Extended Data Fig. 13: Additional cortico-thalamic connectivity of MPF.

a, PT (PHAL/CTB) and RE (FG/AAV-RFP) coinjections (a1-a2). CTB and FG labelled neurons were in DPd and ILA L5/6 (a3). PT axons (PHAL) densely innervated ILA L1-3 but sparsely labelled DPd; RE axons (AAV-RFP) targeted DP layer 1, DPd, and ILA L6 (a4). RE projecting neurons were found in ORBvl, ORBl, ACAv, and AIp (a3-a5) and in SUBd and SUBv, while PT projecting neurons were mainly in SUBv (a10-a11). PT projecting neurons were also in MEApd and hypothalamic nuclei related to reproductive behaviours (MPN, PMd), while RE projecting neurons targeted PMd (a6-a8) linked to defensive behaviour. In brainstem, CTB/FG labelled neurons were in PAG and Kolliker-Fuse nucleus (KF) (a9), involved in respiratory and cardiovascular regulation. b, PHAL/CTB was injected into the RE (b1) while FG into the PVT (b2), revealing distinct bidirectional connections of both nuclei with the DP, ILA, and other MPF regions. Retrograde labelling showed divergent input sources: PVT receives input mainly from visceral sensorimotor-related structures (b3–4), while RE receives input from PMd (b5), mammillary nuclei (MM, LM; b6), SC (b7), ventral lateral geniculate nucleus (LGv) (b8), and SUB (b9). c, Monosynaptic inputs to RE → DP neurons was investigated (schematic). AAVretro-Cre was injected into DP, and a Cre-dependent TVA- and RG-expressing helper virus (AAV8-hSyn-FLEX-TVA-P2A-GFP-2A-oG) and mCherry-expressing G-deleted rabies virus into RE. c1, AAVretro-Cre DP injection. c2, shows RE starter cells via colabeling of Cre, AAV-GFP, and mCherry. c3-c11, Representative images with rabies labelling in (1) cortical areas (ACAv, ACAd, MOs); (2) thalamic nuclei [PVT, rhomboid (RH), medial mediodorsal (MDm), dorsal anteromedial (AMd), ventromedial (VM), and reticular thalamic nuclei (RT)]; (3) BLAa, (4) hippocampal CA1, SUB, ENT; and (5) superior colliculus (SC) and periaqueductal grey (PAG). d, Schematic of RE as a hub integrating and relaying an array of neural inputs to DP. See Supplementary Table 1 for abbreviations.