Fluoxetine counteracts the cognitive and cellular effects of 5-fluorouracil in the rat hippocampus by a mechanism of prevention rather than recovery

Abstract

5-Fluorouracil (5-FU) is a cytostatic drug associated with chemotherapy-induced cognitive impairments that many cancer patients experience after treatment. Previous work in rodents has shown that 5-FU reduces hippocampal cell proliferation, a possible mechanism for the observed cognitive impairment, and that both effects can be reversed by co-administration of the antidepressant, fluoxetine. In the present study we investigate the optimum time for administration of fluoxetine to reverse or prevent the cognitive and cellular effects of 5-FU. Male Lister-hooded rats received 5 injections of 5-FU (25 mg/kg, i.p.) over 2 weeks. Some rats were co-administered with fluoxetine (10 mg/kg/day, in drinking water) for 3 weeks before and during (preventative) or after (recovery) 5-FU treatment or both time periods (throughout). Spatial memory was tested using the novel location recognition (NLR) test and proliferation and survival of hippocampal cells was quantified using immunohistochemistry. 5-FU-treated rats showed cognitive impairment in the NLR task and a reduction in cell proliferation and survival in the subgranular zone of the dentate gyrus, compared to saline treated controls. These impairments were still seen for rats administered fluoxetine after 5-FU treatment, but were not present when fluoxetine was administered both before and during 5-FU treatment. The results demonstrate that fluoxetine is able to prevent but not reverse the cognitive and cellular effects of 5-FU. This provides information on the mechanism by which fluoxetine acts to protect against 5-FU and indicates when it would be beneficial to administer the antidepressant to cancer patients.

Figure 1. Time line showing protocol of drug administration and behavioural testing.

Arrows represent single i.p. injections of BrdU (fine) and 5-FU/saline (thick). Brackets represent the period of time for which fluoxetine was administered in the drinking water. The day after Novel location recognition (NLR) behavioural testing, animals were killed and their brains were removed.

Figure 2. Body weights of rats (a) and their fluid intake (b) during fluoxetine treatment period (mean ± SEM).

Arrows indicate 5-FU (20 mg/kg)/saline injections. Fluoxetine was given in drinking water (10 mg/kg/day) from day 1 to day 20 for the 5-FU+fluoxetine (preventative) group, from day 21 to day 40 for the 5-FU+fluoxetine (recovery) group and from day 0 to day 40 for the 5-FU+fluoxetine (throughout) and the fluoxetine alone groups.

Figure 3. Mean exploration times (mean ± SEM) of the rats for each object in the familiarisation (a) and choice (b) trials.

There was no significant difference in exploration time of either object for any group in the familiarisation trial (p>0.05). In the choice trial, all groups spent significantly longer exploring the object in the novel location (*p<0.05, **p<0.01), except the groups receiving 5-FU alone or 5-FU with fluoxetine in recovery (p>0.05). Preference indices (PI, (c), mean ± SEM) were created by expressing time spent exploring the object in the novel location as a percentage of the sum of exploration time of novel and familiar locations in the choice trial (Bruel-Jungerman et al. 2005). All groups were significantly different from chance (*p<0.05, **p<0.01, ***p<0.001), other than the groups receiving 5-FU alone or 5-FU with fluoxetine in recovery (p>0.05). The total exploration time (mean ± SEM) for both trial combined (d) did not differ significantly between groups (p>0.05).

Figure 4. Mean velocity (mean ± SEM) of rats recorded during the habituation period using EthoVision 4.1.

No significant difference (p>0.05) was found between each group.

Figure 5. Photographs of the nuclei of cells in the SGZ of the dentate gyrus (blue, a), Ki67-positive cells (red, b) and the photos merged (c).

Bar scales indicate 20 µm. Total numbers of Ki67-positive cells (mean ± SEM) in the dentate gyrus were estimated from cell counts (d). Rats receiving 5-FU had significantly fewer Ki67-positive cells (p<0.01) in the subgranular zone (SGZ) and rats receiving fluoxetine had significantly more (p<0.05) than the saline-treated control group. The other treatment groups receiving both 5-FU and fluoxetine did not significantly differ from the controls (p>0.05).

Figure 6. Photographs of the nuclei of cells in the dentate gyrus (blue, a), BrdU-positive cells (green, b) and the photos merged (c).

Bar scales indicate 20 µm. Total numbers of BrdU-positive cells (mean ± SEM) in the dentate gyrus were estimated from cell counts (d). Rats receiving 5-FU had significantly fewer BrdU-positive cells (p<0.001) in the SGZ and rats receiving fluoxetine had significantly more (p<0.01) than the saline-treated control group. Treatment groups receiving both 5-FU and fluoxetine throughout and in prevention did not significantly differ from the controls (p>0.05). The group receiving 5-FU with fluoxetine only in recovery had significantly fewer BrdU-postive cells than the control group (p<0.001).