Neural Cell Adhesion Molecule Deficiency Research Articles

Hyperfunction of the stress response system and novelty-induced hyperactivity correlate with enhanced cocaine-induced conditioned place preference in NCAM-deficient mice.

Several studies in humans and rodents suggest an association between impulsivity and activity of the stress response on the one hand and addiction vulnerability on the other. The neural cell adhesion molecule (NCAM) has been related to several neuropsychiatric disorders in humans. Constitutively NCAM-deficient (-/-) mice display enhanced novelty-induced behavior and hyperfunction of the hypothalamic-pituitary-adrenal axis. Here we hypothesize that NCAM deficiency causes an altered response to cocaine. Cocaine-induced behaviors of NCAM-/- mice and wild-type (+/+) littermates were analyzed in the conditioned place preference (CPP) test. c-fos mRNA levels were investigated by quantitative polymerase chain reaction (qPCR) to measure neural activation after exposure to the cocaine-associated context. NCAM-/- mice showed an elevated cocaine-induced sensitization, enhanced CPP, impaired extinction, and potentiated cocaine-induced hyperlocomotion and CPP after extinction. NCAM-/- showed no potentiated CPP as compared with NCAM+/+ littermates when a natural rewarding stimulus (ie, an unfamiliar female) was used, suggesting that the behavioral alterations of NCAM-/- mice observed in the CPP test are specific to the effects of cocaine. Activation of the prefrontal cortex and nucleus accumbens induced by the cocaine-associated context was enhanced in NCAM-/- compared with NCAM+/+ mice. Finally, cocaine-induced behavior correlated positively with novelty-induced behavior and plasma corticosterone levels in NCAM-/- mice and negatively with NCAM mRNA levels in the hippocampus and nucleus accumbens in wild-type mice. Our findings indicate that NCAM deficiency affects cocaine-induced CPP in mice and support the view that hyperfunction of the stress response system and reactivity to novelty predict the behavioral responses to cocaine.

Neural cell adhesion molecule regulates chondrocyte hypertrophy in chondrogenic differentiation and experimental osteoarthritis.

Chondrocyte hypertrophy‐like change is an important pathological process of osteoarthritis (OA), but the mechanism remains largely unknown. Neural cell adhesion molecule (NCAM) is highly expressed and involved in the chondrocyte differentiation of mesenchymal stem cells (MSCs). In this study, we found that NCAM deficiency accelerates chondrocyte hypertrophy in articular cartilage and growth plate of OA mice. NCAM deficiency leads to hypertrophic chondrocyte differentiation in both murine MSCs and chondrogenic cells, in which extracellular signal‐regulated kinase (ERK) signaling plays an important role. Moreover, NCAM expression is downregulated in an interleukin‐1β‐stimulated OA cellular model and monosodium iodoacetate‐induced OA rats. Overexpression of NCAM substantially inhibits hypertrophic differentiation in the OA cellular model. In conclusion, NCAM could inhibit hypertrophic chondrocyte differentiation of MSCs by inhibiting ERK signaling and reduce chondrocyte hypertrophy in experimental OA model, suggesting the potential utility of NCAM as a novel therapeutic target for alleviating chondrocyte hypertrophy of OA.

Neural Cell Adhesion Molecule Regulates Chondrocyte Hypertrophy in Chondrogenic Differentiation and Experimental Osteoarthritis

Chondrocyte hypertrophy-like change is an important pathological process of osteoarthritis (OA), but the mechanism remains largely unknown. Neural cell adhesion molecule (NCAM) is highly expressed and participate the chondrocyte differentiation of mesenchymal stem cells (MSCs). In this study, we found that NCAM deficiency leads to hypertrophic chondrocyte differentiation in murine MSCs, in which extracellular signal-regulated kinase (ERK) signaling play an important role. Moreover, in an interleukin (IL)-1β induced OA cellular model, NCAM expression is down-regulated. Interestingly, overexpression of NCAM substantially inhibits hypertrophic differentiation in the OA cellular model. Consistently, NCAM deficiency accelerates chondrocyte hypertrophy in growth-plate and articular cartilage of OA mice. In conclusion, NCAM could inhibit hypertrophic chondrocyte differentiation of MSCs by activating ERK signaling and reduce chondrocyte hypertrophy in experimental OA model, suggesting the potential utility of NCAM as a novel therapeutic target for alleviating chondrocyte hypertrophy of OA. Funding Statement: This work was supported by the grants from the National Natural Science Foundation of China (No. 81403161, U1704186, 81402416 and U1404816), Key Scientific Research Project of Colleges and Universities in Henan Province (No. 19A310020 and 16A180015), Henan Collaborative Innovation Center of Molecular Diagnosis and Laboratory Medicine (No. XTCX-2015-ZD3) and Scientific and Technological Research Project of Henan Province (No. 182102311136). Declaration of Interests: The authors declare no competing financial interest. Ethics Approval Statement: All animal experiments were approved by the Ethics Committee of Xinxiang Medical University.

NCAM-deficient mice show prominent abnormalities in serotonergic and BDNF systems in brain – Restoration by chronic amitriptyline

Mood disorders are associated with alterations in serotonergic system, deficient BDNF (brain-derived neurotrophic factor) signaling and abnormal synaptic plasticity. Increased degradation and reduced functions of NCAM (neural cell adhesion molecule) have recently been associated with depression and NCAM deficient mice show depression-related behavior and impaired learning. The aim of the present study was to investigate potential changes in serotonergic and BDNF systems in NCAM knock-out mice. Serotonergic nerve fiber density and SERT (serotonin transporter) protein levels were robustly reduced in the hippocampus, prefrontal cortex and basolateral amygdala of adult NCAM−/− mice. This SERT reduction was already evident during early postnatal development. [3H]MADAM binding experiments further demonstrated reduced availability of SERT in cell membranes of NCAM−/− mice. Moreover, the levels of serotonin and its major metabolite 5-HIAA were down regulated in the brains of NCAM−/− mice. NCAM−/− mice also showed a dramatic reduction in the BDNF protein levels in the hippocampus and prefrontal cortex. This BDNF deficiency was associated with reduced phosphorylation of its receptor TrkB. Importantly, chronic administration of antidepressant amitriptyline partially or completely restored these changes in serotonergic and BDNF systems, respectively. In conclusion, NCAM deficiency lead to prominent and persistent abnormalities in brain serotonergic and BDNF systems, which likely contributes to the behavioral and neurobiological phenotype of NCAM−/− mice.

The Neural Cell Adhesion Molecule Promotes Maturation of the Presynaptic Endocytotic Machinery by Switching Synaptic Vesicle Recycling from Adaptor Protein 3 (AP-3)- to AP-2-Dependent Mechanisms

Newly formed synapses undergo maturation during ontogenetic development via mechanisms that remain poorly understood. We show that maturation of the presynaptic endocytotic machinery in CNS neurons requires substitution of the adaptor protein 3 (AP-3) with AP-2 at the presynaptic plasma membrane. In mature synapses, AP-2 associates with the intracellular domain of the neural cell adhesion molecule (NCAM). NCAM promotes binding of AP-2 over binding of AP-3 to presynaptic membranes, thus favoring the substitution of AP-3 for AP-2 during formation of mature synapses. The presynaptic endocytotic machinery remains immature in adult NCAM-deficient (NCAM-/-) mice accumulating AP-3 instead of AP-2 and its partner protein AP180 in synaptic membranes and vesicles. NCAM deficiency or disruption of the NCAM/AP-2 complex in wild-type (NCAM+/+) neurons by overexpression of AP-2 binding-defective mutant NCAM interferes with efficient retrieval of the synaptic vesicle v-SNARE synaptobrevin 2. Abnormalities in synaptic vesicle endocytosis and recycling may thus contribute to neurological disorders associated with mutations in NCAM.

Neural cell adhesion molecule modulates dopaminergic signaling and behavior by regulating dopamine D2 receptor internalization.

The dopaminergic system plays an important role in the etiology of schizophrenia, and most antipsychotic drugs exert their functions by blocking dopamine D(2) receptors (D(2)Rs). Since the signaling strength mediated by D(2)Rs is regulated by internalization and degradation processes, it is crucial to identify molecules that modulate D(2)R localization at the cell surface. Here, we show that the neural cell adhesion molecule (NCAM) promotes D(2)R internalization/desensitization and subsequent degradation via direct interaction with a short peptide in the third intracellular loop of the D(2)R. NCAM deficiency in mice leads to increased numbers of D(2)Rs at the cell surface and augmented D(2)R signaling as a result of impaired D(2)R internalization. Furthermore, NCAM-deficient mice show higher sensitivity to the psychostimulant apomorphine and exaggerated activity of dopamine-related locomotor behavior. These results demonstrate that, in addition to its classical function in cell adhesion, NCAM is involved in regulating the trafficking of the neurotransmitter receptor D(2)R as well as receptor-mediated signaling and behavior, thus implicating NCAM as modulator of the dopaminergic system and a potential pharmacological target for dopamine-related neurological and psychiatric disorders.

NCAM-mimetic, FGL peptide, restores disrupted fibroblast growth factor receptor (FGFR) phosphorylation and FGFR mediated signaling in neural cell adhesion molecule (NCAM)-deficient mice

Neural cell adhesion molecule (NCAM) is a membrane-bound glycoprotein expressed on the surface of neuronal and glial cells. Previous in vitro studies have demonstrated that NCAM promotes neuronal functions largely via three main interaction partners: the fibroblast growth factor receptor (FGFR), a member of Src family of tyrosine kinases, Fyn and Raf1 kinase which all activate different intracellular signaling pathways. The objective was to clarify, which signaling pathways are being disrupted in NCAM knockout mice and whether FGL peptide is able to restore observed disruptions. Therefore we compared the levels of phosphorylation of FGFR1, Src kinase Fyn, Raf1 kinase, MAP kinases, Akt kinase and calcium/calmodulin-dependent kinases II and IV (CaMKII and CaMKIV) in the hippocampus of NCAM knockout mice to their wild-type littermates. The data of our study show that mice constitutively deficient in all isoforms of NCAM have decreased basal phosphorylation levels of FGFR1 and CaMKII and CaMKIV. Furthermore, NCAM-mimetic, FGL peptide, is found to be able to restore FGFR1, CaMKII and CaMKIV phosphorylation levels and thereby mimic the interactions of NCAM at this receptor in NCAM deficient mice. Also, we found that Fyn(Tyr530), Raf1, MAP kinases and Akt kinase phosphorylation in adult animals is not affected by NCAM deficiency but interestingly, we found an over-expression of another cell adhesion molecule L1. We conclude that in NCAM deficient mice FGFR1-dependent signaling is disrupted and it can be restored by FGL peptide.

Depression‐like behaviour in neural cell adhesion molecule (NCAM)‐deficient mice and its reversal by an NCAM‐derived peptide, FGL

The neural cell adhesion molecule (NCAM) plays a pivotal role in brain plasticity. Brain plasticity itself has a crucial role in the development of depression. The aim of this study was to analyze whether NCAM-deficient (NCAM(-/-)) mice exhibit depression-like behaviour and whether a peptide termed FGL, derived from the NCAM binding site for the fibroblast growth factor (FGF) receptor, is able to reverse the depression-like signs in NCAM(-/-) mice. Our study showed that NCAM(-/-) mice demonstrated increased freezing time in the tail-suspension test and reduced preference for sucrose consumption in the sucrose preference test, reduced adult neurogenesis in the dentate gyrus and reduced levels of the phosphorylated cAMP response element-binding protein (pCREB) in the hippocampus. FGL administered acutely or repeatedly reduced depression-like behaviour in NCAM(-/-) mice without having an effect on their wild-type littermates. Repeated administration of FGL enhanced survival of the newly born neurons in NCAM(-/-) mice and increased the levels of pCREB in both NCAM(+/+) and NCAM(-/-) mice. In conclusion, our data demonstrate that NCAM deficiency in mice results in a depression-like phenotype which can be reversed by the acute or repeated administration of FGL. The results also suggest a role of the deficit in NCAM signalling through the FGF receptor in depression.

Dysregulated CREB signaling pathway in the brain of neural cell adhesion molecule (NCAM)-deficient mice

The neural cell adhesion molecule (NCAM) mediates cell–cell interactions and plays an important role in processes associated with neural plasticity, including learning and memory formation. It has been shown that mice deficient in all isoforms of NCAM (NCAM−/− mice) demonstrate impairment in long-term plasticity at multiple hippocampal synapses, disrupted spatial learning, and impaired contextual and auditory-cued fear conditioning. The formation of long-term memory is associated with activation of transcription factor CREB (cAMP response element binding protein). The aims of this study were to investigate NCAM-mediated signaling transduction pathways and the levels of the phosphorylated (Ser133) active form of the CREB in the brain structures (the pre- and frontal cortex, basolateral amygdala, and hippocampus) involved in the memory formation in NCAM-deficient mice. Immunohistochemical analysis revealed reduced levels of pCREB in the prefrontal cortex (PFC), frontal cortex (FC), CA3 subregion of the hippocampus (CA3) and basolateral nucleus of amygdala (BLA) in NCAM−/− mice. NCAM−/− mice had also reduced levels of the phosphorylated CaMKII and CaMKIV in PFC/FC and the hippocampus, which are the downstream signaling molecules of NCAM. The levels of non-phosphorylated kinases did not differ from those seen in the wild-type mice. These results provide evidence that NCAM deficiency results in the dysregulation of CREB-mediated signaling pathways in the brain regions, which is related to the formation of memory.

Neural Cell Adhesion Molecule-Deficient β-Cell Tumorigenesis Results in Diminished Extracellular Matrix Molecule Expression and Tumour Cell-Matrix Adhesion

To understand by which mechanism neural cell adhesion molecule (N-CAM) limits β tumour cell disaggregation and dissemination, we searched for potential downstream genes of N-CAM during β tumour cell progression by gene expression profiling. Here, we show that N-CAM-deficient β-cell tumorigenesis is associated with changes in the expression of genes involved in cell-matrix adhesion and cytoskeletal dynamics, biological processes known to affect the invasive and metastatic behaviour of tumour cells. The extracellular matrix (ECM) molecules emerged as the primary target, i.e. N-CAM deficiency resulted in down-regulated mRNA expression of a broad range of ECM molecules. Consistent with this result, deficient deposition of major ECM stromal components, such as fibronectin, laminin 1 and collagen IV, was observed. Moreover, N-CAM-deficient tumour cells displayed defective matrix adhesion. These results offer a potential mechanism for tumour cell disaggregation during N-CAM-deficient β tumour cell progression. Prospective consequences of these findings for the role of N-CAM in β tumour cell dissemination are discussed.

Oxytocin neuron activation in NCAM‐deficient mice: anatomical and functional consequences

During stimulated neurosecretion in the rat, oxytocin neurons display a reduced glial coverage and receive an increased number of synapses, changes that are reversed on arrest of stimulation. We identified polysialic acid on the neural cell adhesion molecule (NCAM) as an important mediator of such plasticity. To investigate further the role of this cell surface glycoprotein, we examined the oxytocin system in mice genetically deficient in NCAM. First, ultrastructural analyses revealed that in wild-type mice, the supraoptic nucleus (SON) underwent the same remodelling as in the rat because oxytocin neurons had a diminished astrocytic coverage and increased synaptic input during lactation or chronic salt loading. Surprisingly, the SON displayed this morphology in NCAM-deficient mice as well, whether they were nongestating and hydrated, lactating or dehydrated. The oxytocin system in NCAM-deficient mice was abnormally hyperactive, as illustrated by enhanced plasma and intranuclear concentrations of oxytocin and reduced anxiety-related behaviour. Plasma oxytocin concentrations were also high in lactating NCAM-deficient dams but certain parameters of lactation and maternal behaviour were impaired. NCAM-deficient mice survived ingestion of 2% saline for 7 days and had increased plasma oxytocin but they did not cope with more severe osmotic challenges. Our observations highlight further the remarkable capacity of the adult oxytocin system to undergo neuronal and glial remodelling whenever it is activated. That lack of NCAM did not prevent remodelling indicates that NCAM can be substituted by other molecular mechanisms. Finally, while NCAM deficiency greatly enhanced oxytocin release, it led to impaired oxytocin-dependent physiological and behavioural responses.

Mutation in the Neural Cell Adhesion Molecule Interferes with the Differentiation of Anterior Pituitary Secretory Cells

The neural cell adhesion molecule (NCAM) and its polysialylated isoform (PSA-NCAM) have been shown to influence the proliferation, differentiation and survival of different cell types. Here, we report the pattern of expression of NCAM and PSA-NCAM in the anterior lobe (AL) of the pituitary gland of the adult mouse. We demonstrate that the majority of cells express NCAM, while PSA-NCAM is retained mostly on corticotropes. Analysis of bromodeoxyuridine (BrdU) incorporation shows that the presence of PSA-NCAM on corticotropes is not related to proliferation but most likely to their functional properties. We subsequently analyzed defects induced by NCAM deficiency in adult NCAM knockout mice. In these mice, all secretory cell types in the AL are present and their distribution within the gland is similar to that in wild-type mice. However, proliferation of AL cells is significantly increased. In particular, more BrdU-positive cells are detected among somatotropes and mammotropes in NCAM-deficient mice. In addition, the percentages of secretory cells are changed: somatotropes are more numerous while the number of corticotropes is reduced. These data demonstrate the involvement of NCAM in the proper generation and/or maintenance of the different cell populations in the AL and suggest the importance of PSA in corticotrope functioning.

Consequences of Neural Cell Adhesion Molecule Deficiency on Cell Migration in the Rostral Migratory Stream of the Mouse

In vertebrates, interneurons of the olfactory bulb (OB) are generated postnatally and throughout life at the subventricular zone of the forebrain. The neuronal precursors migrate tangentially through the forebrain using a well defined pathway, the rostral migratory stream (RMS), and a particular mode of migration in a chain-like organization. A severe size reduction of the OB represents the most striking morphological phenotype in neural cell adhesion molecule (NCAM)-deficient mice. This defect has been traced back to a migration deficit of the precursors in the RMS and linked to the lack of the polysialylated form of NCAM. In this study we investigate the morphological alterations and functional properties of the RMS in mice totally devoid of all isoforms of NCAM and polysialic acid (PSA). We show that a morphologically altered, but defined and continuous pathway exists in mutants, and we present in vivo and in vitro evidence that PSA-NCAM in the RMS is not essential for the formation and migration of chains. Instead, we find a massive gliosis associated with the formation of membrane specializations in a heterotypic manner, linking precursors to astrocytes. This finding and the over-representation and defasciculation of axons in the pathway suggest that important interactions between migrating cells and their stationary environment are perturbed in the mutants. Finally, we used transplantation experiments to demonstrate that lack of PSA-NCAM leads to a decrease but not a total blockade of migration and demonstrate that the mutant RMS is functional in transporting normal neuronal precursors to the OB.