8 May 2007. In the May 3 Neuron, Steven Clapcote, David Porteous, and their colleagues report that simple missense mutations in the DISC1 gene lead to impaired behavioral function in mice. The results strengthen the case that variations in the gene play a role in complex neuropsychiatric diseases such as affective disorders and schizophrenia (see SRF live discussion).
DISC1, or disrupted in schizophrenia, was originally discovered by the Porteous group, at the University of Edinburgh, Scotland, when they found that a gross rearrangement of the gene associates with schizophrenia and affective disorders in a large Scottish family (see Millar et al., 2000). Since then, numerous studies have linked DISC1 polymorphisms with the disease in various populations (see SchizophreniaGene data); however, a robust cause-and-effect association has not emerged (see SchizophreniaGene meta-analysis). But this latest finding indicates that in mice, at least, DISC1 mutation is sufficient to cause some of the signs, or endophenotypes, commonly exhibited by people with the disease.
To find DISC1 mutations that might have a neurological impact, first author Clapcote, and colleagues in John Roder's group at Mount Sinai Hospital, Toronto, and elsewhere, screened progeny of mice that had been treated with the chemical mutagen N-ethyl-N-nitrosourea (ENU), which primarily causes simple DNA point mutations. The researchers identified the DISC1 mutations by sequencing exon 2 of the gene in the nearly 1,700 offspring. They focused on exon 2 because it is the largest DISC1 exon and it encodes the head domain of the protein, which interacts with phosphodiesterase 4B, a cyclic AMP degrading enzyme that the Porteous group has also linked to schizophrenia pathology (see SRF related news story). To ensure that other ENU-introduced mutations did not complicate the picture, the researchers back-crossed each mutant strain with the parental strain for five generations, which, according to the authors, should reduce the expected number of additional heterozygous mutations in each mutant line to 0.75.
To screen the mutant mice for neuropsychiatric changes, the authors tested them for prepulse and latent inhibition, two behaviors that are often compromised in patients with schizophrenia. In the former, startle responses elicited by a loud noise are suppressed by a prior stimulus that is insufficient to elicit the startle response. In latent inhibition, pre-exposure to a non-rewarding stimulus reduces subsequent interest in the stimulus even when it is later paired with a reward. Impairments in both are thought to be related to difficulties processing sensory information.
Clapcote and colleagues found that mice with either of two mutations—a leucine for a glutamine at position 31 (Q31L) and a proline for a leucine at position 100 (L100P)—were impaired in both prepulse (PPI) and latent inhibition (LI). The L100P mutation seemed to have a slightly stronger effect, with prepulse inhibiting the startle response in homozygous mutants only about 20 percent of the time, as opposed to 38 percent in Q31L homozygotes and about 60 percent in wild-type mice. Similarly, latent inhibition in L100P animals was about half that in Q31L mice, which in turn was only about one-third that in control mice.
In other behavioral tests, however, differences between the two mutants were not simply a matter of degree. The L100P animals appeared more anxious, moving around much more in an open field test, whereas the Q31L animals were not significantly different from wild-type. These animals also performed poorly in a T-maze test of working memory—both anxiety and impaired working memory are symptoms of schizophrenia. In contrast, the Q31L animals showed experimental behaviors believed to model severe depression. In a forced swim test they gave up more readily, floating for longer than controls and L100P animals, and they showed little interest in sucrose solution or strange mice introduced in a social interaction test.
The authors suggest that the Q31L mutation results in depressive-like behavior, while the L100P mutation exhibited schizophrenic-like behavior. In support of this dichotomy they found that while the antipsychotic drug clozapine restored PPI and LI somewhat in L100P animals, it had no effect on these parameters in Q31L mice. In the latter, PPI and LI impairments were relieved by the antidepressant bupropion, however.
How do these mutations cause such dramatic behavioral problems? The researchers found that both types of mice had significantly reduced brain volume—13 percent loss in the case of L100P and 6 percent for Q31L. These reductions were accompanied by contraction of the cortex, entorhinal cortex, thalamus, and cerebellum. These findings suggest that the mutation causes neurodevelopmental effects, an idea that has been previously postulated. But the researchers also found that levels of four major DISC1 isoforms in the brain appeared normal in the mutant animals, which suggests that the mutations do not affect gene expression but possibly alter activity of the protein.
Since Porteous's group previously showed that DISC1 binds with phosphodiesterase 4B (PDE4B), which may play a key role in regulating working memory (see SRF related news story), they chose to look at this interaction more closely. They found that both mutations reduced DISC1 binding to PDE4B when the proteins were expressed in cultured cells. However, they note that “the degree of binding was variable, particularly with the 31L mutation, suggesting that mutant DISC1 binding to PDE4B is influenced by the effects of unknown fluctuating cellular factors.” Nonetheless, the loss of PDE4B binding suggests that these mutations may impact cAMP signaling in addition to impairing neurodevelopment. “These mouse models support both the neurodevelopmental role for DISC1 and the proposed cAMP signaling role through modulation of PDE4 activity,” write the authors. Indeed, the Q31L mutants also had lower PDE4 activity and, unlike the L100P animals, the PDE4 rolipram had no effect on their PPI or LI.
When Is a Mouse a Model?
Indeed, this variability speaks to the very crux of the DISC1 matter. A clear picture of DISC1 biology and its influence on psychiatric illness has not emerged, possibly because DISC1 variations are responsible for many different phenotypes and are influenced by other genetic or environmental factors. “Our results in the mouse thus emphasize the importance of replicating, validating, and resolving inconsistencies within the current picture of various DISC1 alleles and haplotypes associating with distinct clinical phenotypes and, indeed, normal variation in cognitive function and neurodevelopment,” write the authors.
How valuable mouse models will be for complex human neuropsychiatric disorders remains to be seen. Mice, for example, do not have a prefrontal cortex, a region in the human brain that has been linked to psychotic behavior. And as Nancy Low, McGill University, Montreal, Quebec, and John Hardy, National Institute on Aging, Bethesda, Maryland, write in an accompanying Neuron preview, “Because psychiatric disorders are composed of lists of symptoms, many of which are nonspecific and overlap with those of other psychiatric disorders, a mouse model may be able to mimic some observable behaviors, but the attribution of a specific behavior to a human emotion or cognitive state cannot be made.”
Nonetheless, Low and Hardy suggest that, “These data are generally consistent with the view that genetic modification of the DISC1 locus leads to behavioral outcomes that may correspond to traits or abnormalities observed in psychiatric disorders.” However, they also urge caution in interpreting the data, given that the mutations “have complex biochemical, anatomical and behavioral effects.” They also conclude that “Mouse models, such as these, may be a way forward, but we must use them cautiously and be wary not to only study mice and lose sight of the human condition.”—Tom Fagan.
Clapcote SJ, Lipina TV, Millar JK, Mackie S, Christie S, Ogawa F, Lerch JP, Trimble K, Uchiyama M, Sakuraba Y, Kaneda H, Shiroishi T, Houslay MD, Henkelman RM, Sled JG, Gondo Y, Porteous DJ, Roder JC. Behavioral phenotypes of Disc1 missense mutations in mice. Neuron. 2007, May 3;54:387-402. Abstract
Low NC, Hardy J. What is a schizophrenic mouse? Neuron. 2007, May3;54:348-349. Abstract