Ablation of the Presynaptic Protein Mover Impairs Learning Performance and Decreases Anxiety Behavior in Mice

Abstract

The presynaptic protein Mover/TPRGL/SVAP30 is absent in Drosophila and C. elegans and differentially expressed in synapses in the rodent brain, suggesting that it confers specific functions to subtypes of presynaptic terminals. In order to investigate how the absence of this protein affects behavior and learning, Mover knockout mice (KO) were subjected to a series of established learning tests. To determine possible behavioral and cognitive alterations, male and female 8-week-old KO and C57Bl/6J wildtype (WT) control mice were tested in a battery of memory and anxiety tests. Testing included the cross maze, novel object recognition test (NOR), the Morris water maze (MWM), the elevated plus maze (EPM), and the open field test (OF). Mover KO mice showed impaired recognition memory in the NOR test, and decreased anxiety behavior in the OF and the EPM. Mover KO did not lead to changes in working memory in the cross maze or spatial reference memory in the MWM. However, a detailed analysis of the swimming strategies demonstrated allocentric-specific memory deficits in male KO mice. Our data indicate that Mover appears to control synaptic properties associated with specific forms of memory formation and behavior, suggesting that it has a modulatory role in synaptic transmission.

Keywords: SNARE; behavior; exploratory behavior; hippocampus; knockout mice; mover; spatial learning; synaptic facilitation; synaptic vesicles.

Figure 1

Recognition memory deficits of Mover KO mice in the novel object recognition task. During the training phase, all male (a) and female (e) mice spent equal amounts of time with two similar objects (O1, O2). During the testing phase, only male (b) and female WT animals (f) showed a significant preference for the novel object (N). In contrast, Mover KO mice did not discriminate between the novel (N) and the familiar object (F). Distance traveled did not differ between male (c) or female KO (g) and WT animals. Calculation of the discrimination index (DI) also revealed recognition memory deficits in male (d) and female KO mice (h). Fifty percent chance level is indicated by a dashed line. Paired t-test (a,b,e,f) and unpaired t-test (c,d,g,h); n = 15–17. Data presented as mean ± S.E.M. * p < 0.05; ** p < 0.01.

Figure 2

Working memory and exploration behavior of Mover KO mice in the cross maze. Spontaneous alternation did not differ between male (a) or female (b) KO mice and same-sex WT animals. Male (c) and female (d) KO mice traveled significantly further than WT animals. In addition, female KO mice (f) showed an increased speed compared to WT mice. Mean speed did not differ between male KO mice and WT animals (e). Unpaired t-test, n = 15–17. Data presented as mean ± S.E.M. * p < 0.05.

Figure 3

Spatial learning and spatial reference memory of Mover KO mice in the Morris water maze. Spatial learning of KO mice was not altered as mice improved significantly during acquisition training. Escape latencies did not differ significantly between male (a) or female KO mice (e) and WT animals. Swimming speed did not differ between male (b) or female (f) KO mice and WT animals in the acquisition training. In the probe trial (c,d), all animals displayed a clear preference for the target quadrant. Male KO swam significantly faster than WT animals (d). In contrast, swimming speed was not altered in female KO mice (h) in the probe trial. Chance level is indicated by a dashed line. Two-way (a,b e,f) and one-way (c,g) ANOVA followed by Bonferroni multiple comparisons and unpaired t-test (d,h); n = 13–15. Data presented as mean ± S.E.M. ** p < 0.01, *** p < 0.001.

Figure 4

Qualitative analysis of spatial learning of Mover KO mice in the Morris water maze. Distribution of search strategies used by female (a) and male (b) KO Mover and same-sex WT mice. Animals showed a clear progression towards increasing spatial strategies over the 5 days of acquisition training. (c) Search strategies used by mice to locate the hidden platform in the MWM can be divided into hippocampus-dependent and non-hippocampus-dependent strategies. During the acquisition training, the cognitive scores of male (d) KO mice were significantly lower than those of WT animals. In contrast, cognitive scores did not differ between female KO and WT mice (f). During the probe trial, male (e) and female (g) KO mice showed a similar cognitive score compared to WT mice. Two-way repeated measures analysis of variance (ANOVA) followed by Bonferroni multiple comparisons (d,f) ANOVA and unpaired t-test (e,g); n = 13–15. Data presented as mean ± S.E.M. ANOVA: * p < 0.05, ** p < 0.01 (genotype difference); # p < 0.05 (sex difference).

Figure 5

Exploration and anxiety behavior of Mover KO mice in the open field. Male KO mice spent more time in the center of the arena (a) and were more active than WT animals (c). In contrast, there was no significant difference in the time spent in the center of the box (b) or the distance traveled (d) between female KO and WT animals. The latency to first enter the center of the arena did not differ between KO and WT mice, independent of sex (e,f). Unpaired t-test, n = 15–17. Data presented as mean ± S.E.M. * p < 0.05 (genotype difference); # p < 0.05 (sex difference).

Figure 6

Exploration and anxiety behavior of Mover KO mice in the elevated plus maze. Male KO mice spent significantly less time in the closed arms of the maze (a) and traveled (e) significantly further than WT mice. In contrast, no significant differences in time spent in the closed arms (b) or distance traveled (f) were observed in female mice. The number of arm entries did not differ between male KO (c) or female KO (d) and same-sex WT animals. Unpaired t-test, n = 15–17. Data presented as mean ± S.E.M. * p < 0.05; ** p < 0.01 (genotype difference); ## p < 0.01 (sex difference).