Adenosine deficiency facilitates CA1 synaptic hyperexcitability in the presymptomatic phase of a knock in mouse model of Alzheimer's disease.

The disease's trajectory of Alzheimer's disease (AD) is associated with and worsened by hippocampal hyperexcitability. Here we show that during the asymptomatic stage in a knock in mouse model of Alzheimer's disease (APPNL-G-F/NL-G-F; APPKI), hippocampal hyperactivity occurs at the synaptic compartment, propagates to the soma and is manifesting at low frequencies of stimulation. We show that this aberrant excitability is associated with a deficient adenosine tone, an inhibitory neuromodulator, driven by reduced levels of CD39/73 enzymes, responsible for the extracellular ATP-to-adenosine conversion. Both pharmacologic (adenosine kinase inhibitor) and non-pharmacologic (ketogenic diet) restorations of the adenosine tone successfully normalize hippocampal neuronal activity. Our results demonstrated that neuronal hyperexcitability during the asymptomatic stage of a KI model of Alzheimer's disease originated at the synaptic compartment and is associated with adenosine deficient tone. These results extend our comprehension of the hippocampal vulnerability associated with the asymptomatic stage of Alzheimer's disease.

Transcriptional response of the heart to vagus nerve stimulation.

Heart failure is a major clinical problem, with treatments involving medication, devices, and emerging neuromodulation therapies such as vagus nerve stimulation (VNS). Considering the ongoing interest in using VNS to treat cardiovascular disease, it is important to understand the genetic and molecular changes developing in the heart in response to this form of autonomic neuromodulation. This experimental animal (rat) study investigated the immediate transcriptional response of the ventricular myocardium to selective stimulation of vagal efferent activity using an optogenetic approach. Vagal preganglionic neurons in the dorsal motor nucleus of the vagus nerve were genetically targeted to express light-sensitive chimeric channelrhodopsin variant ChIEF and stimulated using light. RNA sequencing of the left ventricular myocardium identified 294 differentially expressed genes (false discovery rate < 0.05). Qiagen Ingenuity Pathway Analysis (IPA) highlighted 118 canonical pathways that were significantly modulated by vagal activity, of which 14 had a z score of ≥2/≤-2, including EIF-2, IL-2, integrin, and NFAT-regulated cardiac hypertrophy. IPA revealed the effect of efferent vagus stimulation on protein synthesis, autophagy, fibrosis, autonomic signaling, inflammation, and hypertrophy. IPA further predicted that the identified differentially expressed genes were the targets of 50 upstream regulators, including transcription factors (e.g., MYC and NRF1) and microRNAs (e.g., miR-335-3p and miR-338-3p). These data demonstrate that the vagus nerve has a major impact on the myocardial expression of genes involved in the regulation of key biological pathways. The transcriptional response of the ventricular myocardium induced by stimulation of vagal efferents is consistent with the beneficial effect of maintained/increased vagal activity on the heart.NEW & NOTEWORTHY This experimental animal study investigated the immediate transcriptional response of the ventricular myocardium to selective stimulation of vagal efferent activity. Vagal stimulation induced significant transcriptional changes in the heart involving the pathways controlling autonomic signaling, inflammation, fibrosis, and hypertrophy. This study provides the first direct evidence that myocardial gene expression is modulated by the activity of the autonomic nervous system.

Astrocytes produce nitric oxide via nitrite reduction in mitochondria to regulate cerebral blood flow during brain hypoxia.

During hypoxia, increases in cerebral blood flow maintain brain oxygen delivery. Here, we describe a mechanism of brain oxygen sensing that mediates the dilation of intraparenchymal cerebral blood vessels in response to reductions in oxygen supply. In vitro and in vivo experiments conducted in rodent models show that during hypoxia, cortical astrocytes produce the potent vasodilator nitric oxide (NO) via nitrite reduction in mitochondria. Inhibition of mitochondrial respiration mimics, but also occludes, the effect of hypoxia on NO production in astrocytes. Astrocytes display high expression of the molybdenum-cofactor-containing mitochondrial enzyme sulfite oxidase, which can catalyze nitrite reduction in hypoxia. Replacement of molybdenum with tungsten or knockdown of sulfite oxidase expression in astrocytes blocks hypoxia-induced NO production by these glial cells and reduces the cerebrovascular response to hypoxia. These data identify astrocyte mitochondria as brain oxygen sensors that regulate cerebral blood flow during hypoxia via release of nitric oxide.

Royal College of Ophthalmologists' National Ophthalmology Database study of cataract surgery: report 11, techniques and complications of local anesthesia for cataract surgery in the United Kingdom.

PURPOSE: To describe variation in local anesthesia techniques and complications over a 10-year period for cataract surgery in the United Kingdom. SETTING: Reporting centers to the Royal College of Ophthalmologists (RCOphth) National Ophthalmology Database (NOD). DESIGN: Retrospective cross-sectional register-based study. METHODS: Data from the RCOphth NOD were used. Eligible for analysis were 1 195 882 cataract operations performed using local anesthesia between April 1, 2010, and March 31, 2020, in 80 centers. RESULTS: Overall, topical anesthesia alone was used in 152 321 operations (12.7%), combined topical and intracameral in 522 849 (43.7%), sub-Tenon in 461 175 (38.6%), and peribulbar/retrobulbar in 59 537 (5.0%). In National Health Service (NHS) institutions, 48.3% of operations were topical with/without intracameral vs 88.7% in independent sector treatment centers (ISTCs). 45.9% were sub-Tenon in NHS vs 9.6% in ISTCs. 5.8% were peribulbar/retrobulbar in NHS vs 1.7% in ISTCs. Anesthetic complication rates decreased from 2.7% in the 2010 NHS year to 1.5% in the 2019 NHS year (overall, 2.1% for NHS; 0.2% for ISTCs). Overall anesthetic complication rates were 0.3%, 0.3%, 3.5%, and 3.1% for topical alone, combined topical/intracameral, sub-Tenon, and peribulbar/retrobulbar, respectively. Complication rates were higher for sharp-needle anesthesia (peribulbar/retrobulbar) in patients taking warfarin rather than direct oral anticoagulants (4.8% vs 3.1%; P = .024). Considerable variation was observed between centers on anesthetic choices and anesthetic complication rates. CONCLUSIONS: Combined topical and intracameral is the most common choice of anesthesia for cataract surgery in the United Kingdom and is associated with lower anesthetic-related complication rates than sub-Tenon and peribulbar/retrobulbar anesthesia. Variation in the anesthetic choice exists between centers and between NHS and ISTC sectors.

The Royal College of Ophthalmologists' National Ophthalmology Database Study of Cataract Surgery: Report 12, Risk factors for suprachoroidal haemorrhage during cataract surgery.

OBJECTIVE: To establish the incidence of acute intraoperative suprachoroidal haemorrhage (AISH) during cataract surgery and identify the risk factors for this complication. METHODS: Data from the Royal College of Ophthalmologists' National Ophthalmology Database was analysed. During the 11-year study period, from 01/04/2010 to 31/03/2021, 709 083 operations performed on 498 170 patients from 65 centres were eligible for inclusion. RESULTS: AISH occurred in 0.03% (204/709 083, approximately 1 in 3 500) of eligible cataract operations performed during the study period. Posterior capsule rupture was the risk factor most strongly associated with AISH (OR: 17.6, 95% CI: 12.4-24.9, p < 0.001). Other ocular risk factors identified were raised intraocular pressure (IOP) preoperatively (OR: 3.7, 95% CI: 2.5-5.5, p < 0.001), glaucoma (OR: 1.7, 95% CI: 1.2-2.4, p = 0.004). Risk increased with age and patients aged over 90 years were at greatest risk (OR: 6.7, 95% CI: 3.5-12.8, p < 0.001). The addition of intracameral anaesthetic when performing surgery under topical anaesthetic appears to be protective (OR: 0.5, 95% CI: 0.3-0.8, p = 0.003), compared to topical anaesthetic alone. There was a 16-fold increase in the incidence of vision loss when AISH occurred. CONCLUSIONS: The risk of AISH during modern cataract surgery is approximately 1 in 3 500 and is associated with a significant increase in the risk of vision loss should it occur. Posterior capsule rupture is the risk factor most strongly associated with AISH. Preoperative IOP control is a modifiable risk factor. The use of intracameral anaesthesia may reduce the risk of AISH.

Increasing Student Confidence in Writing: Integrating Authentic Manuscript Writing into an Online 8-Week Research Program.

In various formats, students at the secondary and postsecondary levels participate in multiweek authentic science research projects. There have been many papers explaining the operations of such programs, but few have provided explicit instruction on how to incorporate authentic communication practices into the student research process. In this paper, we describe how we integrated primary literature into an 8-week online research program for 8th to 11th graders. Each week, students were introduced to a specific section of a primary research article reflecting different stages of their research project, and they were guided on how to write that specific section for their own research paper. By the end of the program, students had an outline or first draft of a primary research paper based on their research. Following completion of the program, student participants reported greater self-efficacy and confidence in scientific writing. Here, we describe our approach and provide an adaptable framework for integrating primary literature into research projects.

Neural stem cells traffic functional mitochondria via extracellular vesicles.

Neural stem cell (NSC) transplantation induces recovery in animal models of central nervous system (CNS) diseases. Although the replacement of lost endogenous cells was originally proposed as the primary healing mechanism of NSC grafts, it is now clear that transplanted NSCs operate via multiple mechanisms, including the horizontal exchange of therapeutic cargoes to host cells via extracellular vesicles (EVs). EVs are membrane particles trafficking nucleic acids, proteins, metabolites and metabolic enzymes, lipids, and entire organelles. However, the function and the contribution of these cargoes to the broad therapeutic effects of NSCs are yet to be fully understood. Mitochondrial dysfunction is an established feature of several inflammatory and degenerative CNS disorders, most of which are potentially treatable with exogenous stem cell therapeutics. Herein, we investigated the hypothesis that NSCs release and traffic functional mitochondria via EVs to restore mitochondrial function in target cells. Untargeted proteomics revealed a significant enrichment of mitochondrial proteins spontaneously released by NSCs in EVs. Morphological and functional analyses confirmed the presence of ultrastructurally intact mitochondria within EVs with conserved membrane potential and respiration. We found that the transfer of these mitochondria from EVs to mtDNA-deficient L929 Rho0 cells rescued mitochondrial function and increased Rho0 cell survival. Furthermore, the incorporation of mitochondria from EVs into inflammatory mononuclear phagocytes restored normal mitochondrial dynamics and cellular metabolism and reduced the expression of pro-inflammatory markers in target cells. When transplanted in an animal model of multiple sclerosis, exogenous NSCs actively transferred mitochondria to mononuclear phagocytes and induced a significant amelioration of clinical deficits. Our data provide the first evidence that NSCs deliver functional mitochondria to target cells via EVs, paving the way for the development of novel (a)cellular approaches aimed at restoring mitochondrial dysfunction not only in multiple sclerosis, but also in degenerative neurological diseases.

Mechanosensory Signaling in Astrocytes.

Mechanosensitivity is a well-known feature of astrocytes, however, its underlying mechanisms and functional significance remain unclear. There is evidence that astrocytes are acutely sensitive to decreases in cerebral perfusion pressure and may function as intracranial baroreceptors, tuned to monitor brain blood flow. This study investigated the mechanosensory signaling in brainstem astrocytes, as these cells reside alongside the cardiovascular control circuits and mediate increases in blood pressure and heart rate induced by falls in brain perfusion. It was found that mechanical stimulation-evoked Ca2+ responses in astrocytes of the rat brainstem were blocked by (1) antagonists of connexin channels, connexin 43 (Cx43) blocking peptide Gap26, or Cx43 gene knock-down; (2) antagonists of TRPV4 channels; (3) antagonist of P2Y1 receptors for ATP; and (4) inhibitors of phospholipase C or IP3 receptors. Proximity ligation assay demonstrated interaction between TRPV4 and Cx43 channels in astrocytes. Dye loading experiments showed that mechanical stimulation increased open probability of carboxyfluorescein-permeable membrane channels. These data suggest that mechanosensory Ca2+ responses in astrocytes are mediated by interaction between TRPV4 and Cx43 channels, leading to Cx43-mediated release of ATP which propagates/amplifies Ca2+ signals via P2Y1 receptors and Ca2+ recruitment from the intracellular stores. In astrocyte-specific Cx43 knock-out mice the magnitude of heart rate responses to acute increases in intracranial pressure was not affected by Cx43 deficiency. However, these animals displayed lower heart rates at different levels of cerebral perfusion, supporting the hypothesis of connexin hemichannel-mediated release of signaling molecules by astrocytes having an excitatory action on the CNS sympathetic control circuits.SIGNIFICANCE STATEMENT There is evidence suggesting that astrocytes may function as intracranial baroreceptors that play an important role in the control of systemic and cerebral circulation. To function as intracranial baroreceptors, astrocytes must possess a specialized membrane mechanism that makes them exquisitely sensitive to mechanical stimuli. This study shows that opening of connexin 43 (Cx43) hemichannels leading to the release of ATP is the key central event underlying mechanosensory Ca2+ responses in astrocytes. This astroglial mechanism plays an important role in the autonomic control of heart rate. These data add to the growing body of evidence suggesting that astrocytes function as versatile surveyors of the CNS metabolic milieu, tuned to detect conditions of potential metabolic threat, such as hypoxia, hypercapnia, and reduced perfusion.

Combination of In Situ Lcn2 pRNA-RNAi Nanotherapeutics and iNSC Transplantation Ameliorates Experimental SCI in Mice.

Spinal cord injury (SCI) is a debilitating neurological condition characterized by different cellular and molecular mechanisms that interplay in exacerbating the progression of the pathology. No fully restorative therapies are yet available, and it is thus becoming recognized that combinatorial approaches aimed at addressing different aspects of SCI will likely results in greater functional outcomes. Here we employed packaging RNA-mediated RNA interference (pRNA-RNAi) nanotherapeutics to downregulate in situ the expression of lipocalin 2 (Lcn2), a known mediator of neuroinflammation and autocrine mediator of reactive astrogliosis, and to create a more amenable niche for the subsequent transplantation of induced neural stem cells (iNSCs). To our knowledge, this is the first approach that takes advantage of the modular and multifunctional pRNA three-way junction platform in the SCI niche, while also exploiting the therapeutic potential of immune-compatible and feasible iNSC transplants. We show the combination of such treatments in a mouse model of contusion thoracic SCI leads to significant improvement of locomotor function, albeit not better than single pRNA-RNAi treatment, and results in synergistic histopathological effects, such as reduction of glial scar volume, diminished pro-inflammatory response, and promotion of neuronal survival. Our results provide evidence for a novel combinatorial approach for treating SCI.

RNA Nanotherapeutics for the Amelioration of Astroglial Reactivity.

In response to injuries to the CNS, astrocytes enter a reactive state known as astrogliosis, which is believed to be deleterious in some contexts. Activated astrocytes overexpress intermediate filaments including glial fibrillary acidic protein (GFAP) and vimentin (Vim), resulting in entangled cells that inhibit neurite growth and functional recovery. Reactive astrocytes also secrete inflammatory molecules such as Lipocalin 2 (Lcn2), which perpetuate reactivity and adversely affect other cells of the CNS. Herein, we report proof-of-concept use of the packaging RNA (pRNA)-derived three-way junction (3WJ) motif as a platform for the delivery of siRNAs to downregulate such reactivity-associated genes. In vitro, siRNA-3WJs induced a significant knockdown of Gfap, Vim, and Lcn2 in a model of astroglial activation, with a concomitant reduction in protein expression. Knockdown of Lcn2 also led to reduced protein secretion from reactive astroglial cells, significantly impeding the perpetuation of inflammation in otherwise quiescent astrocytes. Intralesional injection of anti-Lcn2-3WJs in mice with contusion spinal cord injury led to knockdown of Lcn2 at mRNA and protein levels in vivo. Our results provide evidence for siRNA-3WJs as a promising platform for ameliorating astroglial reactivity, with significant potential for further functionalization and adaptation for therapeutic applications in the CNS.

Macrophage-Derived Extracellular Succinate Licenses Neural Stem Cells to Suppress Chronic Neuroinflammation.

Neural stem cell (NSC) transplantation can influence immune responses and suppress inflammation in the CNS. Metabolites, such as succinate, modulate the phenotype and function of immune cells, but whether and how NSCs are also activated by such immunometabolites to control immunoreactivity and inflammatory responses is unclear. Here, we show that transplanted somatic and directly induced NSCs ameliorate chronic CNS inflammation by reducing succinate levels in the cerebrospinal fluid, thereby decreasing mononuclear phagocyte (MP) infiltration and secondary CNS damage. Inflammatory MPs release succinate, which activates succinate receptor 1 (SUCNR1)/GPR91 on NSCs, leading them to secrete prostaglandin E2 and scavenge extracellular succinate with consequential anti-inflammatory effects. Thus, our work reveals an unexpected role for the succinate-SUCNR1 axis in somatic and directly induced NSCs, which controls the response of stem cells to inflammatory metabolic signals released by type 1 MPs in the chronically inflamed brain.

High Yield of Adult Oligodendrocyte Lineage Cells Obtained from Meningeal Biopsy.

Oligodendrocyte loss can lead to cognitive and motor deficits. Current remyelinating therapeutic strategies imply either modulation of endogenous oligodendrocyte precursors or transplantation of in vitro expanded oligodendrocytes. Cell therapy, however, still lacks identification of an adequate source of oligodendrocyte present in adulthood and able to efficiently produce transplantable cells. Recently, a neural stem cell-like population has been identified in meninges. We developed a protocol to obtain high yield of oligodendrocyte lineage cells from one single biopsy of adult rat meningeal tissue. From 1 cm2 of adult rat spinal cord meninges, we efficiently expanded a homogenous culture of 10 millions of meningeal-derived oligodendrocyte lineage cells in a short period of time (approximately 4 weeks). Meningeal-derived oligodendrocyte lineage cells show typical mature oligodendrocyte morphology and express specific oligodendrocyte markers, such as galactosylceramidase and myelin basic protein. Moreover, when transplanted in a chemically demyelinated spinal cord model, meningeal-derived oligodendrocyte lineage cells display in vivo-remyelinating potential. This oligodendrocyte lineage cell population derives from an accessible and adult source, being therefore a promising candidate for autologous cell therapy of demyelinating diseases. In addition, the described method to differentiate meningeal-derived neural stem cells into oligodendrocyte lineage cells may represent a valid in vitro model to dissect oligodendrocyte differentiation and to screen for drugs capable to promote oligodendrocyte regeneration.

Comparison of static postural balance between healthy subjects and those with low back pain.

OBJECTIVE: To compare the static postural balance between women suffering from chronic low back pain and healthy subjects, by moving the center of pressure. METHODS: The study included 15 women with low back pain (LBP group) and 15 healthy women (healthy group). They were instructed to remain in standing on the force platform for 30 seconds. We analyzed the area and the speed of displacement of center of pressure of both groups. Data analysis was performed using the Student's t-test, with significance of 5%. RESULTS: Individuals with chronic low back pain showed a larger area of displacement of the center of pressure relative to the healthy ones but there was no significant difference in the speed of displacement of the center of pressure. CONCLUSION: Individuals with chronic low back pain had alterations in static balance with respect to healthy ones. Level of Evidence III, Prognostic Studies.

Comparação do equilíbrio postural estático entre sujeitos saudavéis e lombálgicos / Comparison of static postural balance between healthy subjects and those with low back pain

OBJETIVO: Comparar o equilíbrio postural estático entre portadoras de lombalgia crônica e indivíduos saudáveis, através do deslocamento do centro de pressão. MÉTODOS: Participaram do estudo 15 mulheres com lombalgia (Grupo lombalgia) e 15 mulheres saudáveis (Grupo saudável). Elas foram orientadas a permanecer em posição ortostática sobre a plataforma de força durante 30 segundos. Foram analisadas a área e a velocidade de deslocamento do centro de pressão de ambos os grupos. A análise dos dados foi realizada através do Teste t de Student, com significância de 5%. RESULTADOS: Os indivíduos com lombalgia crônica apresentaram maior área de deslocamento do centro de pressão em relação aos saudáveis, porém não houve diferença significativa em relação à velocidade de deslocamento do centro de pressão. CONCLUSÃO: Indivíduos com lombalgia crônica apresentam alteração do equilíbrio estático em relação a saudáveis. Nível de evidência III, Estudos Prognósticos.

OBJECTIVE: To compare the static postural balance between women suffering from chronic low back pain and healthy subjects, by moving the center of pressure. METHODS: The study included 15 women with low back pain (LBP group) and 15 healthy women (healthy group). They were instructed to remain in standing on the force platform for 30 seconds. We analyzed the area and the speed of displacement of center of pressure of both groups. Data analysis was performed using the Student t-test, with significance of 5%. RESULTS: Individuals with chronic low back pain showed a larger area of displacement of the center of pressure relative to the healthy ones but there was no significant difference in the speed of displacement of the center of pressure. CONCLUSION: Individuals with chronic low back pain had alterations in static balance with respect to healthy ones. Level of Evidence III, Prognostic Studies.

Preventing venous thrombosis in critically ill children: what is the right approach?

BACKGROUND: The incidence of venous thromboembolic (VTE) events in children has increased in recent years (J Neurosurg, 101, 2004, 32; J Thromb Haemost, 1, 2003, 1443) yet there is currently no consensus as to what VTE prophylaxis, if any, should be applied to the pediatric population. OBJECTIVES/AIMS: Our aim was to audit current practice in pediatric VTE prophylaxis across England and Wales and to advocate simple measures for prevention. We illustrate the importance of the condition with a series of cases from the South West Paediatric Burns and Neurosurgical Services based in Bristol. METHODS: Every pediatric intensive care unit (PICU) and burns center admitting children in England and Wales was invited to participate in a structured telephone questionnaire designed to find out how VTE in children were being prevented. We performed a literature review of specific risk factors and management of these factors. RESULTS: Only one of the 24 units surveyed had written guidelines specific for children. Four other units used modified adult guidelines in older children. In the remaining 19 units that had no written guidelines, decisions regarding prophylaxis were based on individual cases and consultant-led. CONCLUSION: There is no consensus in England and Wales as to which VTE prophylactic measures should be applied in patients <18 years of age. The National Institute for Health and Clinical Excellence (NICE) guidelines apply to adults only. Given the rarity of VTE events in children, it is unlikely that randomized controlled trials will provide the answer. We therefore propose that simple empirical measures be formally implemented in critically ill children to reduce the risk of developing this important but under-recognized condition.