Regional-specific effect of fluoxetine on rapidly dividing progenitors along the dorsoventral axis of the hippocampus

Abstract

Hippocampus-dependent cognitive and emotional function appears to be regionally dissociated along the dorsoventral (DV) axis of the hippocampus. Recent observations that adult hippocampal neurogenesis plays a critical role in both cognition and emotion raised an interesting question whether adult neurogenesis within specific subregions of the hippocampus contributes to these distinct functions. We examined the regional-specific and cell type-specific effects of fluoxetine, which requires adult hippocampal neurogenesis to function as an antidepressant, on the proliferation of hippocampal neural stem cells (NSCs). Fluoxetine specifically increased proliferation of NSCs located in the ventral region of the hippocampus while the mitotic index of NSCs in the dorsal portion of the hippocampus remained unaltered. Moreover, within the ventral hippocampus, type II NSC and neuroblast populations specifically responded to fluoxetine, showing increased proliferation; however, proliferation of type I NSCs was unchanged in response to fluoxetine. Activation or inhibition of serotonin receptor 1A (5-HTR1A) recapitulated or abolished the effect of fluoxetine on proliferation of type II NSCs and neuroblast populations in the ventral hippocampus. Our study showed that the effect of fluoxetine on proliferation is dependent upon the type and the position of the NSCs along the DV axis of the hippocampus.

Figure 1. Fluoxetine increases neurogenesis in the ventral part of the hippocampus.

(a) Three different views of the hippocampus in coronal, sagittal, and horizontal planes (left). A: anterior, P: posterior, D: dorsal, V: ventral, M: medial, L: lateral. Coronal blocks showing anatomical boundaries used for defining sub-regions along the DV axis (middle). The hippocampus was divided into ventral (red) and dorsal (blue) segments (right). (b) Representative photos of the DAPI (blue) and BrdU (red) labeled sections in each block along the DV axis. (c) The schedule of the BrdU incorporation experiment of proliferation analysis. Bar graph showing the number of BrdU⁺ cells in each block/segment of the hippocampus (d) or in the whole hippocampus (e) in control and fluoxetine-administered mice. (f) The schedule of the BrdU incorporation experiment of survival analysis. Bar graph showing the number of BrdU⁺ cells in each block/segment of the hippocampus (g) or in the whole hippocampus (h) in control and fluoxetine-administered mice. Data represent the mean ± SEM. *p < 0.05; **p < 0.01; and ***p < 0.001 by two-way ANOVA compared with control mice.

Figure 2. Fluoxetine targets type II NSCs and neuroblasts in the ventral hippocampus.

(a) Schematic defining different types of NSCs in the neuronal differentiation cascade in the dentate gyrus. (b) Representative confocal images of type I cells (b-1 and b-2), type II cells (b-1 and b-3) and neuroblasts (b-4 and b-5). In b1, the red arrow indicates a type I cell with a GFAP-labeled radial process and the purple arrow indicates a type-II cell without a radial process. In b-2 the red arrow indicates dividing type I cells labeled by BrdU antibody. In b-3, the red arrow indicates dividing type II cells labeled by BrdU antibody. In b-4, the red arrow indicates BrdU⁺ neuroblasts not expressing Nestin-GFP. The BrdU⁺ neuroblasts do not have processes and are labeled by DCX in b-5 (red arrow). (c) The schedule of the BrdU incorporation experiment. (d) Representative confocal images showing the state of type I cells, type II cells and neuroblasts in each segment of the dentate gyrus in mice with or without fluoxetine treatment. Red arrows indicate proliferating type II cells. Bar graph showing the effect of fluoxetine on the number of proliferating type I cells (e), type II cells (f), neuroblasts (g), and others (h). (i) Pie charts showing the portion of type I cells, type II cells, neuroblasts, and others in each segment along the DV axis of the dentate gyrus in mice with or without fluoxetine treatment. (j) Bar graph showing the percentage of BrdU⁺ type II cells and neuroblasts among the BrdU⁺ cells in the ventral segment of control and fluoxetine-administeredrated mice. Data represent the mean ± SEM. *p < 0.05; **p < 0.01; and ***p < 0.001 by two-way ANOVA compared with control mice.

Figure 3. Activation of 5-HTR1A increases neurogenesis in the ventral part of the hippocampus.

(a) The schedule of the BrdU incorporation experiment. (b) Representative confocal images showing BrdU⁺ cells in the dentate gyrus in mice treated with/without 8-OH-DPAT. Bar graph showing the number of BrdU⁺ cells in in each block/segment of the hippocampus (c) or in the whole hippocampus (d) in control and 8-OH-DPAT-administrated mice. *p < 0.05; **p < 0.01; and ***p < 0.001 by two-way ANOVA compared with control mice.

Figure 4. 5-HTR1A accounts for the effect of fluoxetine on division of type II cells and neuroblasts in the ventral hippocampus.

(a) The schedule of the BrdU incorporation experiment of proliferation analysis. (b) Representative images showing BrdU⁺ cells in the dentate gyrus in mice treated with fluoxetine alone or fluoxetine and NAN-190. (c) Bar graph showing the number of BrdU⁺ cells in the whole hippocampus and in each block/segment of the hippocampus in control and fluoxetine-administrated mice with/without NAN-190 treatment. Data represent the mean ± SEM. (d) The schedule of the BrdU incorporation experiment in Nestin-GFP mice. (e) Representative confocal images showing the state of type I cells, type II cells and neuroblasts in each segment of the dentate gyrus in control and fluoxetine-administered mice with/without NAN-190 treatment. (f) Bar graph showing the effect of fluoxetine on the number of proliferating type I cells, type II cells, and neuroblasts in mice treated with/without NAN-190. *p < 0.05 by two-way ANOVA compared with control mice.

Figure 5. Regional-specific and cell-type-specific effect of fluoxetine on neurogenesis in the ventral hippocampus.

Fluoxetine promotes division of type II NSCs and neuroblasts specifically in the ventral hippocampus. 5-HTR1A plays the necessary and sufficient role in mediating the effect of fluoxetine on enhanced neurogenesis in the ventral hippocampus.