The Effects of Cyclophosphamide on Hippocampal Cell Proliferation and Spatial Working Memory in Rat

Cyclophosphamide (CP) is a chemotherapy used in combinations that are associated with cognitive impairment. In the present study male Lister-hooded rats (n = 12) were used to investigate the effects of chronic administration of CP (30mg/kg, 7 i.v. doses, or an equivalent volume of saline) on performance in the novel location recognition (NLR) task and on the proliferation and survival of hippocampal cells. The survival of hippocampal cells dividing at the beginning of treatment was significantly reduced by CP. However, no difference was seen between CP treated and control groups for the number of cells proliferating 7 days after the final injection and both groups performed equally well in the NLR task. These results indicate that the given dose of CP acutely reduces the survival of newly born hippocampal cells. However, it does not have a longer term effect on spatial working memory or hippocampal proliferation, suggesting that CP is less neurotoxic than other chemotherapies with which it is used in combination

Fluoxetine Counteracts the Cognitive and Cellular Effects of 5-Fluorouracil in the Rat Hippocampus by a Mechanism of Prevention Rather than Recovery

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

Fluoxetine reverses the memory impairment and reduction in proliferation and survival of hippocampal cells caused by methotrexate chemotherapy

RATIONALE: Adjuvant cancer chemotherapy can cause long-lasting, cognitive deficits. It is postulated that these impairments are due to these drugs targeting neural precursors within the adult hippocampus, the loss of which has been associated with memory impairment. OBJECTIVES: The present study investigates the effects of the chemotherapy, methotrexate (MTX) on spatial working memory and the proliferation and survival of the neural precursors involved in hippocampal neurogenesis, and the possible neuroprotective properties of the antidepressant fluoxetine. METHODS: Male Lister hooded rats were administered MTX (75 mg/kg, two i.v. doses a week apart) followed by leucovorin rescue (i.p. 18 h after MTX at 6 mg/kg and at 26, 42 and 50 h at 3 mg/kg) and/or fluoxetine (10 mg/kg/day in drinking water for 40 days). Memory was tested using the novel location recognition (NLR) test. Using markers, cell proliferation (Ki67) and survival (bromodeoxyuridine/BrdU), in the dentate gyrus were quantified. RESULTS: MTX-treated rats showed a cognitive deficit in the NLR task compared with the vehicle and fluoxetine-treated groups. Cognitive ability was restored in the group receiving both MTX and fluoxetine. MTX reduced both the number of proliferating cells in the SGZ and their survival. This was prevented by the co-administration of fluoxetine, which alone increased cell numbers. CONCLUSIONS: These results demonstrate that MTX induces an impairment in spatial working memory and has a negative long-term effect on hippocampal neurogenesis, which is counteracted by the co-administration of fluoxetine. If translatable to patients, this finding has the potential to prevent the chemotherapy-induced cognitive deficits experienced by many cancer survivors

BrdU and Ki67-positive cell counts.

<p>Total number of BrdU -positive (a) and Ki67-positive (b) cells in the dentate gyrus (mean ± SEM) estimated from cell counts. Rats receiving CP had significantly fewer BrdU-positive cells (<i>p</i><0.05) than the saline-treated control group. No significant difference was found between groups for the total numbers of Ki67-positive cells.</p

Time line showing protocol of drug administration and behavioural testing.

<p>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.</p

Novel location recognition (NLR) task.

<p>Mean exploration times (mean ± SEM) of the rats for each object in the familiarisation (a) and choice (b) trials of the NLR task. There was no significant difference in exploration time of either object for both groups in the familiarisation trial (<i>p</i>>0.05). In the choice trial, both groups spent significantly longer exploring the object in the novel location (<i>p</i><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). Both groups were significantly different from chance (<i>p</i><0.05). The total exploration time (mean ± SEM) for both trial combined (d) did not differ significantly between groups (<i>p</i>>0.05).</p

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

<p>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 (<i>p</i><0.001) in the SGZ and rats receiving fluoxetine had significantly more (<i>p</i><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 (<i>p</i>>0.05). The group receiving 5-FU with fluoxetine only in recovery had significantly fewer BrdU-postive cells than the control group (<i>p</i><0.001).</p

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).

<p>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 (<i>p</i><0.01) in the subgranular zone (SGZ) and rats receiving fluoxetine had significantly more (<i>p</i><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 (<i>p</i>>0.05).</p

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

<p>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.</p

Body weights.

<p>Body weights of rats (mean ± SEM) throughout the study. Arrows indicate CP (30mg/kg)/saline injections.</p