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. 2023 Aug 15;14:4924. doi: 10.1038/s41467-023-40504-6

Fig. 4. Activation of the LPBVglut2 → DMH projection induces strong cold defense responses independent of the POA.

Fig. 4

a Activating the LPBVglut2 → DMH projection via optogenetics. b Expression of ChR2 from LPBVglut2 neural terminals (left) and summary of fiber tracts (shown as blue dots, right) in the DMH. This experiment was repeated at least 10 times independently with similar results. c, d Changes of Tcore (c) and physical activity (d) after photoactivation of LPBVglut2 & ChR2 neural terminals in the DMH (ChR2, n = 10 mice; GFP, n = 8 mice). e Activating the LPBVglut2 → DMH projection while blocking LPB somas with hM4Di. f Representative ChR2 and hM4Di co-expression in LPBVglut2 neurons (n = 5 mice). g Changes in Tcore after activating the LPBVglut2 → DMH projection while blocking LPB neurons (n = 11 mice). (CNO, i.p., 10 mg/kg). h The neurotransmitter properties of LPB-innervating DMH neurons. i The overlap between LPB-innervating DMH neurons (mCherry+) and GABAergic or glutamatergic markers (GFP+). (n = 3 mice each). j, k Schemes for photoactivation (j) and changes in Tcore (k) after photoactivation of the LPBVglut2 → DMH projection while blocking downstream neurons. (n = 5 mice each). l–o Blocking of DMHVglut2 or DMHVgat neurons abolished photoactivation-induced increases in physical activity (l), iBAT thermogenesis (m), nuchal muscle shivering EMG (n), and heart rate (o). Animal numbers were indicated. p–r Changes in Tcore (p), maximum ΔTcore (q) and normalized heat production (r) after photoactivation of the LPBVglut2 → DMH projection under different Ta (n = 6 mice each). Heat production was normalized by the formula (Tcore -Ta) / ΔTcore as reported47. All data are the mean ± sem, and were analyzed by two-way RM ANOVA followed by Bonferroni’s multiple comparisons tests (c, g, k–o), or by uncorrected Fisher’s LSD test (d), (q and r) were analyzed by paired t-test. The p-values are calculated based on statistical tests in Supplementary Table 2. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001; ###p ≤ 0.001; ns, not significant. Light pattern: 473 nm, 12 mW, 10 Hz, 10 ms, 2-s on 2-s off, duration as indicated. Source data are provided as a Source data file.