Fig. 3: Subtype “1” Dat1⁺ neurons in the anterior PeVN innervate the lateral septum.

a Tyrosine hydroxylase (TH) is present in neuronal projections coursing towards the lateral septum (LS). Left: experimental scheme. Right: immunohistochemistry for TH and mCherry in fibers projecting from the PeVN to the LS. Immunohistochemistry was performed for each viral injection (n = 9 independent experiments) with similar results. Scale bar = 10 μm. b TH⁺/Onecut3⁺ neurons situated in the PeVN were labeled retrogradely (left) by stereotaxic injections targeting the LS. Onecut3 localization is shown to the right. Immunohistochemistry was performed for each viral injection (n = 3 independent experiments) and yielded similar results. Scale bars = 20 μm. c Experimental scheme for ex vivo electrophysiological circuit mapping e with optogenetic stimulation (d; n = 12 animals). d Periventricular Dat1⁺ neurons infected with channelrhodopsin-2 (ChR2) generate inward currents upon repetitive light stimulation. e Patch-clamp recordings of neurons in the LS show their activation (11 out of 24 cells) upon light stimulation of ChR2-expressing Dat1⁺ terminals. f Post-hoc reconstruction of a biocytin-filled neuron responding to optogenetic stimulation e confirmed synaptic innervation by Dat1-Cre⁺ (mCherry-filled) afferents originating from the PeVN. Scale bar = 50 μm. 3V third ventricle. Biorender assisted us to draw schemes and anatomical structures (in a–c).