Fluoxetine increases hippocampal neurogenesis and induces epigenetic factors but does not improve functional recovery after traumatic brain injury

Abstract

The selective serotonin reuptake inhibitor fluoxetine induces hippocampal neurogenesis, stimulates maturation and synaptic plasticity of adult hippocampal neurons, and reduces motor/sensory and memory impairments in several CNS disorders. In the setting of traumatic brain injury (TBI), its effects on neuroplasticity and function have yet to be thoroughly investigated. Here we examined the efficacy of fluoxetine after a moderate to severe TBI, produced by a controlled cortical impact. Three days after TBI or sham surgery, mice were treated with fluoxetine (10 mg/kg/d) or vehicle for 4 weeks. To evaluate the effects of fluoxetine on neuroplasticity, hippocampal neurogenesis and epigenetic modification were studied. Stereologic analysis of the dentate gyrus revealed a significant increase in doublecortin-positive cells in brain-injured animals treated with fluoxetine relative to controls, a finding consistent with enhanced hippocampal neurogenesis. Epigenetic modifications, including an increase in histone 3 acetylation and induction of methyl-CpG-binding protein, a transcription factor involved in DNA methylation, were likewise seen by immunohistochemistry and quantitative Western immunoblots, respectively, in brain-injured animals treated with fluoxetine. To determine if fluoxetine improves neurological outcomes after TBI, gait function and spatial learning and memory were assessed by the CatWalk-assisted gait test and Barnes maze test, respectively. No differences in these parameters were seen between fluoxetine- and vehicle-treated animals. Thus while fluoxetine enhanced neuroplasticity in the hippocampus after TBI, its chronic administration did not restore locomotor function or ameliorate memory deficits.

FIG. 1.

Chronic administration of fluoxetine increases the total number of immature neurons in the dentate gyrus. There was a significant reduction in total DCX-immunoreactive cells in the ipsilateral dentate gyrus following unilateral traumatic brain injury compared to sham injury in both vehicle-treated (post-hoc: ***p < 0.005), and in fluoxetine-treated (post-hoc: *p < 0.05) groups. However, chronic administration of fluoxetine increased the total number DCX-immunoreactive cells in the ipsilateral dentate gyrus of the sham (post-hoc: ‡p < 0.05) and TBI (post-hoc: #p < 0.05) groups (TBI, traumatic brain injury; Fluox, fluoxetine; DCX, doublecortin).

FIG. 2.

Chronic administration of fluoxetine increases the number of MBD1-expressing cells in the dentate gyrus. There was a significant reduction in the total number of MBD1-immunoreactive cells in the ipsilateral dentate gyrus following unilateral TBI compared to sham animals only in the vehicle-treated groups (post-hoc: ***p < 0.005). Chronic fluoxetine treatment increased the total number of MBD1-immunoreactive cells in the ipsilateral dentate gyrus of the TBI (post-hoc: ‡p < 0.05) groups (TBI, traumatic brain injury; Fluox: fluoxetine; MBD1, methyl-CpG-binding domain-1).

FIG. 3.

Chronic administration of fluoxetine increases histone 3 acetylation in the hippocampus. (A) There was a significant increase in the ratio of acetylated histone 3 normalized by total histone 3 (AcH3:H3) in brain-injured mice treated with fluoxetine compared to brain-injured mice treated with saline (post-hoc: **p < 0.01). (B) Representative images of Western blots showing the expression of total histone 3 (H3) and acetylated histone 3 (AcH3; Fluox: fluoxetine; TBI, traumatic brain injury).

FIG. 4.

Chronic fluoxetine administration does not reverse TBI-induced impairment in gait spatial parameters. The intensity (A) and maximum areas (B) were reduced at all paws at 5 weeks following TBI. Fluoxetine did not have any effect on either intensity or maximum area in sham or brain-injured mice (NS, not significant between TBI/vehicle-treated and TBI/fluoxetine-treated groups; TBI, traumatic brain injury; Fluox, fluoxetine; RF, right fore; RH, right hind; LF, left fore; LH, left hind; a.u., arbitrary units).

FIG. 5.

Neither TBI nor fluoxetine administration affects depression-like behavior in the forced swimming test. Following the initial 2 min of adjustment in the cylinder, the total time spent in immobility, swimming, and climbing was summed for each behavior. There was no significant difference in time spent in each behavior due to either TBI or fluoxetine (TBI, traumatic brain injury; Fluox: fluoxetine).

FIG. 6.

Chronic fluoxetine treatment does not reduce TBI-induced impairments in spatial learning and memory. Mice subjected to TBI and treated with vehicle on average traveled longer distances to find the escape tunnel than sham-vehicle treated animals (post-hoc: sham-vehicle versus TBI-vehicle: #p < 0.0001). Although brain-injured mice treated with fluoxetine had a tendency to perform better on the Barnes maze test than brain-injured mice treated with vehicle, the difference was not statistically significant (post-hoc: TBI-vehicle versus TBI-fluoxetine: p = 0.13). Shams receiving chronic fluoxetine treatment performed better than sham mice receiving saline or brain-injured mice receiving chronic fluoxetine (post-hoc: sham-vehicle versus sham-fluoxetine: *p < 0.05; sham-fluoxetine versus TBI-fluoxetine: ‡p < 0.0001; TBI, traumatic brain injury; Fluox, fluoxetine).