Biology subjects

Reed, M.

Publications and source records attributed to Reed, M..

18 recordsLinked to original sources

bioRxiv2026

Sex-specific dynamics of activity-dependent regulation of synaptic CaMKII

Similar behavioral outcomes in males and females can mask substantial differences in the mechanisms underlying neuronal function. However, to what extent sex influences the abundance and activity-dependent changes of proteins that govern neuronal plasticity remains poorly understood. Here we investigated sex-specific regulation of calcium/calmodulin-dependent protein kinase II (CaMKII), an evolutionarily conserved regulator of synaptic stability and plasticity in fruit fly Drosophila melanogaster. We find that the steady state levels and activity-driven changes in CaMKII protein differ between female and male at both third instar larval neuromuscular junction (NMJ) and in adult fly mushroom body (MB). These sex-specific differences are downstream of the sex determination pathway, implicating somatic sex identity in the regulation of CaMKII abundance and its activity-dependent dynamics. Despite the differences in CaMKII levels between female and male flies, we observed no impairment of plasticity in either sex. Overall, our findings reveal sex-dependent regulation of both steady-state and activity-dependent levels of a highly conserved neuronal plasticity-associated protein across distinct neuronal contexts and development stages. These results further support the idea that females and males can achieve similar behavioral outcomes through different molecular mechanisms. Growing evidence that sexually dimorphic molecular responses contribute directly to sex differences in vulnerability to stress and disease underscores the importance of identifying these mechanisms and understanding their consequences.

neuroscience↗

bioRxiv2026

Presynaptic Terminals Dynamically Modulate Spontaneous Release Frequency During Early Synaptic Plasticity Through and Entropic Force Framework

Spontaneous synaptic transmission has been established as essential for the maintenance of synaptic weights during action potential-induced transmission. However, spontaneous transmission also changes during synaptic plasticity and has been shown to, in part, mediate changes in synaptic weights. Despite decades of research, a coherent framework for understanding the complex molecular processes that support presynaptic spontaneous transmission during maintenance and plasticity has remained elusive. We show here that presynapses modulate spontaneous transmission frequency during the early time-course of plasticity following entropic force theory. We use live primary hippocampal cultures as a model system and induce plasticity using an established Long-Term Potentiation (LTP) protocol. We then use a combination of electron microscopy, fluorescence microscopy, and computational modeling to show how spontaneous release frequency dynamically changes during early plasticity. We use our entropic force theory to show how the dynamically changing synaptic vesicle pool structure mediates spontaneous release changes. Lastly, we show how these changes are altered in the presence of P301L tau leading to degeneration. The results from this study provide new insights that not only help understand normal synaptic function but also aid in understanding neurodegeneration.

neuroscience↗

bioRxiv2026

Prenatal cannabinoid exposure induces sex-specific alterations in placental growth and lipid metabolism gene expression

BackgroundPrenatal cannabis use is becoming increasingly more commonplace. However, cannabis exposure is linked to adverse pregnancy outcomes, including gestational hypertension, preeclampsia, and preterm birth. The aim of this study was to determine the morphological and molecular effects of prenatal cannabinoid exposure on the placenta. MethodsPregnant Sprague-Dawley rats were exposed daily to vaporized THC (100 mg/mL) starting at gestational day (GD)5 until GD19 when dams were sacrificed and fetuses and placentas collected. Fetuses were genotyped for genetic sex and transcriptomic analysis was performed on male and female THC-exposed and control placentas. ResultsOn GD19, both the fetuses and placentas from the THC group were significantly larger than the control. When separated by sex, both male and female THC fetuses were significantly larger; however, only male THC placentas were significantly larger than male control placentas with no significant difference in placental weight between female control and THC placentas. RNA-sequencing revealed enriched biological processes related to nutrient transport and lipid catabolism, protein-lipid complex formation, and lipoprotein particle remodeling and organization. Further transcriptomic analysis determined that the differentially expressed genes and enriched biological processes related to lipid metabolism were preferentially enriched in the female THC placentas compared to the male, suggesting a sex-specific effect. DiscussionCollectively, these data present sex-specific effects of prenatal cannabinoid exposure on placental growth and global gene expression. These data also suggest that sex influences gene expression of genes related to lipid metabolism in the THC-exposed placentas.

Developmental Biology↗

bioRxiv2026

Potential of HLA-E-targeting diabodies to induce lysis of HIV-1-infected cells by CD8+ T cells

A long-lived reservoir of cells harboring intact HIV-1 provirus persists throughout decades of antiretroviral therapy and can give rise to rapid viral rebound after treatment interruption. Some cure strategies employ cytotoxic T lymphocytes (CTL) to target this reservoir; however, the applicability and efficacy of immunotherapeutic strategies involving MHC class I-restricted CTL is limited by the polymorphic nature of MHC class I molecules and their downregulation by HIV-1 Nef. The non-polymorphic non-classical class I molecule HLA-E is stably expressed on HIV-1-infected CD4+ T cells and presents a potential universal target. We generated a single-chain diabody RLP-13 that redirects CTLs to target cells presenting a well-characterized peptide derived from Mycobacterium tuberculosis in the context of HLA-E. We verified the affinity and specificity of RLP-13. Through co-culture experiments, we confirmed that RLP-13 mediates polyfunctional, HLA-agnostic CTL responses. Using an HIV-1 reporter construct encoding the target peptide, we demonstrated robust and specific elimination of the HIV-1-expressing cell population. This proof-of-concept study shows that HLA-E antigens are promising immunotherapeutic targets that can bypass the limitations of classical MHC class I antigens - allelic variation and downregulation - and that such bispecific antibodies recognizing HIV-1-derived HLA-E binding epitopes could induce elimination of productively infected cells. SummarySengupta, Bachmann et al. utilize a novel HLA-E-restricted CD3-engaging single-chain diabody to induce antigen-specific polyfunctional CTL-responses that are HLA-type-independent. They further show that such biologics have potential to eliminate HIV-1-infected cells by targeting HLA-E-binding epitopes encoded in the HIV-1 provirus.

microbiology↗

bioRxiv2025

Subacute effects of ketamine on neural correlates of reward processing

ObjectivesKetamines prohedonic properties have been linked to enhanced reward-related brain activation during the early post-infusion phase. Its effects during the subacute period ([~]2-24 h post-infusion), when psychotomimetic symptoms fade and neuroplastic adaptations emerge, are less well characterised. This study assessed ketamines subacute effects on reward processing using the Monetary Incentive Delay (MID) task. MethodsIn a randomised, placebo-controlled, crossover study, 28 healthy participants received 0.5 mg/kg racemic ketamine or placebo via 40-minute intravenous infusion. Functional magnetic resonance imaging (fMRI) was acquired [~]5 h post-infusion. Plasma concentrations of ketamine and norketamine were obtained for individual area under the curve (AUC) estimation. Analyses focused on the contrast between expected and actual trial outcomes. ResultsAt five hours post-infusion, ketamine did not significantly modulate MID task-related brain activation, despite pronounced subjective drug effects. Pharmacokinetic modelling confirmed expected ketamine and norketamine profiles, but neither drug exposure (AUC) nor subjective measures correlated with neural activation. ConclusionsProhedonic effects of ketamine may not sufficiently manifest in MID task-related activation in healthy individuals [~]5 hours after infusion. The lack of significant effects provides valuable extension of the existing literature, as ketamines effects might be confined to a more acute time window or differ in clinical populations.

neuroscience↗

bioRxiv2025

Monensin and Its Analogs Exhibit Activity Against Breast Cancer Stem-Like Cells in an Organoid Model

Monensin (MON) is a polyether ionophore antibiotic of natural origin and is an FDA-approved drug for veterinary use. Recent studies have highlighted its potential anti-cancer activity in various in vitro and in vivo models. In this study, we evaluated the anti-breast cancer activity of MON and 37 synthetic analog compounds using cell monolayer and organoid models. Through a mini-ring cell viability assay, several compounds were identified that were more potent and selective against breast cancer cells compared to non-cancerous cells, surpassing the activity of parent MON. MON and these compounds induced significant DNA fragmentation, reduced cell migration, and downregulated SOX2 expression. Furthermore, MON and the most potent analog, compound 12, reduced the percentage of CD44+/CD24-/low stem-like cells and diminished cell self-renewal properties. Proteomics analyses revealed that several pathways, including extracellular matrix organization, were significantly dysregulated by MON and compound 12 in breast cancer cells. Among these, TIMP2, a protein associated with the suppression of tumor growth and metastasis, was identified as one of the most prominently upregulated proteins by MON and compound 12 in MDA-MB-231 cells. This finding was also validated in other breast cancer and melanoma cell lines. To simulate breast cancer metastasis to the brain, a human Hybrid Organoid System: Tumor in Brain Organoid (HOSTBO) model was developed. MON and compound 12 significantly reduced Ki-67 expression within the HOSTBOs, and compound 12 significantly downregulated SOX2 expression. Collectively, MON and compound 12 significantly reduced the proliferation of breast cancer stem-like cells in the organoid models, inhibited their migration, and dysregulated markers associated with stemness, demonstrating their potential as anti-metastatic agents and warranting further clinical development. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=137 SRC="FIGDIR/small/663311v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@388862org.highwire.dtl.DTLVardef@182072dorg.highwire.dtl.DTLVardef@1a55072org.highwire.dtl.DTLVardef@447150_HPS_FORMAT_FIGEXP M_FIG C_FIG

pharmacology and toxicology↗

bioRxiv2025

Dual cholinergic and serotonergic excitatory pathways mediate oxygen sensing in the zebrafish gill

The evolution of oxygen sensing included a transition from a diffuse distribution of respiratory chemoreceptors in the gills of water-breathing vertebrates to chemoreceptor clusters confined to the pulmonary epithelium and carotid body in air-breathers. Since the excitatory neurotransmitters mediating oxygen sensing in anamniotes have never been confirmed, the origins of oxygen sensing in vertebrates have remained controversial. In gills isolated from Tg(elavl3:GCaMP6s) zebrafish expressing a genetically-encoded reporter of intracellular Ca2+ concentration ([Ca2+]i), we demonstrate that acetylcholine (ACh) and nicotine induced a dose-dependent increase in [Ca2+]i in postsynaptic sensory neurons innervating oxygen-chemoreceptive neuroepithelial cells (NECs). Hypoxic stimulation of NECs evoked a similar rise in neuronal [Ca2+]i that was abolished by nicotinic antagonist, hexamethonium. Using immunohistochemistry and RT-qPCR, we identified a novel population of ACh-containing NECs associated with sensory neurons expressing the 2 subunit of nicotinic ACh receptors. In vivo whole-larva Ca2+ imaging showed that cholinergic and hypoxic activation of the gills generated Ca2+ activity in neurons of vagal sensory ganglia with time-dependent characteristics of neurotransmission towards the hindbrain. We identified a second source of hypoxic activity in vagal sensory ganglia operating exclusively through 5-HT3 receptors and dependent upon vesicular monoamine transport (VMAT2) in the gill. We traced expression of 5-HT3 receptors to nerve terminals surrounding a separate population of serotonergic VMAT2-positive NECs. Our investigation reveals independent cholinergic and serotonergic autonomic pathways of oxygen sensing in zebrafish and provides the first physiological evidence that gill chemoreceptors may be homologues of both pulmonary and carotid body chemoreceptors in mammals. Significance statementThe excitatory neurotransmitters mediating oxygen sensing in anamniotes have never been confirmed. Thus, the origins of oxygen sensing in vertebrates have remained controversial. In transgenic zebrafish expressing a genetically-encoded reporter of intracellular Ca2+ concentration, we identified two independent pathways of oxygen sensing in the gill: one involving acetylcholine and interneurons intrinsic to the gill, and the other via serotonin acting directly upon ganglionic neurons. Both pathways resulted in excitation of vagal sensory ganglia that receive hypoxic inputs from the gills and innervate the hindbrain. We argue that gill chemoreceptors are homologues of both pulmonary and carotid body chemoreceptors in mammals.

neuroscience↗

bioRxiv2024

Oxygen chemoreceptor inhibition by dopamine D2 receptors in isolated zebrafish gills

Dopamine is an essential modulator of oxygen sensing and control of ventilation and was the first neurotransmitter described in the carotid body. Little is known of the evolutionary significance of dopamine in oxygen sensing, or whether it plays a similar role in anamniotes. In the model vertebrate, zebrafish (Danio rerio), presynaptic dopamine D2 receptor expression was demonstrated in gill neuroepithelial cells (NECs)--analogues of mammalian oxygen chemoreceptors; however, a mechanism for dopamine and D2 in oxygen sensing in the gills had not been defined. The present study tested the hypothesis that presynaptic D2 receptors provide a feedback mechanism that attenuates the chemoreceptor response to hypoxia. Using an isolated gill preparation from Tg(elavl3:GCaMP6s) zebrafish, we measured hypoxia-induced changes in intracellular Ca2+ concentration ([Ca2+]i) in NECs and postsynaptic neurons. Activation of D2 with dopamine or specific D2 agonist, quinpirole, decreased hypoxic responses in NECs; whereas D2 antagonist, domperidone, had the opposite effect. Addition of SQ22536, an adenylyl cyclase (AC) inhibitor, decreased the effect of hypoxia on [Ca2+]i, similar to dopamine. Activation of AC by forskolin partially recovered the suppressive effect of dopamine on the Ca2+ response to hypoxia. Further, we demonstrate that the response to hypoxia in postsynaptic sensory neurons was dependent upon innervation with NECs, and was subject to modulation by activation of presynaptic D2. Our results provide the first evidence of neurotransmission of the hypoxic signal at the NEC-nerve synapse in the gill and suggest that a presynaptic, modulatory role for dopamine in oxygen sensing arose early in vertebrate evolution. Key pointsO_LIFor the first time, we present an experimental model that permits imaging of intracellular Ca2+ in identified vertebrate oxygen chemoreceptors using GCaMP in a whole/intact sensing organ. C_LIO_LIThe hypoxic response of zebrafish chemoreceptors is attenuated by dopamine through a mechanism involving D2 receptors and adenylyl cyclase. C_LIO_LIZebrafish oxygen chemoreceptors send a hypoxic signal to postsynaptic (sensory) neurons. C_LIO_LIPostsynaptic neuronal responses to hypoxia are modulated by presynaptic D2 receptors, suggesting a link between chemoreceptor inhibition by dopamine and modulation of the hypoxic ventilatory response. C_LIO_LIOur results suggests that a modulatory role for dopamine in oxygen sensing arose early in vertebrate evolution. C_LI

physiology↗

bioRxiv2024

CPI203, a BET inhibitor, down-regulates a consistent set of DNA synthesis genes across a wide array of glioblastoma lines.

IntroductionGlioblastomas utilize malignant gene expression pathways to drive growth. Many of these gene pathways are not directly accessible with molecularly targeted pharmacological agents. Chromatin-modifying compounds can alter gene expression and target glioblastoma growth pathways. In this study, we utilize a systematic screen of chromatin-modifying compounds on a panel of patient-derived glioblastoma lines to identify promising compounds and their associated gene targets. MethodsFive glioblastoma cell lines were subjected to a drug screen of 106 chromatin-modifying compounds representing 36 unique drug classes to determine the twelve most promising drug classes and the best candidate inhibitors in each class. These twelve drugs were then tested with a panel of twelve patient-derived gliomasphere lines to identify growth inhibition and corresponding gene expression patterns. Overlap analysis and weighted co-expression network analysis (WCGNA) were utilized to determine potential target genes and gene pathways. ResultsThe initial drug screen identified twelve candidate pharmacologic agents for further testing. Drug sensitivity testing indicated an overall high degree of variability between gliomasphere lines. However, CPI203 was the most consistently effective compound, and the BET inhibitor class was the most consistently effective class of compounds across the gliomasphere panel. Correspondingly, most of the compounds tested had highly variable effects on gene expression between gliomasphere lines. CPI203 stood out as the only compound to induce a consistent effect on gene expression across different gliomasphere lines, specifically down-regulation of DNA-synthesis genes. Amongst the twelve tested cell lines, high expression of CDKN2A and CDKN2B distinguished more drug sensitive from more drug resistant lines. WCGNA identified two oncogenic gene modules (FBXO5 and MELK) that were effectively downregulated by CPI203 (FBXO5) and ML228 (FBXO5 and MELK). ConclusionsThe bromodomain inhibitor CPI203 induced relatively consistent effects on gene expression and growth across a variety of glioblastoma lines, specifically down-regulating genes associated with DNA replication. We propose that clinically effective BET inhibitors have the potential to induce consistent beneficial effects across a spectrum of glioblastomas.

cancer biology↗

bioRxiv2024

Targeting MDSC-HTR2B to Improve Immune Checkpoint Inhibitors in Breast to Brain Metastasis

Myeloid Derived Suppressor Cells (MDSCs) support breast cancer growth via immune suppression and non-immunological mechanisms. Although 15% of patients with breast cancer will develop brain metastasis, there is scant understanding of MDSCs contribution within the breast-to-brain metastatic microenvironment. Utilizing co-culture models mimicking a tumor-neuron-immune microenvironment and patient tissue arrays, we identified serotonergic receptor, HTR2B, on MDSCs to upregulate pNF-{kappa}B and suppress T cell proliferation, resulting in enhanced tumor growth. In vivo murine models of metastatic and intracranial breast tumors treated with FDA-approved, anti-psychotic HTR2B antagonist, clozapine, combined with immunotherapy anti-PD-1 demonstrated a significant increase in survival and increased T cell infiltration. Collectively, these findings reveal a previously unknown role of MDSC-HTR2B in breast-to-brain metastasis, suggesting a novel and immediate therapeutic approach using neurological drugs to treat patients with metastatic breast cancer.

cancer biology↗

bioRxiv2023

Identification of signalling pathways involved in gill regeneration in zebrafish

The occurrence of regeneration of the organs involved in respiratory gas exchange amongst vertebrates is heterogeneous. In aquatic animals, such as amphibians and fishes, the gills regenerate completely following resection or amputation; whereas in mammals, only partial, facultative regeneration of lung tissue occurs following injury. Given the homology between gills and lungs, the capacity of gill regeneration in aquatic species is of major interest in determining the underlying molecular or signalling pathways involved in respiratory organ regeneration. In the present study, we used adult zebrafish (Danio rerio) to characterize signalling pathways involved in the early stages of gill regeneration. Regeneration of the gills, including the gas exchange surfaces, was induced by resection of the gill filaments and observed over a period of up to 10 days. We screened for the effects of the drugs, SU5402, dorsomorphin, and LY411575, which inhibit FGF, BMP or Notch signalling, respectively, on development of the blastema. Exposure to each drug for 5 days significantly reduced blastema formation in regenerates, compared to unresected controls. In separate experiments, and under normal conditions of regeneration, we used quantitative real-time PCR and observed an increased expression of genes encoding for the bone morphogenetic factor, Bmp2b, fibroblast growth factor, Fgf8a, a transcriptional regulator (Her6) involved in Notch signalling, and Sonic Hedgehog (Shha), in regenerating gills at 10 day post-resection, compared to unresected controls. In situ hybridization confirmed that all four genes were expressed in regenerating gill tissue. This study implicates BMP, FGF, Notch and Shh signalling in gill regeneration in zebrafish. SUMMARY STATEMENTGill regeneration in zebrafish is mediated by multiple signalling pathways, including BMP, FGF, Notch and Sonic Hedgehog.

developmental biology↗

bioRxiv2022

Immunogenicity and Protective Efficacy of a SARS-CoV-2 mRNA Vaccine Encoding Secreted Non-Stabilized Spike Protein in Mice

Establishment of an mRNA vaccine platform in low- and middle-income countries (LMICs) is important to enhance vaccine accessibility and ensure future pandemic preparedness. Here, we describe the preclinical studies of a SARS-CoV-2 mRNA encoding prefusion-unstabilized ectodomain spike protein encapsulated in lipid nanoparticles (LNP) "ChulaCov19". In BALB/c mice, ChulaCov19 at 0.2, 1, 10, and 30 g given 2 doses, 21 days apart, elicited robust neutralizing antibody (NAb) and T cells responses in a dose-dependent relationship. The geometric mean titer (GMT) of micro-virus neutralizing (micro-VNT) antibody against wild-type virus was 1,280, 11,762, 54,047, and 62,084, respectively. Higher doses induced better cross-neutralizing antibody against Delta and Omicron variants. This elicited specific immunogenicity was significantly higher than those induced by homologous prime-boost with inactivated (CoronaVac) or viral vector (AZD1222) vaccine. In heterologous prime-boost study, mice primed with either CoronaVac or AZD1222 vaccine and boosted with 5 g ChulaCov19 generated NAb 7-fold higher against wild-type virus (WT) and was also significantly higher against Omicron (BA.1 and BA.4/5) than homologous CoronaVac or AZD1222 vaccination. AZD1222-prime/mRNA-boost had mean spike-specific IFN{gamma} positive T cells of 3,725 SFC/106 splenocytes, which was significantly higher than all groups except homologous ChulaCov19. Challenge study in human-ACE-2-expressing transgenic mice showed that ChulaCov19 at 1 g or 10 g protected mice from COVID-19 symptoms, prevented SARS-CoV-2 viremia, significantly reduced tissue viral load in nasal turbinate, brain, and lung tissues 99.9-100%, and without anamnestic of Ab response which indicated its protective efficacy. ChulaCov19 is therefore a promising mRNA vaccine candidate either as a primary or a boost vaccination and has entered clinical development.

immunology↗

bioRxiv2022

rTg(TauP301L)4510 mice exhibit increased VGLUT1 in hippocampal presynaptic glutamatergic vesicles and increased extracellular glutamate release

The molecular pathways that contribute to the onset of symptoms in tauopathy models, including Alzheimers Disease (AD), are difficult to distinguish because multiple changes can happen simultaneously at different stages of disease progression. Understanding early synaptic alterations and their supporting molecular pathways is essential in order to develop better pharmacological targets to treat AD. Here we focus on an early onset rTg(TauP301L)4510 tauopathy mouse model that exhibits hyperexcitability in hippocampal neurons of adult mice that is correlated with presynaptic changes and increased extracellular glutamate levels. However, it is not clear if increased extracellular glutamate is caused by presynaptic changes alone, or if presynaptic changes are a contributing factor among other factors. To determine whether pathogenic tau alters presynaptic function and glutamate release, we studied cultured hippocampal neurons at 14-18 DIV from animals of both sexes to measure presynaptic changes in tauP301L positive mice. We observed that presynaptic vesicles exhibit increased vesicular glutamate transporter 1 (VGLUT1) using immunohistochemistry of fixed cells and an established pH-sensitive green fluorescent protein approach. We show that tauP301L positive neurons exhibit a 40% increase in VGLUT1 per vesicle compared to tauP301L negative littermates. Further, we use the extracellular glutamate reporter iGluSnFR to show that increased VGLUT1 per vesicle directly translates into a 40% increase in extracellular glutamate. Together, these results show that increased extracellular glutamate levels observed in tauP301L mice are not caused by increased vesicle exocytosis probability but rather are directly related to increased VGLUT1 transporters per synaptic vesicle.

neuroscience↗

bioRxiv2022

Deletion of CEP164 in mouse photoreceptors post-ciliogenesis interrupts ciliary intraflagellar transport (IFT)

Centrosomal protein 164 (CEP164) is located at the edge of distal appendages in primary cilia and is necessary for basal body (BB) docking to the apical membrane. To investigate the function of CEP164 before and after BB docking in photoreceptors, we deleted CEP164 during retina embryonic development (Six3Cre), in postnatal rod photoreceptors (iCre75) and in mature retina using tamoxifen induction. BBs dock to the cell cortex during postnatal day 6 (P6) and extend a connecting cilium (CC) and an axoneme. P6 retina-specific knockouts (retCep164-/-) are unable to dock BBs, thereby preventing formation of a CC or outer segments (OS). In rod-specific knockouts (rodCep164-/-), Cre expression starts after P9 and CC/OS form. P16 rodCep164-/- rods have nearly normal OS lengths, and maintain OS attachment through P21 despite loss of CEP164. Intraflagellar transport components (IFT88, IFT57 and IFT140) were reduced at P16 rodCep164-/- BBs and CC tips and nearly absent at P21, indicating impaired intraflagellar transport. Nascent OS discs, labeled with a fluorescent dye on P14 and P18 and harvested on P19, showed continued rodCep164-/- disc morphogenesis but absence of P14 discs mid-distally, indicating OS instability. Tamoxifen induction with PROM1ETCre;Cep164F/F (tamCep164-/-) adult mice affected maintenance of both rod and cone OS. The results suggest that CEP164 is key towards recruitment and stabilization of IFT-B particles at BB/CC. Impairment of IFT may be the main driver of ciliary malfunction observed with hypomorphic CEP164 mutations. Author summaryCentrosomal protein of 164 kDa (CEP164) is indispensable for docking of basal bodies to apical membranes and formation of primary cilia. Homozygous truncations of CEP164 are associated with severe ciliopathy whereas missense mutations generate milder forms of NPHP. Recessive mutations of CEP164 are associated with nephronophthisis (NPHP), Meckel-Gruber (MKS) and Bardet-Biedl syndromes (BBS) in which primary cilia fail to form, or do not function correctly. We found that deletion of CEP164 in mouse photoreceptors before docking of the basal body (BB) to the apical inner segment membrane prevents elaboration of connecting cilia (CC, equivalent to transition zones) and outer segments (OSs, modified primary cilia). We also generated mouse models in which deletion of CEP164 occurs after OS are established and observed that the BB/CC and axoneme are initially stable in the absence of CEP164. Loss of CEP164, however, diminished recruitment of IFT-B proteins to the BB/CC affecting intraflagellar transport (IFT), which is required for maintenance of cilia and OSs. We propose that, in ciliopathies caused by missense mutations (e.g., X1460WextX57 associated with LCA) impairment of IFT provides a mechanism for ciliary dysfunction and disease.

neuroscience↗

bioRxiv2021

Effects of Escitalopram and Relearning on Cortical and Subcortical Grey Matter in Healthy Humans

The antidepressant effect of selective serotonin reuptake inhibitors (SSRI) is related to increased neuroplasticity during relearning. Stress-induced dendritic atrophy in key brain areas for learning and memory such as the hippocampus and prefrontal cortex is reversed by SSRI treatment. This finding is accompanied by behavioral stabilization. The aim of this study was to investigated serotonergic modulation effects on structural neuroplasticity (cortical thickness, subcortical volumes) during relearning in healthy subjects. Participants performed daily associative learning tasks over 3 weeks followed by a 3-week relearning phase combined with intake of the SSRI escitalopram or placebo. Evidence suggests that SSRIs promote the brains susceptibility to change on the basis of environment factors. We found no effect of SSRI on grey matter measures during relearning. Here, non-findings might be a consequence of the implemented intensity and duration of study interventions. With sparse literature on healthy participants in this field, future studies will have to further elucidate SSRIs properties on relearning and structural neuroplasticity.

neuroscience↗

bioRxiv2021

Escitalopram Administration, Neuroplastic Effects and Relearning: A Diffusion Tensor Imaging Study in Healthy Individuals

BackgroundNeuroplastic processes are influenced by selective serotonergic reuptake inhibitors, while learning in conjunction with the administration of serotonergic agents alters white matter microstructure in humans. The goal of this double-blind, placebo-controlled imaging study was to investigate the influence of escitalopram on white matter plasticity during (re)learning. MethodsSeventy-one healthy individuals (age = 25.6{+/-}5.0, 43 females) underwent 3 diffusion magnetic resonance imaging sessions: at baseline, after 3-weeks of associative learning (emotional/non-emotional content) and after relearning shuffled associations for an additional 3 weeks. During the relearning phase, subjects received daily escitalopram 10 mg or placebo orally. Statistical analysis was performed with statistical parametric mapping (SPM) and using sandwich estimator. ResultsA three-way and two-way rmANOVA was conducted to analyze the effects of escitalopram on AD, FA, MD and RD during the learning and relearning period. We found no significant three-way or two-way interactions for each DTI metrics (pFDR > 0.05), thus neither after 3 nor after 6 weeks we found significant changes in white matter microstructure. ConclusionWe examined neither an effect of escitalopram nor learning (or relearning) interventions on different DTI metrics. The duration and intensity of study interventions (i.e. administration of escitalopram and learning as the relearning task) might have been chosen insufficiently to induce detectable alterations. Previous studies examining the effects of SSRIs on white matter tracts in humans are underrepresented, but do mainly yield towards non-significant findings. The results implicate that escitalopram does not impact white matter microstructures in healthy subjects.

neuroscience↗

bioRxiv2020

The S1 protein of SARS-CoV-2 crosses the blood-brain barrier: Kinetics, distribution, mechanisms, and influence of ApoE genotype, sex, and inflammation

Evidence strongly suggests that SARS-CoV-2, the cause of COVID-19, can enter the brain. SARS-CoV-2 enters cells via the S1 subunit of its spike protein, and S1 can be used as a proxy for the uptake patterns and mechanisms used by the whole virus; unlike studies based on productive infection, viral proteins can be used to precisely determine pharmacokinetics and biodistribution. Here, we found that radioiodinated S1 (I-S1) readily crossed the murine blood-brain barrier (BBB). I-S1 from two commercial sources crossed the BBB with unidirectional influx constants of 0.287 {+/-} 0.024 L/g-min and 0.294 {+/-} 0.032 L/g-min and was also taken up by lung, spleen, kidney, and liver. I-S1 was uniformly taken up by all regions of the brain and inflammation induced by lipopolysaccharide reduced uptake in the hippocampus and olfactory bulb. I-S1 crossed the BBB completely to enter the parenchymal brain space, with smaller amounts retained by brain endothelial cells and the luminal surface. Studies on the mechanisms of transport indicated that I-S1 crosses the BBB by the mechanism of adsorptive transcytosis and that the murine ACE2 receptor is involved in brain and lung uptake, but not that by kidney, liver, or spleen. I-S1 entered brain after intranasal administration at about 1/10th the amount found after intravenous administration and about 0.66% of the intranasal dose entered blood. ApoE isoform or sex did not affect whole brain uptake, but had variable effects on olfactory bulb, liver, spleen, and kidney uptakes. In summary, I-S1 readily crosses the murine BBB, entering all brain regions and the peripheral tissues studied, likely by the mechanism of adsorptive transcytosis. Graphical Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

neuroscience↗

bioRxiv2020

CRYPTOCHROME suppresses the circadian proteome and promotes protein homeostasis

The daily organisation of most mammalian cellular functions is attributed to circadian regulation of clock-controlled protein expression, driven by daily cycles of CRYPTOCHROME-dependent transcriptional feedback repression. To test this, we compared the circadian proteome and phosphoproteome of wild type and CRY-deficient fibroblast cells. Strikingly, CRY-deficient cells showed a two-fold increase in circadian-regulated proteins, phosphopeptides, and K+ transport. This was accompanied by extensive remodelling of the cellular proteome overall, including reduced phosphatase and proteasome subunit expression. These adaptations rendered CRY-deficient cells more sensitive to stress, which may account for their reduced circadian robustness and contribute to the wide-ranging phenotypes of CRY-deficient mice. We suggest that CRY ultimately functions to suppress, rather than generate, daily rhythms in cellular protein abundance, thereby maintaining protein and osmotic homeostasis.

cell biology↗