Chronic reductions in serotonin transporter function prevent 5-HT1B-induced behavioral effects in mice

Abstract

Background: Obsessive-compulsive disorder (OCD) is characterized by intrusive thoughts, images, or impulses and/or repetitive stereotypical behavior. Obsessive-compulsive disorder patients exhibit reduced prepulse inhibition (PPI) and symptom exacerbation after challenge with 5-HT1B receptor agonists. Recently, gain-of-function alleles of the serotonin transporter (5-HTT) have been associated with OCD. We tested the hypothesis that reducing 5-HTT function chronically, either genetically or via serotonin reuptake inhibitor (SRI) treatment, attenuates PPI deficits and perseverative hyperlocomotion induced by 5-HT1B agonists in mice.

Methods: Mice received subchronic or chronic pretreatment with the SRI fluoxetine and acute treatment with RU24969 (5-HT1A/1B agonist) or 8-OH-DPAT (5-HT1A agonist) and were assessed for PPI, locomotor activity, and spatial patterns of locomotion. The same measures were evaluated in 5-HTT wild-type (WT), heterozygous (HT), and knockout (KO) mice after RU24969 treatment. The effects of WAY100635 (5-HTA antagonist) or GR127935 (5-HT1B/D antagonist) pretreatment on RU24969-induced effects were evaluated. Finally, 5-HT1B binding and functional coupling were assessed in 5-HTT-WT, -HT, and -KO mice, and normal fluoxetine-treated mice.

Results: Chronic, but not subchronic, fluoxetine treatment prevented RU24969-induced PPI deficits and perseverative hyperlocomotion. These RU24969-induced effects were mediated via 5-HT1B and not 5-HT1A receptors. 5-HTT-KO mice showed no effects of RU24969, and 5-HTT-HT mice exhibited intermediate phenotypes. 5-HT1B binding and functional coupling were reduced in the globus pallidus and substantia nigra of 5-HTT-KO mice.

Conclusions: Our results demonstrate that chronic, but not subchronic, fluoxetine treatment and 5-HTT knockout robustly attenuate 5-HT1B agonist-induced PPI deficits and perseverative hyperlocomotion. These results may have implications for the etiology and treatment of OCD.

Figure 1

(A) Percent prepulse inhibition is shown for BALB/cJ mice pretreated subchronically (one week) with 0 (n=15) or 15 (n=15) mg/kg/day fluoxetine. (B) PPI is shown for BALB/cJ mice pretreated chronically (four weeks) with 0 (n=15) or 15 (n=14) mg/kg/day fluoxetine. All mice received saline, RU24969, and 8-OH-DPAT treatment. Chronic, but not subchronic, 15mg/kg/day fluoxetine attenuated the PPI deficits induced by 10mg/kg RU24969. 8-OH-DPAT increased PPI across all pretreatments. Values are means ± SEM. Insets show data with prepulse intensity collapsed. An asterisk (*) indicates a significant difference from saline-treated animals within the same pretreatment group. A pound sign (#) indicates a significant difference from control-pretreated animals within the same treatment group.

Figure 2

(A) Total locomotor activity is shown for BALB/cJ mice pretreated subchronically (one week) with 0 (n=15) or 15 (n=15) mg/kg/day fluoxetine. (B) Total locomotor activity is shown for BALB/cJ mice pretreated chronically (four weeks) with 0 (n=15) or 15 (n=15) mg/kg/day fluoxetine. All mice received saline, RU24969, and 8-OH-DPAT treatment. Chronic, but not subchronic, 15mg/kg/day fluoxetine blocked the locomotor hyperactivity induced by 10mg/kg RU24969. 8-OH-DPAT reduced locomotion across all pretreatments. Values are means ± SEM. An asterisk (*) indicates a significant difference from saline-treated animals within the same pretreatment group. A pound sign (#) indicates a significant difference from control-pretreated animals within the same treatment group.

Figure 3

Locomotor paths taken by BALB/cJ mice receiving chronic fluoxetine (0 or 15mg/kg/day) pretreatment and RU24969 (0 or 10mg/kg) treatment are shown. Paths are representative for spatial d values, but not locomotor activity levels or amount of activity in the center vs. periphery. A control-pretreated mouse receiving RU24969 shows a highly perseverative pattern of locomotion characterized by straight paths along the perimeter of the open field (d=1.2). The remaining three mice show more meandering paths (d=1.4), and varying levels of locomotor activity.

Figure 4

(A) Prepulse inhibition and (B) total locomotor activity of serotonin transporter -WT, -HT, and -KO mice treated with control, RU24969, and 8-OH-DPAT. RU24969 reduced PPI only in 5-HTT-WT mice at the 3dB prepulse intensity. 8-OH-DPAT increased PPI in 5-HTT-WT and -HT, but not -KO mice. RU24969 increased locomotion in 5-HTT-WT and -HT, but not -KO mice. Locomotion was reduced overall in 5-HTT-KO mice. Values are means ± SEM. An asterisk (*) indicates a significant difference from saline-treated animals within the same genotype. A pound sign (#) indicates a significant difference from 5-HTT-WT mice within the drug treatment group.

Figure 5

(A) Prepulse inhibition and (B) total locomotor activity of BALB/cJ mice pretreated with control (n=14), 1mg/kg WAY100635 (n=15), or 5mg/kg GR127935, and treated with 0 or 10mg/kg RU24969 in a counterbalanced fashion. RU24969 treatment reduced PPI compared to control treatment in control- and WAY100635-pretreated, but not GR127935-pretreated mice. 5-HTT-WT mice at the 3dB prepulse intensity. Within RU24969-treatment, WAY100635 pretreatment reduced PPI and GR127935 pretreatment increased PPI at the 6 and 12dB prepulse intensities. RU24969 increased locomotion in control- and WAY100635 pretreated mice, but not GR127935 pretreated mice. Values are means ± SEM. An asterisk (*) indicates a significant difference from saline-treated animals within the same pretreatment group. A pound sign (#) indicates a significant difference from all other pretreatment groups within the same treatment group.

Figure 6

5-HT1B functional coupling as assessed by CP93129-induced [³⁵S]GTP-γ-S binding (A), and 5-HT1B receptor expression as assessed by [³H]5-CT binding (B) is shown for 5-HTT-WT, -HT, and -KO mice in the globus pallidus and substantia nigra. Quantification revealed significantly decreased [³H]5-CT binding in 5-HTT-HT and -KO mice relative to 5-HTT-WT mice. [³⁵S]GTP-γ-S binding was reduced in 5-HTT-KO mice relative to 5-HTT-WT and -HT mice (*p<.05). For representative images, see Suppl. Figs. 1 and 2.