Fig. 4: VMH SF1 neurons project to PVT neurons.
a Schematic depiction of photometry monitoring of PVT neurons in SF1-Cre mice transduced with hM3Dq in VMH SF1 neurons and PVT neurons with GCaMP6f. b–g Acute PVT brain slice photometry experiments show that DREADD activation of SF1 neurons with J60 addition in the circulating ACSF potently increased PVT GCaMP6f signals in the SF1 neuron hM3Dq-transduced mice (n = 4) as compared to control mCherry-transduced mice (n = 3): b representative images of hM3Dq-transduced SF1 neurons in VMH and GCaMP6f-positive PVT neurons; c representative real-time monitoring of GCaMP6f signals before and after J60 addition in an isolated PVT brain slice of hM3Dq-transduced animal; d average PVT GCaMP6f signals from the SF1 neuron hM3Dq-transduced mice (n = 11 sections/4 mice); e average PVT neural GCaMP6f signals from the SF1 neuron mCherry-transduced mice (n = 9 sections/3 mice); f group data of GCaMP6f -positive neurons in the PVT (n = 3 per group; p = 0.0356, two-tailed Student’s t-tests); g representative confocal images of acute brain slice morphological studies show that SF1 neurons project synaptic inputs to GCaMP6f-positive neurons in PVT, as indicated by the hM3Dq-mCherry positive SF1 terminals adjunct on GCaMP6f-positive PVT neurons; 1 h after J60 addition in circulating solutions the brain slices were collected and fixed using PFA. h Representative confocal images of PVT neurons virally transduced with a CaMKII-driven vector carrying EGFP and stained with anti-vGluT2 antibodies. i, j ChR2-assisted circuit mapping in acute PVT brain slices of the SF1 neuron ChR2-transduced mice: i representative light (10 Hz) evoked postsynaptic currents (PSCs) at PVT neurons in the presence of TTX and 4-AP; j addition of CNQX eliminated the light-evoked synaptic currents; blue lines point to the PS. Data represent mean ± s.e.m.; *p < 0.05. Scale bars, 150 μm for b; 20 μm for g, h; 20 pA and 50 ms for i, j. DMH dorsomedial hypothalamus, PVT paraventricular thalamus, V3 third ventricle, VMH ventromedial hypothalamus.
