A septal-ventral tegmental area circuit drives exploratory behavior.

To survive, animals need to balance their exploratory drive with their need for safety. Subcortical circuits play an important role in initiating and modulating movement based on external demands and the internal state of the animal; however, how motivation and onset of locomotion are regulated remain largely unresolved. Here, we show that a glutamatergic pathway from the medial septum and diagonal band of Broca (MSDB) to the ventral tegmental area (VTA) controls exploratory locomotor behavior in mice. Using a self-supervised machine learning approach, we found an overrepresentation of exploratory actions, such as sniffing, whisking, and rearing, when this projection is optogenetically activated. Mechanistically, this role relies on glutamatergic MSDB projections that monosynaptically target a subset of both glutamatergic and dopaminergic VTA neurons. Taken together, we identified a glutamatergic basal forebrain to midbrain circuit that initiates locomotor activity and contributes to the expression of exploration-associated behavior.

A liaison between chaperone-mediated autophagy and exocytotic lysosomes controls the dendritic metastable proteome.

The neuronal metastable proteome includes several aggregation-prone proteins related to neurodegeneration. The complex morphology of neurons with very thin processes and enhanced protein turnover therefore necessitates efficient local machinery to remove excessive protein. In recent work we revealed that chaperone-mediated autophagy (CMA) provides cargo for dendritic exocytic lysosomes, a mechanism that serves in the rapid removal of disease-relevant, supersaturated proteins such as TARDBP/TDP-43 (TAR DNA binding protein) and HTT (huntingtin). We found that lysosomal exocytosis requires docking of the lysosomal protein LAMP2B to the glutamatergic receptor scaffold DLG3/SAP102 and that it is regulated by GRIN/NMDA (N-methyl-D-aspartate)-receptor activity. Thus, the small caliber of dendritic processes might impose a need for local disposal of aggregation-prone proteins like TARDBP and HTT. Moreover, we observed that lysosomal exocytosis might serve in both protein removal and modulation of synaptic processes, and the latter might be an inevitable consequence of the necessity for local disposal of CMA clients in dendrites.

MicroRNA-92a-CPEB3 axis protects neurons against inflammatory neurodegeneration.

Neuroinflammation causes neuronal injury in multiple sclerosis (MS) and other neurological diseases. MicroRNAs (miRNAs) are important modulators of neuronal stress responses, but knowledge about their contribution to neuronal protection or damage during inflammation is limited. Here, we constructed a regulatory miRNA-mRNA network of inflamed motor neurons by leveraging cell type-specific miRNA and mRNA sequencing of mice undergoing experimental autoimmune encephalomyelitis (EAE). We found robust induction of miR-92a in inflamed spinal cord neurons and identified cytoplasmic polyadenylation element-binding protein 3 (Cpeb3) as a key target of miR-92a-mediated posttranscriptional silencing. We detected CPEB3 repression in inflamed neurons in murine EAE and human MS. Moreover, both miR-92a delivery and Cpeb3 deletion protected neuronal cultures against excitotoxicity. Supporting a detrimental effect of Cpeb3 in vivo, neuron-specific deletion in conditional Cpeb3 knockout animals led to reduced inflammation-induced clinical disability in EAE. Together, we identified a neuroprotective miR-92a-Cpeb3 axis in neuroinflammation that might serve as potential treatment target to limit inflammation-induced neuronal damage.

LipidSpace: Simple Exploration, Reanalysis, and Quality Control of Large-Scale Lipidomics Studies.

Lipid analysis gained significant importance due to the enormous range of lipid functions, e.g., energy storage, signaling, or structural components. Whole lipidomes can be quantitatively studied in-depth thanks to recent analytical advancements. However, the systematic comparison of thousands of distinct lipidomes remains challenging. We introduce LipidSpace, a standalone tool for analyzing lipidomes by assessing their structural and quantitative differences. A graph-based comparison of lipid structures is the basis for calculating structural space models and subsequently computing lipidome similarities. When adding study variables such as body weight or health condition, LipidSpace can determine lipid subsets across all lipidomes that describe these study variables well by utilizing machine-learning approaches. The user-friendly GUI offers four built-in tutorials and interactive visual interfaces with pdf export. Many supported data formats allow an efficient (re)analysis of data sets from different sources. An integrated interactive workflow guides the user through the quality control steps. We used this suite to reanalyze and combine already published data sets (e.g., one with about 2500 samples and 576 lipids in one run) and made additional discoveries to the published conclusions with the potential to fill gaps in the current lipid biology understanding. LipidSpace is available for Windows or Linux (https://lifs-tools.org).

Chaperone-mediated autophagy in neuronal dendrites utilizes activity-dependent lysosomal exocytosis for protein disposal.

The complex morphology of neurons poses a challenge for proteostasis because the majority of lysosomal degradation machinery is present in the cell soma. In recent years, however, mature lysosomes were identified in dendrites, and a fraction of those appear to fuse with the plasma membrane and release their content to the extracellular space. Here, we report that dendritic lysosomes are heterogeneous in their composition and that only those containing lysosome-associated membrane protein (LAMP) 2A and 2B fuse with the membrane and exhibit activity-dependent motility. Exocytotic lysosomes dock in close proximity to GluN2B-containing N-methyl-D-aspartate-receptors (NMDAR) via an association of LAMP2B to the membrane-associated guanylate kinase family member SAP102/Dlg3. NMDAR-activation decreases lysosome motility and promotes membrane fusion. We find that chaperone-mediated autophagy is a supplier of content that is released to the extracellular space via lysosome exocytosis. This mechanism enables local disposal of aggregation-prone proteins like TDP-43 and huntingtin.

Integration of nuclear Ca2+ transients and subnuclear protein shuttling provides a novel mechanism for the regulation of CREB-dependent gene expression.

Nuclear Ca2+ waves elicited by NMDAR and L-type voltage-gated Ca2+-channels as well as protein transport from synapse-to-nucleus are both instrumental in control of plasticity-related gene expression. At present it is not known whether fast [Ca2+]n transients converge in the nucleus with signaling of synapto-nuclear protein messenger. Jacob is a protein that translocate a signalosome from N-methyl-D-aspartate receptors (NMDAR) to the nucleus and that docks this signalosome to the transcription factor CREB. Here we show that the residing time of Jacob in the nucleoplasm strictly correlates with nuclear [Ca2+]n transients elicited by neuronal activity. A steep increase in [Ca2+]n induces instantaneous uncoupling of Jacob from LaminB1 at the nuclear lamina and promotes the association with the transcription factor cAMP-responsive element-binding protein (CREB) in hippocampal neurons. The size of the Jacob pool at the nuclear lamina is controlled by previous activity-dependent nuclear import, and thereby captures the previous history of NMDAR-induced nucleocytoplasmic shuttling. Moreover, the localization of Jacob at the nuclear lamina strongly correlates with synaptic activity and [Ca2+]n waves reflecting ongoing neuronal activity. In consequence, the resulting extension of the nuclear residing time of Jacob amplifies the capacity of the Jacob signalosome to regulate CREB-dependent gene expression and will, thereby, compensate for the relatively small number of molecules reaching the nucleus from individual synapses.

The many "Neurofaces" of Prohibitins 1 and 2: Crucial for the healthy brain, dysregulated in numerous brain disorders.

Prohibitin 1 (PHB1) and prohibitin 2 (PHB2) are proteins that are nearly ubiquitously expressed. They are localized in mitochondria, cytosol and cell nuclei. In the healthy CNS, they occur in neurons and non-neuronal cells (oligodendrocytes, astrocytes, microglia, and endothelial cells) and fulfill pivotal functions in brain development and aging, the regulation of brain metabolism, maintenance of structural integrity, synapse formation, aminoacidergic neurotransmission and, probably, regulation of brain action of certain hypothalamic-pituitary hormones.With regard to the diseased brain there is increasing evidence that prohibitins are prominently involved in numerous major diseases of the CNS, which are summarized and discussed in the present review (brain tumors, neurotropic viruses, Alzheimer disease, Down syndrome, Fronto-temporal and vascular dementia, dementia with Lewy bodies, Parkinson disease, Huntington disease, Multiple sclerosis, Amyotrophic lateral sclerosis, stroke, alcohol use disorder, schizophrenia and autism). Unfortunately, there is no PHB-targeted therapy available for any of these diseases.

Golgi satellites are essential for polysialylation of NCAM and expression of LTP at distal synapses.

The complex cytoarchitecture of neurons poses significant challenges for the maturation of synaptic membrane proteins. It is currently unclear whether locally secreted synaptic proteins bypass the Golgi or whether they traffic through Golgi satellites (GSs). Here, we create a transgenic GS reporter mouse line and show that GSs are widely distributed along dendrites and are capable of mature glycosylation, in particular sialylation. We find that polysialylation of locally secreted NCAM takes place at GSs. Accordingly, in mice lacking a component of trans-Golgi network-to-plasma membrane trafficking, we find fewer GSs and significantly reduced PSA-NCAM levels in distal dendrites of CA1 neurons that receive input from the temporoammonic pathway. Induction of long-term potentiation at those, but not more proximal, synapses is severely impaired. We conclude that GSs serve the need for local mature glycosylation of synaptic membrane proteins in distal dendrites and thereby contribute to rapid changes in synaptic strength.

Protein transport from pre- and postsynapse to the nucleus: Mechanisms and functional implications.

The extreme length of neuronal processes poses a challenge for synapse-to-nucleus communication. In response to this challenge several different mechanisms have evolved in neurons to couple synaptic activity to the regulation of gene expression. One of these mechanisms concerns the long-distance transport of proteins from pre- and postsynaptic sites to the nucleus. In this review we summarize current evidence on mechanisms of transport and consequences of nuclear import of these proteins for gene transcription. In addition, we discuss how information from pre- and postsynaptic sites might be relayed to the nucleus by this type of long-distance signaling. When applicable, we highlight how long-distance protein transport from synapse-to-nucleus can provide insight into the pathophysiology of disease or reveal new opportunities for therapeutic intervention.

A Jacob/nsmf gene knockout does not protect against acute hypoxia- and NMDA-induced excitotoxic cell death.

Jacob is a synapto-nuclear messenger protein that encodes and transduces the origin of synaptic and extrasynaptic NMDA receptor signals to the nucleus. The protein assembles a signalosome that differs in case of synaptic or extrasynaptic NMDAR activation. Following nuclear import Jacob docks these signalosomes to the transcription factor CREB. We have recently shown that amyloid-ß and extrasynaptic NMDAR activation triggers the translocation of a Jacob signalosome that results in inactivation of the transcription factor CREB, a phenomenon termed Jacob-induced CREB shut-off (JaCS). JaCS contributes to early Alzheimer's disease pathology and the absence of Jacob protects against amyloid pathology. Given that extrasynaptic activity is also involved in acute excitotoxicity, like in stroke, we asked whether nsmf gene knockout will also protect against acute insults, like oxygen and glucose deprivation and excitotoxic NMDA stimulation. nsmf is the gene that encodes for the Jacob protein. Here we show that organotypic hippocampal slices from wild-type and nsmf-/- mice display similar degrees of degeneration when exposed to either oxygen glucose deprivation or 50 µM NMDAto induce excitotoxicity. This lack of neuroprotection indicates that JaCS is mainly relevant in conditions of low level chronic extrasynaptic NMDAR activation that results in cellular degeneration induced by alterations in gene transcription.

Jacob-induced transcriptional inactivation of CREB promotes Aß-induced synapse loss in Alzheimer's disease.

Synaptic dysfunction caused by soluble ß-amyloid peptide (Aß) is a hallmark of early-stage Alzheimer's disease (AD), and is tightly linked to cognitive decline. By yet unknown mechanisms, Aß suppresses the transcriptional activity of cAMP-responsive element-binding protein (CREB), a master regulator of cell survival and plasticity-related gene expression. Here, we report that Aß elicits nucleocytoplasmic trafficking of Jacob, a protein that connects a NMDA-receptor-derived signalosome to CREB, in AD patient brains and mouse hippocampal neurons. Aß-regulated trafficking of Jacob induces transcriptional inactivation of CREB leading to impairment and loss of synapses in mouse models of AD. The small chemical compound Nitarsone selectively hinders the assembly of a Jacob/LIM-only 4 (LMO4)/ Protein phosphatase 1 (PP1) signalosome and thereby restores CREB transcriptional activity. Nitarsone prevents impairment of synaptic plasticity as well as cognitive decline in mouse models of AD. Collectively, the data suggest targeting Jacob protein-induced CREB shutoff as a therapeutic avenue against early synaptic dysfunction in AD.

Muskelin regulates actin-dependent synaptic changes and intrinsic brain activity relevant to behavioral and cognitive processes.

Muskelin (Mkln1) is implicated in neuronal function, regulating plasma membrane receptor trafficking. However, its influence on intrinsic brain activity and corresponding behavioral processes remains unclear. Here we show that murine Mkln1 knockout causes non-habituating locomotor activity, increased exploratory drive, and decreased locomotor response to amphetamine. Muskelin deficiency impairs social novelty detection while promoting the retention of spatial reference memory and fear extinction recall. This is strongly mirrored in either weaker or stronger resting-state functional connectivity between critical circuits mediating locomotor exploration and cognition. We show that Mkln1 deletion alters dendrite branching and spine structure, coinciding with enhanced AMPAR-mediated synaptic transmission but selective impairment in synaptic potentiation maintenance. We identify muskelin at excitatory synapses and highlight its role in regulating dendritic spine actin stability. Our findings point to aberrant spine actin modulation and changes in glutamatergic synaptic function as critical mechanisms that contribute to the neurobehavioral phenotype arising from Mkln1 ablation.

Organization of Presynaptic Autophagy-Related Processes.

Brain synapses pose special challenges on the quality control of their protein machineries as they are far away from the neuronal soma, display a high potential for plastic adaptation and have a high energy demand to fulfill their physiological tasks. This applies in particular to the presynaptic part where neurotransmitter is released from synaptic vesicles, which in turn have to be recycled and refilled in a complex membrane trafficking cycle. Pathways to remove outdated and damaged proteins include the ubiquitin-proteasome system acting in the cytoplasm as well as membrane-associated endolysosomal and the autophagy systems. Here we focus on the latter systems and review what is known about the spatial organization of autophagy and endolysomal processes within the presynapse. We provide an inventory of which components of these degradative systems were found to be present in presynaptic boutons and where they might be anchored to the presynaptic apparatus. We identify three presynaptic structures reported to interact with known constituents of membrane-based protein-degradation pathways and therefore may serve as docking stations. These are (i) scaffolding proteins of the cytomatrix at the active zone, such as Bassoon or Clarinet, (ii) the endocytic machinery localized mainly at the peri-active zone, and (iii) synaptic vesicles. Finally, we sketch scenarios, how presynaptic autophagic cargos are tagged and recruited and which cellular mechanisms may govern membrane-associated protein turnover in the presynapse.

The needs of a synapse-How local organelles serve synaptic proteostasis.

Synaptic function crucially relies on the constant supply and removal of neuronal membranes. The morphological complexity of neurons poses a significant challenge for neuronal protein transport since the machineries for protein synthesis and degradation are mainly localized in the cell soma. In response to this unique challenge, local micro-secretory systems have evolved that are adapted to the requirements of neuronal membrane protein proteostasis. However, our knowledge of how neuronal proteins are synthesized, trafficked to membranes, and eventually replaced and degraded remains scarce. Here, we review recent insights into membrane trafficking at synaptic sites and into the contribution of local organelles and micro-secretory pathways to synaptic function. We describe the role of endoplasmic reticulum specializations in neurons, Golgi-related organelles, and protein complexes like retromer in the synthesis and trafficking of synaptic transmembrane proteins. We discuss the contribution of autophagy and of proteasome-mediated and endo-lysosomal degradation to presynaptic proteostasis and synaptic function, as well as nondegradative roles of autophagosomes and lysosomes in signaling and synapse remodeling. We conclude that the complexity of neuronal cyto-architecture necessitates long-distance protein transport that combines degradation with signaling functions.

Neddylation-dependent protein degradation is a nexus between synaptic insulin resistance, neuroinflammation and Alzheimer's disease.

BACKGROUND: The metabolic syndrome is a consequence of modern lifestyle that causes synaptic insulin resistance and cognitive deficits and that in interaction with a high amyloid load is an important risk factor for Alzheimer's disease. It has been proposed that neuroinflammation might be an intervening variable, but the underlying mechanisms are currently unknown. METHODS: We utilized primary neurons to induce synaptic insulin resistance as well as a mouse model of high-risk aging that includes a high amyloid load, neuroinflammation, and diet-induced obesity to test hypotheses on underlying mechanisms. RESULTS: We found that neddylation and subsequent activation of cullin-RING ligase complexes induced synaptic insulin resistance through ubiquitylation and degradation of the insulin-receptor substrate IRS1 that organizes synaptic insulin signaling. Accordingly, inhibition of neddylation preserved synaptic insulin signaling and rescued memory deficits in mice with a high amyloid load, which were fed with a 'western diet'. CONCLUSIONS: Collectively, the data suggest that neddylation and degradation of the insulin-receptor substrate is a nodal point that links high amyloid load, neuroinflammation, and synaptic insulin resistance to cognitive decline and impaired synaptic plasticity in high-risk aging.

One-step purification of tag free and soluble lamin B1 from an E. coli bacterial expression system.

Lamin B1 is an intermediate filament protein that is a core component of the nuclear lamina. Structural studies and biochemical characterization of lamin B1 are severely hampered by the tendency of the protein to form inclusion bodies in E. coli bacterial expression systems. Therefore, the purity and consistency of the protein varies from batch to batch. In this work, we have purified a tag-free lamin B1 protein from a soluble fraction following bacterial expression. We also checked the functional properties of the purified as well as of the subsequently lyophilised protein. The current protocol helps to purify functional lamin B1 in a single step.

Molecular Mechanisms of Memory Consolidation That Operate During Sleep.

The role of sleep for brain function has been in the focus of interest for many years. It is now firmly established that sleep and the corresponding brain activity is of central importance for memory consolidation. Less clear are the underlying molecular mechanisms and their specific contribution to the formation of long-term memory. In this review, we summarize the current knowledge of such mechanisms and we discuss the several unknowns that hinder a deeper appreciation of how molecular mechanisms of memory consolidation during sleep impact synaptic function and engram formation.

Jacob, a Synapto-Nuclear Protein Messenger Linking N-methyl-D-aspartate Receptor Activation to Nuclear Gene Expression.

Pyramidal neurons exhibit a complex dendritic tree that is decorated by a huge number of spine synapses receiving excitatory input. Synaptic signals not only act locally but are also conveyed to the nucleus of the postsynaptic neuron to regulate gene expression. This raises the question of how the spatio-temporal integration of synaptic inputs is accomplished at the genomic level and which molecular mechanisms are involved. Protein transport from synapse to nucleus has been shown in several studies and has the potential to encode synaptic signals at the site of origin and decode them in the nucleus. In this review, we summarize the knowledge about the properties of the synapto-nuclear messenger protein Jacob with special emphasis on a putative role in hippocampal neuronal plasticity. We will elaborate on the interactome of Jacob, the signals that control synapto-nuclear trafficking, the mechanisms of transport, and the potential nuclear function. In addition, we will address the organization of the Jacob/NSMF gene, its origin and we will summarize the evidence for the existence of splice isoforms and their expression pattern.

Multiomics of synaptic junctions reveals altered lipid metabolism and signaling following environmental enrichment.

Membrane lipids and their metabolism have key functions in neurotransmission. Here we provide a quantitative lipid inventory of mouse and rat synaptic junctions. To this end, we developed a multiomics extraction and analysis workflow to probe the interplay of proteins and lipids in synaptic signal transduction from the same sample. Based on this workflow, we generate hypotheses about novel mechanisms underlying complex changes in synaptic connectivity elicited by environmental stimuli. As a proof of principle, this approach reveals that in mice exposed to an enriched environment, reduced endocannabinoid synthesis and signaling is linked to increased surface expression of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) in a subset of Cannabinoid-receptor 1 positive synapses. This mechanism regulates synaptic strength in an input-specific manner. Thus, we establish a compartment-specific multiomics workflow that is suitable to extract information from complex lipid and protein networks involved in synaptic function and plasticity.

Transgenic modeling of Ndr2 gene amplification reveals disturbance of hippocampus circuitry and function.

Duplications and deletions of short chromosomal fragments are increasingly recognized as the cause for rare neurodevelopmental conditions and disorders. The NDR2 gene encodes a protein kinase important for neuronal development and is part of a microduplication region on chromosome 12 that is associated with intellectual disabilities, autism, and epilepsy. We developed a conditional transgenic mouse with increased Ndr2 expression in postmigratory forebrain neurons to study the consequences of an increased gene dosage of this Hippo pathway kinase on brain circuitry and cognitive functions. Our analysis reveals reduced terminal fields and synaptic transmission of hippocampal mossy fibers, altered hippocampal network activity, and deficits in mossy fiber-dependent behaviors. Reduced doublecortin expression and protein interactome analysis indicate that transgenic Ndr2 disturbs the maturation of granule cells in the dentate gyrus. Together, our data suggest that increased expression of Ndr2 may critically contribute to the development of intellectual disabilities upon gene amplification.