Fragile X Syndrome (FXS) in Mice

Fragile X syndrome (FXS) is rare, orphan disease caused by a lack of the fragile X mental retardation protein (FMRP).  An expanded trinucleotide repeat on the long (q) arm of the X chromosome was identified as the genetic cause of FXS. The gene, called fragile X mental retardation 1 (FMR1), contains a CGG repeat in its 5’ untranslated region that is normally shorter than 55 repeats. However, this repeat can become unstable upon maternal transmission, resulting in the expansion of the repeat in the next generation. If the CGG repeat reaches >200 repeats, the promoter region will be hypermethylated and the gene will be silenced, resulting in the lack of FMRP expression. FMRP is an important RNA binding protein and plays a role in repressing translation [1]. Therefore, loss of FMRP results in increased levels of protein synthesis.

FMRP deficiency in patients with FXS causes intellectual disability (IQ <70) and several behavioral problems, including hyperactivity, poor social skills and repetitive behaviors. Because the gene causing FXS is located on the X chromosome, males are more likely to be affected (1 in 4000 males vs. 1 in 8000 females [2]). A simple DNA blood test can detect carriers and fully affected individuals.  Although the cause of FXS has been known for decades, no cure exists. When children are diagnosed at an early age, several behavioral, educational and family therapies may reduce some symptoms.

Gross morphological changes have not been identified in the brains of FXS patients. However specific neural systems are altered in fragile X syndrome. Subtle abnormalities in the dendritic spines of neurons have been reported [3-5], and an imbalanced network function between the excitatory glutamatergic and inhibitory GABAergic neurotransmitter systems has been observed [6-8]. Attempts to improve FXS-related symptoms have involved correcting the reduced GABAergic activity or dampening the glutamatergic hyperactivity.  Unfortunately, these approaches have met limited success.

Induction:

The FXS mouse model was developed by deleting the murine version of fragile X mental retardation protein, Fmr1 [9]. Most of the behavioral phenotypes exhibited by Fmr1 knockout mice are similar to those in human FXS patients. These phenotypes include hyperactivity in the open field arena, reduced anxiety in the elevated plus maze, and reductions in learning, memory and sociability [6].

An example study would involve 12 male Fmr1 KO mice and 12 male WT C57Bl/6 mice (wildtype littermates are not commercially available). After arrival at the facility at the age of 8 weeks, the animals are habituated to the facility for another 2 weeks.  Treatments could begin during this habituation period.

Disease parameters & clinical assessment:

After habituation, the animals can be tested in any combination of the following behavioral assays:

  1. Open Field arena (OFA): spontaneous locomotor activity and anxiety
  2. Morris water maze (MWM): learning, memory and cognitive function
  3. Contextual (and cued) fear conditioning: learning (aversive stimulus) with context
  4. Nestlet shredding/nest building: homecage behavior for anxiety and distress
  5. Marble burying: homecage assessment of perseverative and repetitive behaviors
  6. Light-Dark box (LDB): anhedonia and anxiety
  7. Elevated Plus Maze (EPM): anxiety

Optional Endpoints: 

  • GABAA subtype receptor expression
  • GSK3β/AKT phosphorylation and expression

References

  1. Santoro, M.R., S.M. Bray, and S.T. Warren, Molecular mechanisms of fragile X syndrome: a twenty-year perspective. Annu Rev Pathol, 2012. 7: p. 219-45.
  2. Levenga, J., et al., Potential therapeutic interventions for fragile X syndrome. Trends Mol Med, 2010. 16(11): p. 516-27.
  3. de Vrij, F.M., et al., Rescue of behavioral phenotype and neuronal protrusion morphology in Fmr1 KO mice.Neurobiol Dis, 2008. 31(1): p. 127-32.
  4. Levenga, J., et al., Subregion-specific dendritic spine abnormalities in the hippocampus of Fmr1 KO mice.Neurobiol Learn Mem, 2011. 95(4): p. 467-72.
  5. Hall, S.S., et al., Identifying large-scale brain networks in fragile X syndrome. JAMA Psychiatry, 2013. 70(11): p. 1215-23.
  6. Bear, M.F., K.M. Huber, and S.T. Warren, The mGluR theory of fragile X mental retardation. Trends Neurosci, 2004. 27(7): p. 370-7.
  7. D’Hulst, C. and R.F. Kooy, The GABAA receptor: a novel target for treatment of fragile X? Trends Neurosci, 2007. 30(8): p. 425-31.
  8. Kazdoba, T.M., et al., Modeling fragile X syndrome in the Fmr1 knockout mouse. Intractable Rare Dis Res, 2014. 3(4): p. 118-33.
  9. Fmr1 knockout mice: a model to study fragile X mental retardation. The Dutch-Belgian Fragile X Consortium.Cell, 1994. 78(1): p. 23-33.

 

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