Reorganization of inhibitory synapses and increased PSD length of perforated excitatory synapses in hippocampal area CA1 of dystrophin-deficient mdx mice.

Dystrophin is a cytoskeletal membrane-bound protein expressed in both muscle and brain. Brain dystrophin is thought to be involved in the stabilization of gamma-aminobutyric acid (GABA)(A)-receptor (GABA(A)-R)clusters in postsynaptic densities (PSDs) at inhibitory synapses onto pyramidal cells, and its loss has been linked to cognitive impairments in Duchenne muscular dystrophy. Dystrophin-deficient mdx mice have learning deficits and altered synaptic plasticity in cornu ammonis (CA1) hippocampus, but the possibility that altered synapse morphology or distribution may underlie these alterations has not been examined. Here we used in vivo magnetic resonance imaging and histological analyses to assess brain volumetric and cytoarchitectonic abnormalities and quantitative electron microscopy to evaluate the density and ultrastructure of CA1 hippocampal synapses in mdx mice. We found that mdx mice have increased density of axodendritic symmetric inhibitory synapses and larger PSDs in perforated asymmetric excitatory synapses in the proximal, but not distal, CA1 apical dendrites that normally express dystrophin, in the absence of gross brain malformations. Data are discussed in light of the known molecular and neurophysiological alterations in mdx mice. We suggest that increased inhibitory synapse density reflects tenuous compensation of altered clustering of alpha2 subunit-containing GABA(A)-Rs in CA1 dendrites, whereas increased PSD length in perforated synapses suggests secondary alterations in excitatory synapse organization associated with enhanced synaptic excitation.

Deletion of the Coffin-Lowry syndrome gene Rsk2 in mice is associated with impaired spatial learning and reduced control of exploratory behavior.

Coffin-Lowry Syndrome (CLS) is an X-linked syndromic form of mental retardation associated with skeletal abnormalities. It is caused by mutations of the Rsk2 gene, which encodes a growth factor regulated kinase. Gene deletion studies in mice have shown an essential role for the Rsk2 gene in osteoblast differentiation and function, establishing a causal link between Rsk2 deficiency and skeletal abnormalities of CLS. Although analyses in mice have revealed prominent expression of Rsk2 in brain structures that are essential for learning and memory, evidence at the behavioral level for an involvement of Rsk2 in cognitive function is still lacking. Here, we have examined Rsk2-deficient mice in two extensive batteries of behavioral tests, which were conducted independently in two laboratories in Zurich (Switzerland) and Orsay (France). Despite the known reduction of bone mass, all parameters of motor function were normal, confirming the suitability of Rsk2-deficient mice for behavioral testing. Rsk2-deficient mice showed a mild impairment of spatial working memory, delayed acquisition of a spatial reference memory task and long-term spatial memory deficits. In contrast, associative and recognition memory, as well as the habituation of exploratory activity were normal. Our studies also revealed mild signs of disinhibition in exploratory activity, as well as a difficulty to adapt to new test environments, which likely contributed to the learning impairments displayed by Rsk2-deficient mice. The observed behavioral changes are in line with observations made in other mouse models of human mental retardation and support a role of Rsk2 in cognitive functions.

Gene control of synaptic plasticity and memory formation: implications for diseases and therapeutic strategies.

There has been nearly a century of interest in the idea that information is stored in the brain as changes in the efficacy of synaptic connections between neurons that are activated during learning. The discovery and detailed report of the phenomenon generally known as long-term potentiation opened a new chapter in the study of synaptic plasticity in the vertebrate brain, and this form of synaptic plasticity has now become the dominant model in the search for the cellular and molecular bases of learning and memory. Accumulating evidence suggests that the rapid activation of the genetic machinery is a key mechanism underlying the enduring modification of neural networks required for the laying down of memory. Here we briefly review these mechanisms and illustrate with a few examples of animal models of neurological disorders how new knowledge about these mechanisms can provide valuable insights into identifying the mechanisms that go awry when memory is deficient, and how, in turn, characterisation of the dysfunctional mechanisms offers prospects to design and evaluate molecular and biobehavioural strategies for therapeutic prevention and rescue.

Spatial learning and synaptic hippocampal plasticity in type 2 somatostatin receptor knock-out mice.

Somatostatin is implicated in a number of physiological functions in the CNS. These effects are elicited through the activation of at least five receptor subtypes. Among them, sst2 receptors appear the most widely expressed in the cortex and hippocampal region. However, the specific role of this somatostatin receptor subtype in these regions is largely undetermined. In this study, we investigated the role of the sst2 receptor in the hippocampus using mice invalidated for the sst2 gene (sst2 KO mice). Complementary experimental approaches were used. First, mice were tested in behavioral tests to explore the consequences of the gene deletion on learning and memory. Spatial discrimination learning in the radial maze was facilitated in sst2 KO mice, while operant learning of a bar-pressing task was slightly altered. Mice were then processed for electrophysiological study using the ex vivo hippocampal slice preparation. Extracellular recordings in the CA1 area showed an enhancement in glutamatergic (AMPA and NMDA) responses in sst2 KO mice which displayed an increase in the magnitude of the short-term potentiation and long-term depression. In contrast, long-term potentiation was not significantly altered. Taken together, these data demonstrate that somatostatin, acting via sst2 hippocampal receptors, may contribute to a global decrease in glutamate efficiency and consequently alter glutamate-dependent plasticity and spatial learning.

Involvement of sst2 somatostatin receptor in locomotor, exploratory activity and emotional reactivity in mice.

Somatostatin (SRIF) controls many physiological and pathological processes in the central nervous system but the respective roles of the five receptor isotypes (sst1-5) that mediate its effects are yet to be defined. In the present study, we attempted to identify functions of the sst2 receptor using mice with no functional copy of this gene (sst2 KO mice). In contrast with control 129Sv/C57Bl6 mice, sst2 mRNA was no longer detectable in the brain of sst2 KO mice; 125I-labeled Tyr0DTrp8-SRIF14 binding was also greatly reduced in almost all brain structures except for the hippocampal CA1 area, demonstrating that sst2 accounts for most SRIF binding in mouse brain. Invalidation of this subtype generated an increased anxiety-related behaviour in a number of behavioural paradigms, while locomotor and exploratory activity was decreased in stress-inducing situations. No major motor defects could be detected. sst2 KO mice also displayed increased release of pituitary ACTH, a main regulator of the stress response. Thus, somatostatin, via sst2 receptor isotype pathways, appears involved in the modulation of locomotor, exploratory and emotional reactivity in mice.

Behavioral characterization of mdx3cv mice deficient in C-terminal dystrophins.

Cognitive deficits are frequently associated with Duchenne muscular dystrophy (DMD). They might be due to a deficiency in the brain isoforms of the 427 kDa full-length dystrophin, and/or to altered expression of other C-terminal dystrophin-gene products (Dp71, Dp140) also found in brain. Mdx mice, which only lack full-length dystrophin in both muscle and brain, were previously shown to have moderate learning and memory deficits. In the present study, we investigated behavioral responses in mdx3cv mutants, which have altered expression of all the dystrophin-gene products. Contrary to the original mdx mice, mdx3cv mice showed enhanced anxiety-related behaviors and reduced locomotion as compared to control mice. Although those perturbations might be related to the lack in C-terminal dystrophins, they do not seem sufficient to induce strong learning deficits in this mutant. Indeed, we showed that mdx3cv mice may display similar or weaker deficits during the learning of a bar-pressing task, as compared to mdx mice. The relevance of the mdx3cv mutant as a model to study the cognitive deficits associated with DMD is discussed.

Facilitated NMDA receptor-mediated synaptic plasticity in the hippocampal CA1 area of dystrophin-deficient mice.

The contribution of the cytoskeletal membrane-associated protein dystrophin in glutamatergic transmission and related plasticity was investigated in the hippocampal CA1 area of wild-type and dystrophin-deficient (mdx) mice, using extracellular recordings in the ex vivo slice preparation. Presynaptic fiber volleys and field excitatory postsynaptic potentials (fEPSPs) mediated through N-methyl-D-Aspartate receptors (NMDAr) or non-NMDAr were compared in both strains. Comparable synaptic responses were observed in wild-type and mdx mice, suggesting that basal glutamatergic transmission is not altered in the mutants. By contrast, the synaptic strengthening induced by a conditioning stimulation of either 10, 30, or 100 Hz was significantly greater in mdx mice during the first minutes posttetanus. Because the posttetanic potentiation induced in the presence of the NMDAr antagonist D-APV was not affected in the mutants, a critical role of NMDAr in this increase was suggested. The magnitude of the potentiation induced by a 30 Hz stimulation in mdx mice was normalized as compared to wild-type mice by increasing the extracellular magnesium concentration from 1.5 to 3 mM. Moreover, the transitory depression of fEPSPs induced by bath-applied NMDA (50 microM for 30s) was more sensitive to an increased extracellular magnesium concentration in wild-type than in mdx mice. Our results suggest that the absence of dystrophin may facilitate NMDAr activation in the CA1 hippocampal subfield of mdx mice, which may be partly due to a reduction of the voltage-dependent block of this receptor by magnesium.

Spatial discrimination learning and CA1 hippocampal synaptic plasticity in mdx and mdx3cv mice lacking dystrophin gene products.

Duchenne muscular dystrophy is frequently associated with a non-progressive cognitive deficit attributed to the absence of 427,000 mol. wt brain dystrophin, or to altered expression of other C-terminal products of this protein, Dp71 and/or Dp140. To further explore the role of these membrane cytoskeleton-associated proteins in brain function, we studied spatial learning and ex vivo synaptic plasticity in the mdx mouse, which lacks 427,000 mol. wt dystrophin, and in the mdx3cv mutant, which shows a dramatically reduced expression of all the dystrophin gene products known so far. We show that reference and working memories are largely unimpaired in the two mutant mice performing a spatial discrimination task in a radial maze. However, mdx3cv mice showed enhanced emotional reactivity and developed different strategies in learning the task, as compared to control mice. We also showed that both mutants display apparently normal levels of long-term potentiation and paired-pulse facilitation in the CA1 field of the hippocampus. On the other hand, an increased post-tetanic potentiation was shown by mdx, but not mdx3cv mice, which might be linked to calcium-regulatory defects. Otherwise, immunoblot analyses suggested an increased expression of a 400,000 mol. wt protein in brain extracts from both mdx and mdx3cv mice, but not in those from control mice. This protein might correspond to the dystrophin-homologue utrophin. The present results suggest that altered expression of dystrophin or C-terminal dystrophin proteins in brain did not markedly affect hippocampus-dependent spatial learning and CA1 hippocampal long-term potentiation in mdx and mdx3cv mice. The role of these membrane cytoskeleton-associated proteins in normal brain function and pathology remains to be elucidated. Furthermore, the possibility that redundant mechanisms could partially compensate for dystrophins' deficiency in the mdx and mdx3cv models should be further considered.

Influence of dystrophin-gene mutation on mdx mouse behavior. I. Retention deficits at long delays in spontaneous alternation and bar-pressing tasks.

X-linked Duchenne muscular dystrophy (DMD) is frequently associated with a nonprogressive, cognitive defect attributed to the absence of dystrophin in the brain of DMD patients. The mutant mdx mouse, lacking in 427-kDa dystrophin in both muscle and brain tissues, is considered to be a valuable model of human DMD. In the present study, we compared mdx and C57BL/10 control mice and showed that mdx mice had impaired retention in a T-maze, delayed spontaneous alternation task 24 h, but not 6 h, after acquisition. mdx mice were not impaired in acquisition of a bar-pressing task on 4 consecutive days but showed poor retention 22 days after the last training session. Mutants and controls showed similar behavioral responses in free exploration and light/dark choice situations and did not differ in spontaneous locomotor activity or motor coordination. Retention impairments at long delays in mdx mice suggest a role of dystrophin in long-term consolidation processes.