Cytosolic dopamine determines hypersensitivity to blunt force trauma.

The selective vulnerability of dopaminergic neurons to trauma-induced neurodegeneration is conserved across species, from nematodes to humans. However, the molecular mechanisms underlying this hypersensitivity to blunt force trauma remain elusive. We find that extravesicular dopamine, a key driver of Parkinson's disease, extends its toxic role to the acute challenges associated with injury. Ectopic dopamine synthesis in serotonergic neurons sensitizes this resilient neuronal subtype to trauma-induced degeneration. While dopaminergic neurons normally maintain dopamine in a functional and benign state, trauma-induced subcellular redox imbalances elicit dopamine-dependent cytotoxicity. Cytosolic dopamine accumulation, through perturbations to its synthesis, metabolism, or packaging, is necessary and sufficient to drive neurodegeneration upon injury and during aging. Additionally, degeneration is further exacerbated by rapid upregulation of the rate-limiting enzyme in dopamine synthesis, cat-2, via the FOS-1 transcription factor. Fundamentally, our study in C. elegans unravels the molecular intricacies rendering dopaminergic neurons uniquely prone to physical perturbation across evolutionary lines.

Serum protein biomarker profile distinguishes acetylcholine receptor antibody seropositive myasthenia gravis patients from healthy controls.

There is an unmet need for objective disease-specific biomarkers in the heterogeneous autoimmune neuromuscular disorder myasthenia gravis (MG). This cross-sectional study identified a signature of 23 inflammatory serum proteins with proximity extension assay (PEA) that distinguishes acetylcholine receptor antibody seropositive (AChR+) MG patients from healthy controls (HCs). CCL28, TNFSF14, 4E-BP1, transforming growth factor alpha (TGF-α), and ST1A1 ranked top biomarkers. TGF-ß1 and osteoprotegerin (OPG) differed between early- and late-onset MG, whereas CXCL10, TNFSF14, CCL11, interleukin-17C (IL-17C), and TGF-α differed significantly with immunosuppressive treatment. MG patients with moderate to high disease severity had lower uPA. Previously defined MG-associated microRNAs, miR-150-5p, miR-30e-5p, and miR-21-5p, correlated inversely with ST1A1 and TNFSF14. The presented inflammatory proteins that distinguish AChR+ MG are promising serum biomarkers for validation in prospective studies to allow for molecular signatures for patient subgroup stratification and monitoring of treatment response.

Lumbar puncture increases Alzheimer's disease biomarker levels in cerebrospinal fluid of rhesus monkeys.

Cerebrospinal fluid (CSF) samples are commonly collected via lumbar puncture (LP) in both clinical and research settings for measurement of biomarkers of Alzheimer's disease (AD). To determine the effects of LP on CSF AD biomarkers, we collected CSF samples at seven different time points after an LP in rhesus monkeys. We find that amyloid-beta (Aß) and Tau levels increased significantly on day 1, peaked on day 3, and returned to baseline on day 10 after LP. The NFL levels increased significantly on day 5, peaked on day 10, and returned to baseline after day 30. The increased AD biomarker levels were mainly due to CSF outflow and deep intrathecal invasion during LP. Therefore, if LPs are repeated within a short period of time, prior LP can affect Aß and Tau levels within 10 days and NFL levels within 30 days, which may lead to clinical misdiagnosis or incorrect scientific conclusions.

Homozygous ALS-linked mutations in TARDBP/TDP-43 lead to hypoactivity and synaptic abnormalities in human iPSC-derived motor neurons.

Cytoplasmic mislocalization and aggregation of the RNA-binding protein TDP-43 is a pathological hallmark of the motor neuron (MN) disease amyotrophic lateral sclerosis (ALS). Furthermore, while mutations in TARDBP (encoding TDP-43) have been associated with ALS, the pathogenic consequences of these mutations remain poorly understood. Using CRISPR-Cas9, we engineered two homozygous knock-in induced pluripotent stem cell lines carrying mutations in TARDBP encoding TDP-43A382T and TDP-43G348C, two common yet understudied ALS TDP-43 variants. Motor neurons (MNs) differentiated from knock-in iPSCs had normal viability and displayed no significant changes in TDP-43 subcellular localization, phosphorylation, solubility, or aggregation compared with isogenic control MNs. However, our results highlight synaptic impairments in both TDP-43A382T and TDP-43G348C MN cultures, as reflected in synapse abnormalities and alterations in spontaneous neuronal activity. Collectively, our findings suggest that MN dysfunction may precede the occurrence of TDP-43 pathology and neurodegeneration in ALS and further implicate synaptic and excitability defects in the pathobiology of this disease.

Synaptic cell-adhesion molecule latrophilin-2 is differentially directed to dendritic domains of hippocampal neurons.

Hippocampal pyramidal cells possess elaborate dendritic arbors with distinct domains that are targeted with input-specific synaptic sites. This synaptic arrangement is facilitated by synaptic cell-adhesion molecules that act as recognition elements to connect presynaptic and postsynaptic neurons. In this study, we investigate the organization of the synaptic recognition molecule latrophilin-2 at the surface of pyramidal neurons classified by spatial positioning and action potential firing patterns. Surveying two hippocampal neurons that highly express latrophilin-2, late-bursting CA1 pyramidal cells and early-bursting subiculum pyramidal cells, we found the molecule to be differentially positioned on their respective dendritic compartments. Investigating this latrophilin-2 positioning at the synaptic level, we found that the molecule is not present within either the pre- or postsynaptic terminal but rather is tightly coupled to synapses at a perisynaptic location. Together these findings indicate that hippocampal latrophilin-2 distribution patterning is cell-type specific, and requires multiple postsynaptic neurons for its synaptic localization.

Caenorhabditis elegans neuropeptide NLP-27 enhances neurodegeneration and paralysis in an opioid-like manner during fungal infection.

The nervous system of metazoans is involved in host-pathogen interactions to control immune activation. In Caenorhabditis elegans, this includes sleep induction, mediated by neuropeptide-like proteins (NLPs), which increases the chance of survival after wounding. Here we analyzed the role of NLP-27 in the infection of C. elegans with the nematode-trapping fungus Arthrobotrys flagrans. Early responses of C. elegans were the upregulation of nlp-27, the induction of paralysis (sleep), and neurodegeneration of the mechanosensing PVD (Posterior Ventral Process D) neurons. Deletion of nlp-27 reduced neurodegeneration during fungal attack. Induction of nlp-27 was independent of the MAP kinase PMK-1, and expression of nlp-27 in the hypodermis was sufficient to induce paralysis, although NLP-27 was also upregulated in head neurons. NLP-27 contains the pentapeptide YGGYG sequence known to bind the human µ- and κ-type opioid receptors suggesting NLP-27 or peptides thereof act on opioid receptors. The opioid receptor antagonist naloxone shortened the paralysis time like overexpression of NLP-27.

Loss of the polarity protein Par3 promotes dendritic spine neoteny and enhances learning and memory.

The Par3 polarity protein is critical for subcellular compartmentalization in different developmental processes. Variants of PARD3, encoding PAR3, are associated with intelligence and neurodevelopmental disorders. However, the role of Par3 in glutamatergic synapse formation and cognitive functions in vivo remains unknown. Here, we show that forebrain-specific Par3 conditional knockout leads to increased long, thin dendritic spines in vivo. In addition, we observed a decrease in the amplitude of miniature excitatory postsynaptic currents. Surprisingly, loss of Par3 enhances hippocampal-dependent spatial learning and memory and repetitive behavior. Phosphoproteomic analysis revealed proteins regulating cytoskeletal dynamics are significantly dysregulated downstream of Par3. Mechanistically, we found Par3 deletion causes increased Rac1 activation and dysregulated microtubule dynamics through CAMSAP2. Together, our data reveal an unexpected role for Par3 as a molecular gatekeeper in regulating the pool of immature dendritic spines, a rate-limiting step of learning and memory, through modulating Rac1 activation and microtubule dynamics in vivo.

Inhibition of CD44 suppresses the formation of fibrotic scar after spinal cord injury via the JAK2/STAT3 signaling pathway.

Fibrotic scar is one of the main impediments to axon regeneration following spinal cord injury (SCI). In this study, we found that CD44 was upregulated during the formation of fibrotic scar, and blocking CD44 by IM7 caused downregulation of fibrosis-related extracellular matrix proteins at both 2 and 12 weeks post-spinal cord injury. More Biotinylated dextran amine (BDA)-traced corticospinal tract axons crossed the scar area and extended into the distal region after IM7 administration. A recovery of motor and sensory function was observed based on Basso Mouse Scale (BMS) scores and tail-flick test. In vitro experiments revealed that inhibiting CD44 and JAK2/STAT3 signaling pathway decreased the proliferation, differentiation, and migration of fibroblasts induced by the inflammatory supernatant. Collectively, these findings highlight the critical role of CD44 and its downstream JAK2/STAT3 signaling pathway in fibrotic scar formation, suggesting a potential therapeutic target for SCI.

Serum amyloid A and mitochondrial DNA in extracellular vesicles are novel markers for detecting traumatic brain injury in a mouse model.

This study investigates the potential use of circulating extracellular vesicles' (EVs) DNA and protein content as biomarkers for traumatic brain injury (TBI) in a mouse model. Despite an overall decrease in EVs count during the acute phase, there was an increased presence of exosomes (CD63+ EVs) during acute and an increase in microvesicles derived from microglia/macrophages (CD11b+ EVs) and astrocytes (ACSA-2+ EVs) in post-acute TBI phases, respectively. Notably, mtDNA exhibited an immediate elevation post-injury. Neuronal (NFL) and microglial (Iba1) markers increased in the acute, while the astrocyte marker (GFAP) increased in post-acute TBI phases. Novel protein biomarkers (SAA, Hp, VWF, CFD, CBG) specific to different TBI phases were also identified. Biostatistical modeling and machine learning identified mtDNA and SAA as decisive markers for TBI detection. These findings emphasize the importance of profiling EVs' content and their dynamic release as an innovative diagnostic approach for TBI in liquid biopsies.

Vinorelbine causes a neuropathic pain-like state in mice via STING and MNK1 signaling associated with type I interferon induction.

Type I interferons (IFNs) increase the excitability of dorsal root ganglia (DRGs) neurons via MNK-eIF4E signaling to promote pain sensitization in mice. Activation of stimulator of interferon response cGAMP interactor 1 (STING) signaling is pivotal for type I IFN induction. We hypothesized that vinorelbine, a chemotherapeutic and activator of STING, would cause a neuropathic pain-like state in mice via STING signaling in DRG neurons associated with IFN production. Vinorelbine caused tactile allodynia and grimacing in wild-type (WT) mice and increased p-IRF3, type I IFNs, and p-eIF4E in peripheral nerves. Supporting our hypothesis, vinorelbine failed to induce IRF3-IFNs-MNK-eIF4E in StingGt/Gt mice and, subsequently, failed to cause pain. The vinorelbine-elicited increase of p-eIF4E was not observed in Mknk1-/- (MNK1 knockout) mice in peripheral nerves consistent with the attenuated pro-nociceptive effect of vinorelbine in these mice. Our findings show that activation of STING signaling in the periphery causes a neuropathic pain-like state through type I IFN signaling to DRG nociceptors.

Growth-suppressor microRNAs mediate synaptic overgrowth and behavioral deficits in Fragile X mental retardation protein deficiency.

Abnormal neuronal and synapse growth is a core pathology resulting from deficiency of the Fragile X mental retardation protein (FMRP), but molecular links underlying the excessive synthesis of key synaptic proteins remain incompletely defined. We find that basal brain levels of the growth suppressor let-7 microRNA (miRNA) family are selectively lowered in FMRP-deficient mice and activity-dependent let-7 downregulation is abrogated. Primary let-7 miRNA transcripts are not altered in FMRP-deficiency and posttranscriptional misregulation occurs downstream of MAPK pathway induction and elevation of Lin28a, a let-7 biogenesis inhibitor. Neonatal restoration of brain let-7 miRNAs corrects hallmarks of FMRP-deficiency, including dendritic spine overgrowth and social and cognitive behavioral deficits, in adult mice. Blockade of MAPK hyperactivation normalizes let-7 miRNA levels in both brain and peripheral blood plasma from Fmr1 KO mice. These results implicate dysregulated let-7 miRNA biogenesis in the pathogenesis of FMRP-deficiency, and highlight let-7 miRNA-based strategies for future biomarker and therapeutic development.

Cross-species comparative hippocampal transcriptomics in Alzheimer's disease.

Alzheimer's disease (AD) is a multifactorial pathology, with most cases having a sporadic origin. Recently, knock-in (KI) mouse models, such as the novel humanized amyloid-ß (hAß)-KI, have been developed to better resemble sporadic human AD. METHODS: Here, we compared hippocampal publicly available transcriptomic profiles of transgenic (5xFAD and APP/PS1) and KI (hAß-KI) mouse models with early- (EOAD) and late- (LOAD) onset AD patients. RESULTS: The three mouse models presented more Gene Ontology biological processes terms and enriched signaling pathways in common with LOAD than with EOAD individuals. Experimental validation of consistently dysregulated genes revealed five altered in mice (SLC11A1, S100A6, CD14, CD33, and C1QB) and three in humans (S100A6, SLC11A1, and KCNK). Finally, we identified 17 transcription factors potentially acting as master regulators of AD. CONCLUSION: Our cross-species analyses revealed that the three mouse models presented a remarkable similarity to LOAD, with the hAß-KI being the more specific one.

Altering heparan sulfate suppresses cell abnormalities and neuron loss in Drosophila presenilin model of Alzheimer Disease.

We examined the function of heparan-sulfate-modified proteoglycans (HSPGs) in pathways affecting Alzheimer disease (AD)-related cell pathology in human cell lines and mouse astrocytes. Mechanisms of HSPG influences on presenilin-dependent cell loss were evaluated in Drosophila using knockdown of the presenilin homolog, Psn, together with partial loss-of-function of sulfateless (sfl), a gene specifically affecting HS sulfation. HSPG modulation of autophagy, mitochondrial function, and lipid metabolism were shown to be conserved in human cell lines, Drosophila, and mouse astrocytes. RNA interference (RNAi) of Ndst1 reduced intracellular lipid levels in wild-type mouse astrocytes or those expressing humanized variants of APOE, APOE3, and APOE4. Neuron-directed knockdown of Psn in Drosophila produced apoptosis and cell loss in the brain, phenotypes suppressed by reductions in sfl expression. Abnormalities in mitochondria, liposomes, and autophagosome-derived structures in animals with Psn knockdown were also rescued by reduction of sfl. These findings support the direct involvement of HSPGs in AD pathogenesis.

Distinct GABAergic modulation of timing-dependent LTP in CA1 pyramidal neurons along the longitudinal axis of the mouse hippocampus.

Synaptic plasticity in the hippocampus underlies episodic memory formation, with dorsal hippocampus being instrumental for spatial memory whereas ventral hippocampus is crucial for emotional learning. Here, we studied how GABAergic inhibition regulates physiologically relevant low repeat spike timing-dependent LTP (t-LTP) at Schaffer collateral-CA1 synapses along the dorsoventral hippocampal axis. We used two t-LTP protocols relying on only 6 repeats of paired spike-firing in pre- and postsynaptic cells within 10 s that differ in postsynaptic firing patterns. GABAA receptor mechanisms played a greater role in blocking 6× 1:1 t-LTP that recruits single postsynaptic action potentials. 6× 1:4 t-LTP that depends on postsynaptic burst-firing unexpectedly required intact GABAB receptor signaling. The magnitude of both t-LTP-forms decreased along the dorsoventral axis, despite increasing excitability and basal synaptic strength in this direction. This suggests that GABAergic inhibition contributes to the distinct roles of dorsal and ventral hippocampus in memory formation.

Inhibition of 7α,26-dihydroxycholesterol biosynthesis promotes midbrain dopaminergic neuron development.

Dysregulated cholesterol metabolism has been linked to neurodegeneration. We previously found that free, non-esterified, 7α,(25R)26-dihydroxycholesterol (7α,26-diHC), was significantly elevated in the cerebrospinal fluid of patients with Parkinson's disease (PD). In this study we investigated the role of 7α,26-diHC in midbrain dopamine (mDA) neuron development and survival. We report that 7α,26-diHC induces apoptosis and reduces the number of mDA neurons in hESC-derived cultures and in mouse progenitor cultures. Voriconazole, an oxysterol 7α-hydroxylase (CYP7B1) inhibitor, increases the number of mDA neurons and prevents the loss of mDA neurons induced by 7α,26-diHC. These effects are specific since neither 7α,26-diHC nor voriconazole alter the number of Islet1+ oculomotor neurons. Furthermore, our results suggest that elevated 24(S),25-epoxycholesterol, which has been shown to promote mDA neurogenesis, may be partially responsible for the effect of voriconazole on mDA neurons. These findings suggest that voriconazole, and/or other azole CYP7B1 inhibitors may have implications in PD therapy development.

Location of the axon initial segment assembly can be predicted from neuronal shape.

The axon initial segment (AIS) is located at the proximal axon demarcating the boundary between axonal and somatodendritic compartments. The AIS facilitates the generation of action potentials and maintenance of neuronal polarity. In this study, we show that the location of AIS assembly, as marked by Ankyrin G, corresponds to the nodal plane of the lowest-order harmonic of the Laplace-Beltrami operator solved over the neuronal shape. This correlation establishes a coupling between location of AIS assembly and neuronal cell morphology. We validate this correlation for neurons with atypical morphology and neurons containing multiple AnkG clusters on distinct neurites, where the nodal plane selects the appropriate axon showing enriched Tau. Based on our findings, we propose that Turing patterning systems are candidates for dynamically governing AIS location. Overall, this study highlights the importance of neuronal cell morphology in determining the precise localization of the AIS within the proximal axon.

Necl-1/CADM3 regulates cone synapse formation in the mouse retina.

In vertebrates, retinal neural circuitry for visual perception is organized in specific layers. The outer plexiform layer is the first synaptic region in the visual pathway, where photoreceptor synaptic terminals connect with bipolar and horizontal cell processes. However, molecular mechanisms underlying cone synapse formation to mediate OFF pathways remain unknown. This study reveals that Necl-1/CADM3 is localized at S- and S/M-opsin-containing cones and dendrites of type 4 OFF cone bipolar cells (CBCs). In Necl-1-/- mouse retina, synapses between cones and type 4 OFF CBCs were dislocated, horizontal cell distribution became abnormal, and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors were dislocated. Necl-1-/- mice exhibited aberrant short-wavelength-light-elicited signal transmission from cones to OFF CBCs, which was rescued by AMPA receptor potentiator. Additionally, Necl-1-/- mice showed impaired optokinetic responses. These findings suggest that Necl-1 regulates cone synapse formation to mediate OFF cone pathways elicited by short-wavelength light in mouse retina.

Gap junctions fine-tune ganglion cell signals to equalize response kinetics within a given electrically coupled array.

Retinal ganglion cells (RGCs) summate inputs and forward a spike train code to the brain in the form of either maintained spiking (sustained) or a quickly decaying brief spike burst (transient). We report diverse response transience values across the RGC population and, contrary to the conventional transient/sustained scheme, responses with intermediary characteristics are the most abundant. Pharmacological tests showed that besides GABAergic inhibition, gap junction (GJ)-mediated excitation also plays a pivotal role in shaping response transience and thus visual coding. More precisely GJs connecting RGCs to nearby amacrine and RGCs play a defining role in the process. These GJs equalize kinetic features, including the response transience of transient OFF alpha (tOFFα) RGCs across a coupled array. We propose that GJs in other coupled neuron ensembles in the brain are also critical in the harmonization of response kinetics to enhance the population code and suit a corresponding task.

Pleiotropic brain function of whirlin identified by a novel mutation.

Despite some evidence indicating diverse roles of whirlin in neurons, the functional corollary of whirlin gene function and behavior has not been investigated or broadly characterized. A single nucleotide variant was identified from our recessive ENU-mutagenesis screen at a donor-splice site in whirlin, a protein critical for proper sensorineural hearing function. The mutation (head-bob, hb) led to partial intron-retention causing a frameshift and introducing a premature termination codon. Mutant mice had a head-bobbing phenotype and significant hyperactivity across several phenotyping tests. Lack of complementation of head-bob with whirler mutant mice confirmed the head-bob mutation as functionally distinct with compound mutants having a mild-moderate hearing defect. Utilizing transgenics, we demonstrate rescue of the hyperactive phenotype and combined with the expression profiling data conclude whirlin plays an essential role in activity-related behaviors. These results highlight a pleiotropic role of whirlin within the brain and implicate alternative, central mediated pathways in its function.

Non-canonical type 1 cannabinoid receptor signaling regulates night visual processing in the inner rat retina.

Type 1 cannabinoid receptors (CB1Rs) are expressed in major retinal neurons within the rod-pathway suggesting a role in regulating night visual processing, but the underlying mechanisms remain poorly understood. Using acute rat retinal slices, we show that CB1R activation reduces glutamate release from rod bipolar cell (RBC) axon terminals onto AII and A17 amacrine cells through a pathway that requires exchange proteins directly activated by cAMP (EPAC1/2) signaling. Consequently, CB1R activation abrogates reciprocal GABAergic feedback inhibition from A17 amacrine cells. Moreover, the activation of CB1Rs in vivo enhances and prolongs the time course of the dim-light rod-driven visual responses, an effect that was eliminated when both GABAA and GABAC receptors were blocked. Altogether, our findings underscore a non-canonical mechanism by which cannabinoid signaling regulates RBC dyad synapses in the inner retina to regulate dim-light visual responses to fine-tune night vision.