Fig. 5: Kdm4a regulated engram formation in dentate gyrus to facilitate pattern separation task.

a Schematic of Injection of sgKdm4a or sgCtrl lentivirus into the DG of C57BL/6J mice. Scale bar, 500 μm. b RT-qPCR detection of Kdm4a expression level in DG infecting sgKdm4a versus sgCtrl lentivirus. n = 3, Two-tailed unpaired t-test, t₄ = 4.529, * p = 0.0106. Data are presented as mean ± s.e.m. c Knockout of Kdm4a in DG does not affect mouse locomotor activity. sgCtrl, n = 8; sgKdm4a, n = 8. Two-tailed unpaired t-test, t₁₄ = 0.04956, p = 0.9612. Data are presented as mean ± s.e.m. d Kdm4a^DG-KO mice showed normal contextual fear learning when compared with controls. sgCtrl, n = 13; sgKdm4a, n = 12. Two-tailed unpaired t-test, t₂₃ = 0.7862, p = 0.4398. Data are presented as mean ± s.e.m. e Knockout of Kdm4a in DG does not affect mouse fear memory extinction. sgCtrl, n = 6; sgKdm4a, n = 6. (Extinction day 1-6) Two-way ANOVA followed by Sidak test, F_1,10 = 0.2891, p = 0.6026. Data are presented as mean ± s.e.m. f Illustration of contextual fear discrimination task. Mice were allowed to learn to distinguish two slightly different contexts: context A and B. g (Left) There was no difference in freezing levels between Kdm4a^DG-KO mice and control mice in context A. sgCtrl, n = 12, sgKdm4a, n = 9. Two-way ANOVA followed by Sidak test, F_1,20 = 0.5987, p = 0.4481. (Middle) The freezing level of Kdm4a^DG-KO mice in context B was significantly lower than that of control mice. Two-way ANOVA followed by Sidak test, F_1,19 = 5.745, * p = 0.027. (Right) The discrimination index of Kdm4a^DG-KO mice is higher than that of the control mice. Two-way ANOVA followed by Sidak test, F_1,19 = 6.792, * p = 0.0174. Data are presented as mean ± s.e.m. h The chemogenetic virus hM3Dq-mCherry or control vector was delivered into the DG of C57BL/6 J mice. i, CNO was injected 1 h before perfusion and increased Fos expression in the DG with AAV-hM3Dq. Saline, n = 3; CNO, n = 3. Two-tailed unpaired t-test, t₄ = 3.163, * p = 0.0341. Data are presented as mean ± s.e.m. j Schematic of the contextual fear discrimination test. CNO or saline were injected 1 h before exploring of context B each day. k (Left) There was no difference in freezing levels between hM3Dq+CNO mice and other control groups in context A. n = 8 per group. Two-way ANOVA followed by Dunnett test, F_3,28 = 0.07766, p = 0.9716. (Middle) The freezing level of hM3Dq+CNO mice in context B was significantly decreased when compared with other control mice. Two-way ANOVA followed by Dunnett test, F_3,28 = 3.494, * p = 0.0285. (Right) The discrimination index of hM3Dq+CNO mice was significantly higher than that of other control mice. Two-way ANOVA followed by Dunnett test, F_3,28 = 6.885, ** p = 0.0013. Data are presented as mean ± s.e.m. l Characterization of the memory allocation test and context (Imm shock, immediate shock). m Kdm4a-knockdown mice froze significantly lower in context B than that in context A. shCtrl, n = 6, shKdm4a, n = 6. One-way ANOVA followed by Dunnett test, shCtrl: F_2,15 = 23.49, p < 0.0001. ns p = 0.0671, **** p < 0.0001; shKdm4a: F_2,15 = 22.79, p < 0.0001. ** p = 0.0069, **** p < 0.0001. Data are presented as mean ± s.e.m.