Behavioral and neurochemical alterations in mice lacking the RNA-binding protein translin

Abstract

Synapse-specific local protein synthesis is thought to be important for neurodevelopment and plasticity and involves neuronal RNA-binding proteins that regulate the transport and translation of dendritically localized transcripts. The best characterized of these RNA-binding proteins is the fragile X mental retardation protein (FMRP). Mutations affecting the expression or function of FMRP cause fragile X syndrome in humans, and targeted deletion of the gene encoding FMRP results in developmental and behavioral alterations in mice. Translin is an RNA-binding protein that regulates mRNA transport and translation in mouse male germ cells and is proposed to play a similar role in neurons. Like FMRP, translin is present in neuronal dendrites, binds dendritically localized RNA, and associates with microtubules and motor proteins. We reported previously the production of viable homozygous translin knock-out mice, which demonstrate altered expression of multiple mRNA transcripts in the brain and mild motor impairments. Here, we report that translin knock-out mice also exhibit sex-specific differences in tests of learning and memory, locomotor activity, anxiety-related behavior, and sensorimotor gating, as well as handling-induced seizures and alterations in monoamine neurotransmitter levels in several forebrain regions. Similar behavioral and neurochemical alterations have been observed in mice lacking FMRP, suggesting that both proteins may act within the same neuronal systems and signaling pathways. Our results in mice indicate that mutations in translin may contribute to fragile X-like syndromes, mental retardation, attention deficit hyperactivity disorder, epilepsy, and autism spectrum disorders in humans.

Figure 1.

Male translin KO mice exhibit increased escape latencies during training in the Morris water maze, but spatial memory as assessed by probe trials is not impaired. translin KO mice (n = 13, 7 males and 6 females) and WT littermates (n = 17, 9 males and 8 females) were placed in a pool of water made opaque with white paint and trained using four trials per day over multiple days to find a submerged platform at an initial location (Acquisition; days 1–6), at a new location on the opposite side of the pool (Reversal; days 1–3), and at varying locations identified with a visible marker extending from the surface of the pool (Visible; days 1–4). Probe trials in which the platform was removed and mice were allowed to search the pool for 60 s were performed after acquisition and reversal training. A, Male translin KO mice exhibited significantly longer escape latencies than WT littermate controls during acquisition, reversal, and visible platform training. B, Escape latency did not differ significantly between female translin KO mice and WT controls. C, After 6 d of training with the platform in its initial location, translin KO mice and WT littermate controls of both sexes spent a significantly greater percentage of time in the quadrant that previously contained the hidden platform (target) relative to the other quadrants. D, After 3 d of training with the platform in a new location, all groups showed a significant preference for the new target quadrant. Female translin KO mice spent a significantly greater percentage of time in the target quadrant than male KO mice and WT controls. *p < 0.05. Opp, Opposite; Adj, Adjacent.

Figure 2.

Increased escape latency in translin KO mice in the Morris water maze may be related to differences in swim speed, thigmotaxis, or jumping behavior. Swim speed, thigmotaxis, and jumping behavior of translin KO mice (n = 13, 7 males and 6 females) and WT littermates (n = 17, 9 males and 8 females) were recorded throughout acquisition, reversal, and visible platform training in the Morris water maze. A, Relative to male WT controls, the average swim speed of male translin KO mice was significantly greater during hidden platform training and slower during visible platform training. B, Although there was a significant difference in average speed between female translin KO mice and WT littermates overall, post hoc tests did not reveal significant differences on any given day. C, Swimming around the periphery of the pool, a behavior termed thigmotaxis, occurred in a significantly higher proportion of male translin KO mice than male WT littermates at the end of acquisition and during visible platform training. D, Female translin KO mice also exhibited significantly more thigmotaxis during acquisition but not during reversal or visible platform training. E, F, After locating the hidden platform during a given trial, mice are allowed to sit on it for 20 s before being returned to the home cage, but they occasionally jump back into the pool after finding the platform. translin KO mice of both sexes exhibited more of this jumping behavior than WT littermates immediately after finding the platform during acquisition and reversal training (E) and when we attempted to retrieve them from the platform throughout training (F). *p < 0.05; **p < 0.01.

Figure 3.

Female translin KO mice exhibit significantly enhanced freezing after contextual and cued fear conditioning. translin KO mice (n = 23, 13 males and 10 females) and WT littermates (n = 23, 12 males and 11 females) were placed in conditioning chambers in which they received an auditory CS (30 s of 85 dB white noise) coterminating with an US (2 s, 1.5 mA footshock). Freezing behavior was assessed in the chamber before conditioning (Baseline; 2 min), during the period of CS and US presentation (CS–US; 30 s), immediately after conditioning (Immed; 30 s), 24 h later in the same chamber (Context; 5 min), and in a new chamber representing a different context both before (Pre-CS; 2 min) and during (Cued; 3 min) CS presentation. A, All groups exhibited significant freezing responses relative to baseline when reexposed to the context or the CS 24 h after conditioning, but female translin KO mice showed significantly more freezing than female WT mice and males of either genotype. Male translin KO mice demonstrated significantly enhanced freezing immediately after training. B, In a separate experiment with a new group of translin KO mice (n = 18, 5 males and 13 females) and WT littermates (n = 31, 17 males and 14 females), female translin KO mice also exhibited enhanced freezing during presentation of the CS 2 h after conditioning. C, Shock sensitivity, as assessed by the average current level required to produce flinching, vocalizations, or jumping behavior, did not differ significantly between translin KO mice (n = 14, 8 males and 6 females) and WT littermates (n = 17, 8 males and 9 females). *p < 0.05; **p < 0.01.

Figure 4.

translin KO mice exhibit reduced anxiety-related behavior in the elevated zero maze test. translin KO mice (n = 22, 13 males and 9 females) and WT littermates (n = 22, 14 males and 8 females) were placed in the center of a closed quadrant of the elevated zero maze and observed for 5 min. A, Both male and female translin KO mice spent a significantly greater percentage of time in the open quadrants of the zero maze than WT littermates. B, The number of entries into the open quadrants with the head or forepaws did not differ significantly between genotypes, but, of these entries, male translin KO mice ventured completely into the open significantly more often. C, Both male and female translin KO mice traveled significantly farther in the open but not the closed quadrants of the zero maze. D, The average speed of male translin KO mice was significantly greater than WT controls in both the open and closed quadrants. E, Episodes of stretch-attend posture, a stereotyped behavior in which mice tentatively sample the environment by extending their snout and forepaws while keeping their hindpaws in place, were significantly reduced in translin KO mice. F, The number of head dips, an exploratory behavior in which mice extend their snout past the inside or outside edge of the open quadrants, was significantly greater in male mice overall and in male translin KO mice relative to male WT littermates. *p < 0.05; ** p < 0.01; *** p < 0.001.

Figure 5.

translin KO mice exhibit decreased anxiety-related behavior in the light–dark exploration test and normal baseline locomotor activity in the open field. A, B, translin KO mice (n = 15, 9 males and 6 females) and WT littermates (n = 20, 14 males and 6 females) were placed in the center of the open compartment of the light–dark box apparatus and observed for 10 min. A, Male mice spent a greater percentage of time in the open compartment than female mice, and, accounting for sex differences, translin KO mice spent significantly more time in the open than their WT littermates. B, The latency to enter the dark compartment was also significantly longer in translin KO mice, accounting for differences attributable to sex. C, D, translin KO mice (n = 33, 26 males and 7 females) and WT littermates (n = 43, 34 males and 9 females) were placed in the center of the open-field arena, and activity was recorded over 15 min by arrays of photobeams. C, Locomotor activity did not differ significantly between translin KO mice and WT littermates. D, Anxiety-related behavior, as assessed by the percentage of total open-field activity that occurred in the central area of the arena, did not differ significantly between genotypes. *p < 0.05 for WT mice versus KO mice.

Figure 6.

Acoustic startle responses are significantly reduced and prepulse inhibition of startle is significantly increased in translin KO mice. A, Male translin KO mice (n = 16) exhibit significantly reduced whole-body startle amplitudes relative to male WT littermates (n = 20) across a range of stimulus intensities from 100 to 120 dB. Startle responses of female translin KO mice (n = 9) did not differ significantly from female WT littermates (n = 7) or male KO mice. B, translin KO mice exhibit significantly enhanced prepulse inhibition of the startle response relative to WT littermates. There was a significant difference in percentage of prepulse inhibition between male translin KO mice and WT littermates at prepulse intensities of 72, 76, and 84. *p < 0.05, **p < 0.01, ***p < 0.001 for male WT mice versus male translin KO mice.

Figure 7.

translin KO mice exhibit alterations in the level of monoamine neurotransmitters in the forebrain. HPLC was used to determine the concentration of the monoamine neurotransmitters NE, 5-HT, DA, and their metabolites in multiple brain regions in translin KO mice (n = 12, 7 males and 5 females) and WT littermate controls (n = 14, 7 males and 7 females). A, NE is significantly reduced in the hippocampus and cortex in male translin KO mice and in the amygdala, hippocampus, and cortex in female translin KO mice. B, 5-HT levels are significantly decreased in the hippocampus, cortex, and striatum in male translin KO mice and in the amygdala in female translin KO mice. C, Levels of DA in the amygdala do not differ between genotypes, but levels of the metabolites DOPAC and HVA are significantly increased in female translin KO mice. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.