Supplementary Information

Supplementary Figures and Legends

Supplementary Figure S1

htr3a-/- mice show normal cell proliferation and neurogenesis in the dorsal and ventral parts of the hippocampal dentate gyrus at baseline.

Supplementary Figure S2

The 5-HT3 receptor is required for exercise-induced cell proliferation in the dorsal and ventral parts of the hippocampal dentate gyrus.

Supplementary Figure S3

Stimulation of the 5-HT3 receptor promotes neurogenesis in both dorsal and ventral parts of the hippocampal dentate gyrus.

Supplementary Figure S4

The 5-HT3 receptor is required for exercise-induced neurogenesis in the dorsal and ventral parts of the hippocampal dentate gyrus.

Supplementary Figure S1

htr3a-/- mice show normal cell proliferation and neurogenesis in the dorsal and ventral parts of the hippocampal dentate gyrus at baseline.

(a–d) Analysis of baseline cell proliferation and neurogenesisin the dorsal and ventral parts of the hippocampal dentate gyrus. Quantification of the BrdU-labeled cells (a and b) and the BrdU/NeuN-double-labeled cells (c and d) in the dorsal (a and c) and ventral (b and d) parts of the hippocampal dentate gyrus(BrdU-labeled cells, WT, n = 9 mice; KO, n = 8 mice; BrdU/NeuN-double-labeled cells, n = 5 mice). The number of BrdU-positive cells (cell proliferation) in htr3a-/- mice on Day 5 did not differ from that in wild-type mice (wild-type vs. htr3a-/-: dorsal part, 2310 ± 81.97 vs. 2316 ± 185.0, p = 0.9767; ventral part, 1028 ± 27.43 vs. 1036 ± 71.21, p = 0.9143). In addition, there were no significant differences in the number of BrdU/NeuN-double-positive cells on Day 29 (neurogenesis) between wild-type and htr3a-/-mice (wild-type vs. htr3a-/-: dorsal part, 713.6 ± 27.73 vs.741.6 ± 20.02, p = 0.4367; ventral part, 317.6 ± 26.46 vs. 289.6 ± 14.18, p = 0.3782). ns, not significant (two-tailed t test).Means ± s.e.m. are shown in all histograms.

Supplementary Figure S2

The 5-HT3 receptor is required for exercise-induced cell proliferation in the dorsal and ventral parts of the hippocampal dentate gyrus.

(a–d) Analysis of exercise-induced cell proliferation in the dorsal and ventral parts of the hippocampal dentate gyrus. Quantification of the BrdU-labeled cells (a and b)and the BrdU/DCX-double-labeled cells (c and d) in the dorsal (a and c) and ventral (b and d) parts of the hippocampal dentate gyrus (BrdU-labeled cells, n = 8 mice; BrdU/DCX-double-labeled cells, n = 4 mice).Exercise for 6 days significantly increased the number of BrdU-positive cells (non-exercise vs. exercise: dorsal part, 2075± 79.18 vs. 3161±79.93, p < 0.0001; ventral part, 999.0± 36.27 vs. 1443±48.99, p < 0.0001)and BrdU/DCX-double-positive cells (non-exercise vs. exercise: dorsal part, 664.0±27.90 vs. 1038±62.60, p = 0.0055; ventral part, 324.0±25.82 vs. 460.0±18.62, p = 0.0052) in the subgranular zone of the dorsal and ventral parts of the dentate gyrus in wild-type mice.In contrast to wild-type mice, no significant increase in the number of BrdU-positive cells (non-exercise vs. exercise: dorsal part, 2077± 106.9 vs. 2279±102.5, p = 0.1940; ventral part, 1045 ± 54.53 vs. 1031± 56.38, p = 0.8609)norBrdU/DCX-double-positive cells(non-exercise vs. exercise: dorsal part, 660.0± 59.19 vs. 746.0±74.86, p = 0.4022; ventral part, 368.0±33.15 vs. 346.0±39.65, p = 0.6852)was found in htr3a-/- mice after exercise for 6 days.**p < 0.01, ***p < 0.001; ns, not significant (two-tailed t test). Means ± s.e.m. are shown in all histograms.

Supplementary Figure S3

Stimulation of the 5-HT3 receptor promotes neurogenesis in both dorsal and ventral parts of the hippocampal dentate gyrus.

(a–d) Analysis of the effects of 5-HT3 receptor stimulation on cell proliferation in the dorsal and ventral parts of the hippocampal dentate gyrus. Quantification of the BrdU-labeled cells (a and b)and the BrdU/DCX-double-labeled cells (c and d) in the dorsal (a and c) and ventral (b and d) parts of the hippocampal dentate gyrus(WT, n = 5 mice; KO, n = 4 mice). Treatment with SR 57227A significantly increased the numbers of BrdU-positive cells (dorsal part: saline, 760.0±59.06; SR 57227A: 1 mg/kg, 788.8±92.18, p = 0.7991; 5 mg/kg, 1034± 74.76, p = 0.0208; 10 mg/kg, 998.4±49.60, p = 0.0149; ventral part:saline, 480.0 ± 30.88; SR 57227A: 1 mg/kg, 476.8 ± 64.13, p = 0.9652; 5 mg/kg, 624.0 ± 50.85, p = 0.0418; 10 mg/kg, 574.4 ± 26.34, p = 0.0485) and BrdU/DCX-double-positive cells (saline vs. 5 mg/kg SR 57227A: dorsal part, 308.8±17.27 vs. 440.0±23.32, p = 0.0019; ventral part, 203.2± 10.31 vs. 267.2± 24.34, p = 0.0418)in both dorsal and ventral parts of the dentate gyrus in wild-type mice. In contrast, no significant increasewas observed in SR 57227A-treated htr3a-/- mice compared with saline-treated htr3a-/- mice (saline vs. 5 mg/kg SR 57227A: BrdU-positive: dorsal part, 764.0±48.61 vs. 822.0± 38.14, p = 0.3841; ventral part, 556.0±83.43 vs. 470.0± 26.81, p = 0.3643; BrdU/DCX-double-positive: dorsal part, 320.0± 13.47 vs. 302.0 ± 5.033, p = 0.2571; ventral part, 204.0±26.43 vs. 184.0±13.47, p = 0.5253).*p < 0.05,**p < 0.01; ns, not significant (two-tailed t test). Means ± s.e.m. are shown in all histograms.

Supplementary Figure S4

The 5-HT3 receptor is required for exercise-induced neurogenesis in the dorsal and ventral parts of the hippocampal dentate gyrus.

(a and b) Analysis of exercise-induced hippocampal neurogenesis. Quantification of the BrdU/NeuN-double-labeled cells in the granule cell layer of the dorsal (a) and ventral (b) parts of the hippocampal dentate gyrus (n = 5 mice). Three weeks of exercise significantly increased the number of BrdU/NeuN-double-positive cells in both dorsal and ventral parts of the dentate gyrus of wild-type mice (non-exercise vs. exercise: dorsal part, 1045± 62.41 vs. 2029± 161.7, p = 0.0005; ventral part, 513.6±54.34 vs. 939.2±113.0, p = 0.0095). In contrast, there was no significant increase inhtr3a-/- mice after 3 weeks of exercise (non-exercise vs. exercise: dorsal part, 1101± 50.82 vs. 1181±105.5, p = 0.5137; ventral part, 606.4±27.41 vs. 651.2±41.00, p = 0.3902). **p < 0.01, ***p < 0.001; ns, not significant (two-tailed t test). Means ± s.e.m. are shown in all histograms.

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