进行活跃翻译mRNA谱分析，并通过Illumina NextSeq平台完成深度测序。本研究对经链霉素或磷酸盐缓冲液（PBS）处理的C57BL/6J小鼠的盲肠样本进行16S RNA谱分析，并通过Illumina MiSeq平台完成深度测序。