Reelin differentially shapes dendrite morphology of medial entorhinal cortical ocean and island cells.

The function of medial entorhinal cortex layer II (MECII) excitatory neurons has been recently explored. MECII dysfunction underlies deficits in spatial navigation and working memory. MECII neurons comprise two major excitatory neuronal populations, pyramidal island and stellate ocean cells, in addition to the inhibitory interneurons. Ocean cells express reelin and surround clusters of island cells that lack reelin expression. The influence of reelin expression by ocean cells and interneurons on their own morphological differentiation and that of MECII island cells has remained unknown. To address this, we used a conditional reelin knockout (RelncKO) mouse to induce reelin deficiency postnatally in vitro and in vivo. Reelin deficiency caused dendritic hypertrophy of ocean cells, interneurons and only proximal dendritic compartments of island cells. Ca2+ recording showed that both cell types exhibited an elevation of calcium frequencies in RelncKO, indicating that the hypertrophic effect is related to excessive Ca2+ signalling. Moreover, pharmacological receptor blockade in RelncKO mouse revealed malfunctioning of GABAB, NMDA and AMPA receptors. Collectively, this study emphasizes the significance of reelin in neuronal growth, and its absence results in dendrite hypertrophy of MECII neurons.

Caudally pronounced deficiencies in preplate splitting and migration underly a rostro-caudal progression of cortical lamination defects in the reeler brain.

In mammalian neocortex development, every cohort of newborn neurons is guided toward the marginal zone, leading to an "inside-out" organization of the 6 neocortical layers. This migratory pattern is regulated by the extracellular glycoprotein Reelin. The reeler mouse shows a homozygous mutation of the reelin gene. Using RNA in situ hybridization we could demonstrate that the Reelin-deficient mouse cortex (male and female) displays an increasing lamination defect along the rostro-caudal axis that is characterized by strong cellular intermingling, but roughly reproduces the "inside-out" pattern in rostral cortex, while caudal cortex shows a relative inversion of neuronal positioning ("outside-in"). We found that in development of the reeler cortex, preplate-splitting is also defective with an increasing severity along the rostro-caudal axis. This leads to a misplacement of subplate neurons that are crucial for a switch in migration mode within the cortical plate. Using Flash Tag labeling and nucleoside analog pulse-chasing, we found an according migration defect within the cortical plate, again with a progressive severity along the rostro-caudal axis. Thus, loss of one key player in neocortical development leads to highly area-specific (caudally pronounced) developmental deficiencies that result in multiple roughly opposite rostral versus caudal adult neocortical phenotypes.

Reelin restricts dendritic growth of interneurons in the neocortex.

Reelin is a large secreted glycoprotein that regulates neuronal migration, lamination and establishment of dendritic architecture in the embryonic brain. Reelin expression switches postnatally from Cajal-Retzius cells to interneurons. However, reelin function in interneuron development is still poorly understood. Here, we have investigated the role of reelin in interneuron development in the postnatal neocortex. To preclude early cortical migration defects caused by reelin deficiency, we employed a conditional reelin knockout (RelncKO) mouse to induce postnatal reelin deficiency. Induced reelin deficiency caused dendritic hypertrophy in distal dendritic segments of neuropeptide Y-positive (NPY+) and calretinin-positive (Calr+) interneurons, and in proximal dendritic segments of parvalbumin-positive (Parv+) interneurons. Chronic recombinant Reelin treatment rescued dendritic hypertrophy in Relncko interneurons. Moreover, we provide evidence that RelncKO interneuron hypertrophy is due to presynaptic GABABR dysfunction. Thus, GABABRs in RelncKO interneurons were unable to block N-type (Cav2.2) Ca2+ channels that control neurotransmitter release. Consequently, the excessive Ca2+ influx through AMPA receptors, but not NMDA receptors, caused interneuron dendritic hypertrophy. These findings suggest that reelin acts as a 'stop-growth-signal' for postnatal interneuron maturation.

Thrombospondin 1 and Reelin act through Vldlr to regulate cardiac growth and repair.

Adult mammalian cardiomyocytes have minimal cell cycle capacity, which leads to poor regeneration after cardiac injury such as myocardial infarction. Many positive regulators of cardiomyocyte cell cycle and cardioprotective signals have been identified, but extracellular signals that suppress cardiomyocyte proliferation are poorly understood. We profiled receptors enriched in postnatal cardiomyocytes, and found that very-low-density-lipoprotein receptor (Vldlr) inhibits neonatal cardiomyocyte cell cycle. Paradoxically, Reelin, the well-known Vldlr ligand, expressed in cardiac Schwann cells and lymphatic endothelial cells, promotes neonatal cardiomyocyte proliferation. Thrombospondin1 (TSP-1), another ligand of Vldlr highly expressed in adult heart, was then found to inhibit cardiomyocyte proliferation through Vldlr, and may contribute to Vldlr's overall repression on proliferation. Mechanistically, Rac1 and subsequent Yap phosphorylation and nucleus translocation mediate the regulation of the cardiomyocyte cell cycle by TSP-1/Reelin-Vldlr signaling. Importantly, Reln mutant neonatal mice displayed impaired cardiomyocyte proliferation and cardiac regeneration after apical resection, while cardiac-specific Thbs1 deletion and cardiomyocyte-specific Vldlr deletion promote cardiomyocyte proliferation and are cardioprotective after myocardial infarction. Our results identified a novel role of Vldlr in consolidating extracellular signals to regulate cardiomyocyte cell cycle activity and survival, and the overall suppressive TSP-1-Vldlr signal may contribute to the poor cardiac repair capacity of adult mammals.

Different activity patterns control various stages of Reelin synthesis in the developing neocortex.

Reelin is a large extracellular matrix protein abundantly expressed in the developing neocortex of mammals. During embryonic and early postnatal stages in mice, Reelin is secreted by a transient neuronal population, the Cajal-Retzius neurons (CRs), and is mostly known to insure the inside-out migration of neurons and the formation of cortical layers. During the first 2 postnatal weeks, CRs disappear from the neocortex and a subpopulation of GABAergic neurons takes over the expression of Reelin, albeit in lesser amounts. Although Reelin expression requires a tight regulation in a time- and cell-type specific manner, the mechanisms regulating the expression and secretion of this protein are poorly understood. In this study, we establish a cell-type specific profile of Reelin expression in the marginal zone of mice neocortex during the first 3 postnatal weeks. We then investigate whether electrical activity plays a role in the regulation of Reelin synthesis and/or secretion by cortical neurons during the early postnatal period. We show that increased electrical activity promotes the transcription of reelin via the brain-derived neurotrophic factor/TrkB pathway, but does not affect its translation or secretion. We further demonstrate that silencing the neuronal network promotes the translation of Reelin without affecting the transcription or secretion. We conclude that different patterns of activity control various stages of Reelin synthesis, whereas its secretion seems to be constitutive.

The ZSWIM8 ubiquitin ligase regulates neurodevelopment by guarding the protein quality of intrinsically disordered Dab1.

Protein quality control (PQC) is essential for maintaining protein homeostasis and guarding the accuracy of neurodevelopment. Previously, we found that a conserved EBAX-type CRL regulates the protein quality of SAX-3/ROBO guidance receptors in Caenorhabditis elegans. Here, we report that ZSWIM8, the mammalian homolog of EBAX-1, is essential for developmental stability of mammalian brains. Conditional deletion of Zswim8 in the embryonic nervous system causes global cellular stress, partial perinatal lethality and defective migration of neural progenitor cells. CRISPR-mediated knockout of ZSWIM8 impairs spine formation and synaptogenesis in hippocampal neurons. Mechanistic studies reveal that ZSWIM8 controls protein quality of Disabled 1 (Dab1), a key signal molecule for brain development, thus protecting the signaling strength of Dab1. As a ubiquitin ligase enriched with intrinsically disordered regions (IDRs), ZSWIM8 specifically recognizes IDRs of Dab1 through a "disorder targets misorder" mechanism and eliminates misfolded Dab1 that cannot be properly phosphorylated. Adult survivors of ZSWIM8 CKO show permanent hippocampal abnormality and display severely impaired learning and memory behaviors. Altogether, our results demonstrate that ZSWIM8-mediated PQC is critical for the stability of mammalian brain development.

Critical functions of extracellular matrix in brain metastasis seeding.

Human brain is characterized by extremely sparse extracellular matrix (ECM). Despite its low abundance, the significance of brain ECM in both physiological and pathological conditions should not be underestimated. Brain metastasis is a serious complication of cancer, and recent findings highlighted the contribution of ECM in brain metastasis development. In this review, we provide a comprehensive outlook on how ECM proteins promote brain metastasis seeding. In particular, we discuss (1) disruption of the blood-brain barrier in brain metastasis; (2) role of ECM in modulating brain metastasis dormancy; (3) regulation of brain metastasis seeding by ECM-activated integrin signaling; (4) functions of brain-specific ECM protein reelin in brain metastasis. Lastly, we consider the possibility of targeting ECM for brain metastasis management.

Reelin regulates the migration of late-born hippocampal CA1 neurons via cofilin phosphorylation.

Reelin, a large secreted glycoprotein, plays an important role in neuronal migration during brain development. The C-terminal region (CTR) of Reelin is involved in the efficient activation of downstream signaling and its loss leads to abnormal hippocampal layer formation. However, the molecular mechanism by which Reelin CTR regulates hippocampal development remains unknown. Here, we showed that the migration of late-born, but not early-born, neurons is impaired in the knock-in mice in which Reelin CTR is deleted (ΔC-KI mice). The phosphorylation of cofilin, an actin-depolymerizing protein, was remarkably decreased in the hippocampus of the ΔC-KI mice. Exogenous expression of pseudo-phosphorylated cofilin rescued the ectopic positioning of neurons in the hippocampus of ΔC-KI mice. These results suggest that Reelin CTR is required for the migration of late-born neurons in the hippocampus and that this event involves appropriate phosphorylation of cofilin.

A key requirement for synaptic Reelin signaling in ketamine-mediated behavioral and synaptic action.

Ketamine is a noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist that produces rapid antidepressant action in some patients with treatment-resistant depression. However, recent data suggest that ∼50% of patients with treatment-resistant depression do not respond to ketamine. The factors that contribute to the nonresponsiveness to ketamine's antidepressant action remain unclear. Recent studies have reported a role for secreted glycoprotein Reelin in regulating pre- and postsynaptic function, which suggests that Reelin may be involved in ketamine's antidepressant action, although the premise has not been tested. Here, we investigated whether the disruption of Reelin-mediated synaptic signaling alters ketamine-triggered synaptic plasticity and behavioral effects. To this end, we used mouse models with genetic deletion of Reelin or apolipoprotein E receptor 2 (Apoer2), as well as pharmacological inhibition of their downstream effectors, Src family kinases (SFKs) or phosphoinositide 3-kinase. We found that disruption of Reelin, Apoer2, or SFKs blocks ketamine-driven behavioral changes and synaptic plasticity in the hippocampal CA1 region. Although ketamine administration did not affect tyrosine phosphorylation of DAB1, an adaptor protein linked to downstream signaling of Reelin, disruption of Apoer2 or SFKs impaired baseline NMDA receptor-mediated neurotransmission. These results suggest that maintenance of baseline NMDA receptor function by Reelin signaling may be a key permissive factor required for ketamine's antidepressant effects. Taken together, our results suggest that impairments in Reelin-Apoer2-SFK pathway components may in part underlie nonresponsiveness to ketamine's antidepressant action.

Molecular Mechanisms of Reelin in the Enteric Nervous System and the Microbiota-Gut-Brain Axis: Implications for Depression and Antidepressant Therapy.

Current pharmacological treatments for depression fail to produce adequate remission in a significant proportion of patients. Increasingly, other systems, such as the microbiome-gut-brain axis, are being looked at as putative novel avenues for depression treatment. Dysbiosis and dysregulation along this axis are highly comorbid with the severity of depression symptoms. The endogenous extracellular matrix protein reelin is present in all intestinal layers as well as in myenteric and submucosal ganglia, and its receptors are also present in the gut. Reelin secretion from subepithelial myofibroblasts regulates cellular migration along the crypt-villus axis in the small intestine and colon. Reelin brain expression is downregulated in mood and psychotic disorders, and reelin injections have fast antidepressant-like effects in animal models of depression. This review seeks to discuss the roles of reelin in the gastrointestinal system and propose a putative role for reelin actions in the microbiota-gut-brain axis in the pathogenesis and treatment of depression, primarily reflecting on alterations in gut epithelial cell renewal and in the clustering of serotonin transporters.

A novel Reelin construct, R36, recovered behavioural deficits in the heterozygous reeler mouse.

Reelin, a large extracellular glycoprotein, plays a critical role in prenatal brain development and postnatally in synaptic plasticity, learning and memory. Dysregulation of Reelin signalling has been implicated in several neuropsychiatric disorders including schizophrenia, autism, depression and Alzheimer's disease. Previous studies have demonstrated that Reelin's central fragment, R3456, binds to ApoER2, inducing ApoER2 clustering and subsequent intracellular signalling. We previously reported the development of a novel luciferase complementation assay, which we used to demonstrate that R3456 can lead to ApoER2 receptor dimerization. Using this same assay, we explored various smaller fragments and combinations from R3456, and we identified a construct of repeats 3 and 6 (R36), which could still elicit equivalent receptor dimerization. The purpose of this study was to test R36 for biological effects in vitro and in vivo. We show that R36 was capable of initiating intracellular signalling in primary neuronal cultures. In addition, we demonstrate that a single intracerebroventricular injection of R36 protein into a model of Reelin deficiency, the heterozygous reeler mice, can significantly improve cognition. These data support a role for the new construct R36 to enhance the Reelin pathway, and the future possibility of exploring gene therapy approaches with R36 in diseases characterized by reduced levels of Reelin.

TRIM40 ameliorates diabetic retinopathy through suppressing inflammation via Reelin/DAB1 signaling disruption: A mechanism by proteasomal degradation of DAB1.

Diabetic retinopathy (DR) is a common microvascular complication of diabetes mellitus. Reelin, an extracellular matrix protein, and its effector protein Disabled1 (DAB1) have been linked to cellular events and retinal development. However, whether and how Reelin/DAB1 signaling causes DR remains to be investigated. In our study, significantly increased expression of Reelin, very low density lipoprotein receptor (VLDLR), ApoE receptor 2 (ApoER2) and phosphorylated DAB1 in retinas of streptozotocin (STZ)-induced DR mouse model was observed, along with enhanced expression of proinflammatory factors. Similar results are confirmed in high glucose (HG)-treated human retinal pigment epithelium cell line ARPE-19. Surprisingly, dysregulated tripartite motif-containing 40 (TRIM40), an E3 ubiquitin ligase, is found to be involved in DR progression by bioinformatic analysis. We observe a negative correlation between TRIM40 and p-DAB1 protein expression levels under HG conditions. Importantly, we find that TRIM40 over-expression markedly ameliorates HG-induced p-DAB1, PI3K, p-protein B kinase (AKT) and inflammatory response in HG-treated cells, but dose not affect Reelin expression. Of note, Co-IP and double immunofluorescence identify an interaction between TRIM40 and DAB1. Furthermore, we show that TRIM40 enhances K48-linked polyubiquitination of DAB1, thereby promoting DAB1 degradation. Finally, promoting TRIM40 expression by intravenous injection of the constructed adeno-associated virus (AAV-TRIM40) markedly ameliorates DR phenotypes in STZ-treated mice, as indicated by the decreased blood glucose and glycosylated hemoglobin (HbAlc) levels, and increased hemoglobin contents. Additionally, diabetes-related elevation of acellular capillaries was also meliorated in mice over-expressing TRIM40. The electroretinogram (ERG) deficits were strongly rescued in mice receiving AAV-TRIM40 injection. Moreover, AAV-TRIM40 attenuates the inflammation and p-DAB1 expression in retinal tissues of STZ-treated mice. Collectively, our findings disclose a mechanism through which TRIM40 limits DAB1 stability under physiological conditions and reveals TRIM40 as a potential therapeutic target for the intervention of Reelin/DAB1 signaling, contributing to DR treatment.

VLDLR is not essential for reelin-induced neuronal aggregation but suppresses neuronal invasion into the marginal zone.

In the developing neocortex, radially migrating neurons stop migration and form layers beneath the marginal zone (MZ). Reelin plays essential roles in these processes via its receptors, apolipoprotein E receptor 2 (ApoER2) and very low density lipoprotein receptor (VLDLR). Although we recently reported that reelin causes neuronal aggregation via ApoER2, which is thought to be important for the subsequent layer formation, it remains unknown what effect reelin exerts via the VLDLR. Here, we found that ectopic reelin overexpression in the Vldlr-mutant mouse cortex causes neuronal aggregation, but without an MZ-like cell-sparse central region that is formed when reelin is overexpressed in the normal cortex. We also found that both the early-born and late-born Vldlr-deficient neurons invade the MZ and exhibit impaired dendrite outgrowth from before birth. Rescue experiments indicate that VLDLR suppresses neuronal invasion into the MZ via a cell-autonomous mechanism, possibly mediated by Rap1, integrin and Akt. These results suggest that VLDLR is not a prerequisite for reelin-induced neuronal aggregation and that the major role of VLDLR is to suppress neuronal invasion into the MZ during neocortical development.

Reelin Plasma Levels Identify Cognitive Decline in Alcohol Use Disorder Patients During Early Abstinence: The Influence of APOE4 Expression.

BACKGROUND: Apolipoprotein E (APOE)-4 isoform, reelin, and clusterin share very-low-density liporeceptor and apolipoprotein E receptor 2 receptors and are related to cognition in neuropsychiatric disorders. These proteins are expressed in plasma and brain, but studies involving plasma expression and cognition are scarce. METHODS: We studied the peripheral expression (plasma and peripheral blood mononuclear cells) of these proteins in 24 middle-aged patients with alcohol use disorder (AUD) diagnosed at 4 to 12 weeks of abstinence (t = 0) and 34 controls. Cognition was assessed using the Test of Detection of Cognitive Impairment in Alcoholism. In a follow-up study (t = 1), we measured reelin levels and evaluated cognitive improvement at 6 months of abstinence. RESULTS: APOE4 isoform was present in 37.5% and 58.8% of patients and controls, respectively, reaching similar plasma levels in ε4 carriers regardless of whether they were patients with AUD or controls. Plasma reelin and clusterin were higher in the AUD group, and reelin levels peaked in patients expressing APOE4 (P < .05, η2 = 0.09), who showed reduced very-low-density liporeceptor and apolipoprotein E receptor 2 expression in peripheral blood mononuclear cells. APOE4 had a negative effect on memory/learning mainly in the AUD group (P < .01, η2 = 0.15). Multivariate logistic regression analyses identified plasma reelin as a good indicator of AUD cognitive impairment at t = 0. At t = 1, patients with AUD showed lower reelin levels vs controls along with some cognitive improvement. CONCLUSIONS: Reelin plasma levels are elevated during early abstinence in patients with AUD who express the APOE4 isoform, identifying cognitive deterioration to a great extent, and it may participate as a homeostatic signal for cognitive recovery in the long term.

Influence of Chronic Electroconvulsive Seizures on Plasticity-Associated Gene Expression and Perineuronal Nets Within the Hippocampi of Young Adult and Middle-Aged Sprague-Dawley Rats.

BACKGROUND: Electroconvulsive seizure therapy is often used in both treatment-resistant and geriatric depression. However, preclinical studies identifying targets of chronic electroconvulsive seizure (ECS) are predominantly focused on animal models in young adulthood. Given that putative transcriptional, neurogenic, and neuroplastic mechanisms implicated in the behavioral effects of chronic ECS themselves exhibit age-dependent modulation, it remains unknown whether the molecular and cellular targets of chronic ECS vary with age. METHODS: We subjected young adult (2-3 months) and middle-aged (12-13 months), male Sprague Dawley rats to sham or chronic ECS and assessed for despair-like behavior, hippocampal gene expression, hippocampal neurogenesis, and neuroplastic changes in the extracellular matrix, reelin, and perineuronal net numbers. RESULTS: Chronic ECS reduced despair-like behavior at both ages, accompanied by overlapping and unique changes in activity-dependent and trophic factor gene expression. Although chronic ECS had a similar impact on quiescent neural progenitor numbers at both ages, the eventual increase in hippocampal progenitor proliferation was substantially higher in young adulthood. We noted a decline in reelin⁺ cell numbers following chronic ECS only in young adulthood. In contrast, an age-invariant, robust dissolution of perineuronal net numbers that encapsulate parvalbumin⁺ neurons in the hippocampus were observed following chronic ECS. CONCLUSION: Our findings indicate that age is a key variable in determining the nature of chronic ECS-evoked molecular and cellular changes in the hippocampus. This raises the intriguing possibility that chronic ECS may recruit distinct, as well as overlapping, mechanisms to drive antidepressant-like behavioral changes in an age-dependent manner.

Monoallelic and biallelic mutations in RELN underlie a graded series of neurodevelopmental disorders.

Reelin, a large extracellular protein, plays several critical roles in brain development and function. It is encoded by RELN, first identified as the gene disrupted in the reeler mouse, a classic neurological mutant exhibiting ataxia, tremors and a 'reeling' gait. In humans, biallelic variants in RELN have been associated with a recessive lissencephaly variant with cerebellar hypoplasia, which matches well with the homozygous mouse mutant that has abnormal cortical structure, small hippocampi and severe cerebellar hypoplasia. Despite the large size of the gene, only 11 individuals with RELN-related lissencephaly with cerebellar hypoplasia from six families have previously been reported. Heterozygous carriers in these families were briefly reported as unaffected, although putative loss-of-function variants are practically absent in the population (probability of loss of function intolerance = 1). Here we present data on seven individuals from four families with biallelic and 13 individuals from seven families with monoallelic (heterozygous) variants of RELN and frontotemporal or temporal-predominant lissencephaly variant. Some individuals with monoallelic variants have moderate frontotemporal lissencephaly, but with normal cerebellar structure and intellectual disability with severe behavioural dysfunction. However, one adult had abnormal MRI with normal intelligence and neurological profile. Thorough literature analysis supports a causal role for monoallelic RELN variants in four seemingly distinct phenotypes including frontotemporal lissencephaly, epilepsy, autism and probably schizophrenia. Notably, we observed a significantly higher proportion of loss-of-function variants in the biallelic compared to the monoallelic cohort, where the variant spectrum included missense and splice-site variants. We assessed the impact of two canonical splice-site variants observed as biallelic or monoallelic variants in individuals with moderately affected or normal cerebellum and demonstrated exon skipping causing in-frame loss of 46 or 52 amino acids in the central RELN domain. Previously reported functional studies demonstrated severe reduction in overall RELN secretion caused by heterozygous missense variants p.Cys539Arg and p.Arg3207Cys associated with lissencephaly suggesting a dominant-negative effect. We conclude that biallelic variants resulting in complete absence of RELN expression are associated with a consistent and severe phenotype that includes cerebellar hypoplasia. However, reduced expression of RELN remains sufficient to maintain nearly normal cerebellar structure. Monoallelic variants are associated with incomplete penetrance and variable expressivity even within the same family and may have dominant-negative effects. Reduced RELN secretion in heterozygous individuals affects only cortical structure whereas the cerebellum remains intact. Our data expand the spectrum of RELN-related neurodevelopmental disorders ranging from lethal brain malformations to adult phenotypes with normal brain imaging.

Synaptic and Genetic Bases of Impaired Motor Learning Associated with Modified Experience-Dependent Cortical Plasticity in Heterozygous Reeler Mutants.

Patients with neurodevelopmental disorders show impaired motor skill learning. It is unclear how the effect of genetic variation on synaptic function and transcriptome profile may underlie experience-dependent cortical plasticity, which supports the development of fine motor skills. RELN (reelin) is one of the genes implicated in neurodevelopmental psychiatric vulnerability. Heterozygous reeler mutant (HRM) mice displayed impairments in reach-to-grasp learning, accompanied by less extensive cortical map reorganization compared with wild-type mice, examined after 10 days of training by intracortical microstimulation. Assessed by patch-clamp recordings after 3 days of training, the training induced synaptic potentiation and increased glutamatergic-transmission of cortical layer III pyramidal neurons in wild-type mice. In contrast, the basal excitatory and inhibitory synaptic functions were depressed, affected both by presynaptic and postsynaptic impairments in HRM mice; and thus, no further training-induced synaptic plasticity occurred. HRM exhibited downregulations of cortical synaptophysin, immediate-early gene expressions, and gene enrichment, in response to 3 days of training compared with trained wild-type mice, shown using quantitative reverse transcription polymerase chain reaction, immunohistochemisty, and RNA-sequencing. We demonstrated that motor learning impairments associated with modified experience-dependent cortical plasticity are at least partially attributed by the basal synaptic alternation as well as the aberrant early experience-induced gene enrichment in HRM.

A frameshift-deletion mutation in Reelin causes cerebellar hypoplasia in White Swiss Shepherd dogs.

Cerebellar hypoplasia is a heterogeneous neurological condition in which the cerebellum is smaller than usual or not completely developed. The condition can have genetic origins, with Mendelian-effect mutations described in several mammalian species. Here, we describe a genetic investigation of cerebellar hypoplasia in White Swiss Shepherd dogs, where two affected puppies were identified from a litter with a recent common ancestor on both sides of their pedigree. Whole genome sequencing was conducted for 10 dogs in this family, and filtering of these data based on a recessive transmission hypothesis highlighted five protein-altering candidate variants - including a frameshift-deletion of the Reelin (RELN) gene (p.Val947*). Given the status of RELN as a gene responsible for cerebellar hypoplasia in humans, sheep and mice, these data strongly suggest the loss-of-function variant as underlying these effects. This variant has not been found in other dog breeds nor in a cohort of European White Swiss Shepherds, suggesting a recent mutation event. This finding will support the genotyping of a more diverse sample of dogs, and should aid future management of the harmful allele through optimised mating schemes.

Lipophorin receptors regulate mushroom body development and complex behaviors in Drosophila.

BACKGROUND: Drosophila melanogaster lipophorin receptors (LpRs), LpR1 and LpR2, are members of the LDLR family known to mediate lipid uptake in a range of organisms from Drosophila to humans. The vertebrate orthologs of LpRs, ApoER2 and VLDL-R, function as receptors of a glycoprotein involved in development of the central nervous system, Reelin, which is not present in flies. ApoER2 and VLDL-R are associated with the development and function of the hippocampus and cerebral cortex, important association areas in the mammalian brain, as well as with neurodevelopmental and neurodegenerative disorders linked to those regions. It is currently unknown whether LpRs play similar roles in the Drosophila brain. RESULTS: We report that LpR-deficient flies exhibit impaired olfactory memory and sleep patterns, which seem to reflect anatomical defects found in a critical brain association area, the mushroom bodies (MB). Moreover, cultured MB neurons respond to mammalian Reelin by increasing the complexity of their neurite arborization. This effect depends on LpRs and Dab, the Drosophila ortholog of the Reelin signaling adaptor protein Dab1. In vitro, two of the long isoforms of LpRs allow the internalization of Reelin, suggesting that Drosophila LpRs interact with human Reelin to induce downstream cellular events. CONCLUSIONS: These findings demonstrate that LpRs contribute to MB development and function, supporting the existence of a LpR-dependent signaling in Drosophila, and advance our understanding of the molecular factors functioning in neural systems to generate complex behaviors in this model. Our results further emphasize the importance of Drosophila as a model to investigate the alterations in specific genes contributing to neural disorders.

Analysis of distribution and localization of proteins of the reelin signalling pathway in mesial temporal lobe epilepsy.

INTRODUCTION: Granule cell dispersion (GCD) is pathognomonic of hippocampal sclerosis seen in the mesial temporal lobe epilepsy (MTLE). Current animal studies indicate deficiency of Reelin is associated with abnormal granule cell migration leading to GCD. The present study aimed to evaluate complete Reelin signalling pathway to assess whether Reelin deficiency is related to MTLE. MATERIALS AND METHODS: Hippocampal sclerosis was confirmed by H and E stain. To explore the amount and cellular location of the Reelin cascade molecules, the hippocampal tissues from MTLE surgery and controls (n = 15 each) were studied using Immuno-histochemistry (IHC). Additionally, confocal imaging was used to validate the IHC findings by co-localization of different proteins. Quantification of IHC images was performed using histo-score and confocal images by Image J software. RESULTS: Immune expression of active Reelin was significantly reduced in patients. Reelin receptors were deranged, apolipoprotein E receptor 2 was increased while very low-density lipoprotein receptor was reduced. Disabled-1, a downstream molecule was significantly reduced in MTLE. Its ultimate target, cofilin was thus disinhibited and expressed more in MTLE. Reelin cleaving protease, matrix metalloprotease-9 (MMP-9) and MMP-9 inhibitor, tissue inhibitor of matrix protease-1, showed reduced expression in extracellular matrix. Semi-quantification of immunohistochemistry was done using Histo (H) score. H score of Reelin in diseased patients was 15 against 125 for control patients. These results were validated by confocal fluorescence microscopy. CONCLUSIONS: Reelin signalling cascade was deranged in chronic MTLE. Pharmacological manipulation of Reelin cascade can be done at various levels and it may provide novel treatment options for MTLE.