Neonatal exposure to fluoxetine and fluvoxamine alteres spine density in mouse hippocampal CA1 pyramidal neurons

Abstract

Some women in childbearing ages take selective serotonin reuptake inhibitors (SSRIs) such as fluoxetine and fluvoxamine for treating psychiatric symptoms. However, these compounds may cause some side effects to their children. It has been identified that early life exposure to SSRIs increased the chance of developing mood disorders and the biological basis is still unclear. Here, we studied the effects of neonatal exposure to SSRIs on neuronal morphology. We used GFP-transgenic mice to investigate the acute and long-lasting effects of early life exposure to SSRIs on dendritic spine density of CA1 neurons. We found that 18-day drug applications of fluoxetine and fluvoxamine significantly reduced spine density of basal dendrites at postnatal day 22 (P22), but only fluvoxamine caused a reduction of spine density of apical dendrites. Interestingly, compared with the control group, the spine densities of basal dendrites after fluoxetine and fluvoxamine exposure and the spine density of apical dendrites after fluoxetine exposure increased in adult mice at the age of P90. We also observed impaired locomotor activity in adult mice after exposure to SSRIs. Our findings demonstrated that neonatal exposure to SSRIs was capable of influencing the morphological plasticity of excitatory synapses. It raised the caution for clinical use of SSRIs.

Keywords: SSRI; dendritic spine density; fluoxetine; fluvoxamine; neuronal development.

Figure 1

Schematic diagram of experimental design and morphological analysis. (A) GFP transgenic mice received daily injection (i.p.) of SSRIs or vehicle from P4 to P21. The drug dose is 10 mg/kg for both of the two SSRIs, fluoxetine and fluvoxamine, while control group received similar volume of saline. For acute morphological analysis, mice were sacrificed at P22, 20-24 h after the last injection. For behavioral tests, mice were separated by sex on P30 and housed under 12:12 h light/dark cycle. At the age of P90, part of the male mice was used for open field test, while others were sacrificed for evaluation of long-term consequence of SSRI treatment on neuronal morphology. (B) Image data were collected byZeiss confocal microscope. upper panel: hippocampal CA1 pyramidal neurons of GFP-O transgenic mice in low magnification (10X, NA 0.30, 488 nm laser, LSM 510); lower panel: dendritic spines under high magnification (100X, NA 1.40, 488 nm laser, LSM 510) on secondary dendrites. Scale bar, 5 μm. (C) Neurites and dendritic spines were traced with software Neurolucida (version 6.0) and the spine density was analyzed by NeuroEx-plorer (version 4.0).

Figure 2

Spine density of CA1 pyramidal neurons after early life exposure to SSRIs. (A, B) Spine density of both apical and basal dendrites were examined at the age of P22. Both of the two groups given SSRIs showed a decreased spine density in basal dendrites (saline, n = 38; flx, n = 30; fluvo, n = 37) compared with saline group, while only fluvoxamine treated group showed a significant decrease of spine density of apical dendrites (saline, n = 33; flx, n = 27; fluvo, n = 33) in CA1 pyramidal neurons. (C, D) Spine density were also examined at the age of P90. Compared with the control group (n=26), the spine density of basal dendrites after fluoxetine (n=30) and fluvoxamine (n=22) exposure and the spine density of apical dendrites after fluoxetine (n=21) exposure were higher in mice at the age of P90. n = number of dendrites. Data are expressed by mean ± SEM and analyzed by LSD test. (*, p<0.05; **, p < 0.01, compared with saline group.)

Figure 3

Open field test. (A) Impaired locomotor activities in adult mice after early life exposure to SSRIs. Both of the two groups given SSRIs exhibited a significant decrease of traveling distance during 60 min in the open field. (B) Reduced moving time in fluoxetine group. Total moving time in 60 min was recorded and only fluoxetine group showed a significant decrease of moving time compared with saline group. (C) Normal center time. All three groups showed the similar amount of time spent in center time, which indicated that there is no major difference of exploratory activity between saline- and drug-treated mice. Each plot represents data collected from 8 mice per group (saline, flx or fluvo). Data are expressed by mean ± SEM and analyzed by Student's t test, *, p < 0.05, **, p < 0.01.