Oxidized Carbon Nanoparticles Enhance Cellular Energetics With Application to Injured Brain.

Pro-energetic effects of functionalized, oxidized carbon nanozymes (OCNs) are reported. OCNs, derived from harsh acid oxidation of single-wall carbon nanotubes or activated charcoal are previously shown to possess multiple nanozymatic activities including mimicking superoxide dismutase and catalyzing the oxidation of reduced nicotinamide adenine dinucleotide (NADH) to NAD+. These actions are predicted to generate a glycolytic shift and enhance mitochondrial energetics under impaired conditions. Impaired mitochondrial energy metabolism is increasingly recognized as an important facet of traumatic brain injury (TBI) pathophysiology and decreases the efficiency of electron transport chain (ETC)-coupled adenosine triphosphate (ATP) and NAD+ regeneration. In vitro, OCNs promote a pro-aerobic shift in energy metabolism that persists through ETC inhibition and enhances glycolytic flux, glycolytic ATP production, and cellular generation of lactate, a crucial auxiliary substrate for energy metabolism. To address specific mechanisms of iron injury from hemorrhage, OCNs with the iron chelator, deferoxamine (DEF), covalently-linked were synthesized. DEF-linked OCNs induce a glycolytic shift in-vitro and in-vivo in tissue sections from a rat model of TBI complicated by hemorrhagic contusion. OCNs further reduced hemorrhage volumes 3 days following TBI. These results suggest OCNs are promising as pleiotropic mediators of cell and tissue resilience to injury.

Enhanced presynaptic mitochondrial energy production is required for memory formation.

Some of the prominent features of long-term memory formation include protein synthesis, gene expression, enhanced neurotransmitter release, increased excitability, and formation of new synapses. As these processes are critically dependent on mitochondrial function, we hypothesized that increased mitochondrial respiration and dynamics would play a prominent role in memory formation. To address this possibility, we measured mitochondrial oxygen consumption (OCR) in hippocampal tissue punches from trained and untrained animals. Our results show that context fear training significantly increased basal, ATP synthesis-linked, and maximal OCR in the Shaffer collateral-CA1 synaptic region, but not in the CA1 cell body layer. These changes were recapitulated in synaptosomes isolated from the hippocampi of fear-trained animals. As dynamin-related protein 1 (Drp1) plays an important role in mitochondrial fission, we examined its role in the increased mitochondrial respiration observed after fear training. Drp1 inhibitors decreased the training-associated enhancement of OCR and impaired contextual fear memory, but did not alter the number of synaptosomes containing mitochondria. Taken together, our results show context fear training increases presynaptic mitochondria respiration, and that Drp-1 mediated enhanced energy production in CA1 pre-synaptic terminals is necessary for context fear memory that does not result from an increase in the number of synaptosomes containing mitochondria or an increase in mitochondrial mass within the synaptic layer.

Using a Patient Educational Video to Improve Knowledge of Maternal Mortality Warning Signs: A Randomized Controlled Trial.

OBJECTIVE: To determine whether a community-informed, language-concordant postpartum video education campaign, developed with community input, improves patients' knowledge of warning signs for postpartum maternal mortality (infection, hemorrhage, hypertensive disorders, and postpartum depression) compared with routine discharge procedures. METHODS: A single-center, investigator-blinded, parallel-group randomized controlled trial of postpartum individuals who delivered at a large, urban, tertiary care hospital. Eligible participants were enrolled and completed a baseline knowledge questionnaire. After delivery, they were randomized to routine discharge education (control) or routine education plus video education (intervention). After discharge education, patient knowledge was again assessed in both groups before participants left the hospital. The primary outcome was the percentage of participants who showed improvement in their knowledge, measured by the number of correct questionnaire responses after education compared with their baseline, assessed as a binary outcome. A sample size of 150 (75 per group) was planned to detect a 25% absolute increase in the frequency of the primary outcome. RESULTS: From July to August 2022, 296 participants were screened and 200 were randomized (100 per group). Eighty-two percent of participants had college or graduate education, and 71.5% had commercial insurance. There was no significant difference in baseline characteristics. There was no statistically significant difference in the percentage of participants who improved their scores between the baseline and posteducation questionnaires (64.5% vs 50.0%, P =.09). However, the median posteducation questionnaire total score was significantly higher in the video education group (14 [interquartile range 12-15] vs 13 [interquartile range 12-14], P =.003). In addition, they more frequently reported that video education was "very helpful" (83.9% vs 72.5%, P =.23) and that they were "very satisfied" with their education (86.1% vs 75.5%, P =.29). CONCLUSION: Enhanced postpartum education through a novel video did not result in a statistically significant difference in frequency of improved score on the posteducation questionnaires but was associated with increased satisfaction with care. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov , NCT05159726.

Impaired Experience-Dependent Refinement of Place Cells in a Rat Model of Alzheimer's Disease.

BACKGROUND: Hippocampal place cells play an integral role in generating spatial maps. Impaired spatial memory is a characteristic pathology of Alzheimer's disease (AD), yet it remains unclear how AD influences the properties of hippocampal place cells. OBJECTIVE: To record electrophysiological activity in hippocampal CA1 neurons in freely-moving 18-month-old male TgF344-AD and age-matched wild-type (WT) littermates to examine place cell properties. METHODS: We implanted 32-channel electrode arrays into the CA1 subfield of 18-month-old male WT and TgF344-AD (n = 6/group) rats. Ten days after implantation, single unit activity in an open field arena was recorded across days. The spatial information content, in-field firing rate, and stability of each place cell was compared across groups. Pathology was assessed by immunohistochemical staining, and a deep neural network approach was used to count cell profiles. RESULTS: Aged TgF344-AD rats exhibited hippocampal amyloid-ß deposition, and a significant increase in Iba1 immunoreactivity and microglia cell counts. Place cells from WT and TgF344-AD rat showed equivalent spatial information, in-field firing rates, and place field stability when initially exposed to the arena. However, by day 3, the place cells in aged WT rats showed characteristic spatial tuning as evidenced by higher spatial information content, stability, and in-field firing rates, an effect not seen in TgF344-AD rats. CONCLUSION: These findings support the notion that altered electrophysiological properties of place cells may contribute to the learning and memory deficits observed in AD.

Metformin Reduces Repeat Mild Concussive Injury Pathophysiology.

Mild traumatic brain injury (mTBI) can initiate complex pathophysiological changes in the brain. Numerous cellular and molecular mechanisms underlying these pathologic changes are altered after injury, including those involved in energy utilization, excitotoxicity, ionic disturbances, and inflammation. It is thought that targeting multiple mechanisms may be necessary to produce meaningful reductions in brain pathology and to improve outcome. Previous studies have reported that the anti-diabetic drug metformin can also affect inflammatory, cell survival, and metabolic outcomes, possibly by acting on multiple targets and/or pathways. We therefore questioned whether metformin treatment can reduce pathology after repeat mild closed head injury (rmCHI) in male C57Bl/6 mice. We found that metformin, administered acutely after each head impact, resulted in markedly reduced white matter damage, astrogliosis, loss of hippocampal parvalbumin neurons, and improved mitochondrial function. In addition, both motor and cognitive functions were significantly improved when tested after discontinuation of the treatment. These studies suggest that metformin may be beneficial as a treatment for repeat concussion.

Patient-derived iPSCs link elevated mitochondrial respiratory complex I function to osteosarcoma in Rothmund-Thomson syndrome.

Rothmund-Thomson syndrome (RTS) is an autosomal recessive genetic disorder characterized by poikiloderma, small stature, skeletal anomalies, sparse brows/lashes, cataracts, and predisposition to cancer. Type 2 RTS patients with biallelic RECQL4 pathogenic variants have multiple skeletal anomalies and a significantly increased incidence of osteosarcoma. Here, we generated RTS patient-derived induced pluripotent stem cells (iPSCs) to dissect the pathological signaling leading to RTS patient-associated osteosarcoma. RTS iPSC-derived osteoblasts showed defective osteogenic differentiation and gain of in vitro tumorigenic ability. Transcriptome analysis of RTS osteoblasts validated decreased bone morphogenesis while revealing aberrantly upregulated mitochondrial respiratory complex I gene expression. RTS osteoblast metabolic assays demonstrated elevated mitochondrial respiratory complex I function, increased oxidative phosphorylation (OXPHOS), and increased ATP production. Inhibition of mitochondrial respiratory complex I activity by IACS-010759 selectively suppressed cellular respiration and cell proliferation of RTS osteoblasts. Furthermore, systems analysis of IACS-010759-induced changes in RTS osteoblasts revealed that chemical inhibition of mitochondrial respiratory complex I impaired cell proliferation, induced senescence, and decreased MAPK signaling and cell cycle associated genes, but increased H19 and ribosomal protein genes. In summary, our study suggests that mitochondrial respiratory complex I is a potential therapeutic target for RTS-associated osteosarcoma and provides future insights for clinical treatment strategies.

A method for assessing tissue respiration in anatomically defined brain regions.

The survival and function of brain cells requires uninterrupted ATP synthesis. Different brain structures subserve distinct neurological functions, and therefore have different energy production/consumption requirements. Typically, mitochondrial function is assessed following their isolation from relatively large amounts of starting tissue, making it difficult to ascertain energy production/failure in small anatomical locations. In order to overcome this limitation, we have developed and optimized a method to measure mitochondrial function in brain tissue biopsy punches excised from anatomically defined brain structures, including white matter tracts. We describe the procedures for maintaining tissue viability prior to performing the biopsy punches, as well as provide guidance for optimizing punch size and the drug doses needed to assess various aspects of mitochondrial respiration. We demonstrate that our method can be used to measure mitochondrial respiration in anatomically defined subfields within the rat hippocampus. Using this method, we present experimental results which show that a mild traumatic brain injury (mTBI, often referred to as concussion) causes differential mitochondrial responses within these hippocampal subfields and the corpus callosum, novel findings that would have been difficult to obtain using traditional mitochondrial isolation methods. Our method is easy to implement and will be of interest to researchers working in the field of brain bioenergetics and brain diseases.

Traumatic brain injury and hippocampal neurogenesis: Functional implications.

In the adult brain, self-renewing radial-glia like (RGL) progenitor cells have been shown to reside in the subventricular zone and the subgranular zone of the hippocampus. A large body of evidence shows that experiences such as learning, enriched environment and stress can alter proliferation and differentiation of RGL progenitor cells. The progenitor cells present in the subgranular zone of the hippocampus divide to give rise to newborn neurons that migrate to the dentate gyrus where they differentiate into adult granule neurons. These newborn neurons have been found to have a unique role in certain types of hippocampus-dependent learning and memory, including goal-directed behaviors that require pattern separation. Experimental traumatic brain injury (TBI) in rodents has been shown to alter hippocampal neurogenesis, including triggering the acute loss of newborn neurons, as well as progenitor cell hyper-proliferation. In this review, we discuss the role of hippocampal neurogenesis in learning and memory. Furthermore, we review evidence for the molecular mechanisms that contribute to newborn neuron loss, as well as increased progenitor cell proliferation after TBI. Finally, we discuss strategies aimed at enhancing neurogenesis after TBI and their possible therapeutic benefits.

Loss of PTEN-induced kinase 1 (Pink1) reduces hippocampal tyrosine hydroxylase and impairs learning and memory.

Phosphatase and tensin homolog (PTEN)-induced kinase 1 (Pink1) is involved in mitochondrial quality control, which is essential for maintaining energy production and minimizing oxidative damage from dysfunctional/depolarized mitochondria. Pink1 mutations are the second most common cause of autosomal recessive Parkinson's disease (PD). In addition to characteristic motor impairments, PD patients also commonly exhibit cognitive impairments. As the hippocampus plays a prominent role in cognition, we tested if loss of Pink1 in mice influences learning and memory. While wild-type mice were able to perform a contextual discrimination task, age-matched Pink1 knockout (Pink1-/-) mice showed an impaired ability to differentiate between two similar contexts. Similarly, Pink1-/- mice performed poorly in a delayed alternation task as compared to age-matched controls. Poor performance in these cognitive tasks was not the result of overt hippocampal pathology. However, a significant reduction in hippocampal tyrosine hydroxylase (TH) protein levels was detected in the Pink1-/- mice. This decrease in hippocampal TH levels was also associated with reduced DOPA decarboxylase and dopamine D2 receptor levels, but not post-synaptic dopamine D1 receptor levels. These presynaptic changes appeared to be selective for dopaminergic fibers as hippocampal dopamine beta hydroxylase, choline acetyltransferase, and tryptophan hydroxylase levels were unchanged in Pink1-/- mice. Administration of the dopamine D1 receptor agonist SKF38393 to Pink1-/- mice was found to improve performance in the context discrimination task. Taken together, our results show that Pink1 loss may alter dopamine signaling in the hippocampus, which could be a contributing mechanism for the observed learning and memory impairments.

Carnosic Acid Improves Outcome after Repetitive Mild Traumatic Brain Injury.

In the majority of cases, the cognitive and behavioral impairments resulting from a mild traumatic brain injury (TBI) (also referred to as concussion) wane within days to weeks. In contrast, these impairments can persist for months to years after repetitive mild TBI (rmTBI). The cellular and molecular mechanisms underlying these impairments are not well understood. In the present study, we examined the consequences of rmTBI (three weight drops each separated by 72 h) on brain tissue respiration, pathology, and cognitive performance in mice. The transcription factor nuclear factor-erythroid 2-realted factor 2 (Nrf2) has been demonstrated to enhance the expression of numerous cytoprotective genes. Carnosic acid (CA) has been shown to activate Nrf2 and suppress the proinflammatory transcription factor nuclear factor kappa B (NF-κB). Because contemporaneous activation of cytoprotective genes and inhibition of proinflammatory genes can be beneficial, we questioned whether CA can be used to mitigate the pathobiology of rmTBI. The rmTBI increased hippocampal adenosine triphosphate-linked tissue respiration and proton leak that were unaffected by CA treatment. The rmTBI also caused significant motor and cognitive dysfunction, as tested using the foot fault, Barnes maze, and novel object recognition tasks. These impairments occurred in the absence of visible neuronal or dendritic loss. Post-rmTBI administration of CA significantly improved motor and cognitive function, and decreased Gfap and Iba1 immunoreactivities within white matter tracks. Taken together, these results show that rmTBI can cause cognitive impairments in the absence of overt neuronal pathologies, and post-injury treatment with CA can lessen some of these impairments.

Cell type-dependent effects of ellagic acid on cellular metabolism.

Ellagic acid is a botanical polyphenol which has been shown to have numerous effects on cellular function. Ellagic acid can induce apoptosis and inhibit the proliferation of various cancer cell types in vitro and in vivo. As such, ellagic acid has attracted significant interest as a potential chemotherapeutic compound. One mechanism by which ellagic acid has been proposed to affect cellular physiology is by regulating metabolic pathways. Here we show the dose-dependent effects of ellagic acid on cellular energy production and downstream induction of the apoptotic program in HEK293, HeLa, MCF7, and HepG2 cells. At physiologically relevant doses, ellagic acid has pleiotropic and cell-type specific effects on mitochondrial function. At high doses ellagic acid can also influence glycolytic pathways and induce cell death. Our results demonstrate that ellagic acid can influence mitochondrial function at therapeutically relevant concentrations. The observed effects of ellagic acid on cellular respiration are complex and cell type-specific, which may limit the chemotherapeutic utility of this compound.

High-fat diet impairs spatial memory and hippocampal intrinsic excitability and sex-dependently alters circulating insulin and hippocampal insulin sensitivity.

BACKGROUND: High-fat diets promoting obesity/type-2 diabetes can impair physiology and cognitive performance, although sex-dependent comparisons of these impairments are rarely made. Transient reductions in Ca(2+)-dependent afterhyperpolarizations (AHPs) occur during memory consolidation, enhancing intrinsic excitability of hippocampal CA1 pyramidal neurons. In rats fed standard diets, insulin can enhance memory and reduce amplitude and duration of AHPs. METHODS: Effects of chronic high-fat diet (HFD) on memory, circulating insulin, and neuronal physiology were compared between young adult male and female Long-Evans rats. Rats fed for 12 weeks (from weaning) a HFD or a control diet (CD) were then tested in vivo prior to in vitro recordings from CA1 pyramidal neurons. RESULTS: The HFD significantly impaired spatial memory in both males and females. Significant sex differences occurred in circulating insulin and in the insulin sensitivity of hippocampal neurons. Circulating insulin significantly increased in HFD males but decreased in HFD females. While the HFD significantly reduced hippocampal intrinsic excitability in both sexes, CA1 neurons from HFD females remained insulin-sensitive but those from HFD males became insulin-insensitive. CONCLUSIONS: Findings consistent with these have been characterized previously in HFD or senescent males, but the effects observed here in young females are unique. Loss of CA1 neuronal excitability, and sex-dependent loss of insulin sensitivity, can have significant cognitive consequences, over both the short term and the life span. These findings highlight needs for more research into sex-dependent differences, relating systemic and neural plasticity mechanisms in metabolic disorders.

A High-Fat Diet Causes Impairment in Hippocampal Memory and Sex-Dependent Alterations in Peripheral Metabolism.

While high-fat diets are associated with rising incidence of obesity/type-2 diabetes and can induce metabolic and cognitive deficits, sex-dependent comparisons are rarely systematically made. Effects of exclusive consumption of a high-fat diet (HFD) on systemic metabolism and on behavioral measures of hippocampal-dependent memory were compared in young male and female LE rats. Littermates were fed from weaning either a HFD or a control diet (CD) for 12 wk prior to testing. Sex-different effects of the HFD were observed in classic metabolic signs associated with type-2 diabetes. Males fed the HFD became obese, and had elevated fasted blood glucose levels, elevated corticosterone, and impaired glucose-tolerance, while females on the HFD exhibited only elevated corticosterone. Regardless of peripheral metabolism alteration, rats of both sexes fed the HFD were equally impaired in a spatial object recognition memory task associated with impaired hippocampal function. While the metabolic changes reported here have been characterized previously in males, the set of diet-induced effects observed here in females are novel. Impaired memory can have significant cognitive consequences, over the short-term and over the lifespan. A significant need exists for comparative research into sex-dependent differences underlying obesity and metabolic syndromes relating systemic, cognitive, and neural plasticity mechanisms.