No abstract

Genetic mutations in nitrogen permease regulator-like 2 (NPRL2) are associated with a wide spectrum of familial focal epilepsies, autism, and sudden unexpected death of epileptics (SUDEP), but the mechanisms by which NPRL2 contributes to these effects are not well known. NPRL2 is a requisite subunit of the Gap Activity TOward Rags 1 (GATOR1) complex, which functions as a negative regulator of mammalian Target Of Rapamycin Complex 1 (mTORC1) kinase when intracellular amino acids are low.Here we show that loss of NPRL2 expression in mouse excitatory glutamatergic neurons causes seizures prior to death, consistent with SUDEP in humans with epilepsy. Additionally, the absence of NPRL2 expression increases mTORC1-dependent signal transduction and significantly alters amino acid homeostasis in the brain. Loss of NPRL2 reduces dendritic branching and increases the strength of electrically stimulated action potentials in neurons. The increased action potential strength is consistent with elevated expression of epilepsy-linked, voltage-gated sodium channels in the NPRL2-deficient brain. Targeted deletion of NPRL2 in primary neurons increases the expression of sodium channel Scn1A, whereas treatment with the pharmacological mTORC1 inhibitor called rapamycin prevents Scn1A upregulation. These studies demonstrate a novel role of NPRL2 and mTORC1 signaling in the regulation of sodium channels, which can contribute to seizures and early lethality.Significance Statement: NPRL2 is a requisite subunit of the epilepsy-linked GATOR1 complex that functions as a negative regulator of mTORC1 kinase when intracellular amino acids are limited. Here we report the generation and characterization of a new neurological model of GATOR1-dependent mTORopathy, caused by the loss of NPRL2 function in glutamatergic neurons. Loss of NPRL2 increases mTORC1 signal transduction, significantly alters amino acid homeostasis in the brain, and causes SUDEP.In addition, loss of NPRL2 increases the strength of electrically stimulated action potentials and the expression of epilepsy-linked sodium channels. These data reveal an unanticipated link between 3 intracellular amino acid signaling by NPRL2 and a novel mTORC1-dependent regulation of sodiumchannel expression in epilepsy.

Frontotemporal dementia (FTD) is a heterogeneous clinical disorder characterized by progressive abnormalities in behavior, executive functions, personality, language and/or motricity. A neuropathological subtype of FTD, frontotemporal lobar degeneration (FTLD)-FET, is characterized by protein aggregates consisting of the RNA-binding protein fused in sarcoma (FUS). The cause of FTLD-FET is not well understood and there is a lack of genetic evidence to aid in the investigation of mechanisms of the disease. The goal of this study was to identify genetic variants contributing to FTLD-FET and to investigate their effects on FUS pathology. We performed whole-exome sequencing on a 50-year-old FTLD patient with ubiquitin and FUS-positive neuronal inclusions and unaffected parents, and identified a de novo postzygotic nonsense variant in the NCDN gene encoding Neurochondrin (NCDN), NM_014284.3:c.1206G > A, p.(Trp402*). The variant was associated with a ~ 31% reduction in full-length protein levels in the patient’s brain, suggesting that this mutation leads to NCDN haploinsufficiency. We examined the effects of NCDN haploinsufficiency on FUS and found that depleting primary cortical neurons of NCDN causes a reduction in the total number of FUS-positive cytoplasmic granules. Moreover, we found that these granules were significantly larger and more highly enriched with FUS. We then examined the effects of a loss of FUS function on NCDN in neurons and found that depleting cells of FUS leads to a decrease in NCDN protein and mRNA levels. Our study identifies the NCDN protein as a likely contributor of FTLD-FET pathophysiology. Moreover, we provide evidence for a negative feedback loop of toxicity between NCDN and FUS, where loss of NCDN alters FUS cytoplasmic dynamics, which in turn has an impact on NCDN expression.

Background. The medial prefrontal cortex (mPFC) is part of a complex circuit controlling stress responses by sending projections to different limbic structures including the nucleus accumbens (NAc) and ventral tegmental area (VTA). However, the impact of chronic stress on NAc- and VTA-projecting mPFC neurons is still unknown and the distinct contribution of these pathways to stress responses in males and females is unclear. Methods. Behavioral stress response in both sexes was induced by 21 days of chronic variable stress (CVS). We used an inter-sectional viral approach to label both pathways and assess the functional, morphological, and transcriptional adaptations in NAc- and VTA-projecting mPFC neurons in stressed males and females. Using chemogenetic approaches, we modified neuronal activity of NAc-projecting mPFC neurons to decipher their contribution to stress phenotypes in both sexes. Results. CVS induced depressive-like behaviors in males and females. NAc- and VTA-projecting mPFC neurons exhibited sex-specific functional, morphological, and transcriptional alterations. The functional changes were more important in females in NAc-projecting mPFC neurons while males exhibited a more drastic reduction in dendritic complexity in VTA-projecting mPFC neurons after CVS. Finally, chemogenetic overactivation of the cortico-accumbal pathway triggered anxiety and behavioral despair in both sexes while its inhibition rescued the phenotype only in females. Conclusions. Our results suggest that chronic stress impacts the cortico-accumbal and cortico-tegmental pathways differently in males and females through sex-specific functional, morphological, and transcriptional alterations in mPFC neurons. Furthermore, our results suggest that the cortico-accumbal pathway contributes to the expression of anxiety and behavioral despair in females.