Elsevier

Neurobiology of Disease

Volume 21, Issue 1, January 2006, Pages 217-227
Neurobiology of Disease

Hippocampal synaptic plasticity is impaired in the Mecp2-null mouse model of Rett syndrome

https://doi.org/10.1016/j.nbd.2005.07.005Get rights and content

Abstract

Rett syndrome is an X-linked neurodevelopmental disorder caused by mutations in the gene encoding the transcriptional repressor methyl-CpG-binding protein 2 (MeCP2). Here we demonstrate that the Mecp2-null mouse model of Rett syndrome shows an age-dependent impairment in hippocampal CA1 long-term potentiation induced by tetanic or theta-burst stimulation. Long-term depression induced by repetitive low-frequency stimulation is also absent in behaviorally symptomatic Mecp2-null mice. Immunoblot analyses from behaviorally symptomatic Mecp2-null mice reveal altered expression of N-methyl-d-aspartate receptor subunits NR2A and NR2B. Presynaptic function is also affected, as demonstrated by a significant reduction in paired-pulse facilitation. Interestingly, the properties of basal neurotransmission are normal in the Mecp2-null mice, consistent with our observations that the levels of expression of synaptic and cytoskeletal proteins, including glutamate receptor subunits GluR1 and GluR2, PSD95, synaptophysin-1, synaptobrevin-2, synaptotagmin-1, MAP2, βIII-tubulin and NF200, are not significantly altered. Together, these data provide the first evidence that the loss of Mecp2 expression is accompanied by age-dependent alterations in excitatory synaptic plasticity that are likely to contribute to the cognitive and functional deficits underlying Rett syndrome.

Introduction

Rett syndrome is an X-linked neurodevelopmental disorder and a leading genetic cause of severe mental retardation in girls, affecting approximately 1 in 10,000–20,000 female births worldwide (Hagberg et al., 1985, Kozinetz et al., 1993). First described in 1966 by an Austrian pediatrician, Andreas Rett, patients with Rett syndrome are born healthy and appear to develop normally for 6 to 18 months, achieving the usual motor, language and social milestones. Affected children undergo stereotyped patterns of neurological and behavioral regression with varying degrees of severity (Hagberg and Witt-Engerstrom, 1986); characteristic features of Rett syndrome include loss of purposeful hand use and communication skills, development of irregular breathing patterns and motor stereotypes including hand wringing and development in many patients of seizures and autistic features such as emotional withdrawal and anxiety.

The neuropathology of Rett syndrome points to arrested neuronal development rather than neurodegeneration or severe malformation of nervous tissue. Simplified neuronal branching patterns have been observed in the frontal, motor, and inferior temporal lobes of children (Armstrong, 2001). In addition, immunocytochemical studies have demonstrated reductions in dendritic branching and areas of reduced spine density, or “naked” dendrites in affected children (Belichenko et al., 1994). Together, these observations have led to the suggestion that there is an overall reduction in the number of synaptic inputs to neurons in the Rett brain. Consistent with the hypothesis that Rett syndrome is a disorder of abnormal neuronal maturation, additional studies of postmortem cortices of Rett syndrome patients found, in addition to a generalized reduction in dendritic arborization, a reduction in cholinergic markers and immature expression of the dendritic cytoskeletal proteins, including MAP2 (Kaufmann et al., 1995, Kaufmann et al., 1997, Kaufmann et al., 2000). Early autoradiographic studies on the expression of neurotransmitter receptors, including NMDA-(N-methyl-d-aspartate), AMPA-(α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid), kainate- and metabotropic-type glutamate receptors (GluRs) and GABA receptors reveal complex, age-related abnormalities in receptor densities in the frontal cortex and basal ganglia (Blue et al., 1999a, Blue et al., 1999b). AMPA and NMDA receptor densities in the young (<8 years old) Rett brain were significantly higher than controls, while in older Rett brain, receptor densities were dramatically reduced. While highly suggestive, it is also important to consider that these studies were limited by the relatively small number of human samples, and that the disturbances in excitatory neurotransmitter levels may be related to some of the clinical manifestations of Rett syndrome, such as seizures and abnormal EEG activity. Indeed, it remains entirely an open question whether the morphological abnormalities observed in postmortem human tissue are the cause or consequence of neurodevelopmental defects in Rett syndrome.

Mutations in the X-linked gene encoding the transcriptional repressor methyl-CpG-binding protein 2 (MeCP2) are responsible for a majority (up to 90%) of Rett syndrome cases (Amir et al., 1999). MeCP2 binds specifically to CpG-methylated DNA templates (Free et al., 2001) and is thought to inhibit gene transcription by recruiting co-repressor and histone deacetylase complexes and altering the organizational structure of genomic DNA (Nan et al., 1998). Genetically engineered mice that lack Mecp2, or express a truncated form of Mecp2 that very closely mimic a common mutation in humans, recapitulate many key features of the clinical Rett syndrome phenotype. Targeted Mecp2 gene deletion only in neurons leads to a full neurological phenotype, suggesting that the behavioral phenotype of Rett syndrome could be explained solely by MeCP2 dysfunction in neurons (Chen et al., 2001).

Since the very recent availability of the Mecp2-null mice, studies focused on molecular, anatomical or behavioral studies of the knockout mouse model have provided key insights to the consequences of the loss of Mecp2 expression, yet none have indicated what functional aspects of neurotransmission and synaptic plasticity are affected by such a loss. Indeed, given the widespread speculation or even the assumption that the molecular, biochemical or morphological changes would lead to alterations in synaptic transmission and plasticity, it is a critical missing piece in understanding the puzzle of Rett syndrome to elucidate what functional changes in fact result from the loss of Mecp2. In fact, neurons have been shown to be exquisitely capable of homeostatically compensating or regulating the efficacy of neuronal communication in the face of drastic changes in morphology or activity (for review, see Turrigiano and Nelson, 2004), and so, in the absence of electrophysiological experiments, it would not be straightforward to predict what changes in synaptic properties would result in the Mecp2 knockout brain.

Because excitatory neurotransmission and plasticity in the CA1 region of the rodent hippocampus has been extremely well studied due to historical interests in its potentially central role in learning and memory, it provides an invaluable general framework for examining and understanding the role of Mecp2 in glutamatergic synapses in virtually all central nervous system networks. Disruptions in activity-dependent functions of glutamatergic neurotransmission are likely to play a central role in the pathophysiology of epilepsy, movement disorders, brainstem mechanisms that control respiration, as well as in cognitive deficits—all present to varying degrees in patients with Rett syndrome. In the present study, we sought to determine whether the loss of Mecp2 gives rise to impairment of synaptic function and plasticity in the CA1 region of the hippocampus. We found age-dependent changes in the maintenance of both long-term potentiation and depression (LTP and LTD) consistent with our findings of altered expression of NMDA-type glutamate receptors in the Mecp2-null mouse model of Rett syndrome. In contrast to these changes, we found basal synaptic properties and markers to be normal. Thus, disruption of activity-dependent synaptic plasticity, but surprisingly not basal neurotransmission, may contribute to abnormal functions of neural circuits throughout the brain subserving the myriad of autonomic and motor functions that are so severely affected in Rett syndrome. Moreover, since the neuroanatomical deficits observed in Rett syndrome are shared by a host of other mental retardation syndromes with genetic bases, our observations on the functional consequences of morphological reorganization and neurodevelopmental arrest in Rett syndrome may provide useful insight or comparison for syndromes such as Coffin-Lowry, Rubenstein-Taybi, Fragile X and Down's syndromes.

Section snippets

Mecp2-null mice

We used two available Mecp2-null mouse models generated by the Cre LoxP recombination system to delete only exon 3 of Mecp2 (Chen et al., 2001) (obtained from the Mutant Mouse Resource Center at University of California, Davis) or to delete exons 3 and 4 of Mecp2 (Guy et al., 2001) (obtained from Jackson Laboratories, Bar Harbor, ME). Both lines were back-crossed for at least 8 generations to a C57Bl/6 background to eliminate potential strain-dependent differences. Statistical comparisons of

Altered hippocampal NMDA receptor expression in symptomatic Mecp2-null mice

Observations of significant alterations in the expression of NMDA- and AMPA-type glutamate receptors from postmortem brain samples from Rett syndrome patients (Blue et al., 1999a, Blue et al., 1999b), as well as elevations in CSF glutamate levels in Rett patients compared to autistic controls (Hamberger et al., 1992, Lappalainen and Riikonen, 1996) support the prevalent hypothesis that dysfunction of excitatory neurotransmission plays a central role in the pathophysiology of Rett syndrome.

Discussion

We have used the Mecp2-null mouse model to provide functional evidence in support of the hypothesis that Rett syndrome is a “clinical disorder of synaptic plasticity” (Johnston, 2004). Two principal findings on the functional consequences of Mecp2 loss emerge from our study. First, we demonstrate that NMDA-receptor-dependent LTP and LTD in the CA1 region of the symptomatic Mecp2-null mouse hippocampus are significantly attenuated compared to age-matched controls. Data presented here suggest

Acknowledgments

This work was supported by the Rett Syndrome Research Foundation and NIH (MH59800) grants to RMF, and grants to JHE from the Canadian Institutes of Health Research (MOP57765), and the Heart and Stroke Foundation of Canada (NA-5110). DGMJ is the recipient of an Epilepsy Canada postdoctoral fellowship. We thank members of the Fitzsimonds and Eubanks laboratories for their valuable comments on the manuscript, and Ms. M. Conraads for her insights and perseverance.

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    The first two authors contributed equally to this work.

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