Mice Mutated in the First Fibronectin Domain of Adhesion Molecule L1 Show Brain Malformations and Behavioral Abnormalities.

The X-chromosome-linked cell adhesion molecule L1 (L1CAM), a glycoprotein mainly expressed by neurons in the central and peripheral nervous systems, has been implicated in many neural processes, including neuronal migration and survival, neuritogenesis, synapse formation, synaptic plasticity and regeneration. L1 consists of extracellular, transmembrane and cytoplasmic domains. Proteolytic cleavage of L1's extracellular and transmembrane domains by different proteases generates several L1 fragments with different functions. We found that myelin basic protein (MBP) cleaves L1's extracellular domain, leading to enhanced neuritogenesis and neuronal survival in vitro. To investigate in vivo the importance of the MBP-generated 70 kDa fragment (L1-70), we generated mice with an arginine to alanine substitution at position 687 (L1/687), thereby disrupting L1's MBP cleavage site and obliterating L1-70. Young adult L1/687 males showed normal anxiety and circadian rhythm activities but enhanced locomotion, while females showed altered social interactions. Older L1/687 males were impaired in motor coordination. Furthermore, L1/687 male and female mice had a larger hippocampus, with more neurons in the dentate gyrus and more proliferating cells in the subgranular layer, while the thickness of the corpus callosum and the size of lateral ventricles were normal. In summary, subtle mutant morphological changes result in subtle behavioral changes.

CHL1 depletion affects dopamine receptor D2-dependent modulation of mouse behavior.

Introduction: The dopaminergic system plays a key role in the appropriate functioning of the central nervous system, where it is essential for emotional balance, arousal, reward, and motor control. The cell adhesion molecule close homolog of L1 (CHL1) contributes to dopaminergic system development, and CHL1 and the dopamine receptor D2 (D2R) are associated with mental disorders like schizophrenia, addiction, autism spectrum disorder and depression. Methods: Here, we investigated how the interplay between CHL1 and D2R affects the behavior of young adult male and female wild-type (CHL+/+) and CHL1-deficient (CHL1-/-) mice, when D2R agonist quinpirole and antagonist sulpiride are applied. Results: Low doses of quinpirole (0.02 mg/kg body weight) induced hypolocomotion of CHL1+/+ and CHL1-/- males and females, but led to a delayed response in CHL1-/- mice. Sulpiride (1 mg/kg body weight) affected locomotion of CHL1-/- females and social interaction of CHL1+/+ females as well as social interactions of CHL1-/- and CHL1+/+ males. Quinpirole increased novelty-seeking behavior of CHL1-/- males compared to CHL1+/+ males. Vehicle-treated CHL1-/- males and females showed enhanced working memory and reduced stress-related behavior. Discussion: We propose that CHL1 regulates D2R-dependent functions in vivo. Deficiency of CHL1 leads to abnormal locomotor activity and emotionality, and to sex-dependent behavioral differences.

Mice Mutated in the Third Fibronectin Domain of L1 Show Enhanced Hippocampal Neuronal Cell Death, Astrogliosis and Alterations in Behavior.

Adhesion molecules play major roles in cell proliferation, migration, survival, neurite outgrowth and synapse formation during nervous system development and in adulthood. The neural cell adhesion molecule L1 contributes to these functions during development and in synapse formation and synaptic plasticity after trauma in adulthood. Mutations of L1 in humans result in L1 syndrome, which is associated with mild-to-severe brain malformations and mental disabilities. Furthermore, mutations in the extracellular domain were shown to cause a severe phenotype more often than mutations in the intracellular domain. To explore the outcome of a mutation in the extracellular domain, we generated mice with disruption of the dibasic sequences RK and KR that localize to position 858RKHSKR863 in the third fibronectin type III domain of murine L1. These mice exhibit alterations in exploratory behavior and enhanced marble burying activity. Mutant mice display higher numbers of caspase 3-positive neurons, a reduced number of principle neurons in the hippocampus, and an enhanced number of glial cells. Experiments suggest that disruption of the dibasic sequence in L1 results in subtle impairments in brain structure and functions leading to obsessive-like behavior in males and reduced anxiety in females.

Mitochondrial and Neuronal Dysfunctions in L1 Mutant Mice.

Adhesion molecules regulate cell proliferation, migration, survival, neuritogenesis, synapse formation and synaptic plasticity during the nervous system's development and in the adult. Among such molecules, the neural cell adhesion molecule L1 contributes to these functions during development, and in synapse formation, synaptic plasticity and regeneration after trauma. Proteolytic cleavage of L1 by different proteases is essential for these functions. A proteolytic fragment of 70 kDa (abbreviated L1-70) comprising part of the extracellular domain and the transmembrane and intracellular domains was shown to interact with mitochondrial proteins and is suggested to be involved in mitochondrial functions. To further determine the role of L1-70 in mitochondria, we generated two lines of gene-edited mice expressing full-length L1, but no or only low levels of L1-70. We showed that in the absence of L1-70, mitochondria in cultured cerebellar neurons move more retrogradely and exhibit reduced mitochondrial membrane potential, impaired Complex I activity and lower ATP levels compared to wild-type littermates. Neither neuronal migration, neuronal survival nor neuritogenesis in these mutants were stimulated with a function-triggering L1 antibody or with small agonistic L1 mimetics. These results suggest that L1-70 is important for mitochondrial homeostasis and that its absence contributes to the L1 syndrome phenotypes.

Amelioration of the abnormal phenotype of a new L1 syndrome mouse mutation with L1 mimetics.

L1 syndrome is a rare developmental disorder characterized by hydrocephalus of varying severity, intellectual deficits, spasticity of the legs, and adducted thumbs. Therapy is limited to symptomatic relief. Numerous gene mutations in the L1 cell adhesion molecule (L1CAM, hereafter abbreviated L1) were identified in L1 syndrome patients, and those affecting the extracellular domain of this transmembrane type 1 glycoprotein show the most severe phenotypes. Previously analyzed rodent models of the L1 syndrome focused on L1-deficient animals or mouse mutants with abrogated cell surface expression of L1, making it difficult to test L1 function-triggering mimetic compounds with potential therapeutic value. To overcome this impasse, we generated a novel L1 syndrome mouse with a mutation of aspartic acid at position 201 in the extracellular part of L1 (p.D201N, hereafter termed L1-201) that displays a cell surface-exposed L1 accessible to the L1 mimetics. Behavioral assessment revealed an increased neurological deficit score and increased locomotor activity in male L1-201 mice carrying the mutation on the X-chromosome. Histological analyses of L1-201 mice showed features of the L1 syndrome, including enlarged ventricles and reduced size of the corpus callosum. Expression levels of L1-201 protein as well as extent of cell surface biotinylation and immunofluorescence labelling of cultured cerebellar neurons were normal. Importantly, treatment of these cultures with the L1 mimetic compounds duloxetine, crotamiton, and trimebutine rescued impaired cell migration and survival as well as neuritogenesis. Altogether, the novel L1 syndrome mouse model provides a first experimental proof-of-principle for the potential therapeutic value of L1 mimetic compounds.