A study of juvenile-to-adult refinement of thalamic reticular circuits via LRRTM3 enables high-resolution sensory encoding. Dongsu Lee and Kyung Ah Han et al.

In the study, we identified LRRTM3, as an excitatory synapse-specific cell-adhesion molecules, that regulates the TRN circuit maturation. Using single-nucleus RNA sequencing, ex vivo and in vivo electrophysiology, immunohistochemistry, and behavioral assays with TRN-specific Lrrtm3 conditional knockout mice, we show that TRN-LRRTM3 modulates CT-TRN synaptic connectivity in a developmentally dynamic manner, maintains the excitation-inhibition (E/I) balance required for sensory processing, and thalamic sensory tuning for tactile discrimination. Also, we uncover a post-critical period refinement by distinct extracellular interactions of LRRTM3 across post-juvenile stages. Our findings redefine the TRN dynamically tuned circuit after the classical critical period. These results offer a framework for understanding how inhibitory circuit maturation may contribute to sensory processing after early development stage. We performed single nucleus RNA-sequencing (snRNA-seq) to profile the thalamus of 6-and 15-weeks-old C57BL/6 mice. Single-nucleus transcriptomes of murine thalamic nuclei were generated using a fixative-exchange sequencing (FX-seq) workflow combined with sci-RNA-seq3 combinatorial barcoding. Thalamic tissues from 6- and 15-week-old C57BL/6 mice were perfusion-fixed, cryosectioned, crosslinked with a regioselective nucleobase crosslinker, enzymatically digested, and processed to isolate intact nuclei.本研究中，我们鉴定发现LRRTM3作为兴奋性突触特异性细胞黏附分子（excitatory synapse-specific cell-adhesion molecules），可调控丘脑网状核（Thalamic Reticular Nucleus, TRN）环路的成熟过程。我们采用单细胞核RNA测序（single-nucleus RNA sequencing, snRNA-seq）、离体（ex vivo）与在体（in vivo）电生理记录、免疫组织化学染色技术，以及针对丘脑网状核特异性Lrrtm3条件性敲除小鼠的行为学实验，证实TRN-LRRTM3以发育动态的方式调节皮层-丘脑网状核（cortico-thalamic reticular nucleus, CT-TRN）突触连接，维持感觉加工所需的兴奋-抑制（excitation-inhibition, E/I）平衡，以及介导触觉辨别所需的丘脑感觉调谐功能。此外，我们还揭示了LRRTM3在幼年后不同阶段通过独特的细胞外相互作用，实现关键期后的环路精细化调控。本研究的发现重新定义了经典关键期后丘脑网状核动态调控的环路机制。上述研究结果为理解抑制性环路成熟如何在早期发育阶段后参与感觉加工提供了理论框架。 我们对6周龄和15周龄的C57BL/6小鼠的丘脑组织开展了单细胞核RNA测序（snRNA-seq）以解析其转录组特征。本研究通过固定置换测序（fixative-exchange sequencing, FX-seq）流程结合sci-RNA-seq3组合条形码技术，构建了小鼠丘脑核团的单细胞核转录组图谱。具体实验流程如下：对6周龄和15周龄的C57BL/6小鼠的丘脑组织进行灌注固定、冰冻切片、区域选择性核碱基交联剂交联、酶解消化，最终分离得到完整的细胞核。