Comprehensive behavioral analysis of ENU-induced Disc1-Q31L and -L100P mutant mice

Abstract

Background: Disrupted-in-Schizophrenia 1 (DISC1) is considered to be a candidate susceptibility gene for psychiatric disorders, including schizophrenia, bipolar disorder, and major depression. A recent study reported that N-ethyl-N-nitrosourea (ENU)-induced mutations in exon 2 of the mouse Disc1 gene, which resulted in the amino acid exchange of Q31L and L100P, caused an increase in depression-like behavior in 31 L mutant mice and schizophrenia-like behavior in 100P mutant mice; thus, these are potential animal models of psychiatric disorders. However, remaining heterozygous mutations that possibly occur in flanking genes other than Disc1 itself might induce behavioral abnormalities in the mutant mice. Here, to confirm the effects of Disc1-Q31L and Disc1-L100P mutations on behavioral phenotypes and to investigate the behaviors of the mutant mice in more detail, the mutant lines were backcrossed to C57BL/6JJcl through an additional two generations and the behaviors were analyzed using a comprehensive behavioral test battery.

Results: Contrary to expectations, 31 L mutant mice showed no significant behavioral differences when compared with wild-type control mice in any of the behavioral tests, including the Porsolt forced swim and tail suspension tests, commonly used tests for depression-like behavior. Also, 100P mutant mice exhibited no differences in almost all of the behavioral tests, including the prepulse inhibition test for measuring sensorimotor gating, which is known to be impaired in schizophrenia patients; however, 100P mutant mice showed higher locomotor activity compared with wild-type control mice in the light/dark transition test.

Conclusions: Although these results are partially consistent with the previous study in that there was hyperactivity in 100P mutant mice, the vast majority of the results are inconsistent with those of the previous study; this discrepancy may be explained by differences in the genetic background of the mice, the laboratory environment, experimental protocols, and more. Further behavioral studies under various experimental conditions are necessary to determine whether these Disc1 mutant mouse lines are suitable animal models of schizophrenia and major depression.

Figure 1

General health and neurological screen. (A) Body weight (g), (B) body temperature, (C) grip strength score, and (D) wire hang latency were recorded. 31 L and 100P mutant mice showed normal physical characteristics, except 31 L mutant mice had a lower grip strength score compared with wild-type control mice. Data represent the mean ± SEM. The p values indicate a genotype effect in a t-test.

Figure 2

Locomotor activity and anxiety-like behavior. (A-H) Open field test: (A, E) total distance traveled, (B, F) vertical activity, (C, G) time spent in the center area, and (D, H) stereotypic counts were recorded. In each strain, there were no significant differences in the behavioral indices of the open field test between the genotypes. (I-L) Light/dark transition test: (I) distance traveled in the light/dark compartments, (J) time spent in the light compartment, (K) latency to enter the light compartment, and (L) number of light/dark transitions were recorded. 100P mutants traveled a higher distance in the light compartment and showed a higher number of transitions compared with the wild-type mice. (M-P) Elevated plus maze test: (M) distance traveled, (N) number of arm entries, (O) percentage of time spent in open arms, and (P) percentage of entries into the open arms were calculated. No genotype effects were found in this test. Data represent the mean ± SEM. The p values indicate a genotype effect in a t-test.

Figure 3

Pain sensitivity and motor coordination/motor learning. (A) Latency (sec) to the first fore- or hind-paw response in the hot plate test was measured. The latencies of 31 L and 100P mutant mice were similar to those of wild-type mice. (B, C) Latency (sec) to fall in the rotarod test was recorded. In each strain, there were no significant differences in the latency to fall between mutant and wild-type mice. Data represent the mean ± SEM. The p values indicate a genotype effect in a t-test (A) or a two-way repeated measures ANOVA (B, C).

Figure 4

Sociability and social novelty preference. (A-E) Social interaction test in a novel environment: (A) total duration of contacts, (B) number of contacts, (C) total duration of active contacts, (D) mean duration of each contact, and (E) total distance traveled were analyzed. (F, G) Crawley's sociability and social novelty preference test: (F) time spent around the cage with stranger 1 or around the empty cage, and (G) time spent around the cage with stranger 1 and stranger 2 were measured. No significant differences in the behavioral indices between mutant and wild-type mice were found in each strain. Data represent the mean ± SEM. The p values indicate a genotype effect in a t-test.

Figure 5

Depression-like behavior and sensorimotor gating. (A, B) Porsolt forced swim test: (A) immobility time (sec) on day 1 and day 2 in 31 L mutant and control mice and (B) in 100P mutant and control mice were recorded. On day 1, 100P mutants showed a significantly higher immobility time. (C, D) Tail suspension test: (C) immobility time (sec) in 31 L mutant and control mice and also (D) immobility time (sec) in 100P mutant and control mice were recorded. No significant differences in immobility time were found between the genotypes in each strain. (E, F) Startle response/prepulse inhibition test: (E) amplitudes of the startle response and (F) percentage of prepulse inhibition were recorded; there were no significant genotype effects on the behavioral indices in each strain. Data represent the mean ± SEM. The p values indicate a genotype effect in a t-test (A-D) or a two-way repeated measures ANOVA (E, F).

Figure 6

Learning and memory. (A-D) T-maze test (forced alternation task): (A, B) percent correct responses in the training session and delay session in 31 L mutant and control mice and (C, D) in 100P mutant and control mice were calculated. (E-J) Contextual and cued fear conditioning test: (E, H) the percentage of freezing time in conditioning, (F, I) in the context testing, and (G, J) in the cued test was recorded. In each strain, no significant genotype effects on freezing time were found in any of the tests. Data represent the mean ± SEM. Data were analyzed using two-way repeated measures ANOVAs and the p values indicate a genotype effect.