Glioblastoma (GBM) is an aggressive brain tumor, characterized by infiltrative cells at the tumor border, for which radiotherapy is the first-line therapy. Although radiotherapy helps with tumor control in the short term, there is evidence that it alters various biophysical aspects of the tumor microenvironment and contributes to increased invasiveness in the long run. Radiation causes strand breaks in hyaluronan (HA), the most common component of the brain extracellular matrix (ECM), leading to the formation of low molecular weight (MW) HA fragments (HA-F). This is relevant because literature indicates that HA-F can act as a chemoattractant for cancerous cells. Furthermore, radiotherapy also alters fluid flow of the brain, which has also been implicated in invasion of GSCs. Thus, the objective of this work is to test the effect of interstitial fluid flow (IFF) on migration of patient derived glioma stem-like cells (GSCs) towards radiation-induced HA-F and standard HA-F. Under static conditions, GSC lines with high expression of CD44 generally showed increased migration with increasing HA-F concentrations and HA-F MWs. Chemotaxis of GSCs towards irradiation-induced HA-F was dose dependent, with varying trends depending on the cell line. GSCs with low expression of CD44 did not exhibit a dose dependent increase in migration. When comparing invasion under static and flow conditions, it was found that HA-Fs mitigated the increased invasion normally observed under flow. Further work is needed to better understand the link between irradiation-mediated HA degradation, IFF, and GSC invasion. This work will have implications for improving GBM response to radiotherapy.

Arc (activity-regulated cytoskeleton-associated protein) is a hub protein known for regulating synaptic plasticity, such as long-term depression (LTD), long-term potentiation (LTP) and homeostatic scaling. Although Arc has been well investigated for modulating these processes, the cell biological mechanisms and control of Arc function are little understood. In that sense, nanobodies present a new possibility for probing and interfering in those dynamics, due to its unique properties when compared to conventional antibodies. Those characteristics include a higher chemical stability and efficiency, possibility of being recombinantly expressed in different cells and also can reach hidden or hard access epitopes, favoring its use as an intracellular tool. For that purpose, 6 nanobodies clones were developed and validated by Ishizuka and collaborators, showing their ability to bind to native and purified Arc protein, and had their binding sites characterized. Two of those nanobodies bound to Arc C-terminal domain (CTD), with homology to retroviral Gag capsid domain. The Arc CTD is has two lobes, the N-lobe and C-lobe. The N-lobe harbors a ligand binding pocket for several postsynaptic Arc binding partners, including the transmembrane AMPA receptor regulatory protein gamma 2 (TARPγ2), also known as Stargazin. This auxiliary protein acts on AMPA receptor trafficking and anchoring in the cellular membrane. Thus, the current study shows the process of development, validation and preliminary data in neuronal transfection aiming functional studies of Arc protein within AMPA receptor dynamics, using nanobodies as functional blockers.

Exposure to high ambient temperature (HAT) is associated with increased mortality, weight loss, and metabolic function in birds, all of which are likely downstream effects of reduced food intake. While the mechanisms mediating the physiological responses to heat stress are documented, the neural mechanisms mediating behavioral responses are poorly understood. The aim of the present study was thus to investigate the hypothalamic mechanisms mediating HAT-induced anorexia in four-day old chickens. In the following experiments, chicks were exposed to either a thermoneutral, or HAT environment. The purpose of these experiments were to evaluate the effect of HAT on food intake, on the efficacy of appetite-associated factors, and to determine hypothalamic sites of action and changes in gene expression in response to HAT. In summary, our results demonstrated that HAT causes a reduction in food intake that is likely mediated by downregulation of orexigenic factors via activation of the stress response.

The aim of our study was to determine the best combination of clinical factors in predicting post-concussion syndrome (PCS) – assessed with the British Columbia Post-Concussion Symptom Inventory (BC-PSI) – at 3 months following mild TBI. Patients with mTBI (16-60 years) were defined as Glasgow Coma Scale (GCS) score between 13-15, loss of consciousness (LOC)<30 min and post-traumatic amnesia (PTA)<24 hours. Clinical/demographic factors were age, sex, GCS, LOC, PTA, traumatic intracranial finding on MRI (within 72 h), other somatic injuries, plasma-derived proteins associated with CNS damage (Tau, GFAP and NFL) and inflammation (IFN, IL-8, Eotaxin, MIP-1, MCP-1, IP-10, IL-17A, IL-9, TNF, FGF-basic PDGF and IL-1ra). Blood was drawn 24-72 hours, 2 weeks, and 3 months after injury. The discriminatory value (AUC analysis) of each individual blood biomarker predictor was at chance level. For each timepoint, combining all blood biomarkers also did not predict PCS at clinically relevant levels (AUCs 0.66-0.71). At 24-72 hours, the blood biomarkers included in the best-subset model predicting PCS were TNF, IL-9, IL-17A and IL-8. At both 2-weeks and 3 weeks, a combination of PDGF, IL-8 and IFN, best predicted PCS. The baseline clinical injuries predicting PCS together with 24-72 hours blood were woman, longer PTA, traumatic intracranial findings, lower GCS, and with 2-week and 3-month blood: woman, longer PTA, and traumatic intracranial findings. In conclusion, our study shows that blood biomarker level in both acute, subacute and chronic phase of MTBI were associated with PCS, but these biomarkers did not have clinical specificity/sensitivity to reliably predict PCS.

An intact BBB maintains homeostasis by regulating the passage of macromolecules into the brain and preventing the passage of neurotoxins and bacteria. Additionally, it limits peripheral immune cell infiltration and regulates cerebral blood flow. Traumatic brain injury (TBI) involves a complex cascade of events, starting with an initial insult followed by secondary injury such as blood brain barrier (BBB) breakdown. In the context of traumatic brain injury (TBI), disruption of the BBB is linked to worse clinical outcomes in patients in both the acute and chronic phases. The underlying mechanisms responsible for regulating BBB permeability following TBI remain to be elucidated. EphA4, a membrane bound receptor tyrosine kinase, has become of interest in many neurological disorders such as TBI, ALS, Alzheimer’s disease, and Parkinson’s disease. Our lab’s findings indicate that conditional deletion of EphA4 on endothelial cells reduced BBB disruption following the cortical controlled impact (CCI) model of TBI. These data correlate with improved CBF, behavioral recovery, junctional protein expression and reduced lesion volume compared to WT mice. Additionally, infiltrating peripheral immune cell quantification and analysis revealed significant differences in peripheral immune cell recruitment and gene expression by RNA sequencing of CD45+ cells, indicating EphA4 may mediate endothelial-immune crosstalk in the BBB niche. These data improve our understanding of the effect of endothelial EphA4 on BBB breakdown following TBI injury.

Hippocampal CA2 neurons exhibit different synaptic plasticity responses to distal (perforant path, PP) verses proximal (Schaffer collateral, SC) dendritic inputs. Strong tetanization of distal PP inputs readily induces long-term potentiation (LTP) in CA2 neurons, while strong tetanization of proximal SC inputs does not. The molecular mechanisms underlying this functional heterogeneity are unknown. Recently, our laboratory reported a heterogeneity in the spatial distribution of mitochondria in CA2 neuron dendrites. Specifically, distal dendrites harbor larger mitochondria with enriched expression of the mitochondrial calcium uniporter (MCU). Here we test the hypothesis that the distinct mitochondrial properties of CA2 neuron distal dendrites promote the expression of LTP, and conversely, the lack thereof in the proximal dendrites results in LTP-resistant responses. Using CA2-specific MCU knockout mice (MCUfl/fl; Amigo2-cre+) and control mice (MCUfl/fl; Amigo2-cre-), we compared the plasticity profiles of distal PP and proximal SC synaptic inputs in acute hippocampal slices. Extracellular field potential recordings of CA2 neurons were made in response to strong tetanization of distal PP or proximal SC inputs. Consistent with observations in wildtype mice, preliminary results in cre- control mice, show distal PP stimulation results in robust LTP (163±24%, n=5) while proximal SC stimulation does not (108±7%, n=4). However, in CA2 MCU KO mice the plasticity profile was flipped, with proximal SC stimulation producing LTP (123±9%, n=5) and distal PP stimulation yielding reduced LTP (142±10%, n=2). These preliminary results suggest that molecularly distinct populations of mitochondria confer unique synaptic properties essential to CA2 neuronal function.

Glioblastoma (GBM) is characterized by exceptionally low survival and resistance to treatment that is driven by a propensity of glioma stem cells (GSCs) to invade the brain parenchyma and reseed tumors along flow paths. Pathological interstitial fluid flow (IFF) arising from the pressure gradient between the GBM tumor and the surrounding tissue drives this invasion and is dependent on distinct molecular mechanisms in the tumor microenvironment (TME). One mechanism anticipated to underlie IFF-enhanced invasion is dependent on the response of sphingosine-1-phosphate receptors (S1PRs) to sphingosine-1-phosphate delivered by flow. S1PR3 directs migration and physiological responses of various cell types throughout the CNS and may be a viable target. However, the utility of S1PR3 targeting is dependent on interpatient heterogeneity and patient-specific factors like cellular composition of the tumor microenvironment. Thus, we used immunohistochemistry to correlate S1PR3 protein expression at the GBM invasive front with overall survival (OS) across a heterogeneous group of patients. Then, we localized S1PR3 to GSCs in xenografts and ALDH1L1+ astrocytes and IBA1+ microglia in patient samples at the invasive front. We plan to investigate the efficacy of S1PR3 inhibition across patient-specific models that incorporate these cell types within the tumor microenvironment.

This tool is intended to help patients with Parkinson’s disease visualize the complex, subjective symptoms they experience. Non-motor symptoms such as pain, constipation, depression, and lethargy are common in Parkinson’s disease patients, as are motor symptoms such as rigidity, slowness of movement, and tremor. All patients do not exhibit consistent symptoms and complain of a variety of pains and conditions in sporadic locations. There is currently no cure for Parkinson’s disease, and various medications can only relieve symptoms. Because the response to anti-Parkinsonian drugs varies from patient to patient, it is critical to observe and comprehend the drug response following administration. Using this tool, we can quickly identify Parkinson’s disease symptoms that need to be checked in a variety of ways. First, the patient can express his or her symptoms using a color scale ranging from 0 to 3 on a human body figure. Any symptoms that the patient finds inconvenient, including motor and non-motor symptoms, can be depicted in the figure. Following that, emotions such as depression, helplessness, anger, and happiness can be expressed by rating them on a scale of 0 to 100 points. Finally, patients can check the time of day they take the drug on a 24-hour scale and express the change that occurs after taking the drug with a color on a 0 to 3-point scale. Because patients’ symptoms can vary according to the circadian cycle, they are allowed to rate their symptoms on a 24-hour scale. This tool has the advantage of allowing for more efficient and detailed communication between patients and medical personnel. More research and development are required.

Over 100,000 US citizens died from overdoses April 2020-May 2021. The Emergency Department (ED) Bridge to Treatment study aims were to identify rate of successful bridging from ED to outpatient treatment for patients screened with an opioid use disorder (OUD) in the ED and referred into care, following either buprenorphine initiation or home induction with a prescription. Factors studied include linkage with peer specialists, prescribing of buprenorphine, rapidity of access to outpatient treatment, and retention in treatment. During the seventeen-month study 41 ED physicians completed required waiver. April 2019 – August 2020, 401 ED patients were identified with OUD, 202 screened as appropriate for buprenorphine outpatient care. Buprenorphine was initiated in ED for 158/202 (78.2%) patients based on history and/or a withdrawal scale. Initiation on buprenorphine predicted successful appearance in ambulatory clinic (OR: 2.22, p= .036). Of 158 started on buprenorphine in the ED, 115 (72.9%) transitioned to follow up. Overall, 139 patients (69%) of 202 patients appropriate for ambulatory treatment successfully arrived at the clinic (in person or virtually). Among 132 patients admitted into the clinic, retention for one month or more was 82.6% (109/132). Patients linked with a peer were twice as likely to cross bridge than patients without a peer (OR=2.103; p=.09). Study was repeated during the pandemic, September 2020 – August 2021: 71% of patients discharged from the ED and referred to ambulatory treatment were seen at clinic (108/152 patients) and peer linkages significantly affected crossing bridge from the ED to their outpatient prescriber visit.

Current HIV antiretroviral therapies are unable to cross the blood-brain barrier and target viral reservoirs in the brain, including perivascular macrophages (PVMs). Previously, we and other groups have demonstrated the upregulation of colony-stimulating factor 1 receptor (CSF1R) in the PVMs of simian immunodeficiency virus (SIV)-infected rhesus macaques. Here we investigate the ability of BLZ945, a CSF1R inhibitor, to target SIV-infected PVMs and reduce brain viral burden during acute SIV infection. Nine adult male rhesus macaques were infected with SIVmac251 and received either no treatment (n=3), low dose (10mg/kg; n=3), or high dose BLZ945 (30mg/kg; n=3) orally daily from 10 days post infection until study termination at 30 days. Post-mortem brain tissue and liver tissue were examined for changes in PVM populations, microglia, neuroimmune mRNA profile, or pathological evidence of liver toxicity. In the brain, high and low dose BLZ945 treated animals showed a significant decrease in the number of CD163 and CD206 PVM cells which correlated with decreased viral loads, but no change in P2RY12+ microglia, showing that BLZ945 selectively ablates brain PVMs. Analysis of the mRNA neuroimmune profile further showed down regulation of key disease associated inflammatory pathways with BLZ945 treatment. No significant liver pathology indicating drug toxicity was associated with BLZ945. Our results indicate the ability of BLZ945 to selectively kill PVMs and reduce brain viral loads in acute SIV infection without observed liver toxicity. Future studies will examine the ability of BLZ945 to treat chronic SIV/HIV infection and prevent viral rebound after treatment cessation.

Although glioblastoma (GBM) is the most common malignant brain cancer, and of the most aggressive, there is no cure, and only 5% of patients live 5 years following diagnosis. Standard treatment for a GBM is surgery, followed by radiation and chemotherapy. However, nearly all GBM patients will experience tumor recurrence, leading to a 5% average survival after 5 years. Recent findings from the Sheng Laboratory have shown that phosphoinositide 3-kinase (PI3K) signaling, particularly the PI3K catalytic subunit beta (PIK3CB), confers a prognostic significance in recurrent GBM. Blocking PIK3CB inactivates PI3K, inhibits viability, and slows down growth of PI3K-dependent GBM (expressing high levels of PIK3CB), while having no effect on PI3K-independent GBM (expressing low levels of PIK3CB). This has raised a critical question of how to target PI3K-independent GBM, particularly those expressing low levels of PIK3CB (PIK3CB-low). Given that PIK3CB-low GBM is independent of PI3K and lacks therapeutic targets, our objective in this study is to identify potential drug targets that can be used to effectively treat PIK3CB-low GBM. Specifically, we aim to retrospectively analyze data using Dep Map to locate a list of potential genes essential for survival of GBM cells that lowly express PIK3CB. Using this list, we aim to identify key gene candidates that will then be selectively knocked out in the GBM cell line. If these are viable candidates, then targeting of these candidates will induce significant cell death and inhibition of growth in GBM independent of PI3K, thus identifying a novel therapeutic option for these patients.

Eukaryotic initiation factor 4E (eIF4E) is crucial for cap-dependent mRNA translation, and phosphorylation of eIF4E on Ser209 regulates function in brain. However, the function of eIF4E phosphorylation in translation and long-term synaptic plasticity is undetermined. Here we show that phospho-ablated eIF4ES209A knockin mice are profoundly impaired in dentate gyrus LTP maintenance in vivo, while basal perforant path-evoked synaptic efficacy and LTP induction is intact. mRNA cap-pulldown assays showed that phosphorylation is required for synaptic activity-induced discharge of translational repressors (CYFIP1/FMRP) from eIF4E and enhanced initiation complex formation. Using global ribosomal profiling, we identified phospho-eIF4E-dependent translation of the Wnt pathway in LTP. Surprisingly, -catenin massively associated with cap-bound eIF4E following LTP induction in wildtype, but not eIF4ES209A mice. These results demonstrate a critical role for stimulus-evoked eIF4E phosphorylation in dentate gyrus LTP maintenance, remodeling of the cap-binding complex, and specific Wnt translation.

Identification of peripheral biomarkers for diagnosis, prognosis, and basic research would allow for access to information only available through invasive methods. Alterations in the whole blood RNA transcriptome could inform biomarkers of Parkinson’s disease and its progression. We utilized 4,871 longitudinal whole blood RNA-seq samples from 1,603 participants from the Parkinson’s Progression Marker Initiative cohort. Samples have an average of 100 million read pairs and have significantly altered expression (2,019 differentially expressed genes). Here, we propose the use of feature reduction and machine learning for the classification of disease and healthy control participants using the PPMI data. We use differentially expressed genes as a filter for feature selection and evaluated six different classification models using RNA-seq data alone and a separate analysis that included the sex of participants. The inclusion of participant age with RNA sequencing of whole blood improves machine learning accuracy in Parkinson’s disease. Additionally, we identify misclassified cases as patients whose primary diagnosis was a neurological symptoms or the diagnosis changed midtrial. Further research will investigate the progression markers that are linked to these misclassifications.

High-frequency irreversible electroporation (H-FIRE) utilizes pulsed electric fields to precisely and nonthermally ablate brain tumors while transiently disrupting the peritumoral blood-brain barrier (pBBB), which can enhance therapeutic delivery to invasive tumor margins. Mechanisms of H-FIRE-induced pBBB disruption remain incompletely characterized. We hypothesize that bystander effects of H-FIRE tumor cell ablation, specifically the release of tumor-derived extracellular vesicles (TDEVs), mediate pBBB endothelium disruption. F98 glioma, LL/2 Lewis lung carcinoma, and bEnd.3 cerebral endothelial cell lines modelled primary and metastatic brain cancer and BBB endothelium, respectively. BEnd.3 monolayers were exposed to supernatants of H-FIRE-treated cancer cells to model bystander effects of treatment, and endothelial cell response was assessed via microscopic observation and apoptosis quantification. TDEVs were isolated from supernatants of cancer cells treated with 0, 1,500, or 3,000 V/cm H-FIRE, and characterized via nanoparticle tracking analysis and transmission electron microscopy. Transwell® BBB endothelium models were exposed to TDEVs to evaluate the effect of H-FIRE-induced TDEVs on pBBB endothelium permeability. Supernatants of H-FIRE-treated cancer cells induced rapid bEnd.3 morphologic changes and monolayer disruption in a dose-dependent manner, without biologically significant apoptosis, suggesting that bystander effects mediate pBBB endothelium disruption following H-FIRE brain tumor ablation. Release of TDEVs consistent with exosomes increased following 1,500 V/cm H-FIRE treatment and decreased following 3,000 V/cm treatment for both tumor cell lines. TDEVs isolated from supernatants of 3,000 V/cm-treated tumor cells increased endothelium permeability in the Transwell® model, suggesting that H-FIRE modulates TDEV cargo in a way that disrupts the function of the BBB endothelium.

Zfp189 is one CREB-target gene which itself encodes an unstudied NAc neuronal transcription factor that has been demonstrated to regulate transcriptional networks in neuropsychiatric disorders. Preliminary data reveals that using the CRISPR/dCas9-mediated CREB delivery to the Zfp189 CRE site increases Zfp189 mRNA levels in the NAc and decreases reward associations for mild doses of cocaine, indicative of tolerance. To examine the downstream relationship between ZFP189 and physiological drug response, three re-engineered ZFP189 variants were used. The ZFP189 variants used were: ZFP189WT (the endogenous gene with transcriptionally inhibitory KRAB domain), ZFP189DN (maintains binding domain but lacks KRAB domain), and ZFP189VPR (KRAB domain replaced with the powerful transcriptional activator VP64-p65-Rta (VPR)). Mice received one of these ZFP variants to the NAc via viral-mediated gene transfer. We then performed a drug locomotor sensitization assay with saline, cocaine or morphine. In response to cocaine, the ZFP189VPR moved significantly more than the ZFP189WT group. Even following dissipation of viral expression, the ZFP189VPR group maintains heightened cocaine locomotor sensitization. More interestingly, this increased locomotion appears to be unique to cocaine, as there is no difference in locomotor activity between the ZFP variant groups in response to saline or morphine administration. RNA sequencing revealed that the differences in behavioral response to cocaine across the variant groups coincided with differences transcriptional changes. These results could suggest a drug specific effect of ZFP189 with cocaine and possibly other stimulants. Combined with the CRISPR/dCas9 results, CREB may be induced to interact at the Zfp189 site in response to cocaine use.

Alzheimer’s Disease (AD) is a neurodegenerative disease process manifesting clinically with cognitive impairment and diverse pathological symptoms including neuroinflammation. IL6 is a multifaceted cytokine involved in inflammation. IL6 signals classically through the membrane bound receptor or by transsignaling with the IL6/soluble IL6R (sIL6R) complex and membrane bound glycoprotein 130 (gp130). Transsignaling has been demonstrated as the primary mechanism of IL6 mediated neurodegeneration. The IL6R rs2228145 coding polymorphism (Asp358Ala) (C allele) has been demonstrated to increase CSF and plasma sIL6R levels. To determine whether IL6 trans-signaling has a role in AD, we genotyped the C allele and assayed IL6/IL6r concentrations in paired plasma and CSF samples obtained from 120 participants, 66 with normal cognition, 46 mild cognitive impairment, and 8 probable AD. Inheritance of the C allele and measures of plasma IL6 and sIL6R were correlated with cognitive status as well as clinical data, including the: Montreal Cognitive Assessment (MoCA), modified Preclinical Alzheimer’s Cognitive Composite (mPACC), cognitive domain scores, and CSF concentrations of ptau181, β-amyloid (Aβ) Aβ40 and Aβ42 to determine trans-signaling associations with indices of AD symptoms and pathology. We found that inheritance of the C allele and elevated plasma/CSF sIL6r levels were correlated with poorer scores on mPACC, MoCA and memory domain, higher CSF ptau and lower CSF Aβ42/40. Our results suggest IL6 trans-signaling and the possession of the C allele may be modifiers of AD symptoms and pathology. These results warrant further study, as this subgroup of patients may be ideally responsive to IL6 receptor blocking therapies.

Brain development is orchestrated by fine but complex spatiotemporal cell-to-cell communication, one of the most important being Wnt signaling. Besides other functions, Wnt/Planar Cell Polarity pathway (Wnt/PCP) is crucial for midbrain morphogenesis and cell fate decisions of midbrain dopaminergic (mDA) neurons. This lineage is affected for example in patients with Parkinson’s disease, Attention deficit hyperactivity disorder (ADHD), Autism spectrum disorder, or Tourette’s syndrome. Nevertheless, Wnt/PCP signaling and its physiological consequences are still poorly understood. We previously showed that proneurotrophin receptor SorCS2 is required for dopaminergic innervation and wiring, while SorCS2 loss of function causes an ADHD-like phenotype in mice (Neuron 2014; Transl Psychiatry 2021). Indeed, SorCS2 was recently identified as a risk gene for ADHD. By combining proteomics, RNA sequencing, biochemical methods, high-resolution imaging, and functional experiments in frog, fish, and mice, this comprehensive study identifies novel signaling crosstalk where SorCS2 interacts with the core Wnt/PCP receptor Ror2 to regulate its correct activity during mDA lineage development. Moreover, we formulate a molecular mechanism of how the Ror2-SorCS2 signaling complex directs embryogenesis across vertebrates emphasizing its conserved function. These observations are highly relevant for future clinical applications for patients with abnormal Wnt or SorCS2 signaling activity.

Anti N-methyl-d-aspartate receptor (NMDAR) encephalitis is a specific autoimmune CNS disorder that decouples electrochemical synapses from their neuronal network, causing seizures, neuropsychiatric symptoms, movement problems, and autonomic dysfunction. Although well studied in the adult population, the clinical characteristics and potential triggers in pediatric cases of anti-NMDAR encephalitis are not well understood. Our objective was to characterize the patients with anti-NMDAR encephalitis and identify the most common presenting symptoms and etiologies. We retrospectively analyzed patients with anti-NMDAR at Texas Children’s Hospital (TCH) between 2010 and 2021. Of the 65 pediatric cases at TCH, our cohort is 65% female and 62% Hispanic, which is 1.6 times higher than the demographics of our TCH patient population and that of the Houston Metropolitan area at large (Harris County, 39% Hispanic). The average age of onset in our pediatric cohort was 7.2 years (range 3 months to 17.9 years). Post-herpetic NMDAR encephalitis and ovarian teratoma associated encephalitis made up 12.3% and 4.6% respectively. Among the idiopathic NMDAR encephalitis group, the most common presenting symptom was focal weakness associated with altered gait and speech regression. Within our cohort, 100% had behavioral/cognitive symptoms, 79% had seizures, 73% had speech problems, 67% had movement disorder, and 61% had memory deficits. Our study describes the clinical characteristics which help define the presenting symptoms and potential etiologies in a heterogenous population from the largest single center pediatric cohort of anti-NMDAR encephalitis to date.

CA2 is an understudied subregion of the hippocampus that is essential for social memory and is enriched in receptors for the social neuropeptides oxytocin and vasopressin. Bath application of either neuropeptide induces synaptic plasticity selectively at CA2 synapses. However, the downstream molecular mechanisms supporting this plasticity remain unclear. Previously, our lab generated subregion-specific dendritic transcriptomes to identify RNAs enriched in CA2 dendrites that may contribute to plasticity. We hypothesize that social neuropeptide-receptor mediated signaling selectively engages local translation of CA2-enriched dendritic RNAs to promote synaptic plasticity. To first optimize cell-specific visualization of de novo protein translation, we bath-applied the t-RNA analog, puromycin, to label newly synthesized proteins in live slices. Slices were fixed after 30 minutes and processed for puromycin immunohistochemistry. Puromycin fluorescence was highest in DG compared to other regions, followed by CA3 and CA1/CA2, indicating that baseline levels of translation differ by cell-type. Simultaneously, we treated slices with the group 1 mGluR agonist, DHPG, to induce translation-dependent plasticity to determine our protocol’s sensitivity in detecting increases in receptor-mediated translation. DHPG-induced puromycin fluorescence varied by region and condition suggesting that receptor-mediated translation is also regulated in a cell-type specific manner. In future experiments, changes in de novo translation of candidate dendritic RNAs will be measured using puromycin combined with the proximity ligation assay (puro-PLA). Visualizing what, when, and where dendritic proteins are actively synthesized is fundamental to our understanding of the regulatory mechanisms supporting the synaptic changes underlying learning.

A threat to safety in a predator-prey interaction is a psychological stressor that induces physiological/hormonal changes, altering overall food intake and appetite preferences. The increased propensity to eat high-calorie “palatable” food particularly appears when the stressful situation ends. However, little is known about specific neural mechanisms that promote high-fat diet (HFD) overconsumption after a threat. Using in vivo Ca2+ imaging, we found that proenkephalin (Penk)-expressing-lateral hypothalamic (LH) neurons of mice are highly activated in the presence of cat urine, a predator scent stimuli (PSS). A day after exposure to PSS, the Penk-expressing LH (LH Penk) neurons show more potentiated activity during the first eating bout of HFD, whereas chronic inhibition of the same neurons normalizes the PSS-induced HFD overconsumption. PSS elevates corticosterone levels, which induces long-term activation of LHPenk neurons. Indeed, pharmacological treatment with corticosterone promotes HFD consumption the next day and elicits a corresponding increase in the activity of LH Penk neurons in response to HFD. Thus, we identify LH Penk neurons as a critical neural substrate comprising threat-induced neuronal adaptation to induce emotional overconsumption of palatable foods following a threat.

Positron emission tomography (PET) imaging with prostate specific membrane antigen (PSMA) binding tracers has been found incidentally to demonstrate uptake in CNS tumors. Following the encouraging findings of several such case reports, there is a growing interest in the potential application of PSMA-targeted PET imaging for diagnostics, theranostics, and monitoring of CNS tumors. This is a systematic literature review on PSMA binding tracers in CNS tumors. A PubMed search was conducted, including preclinical and clinical reports. One hundred and twelve records were identified, and after screening, 56 were included in the final report. Tissue studies demonstrated PSMA expression in tumor vascular endothelial cells, without expression in normal brain tissue, though the extent and intensity of staining varied by anti-PSMA antibody and methodology. Most included studies reported on gliomas, which showed strong PSMA ligand uptake and more favorable tumor to background ratios than other PET tracers. There are also case reports demonstrating PSMA ligand uptake in prostate cancer brain metastases, non-prostate cancer brain metastases, and meningiomas. We also review the properties of the various PSMA-binding radiotracers available. Therapeutic and theranostic applications of PSMA binding tracers have been studied, including labeled alpha- and beta-ray emitting isotopes, as well as PSMA targeting in directing MRI-guided focused ultrasound. There is a potential application for PSMA-targeted PET in neuro-oncology as a combination of diagnostic and therapeutic use, as a theranostic modality for managing CNS tumors. Further research is needed regarding the mechanism(s) of PSMA expression in CNS tumors and its differential performance by tumor type.

Perisynaptic astrocyte processes (PAPs) are fine terminal structures that are located on the distal processes of astrocytes. These PAPs contain numerous neurotransmitter receptors, ion channels, cell-adhesion molecules, and synaptogenic molecules, allowing astrocytes to contribute to synapse homeostasis, development, and stabilization. Despite decades of research indicating astrocytes enwrap or contact synaptic elements, little known regarding the molecular mechanisms that trigger PAP arrival to a synapse. Brain derived neurotrophic factor (BDNF) has been highly implicated as a molecule underlying both synapse development and plasticity in neurons. In both developing and highly active synapses, BDNF release from pre- or post-synaptic elements binds to its receptor TrkB to enhance synapse maturation. Our recently published work demonstrates the surprising finding that astrocytes express high levels of TrkB. We identified the truncated TrkB receptor, TrkB.T1, as the predominant TrkB receptor in cortex that is predominantly expressed in astrocytes relative to other cell populations. Direct genetic manipulation of TrkB.T1 globally and in astrocytes alone is sufficient to reduce astrocyte morphological maturation both in vitro and in vivo, alters mature astrocyte gene expression in vivo, and reduces excitatory synapse formation in vitro. Utilizing a magnetic cell-sorting approach to develop pure astrocyte neuron co-cultures, our preliminary immunohistochemistry data reveal that TrkB.T1 knockout (KO) astrocytes fail to enwrap glutamatergic synapses in vitro. Ongoing studies using astrocyte neuron co-cultures and experience dependent plasticity in the mouse barrel cortex in vivo are aimed at examining BDNF/TrkB.T1 signaling as a mechanism underlying PAP arrival to developing synapses. Alterations in synaptic function and development have been implicated in multiple neurodevelopmental disorders. This work will advance the understanding of the role of astrocytes in synaptic development and offer new therapeutic targets where synapse development is implicated.

Epilepsy is a neurological condition characterized by recurring, spontaneous seizures, often attributed to underlying imbalances in excitation and inhibition in the brain. Many cellular dysfunctions have been linked to epileptic brain activity and a variety of acquired epilepsy models, including alterations in inhibitory interneurons and extracellular matrix (ECM) structures. Perineuronal nets (PNNs) are highly negatively charged, lattice-like formations of the ECM which typically surround parvalbumin-positive interneurons in the cerebral cortex and assist in the persistent firing of those cells. Of particular interest are the functional differences of PNNs in healthy versus epileptic brains, as well as how PNNs may be altered in various acquired epilepsy models. In the present study, we have characterized PNNs in postmortem human temporal lobe epilepsy tissue and compared their structures and cellular markers to non-epileptic tissue. We examined our pilocarpine mouse model of acquired epilepsy to recapitulate these findings, and in doing so have observed that a population of cells in the hippocampal CA1 region displays PNNs post-onset of status epilepticus. We sought to identify these cells and investigate their properties. We have observed that they are in fact interneurons, likely basket cells, via immunohistochemical staining, and furthermore show that the electrophysiological activity of the interneurons from pilocarpine-treated mice that developed status epilepticus indeed differs from that of control mice. We propose that our findings signify a functional difference in these cells triggered by pilocarpine-induced status epilepticus, as indicated by the development of surrounding PNN structures.

Folate is a key methyl donor with important biological functions including DNA methylation regulation. Recently, folate has been linked to tRNA cytosine-5 methylation (m5C) and translation in mammalian mitochondria. However, the influence of folate intake on mRNA m5C modification and translation remains largely unknown. Here, we provide transcriptome-wide landscapes of m5C modification in poly(A)-enriched RNAs together with mRNA transcription and translation profiles for mouse neural stem cells (NSCs) cultured in three different concentrations of folate. NSCs cultured in these conditions were found to have distinct mRNA methylation profiles. Despite uncovering only a few differentially expressed genes, hundreds of differentially translated genes were identified in NSCs with folate deficiency or supplementation. The differentially translated genes induced by low folate are associated with cytoplasmic translation and mitochondrial function, while the differentially translated genes induced by high folate are associated with increased neural stem cell proliferation. Interestingly, compared to total mRNAs, polysome mRNAs contained high levels of m5C. Furthermore, integrative analysis indicated a transcript-specific relationship between RNA m5C methylation and mRNA translation efficiency. Altogether, our study reports a transcriptome-wide influence of folate on mRNA m5C methylation and translation in NSCs and reveals a potential link between mRNA m5C methylation and mRNA translation.

Physiological barriers are found throughout the body, and their integrity is key for maintaining healthy functions. These barriers, established by monolayers of endothelial and epithelial cells, restrict transport to enable different microenvironments on either side of the monolayer. Due to the importance of physiological barriers, many in vitro models have been developed for investigating barrier properties, most using track-etched membranes. These models rely on endothelial or epithelial monolayers cultured on engineered basement membrane (BM) constructs that are designed to mimic the in vivo BM. Current BM mimics do not faithfully recapitulate key in vivo physiological properties such as BM thickness, porosity, stiffness, and fibrous composition. Here, we use networks of precisely arranged nanofibers to form physiologically relevant basement membranes for a microfluidic blood-brain barrier model. Our nanofiber networks form ultra-thin (1 – 3 μm), ultra-porous (~88%) fibrous basement-membrane mimics. We show that our nanofibers networks enable close contact between endothelial monolayers and supporting cell types such as pericytes and astrocytes across the membrane, which are known to regulate barrier tightness through direct contact and signaling. Cytoskeletal staining reveals barrier formation on the nanofiber membranes with ubiquitous localization of tight junction proteins at areas of cell-cell contact. The nanofiber mimics can be easily incorporated in both transwell and microfluidic devices, allowing for adoption in areas of barrier models and offer new opportunities in modeling human physiology, drug discovery, cell migration studies, and barrier dysfunction studies.

Disruptions in frontostriatal dopaminergic-mediated reward learning and valuation processes may be central to the etiology and maintenance of depression (Heller et al., 2009; Pizzagalli, 2010). These disruptions have neural and behavioral correlates that have been shown to predict symptom improvement following established treatment (Brown et al., 2021; Huys et al., 2020). Previous work suggests that directly targeting this dysfunction with a single session of a directed learning paradigm can alter learning at a short timescale, but it is not yet known if such treatment leads to long-lasting changes in learning (Brown, in prep). Accordingly, we implemented a multi-session behavioral learning paradigm that trains individuals to attend to specific cues in a reinforcement learning (RL) task in aims of producing specific and long-lasting changes in learning patterns. We hypothesized that our learning paradigm would produce enduring and generalizable recovery of deficits in computationally-based learning components important to depression. 947 participants were recruited from Amazon Mechanical Turk to complete one of 14 different versions of our modified RL task for up to 8 subject visits. In addition to the modified task, participants completed the Positive and Negative Affect Schedule, the Depression, Stress, and Anxiety Subscales, and demographic questionnaires. Results indicate decreased depression scores across visits for two conditions and alterations in learning rate and reward sensitivity associated with decreased symptom scores for an additional condition. These results suggest the potential of noninvasive and cost-effective behavioral learning paradigms that directly target disrupted learning processes and produce changes in symptoms implicated in depression.

Social reintegration can be a significant difficulty for Veterans with posttraumatic stress disorder (PTSD), and associated symptoms of anger and aggression can be a barrier to treatment. Prediction error-like signals encoded in the striatum have been implicated in social learning, and recent studies have related impairments in prediction-error signaling with various types of stress, including acute, chronic, and early-life stressors. Our study seeks to understand the impact of PTSD and its treatment on social cooperation and learning among Veterans. Thirty Veterans, enrolled in a six week residential treatment program for PTSD at the Salem VA Medical Center, performed the Multi-Round Trust (MRT) task before and after treatment. The MRT task can be used to assess cooperation and learning over ten sequential exchanges of trust. Within each round, Veterans can send up to $20 to an anonymous partner. The amount sent is tripled, and the partner can then return any portion of the tripled amount. Prior work suggests that mutually beneficial cooperation emerges when individuals adjust investments and repayments based on deviations in reciprocity, a type of social prediction error. In preliminary results, we find that symptoms of PTSD are negatively related to cooperation, operationalized as the amount invested, and improvements in PTSD symptoms following treatment are associated with increases in cooperation. These findings are consistent with the hypothesis that PTSD disrupts Veterans’ ability to cooperate and learn within social exchanges, and ongoing analyses will examine treatment-related changes in neural correlates of social learning.