Epilepsy phenotype and its reproducibility after lateral fluid percussion-induced traumatic brain injury in rats: Multicenter EpiBioS4Rx study project 1.

OBJECTIVE: This study was undertaken to assess reproducibility of the epilepsy outcome and phenotype in a lateral fluid percussion model of posttraumatic epilepsy (PTE) across three study sites. METHODS: A total of 525 adult male Sprague Dawley rats were randomized to lateral fluid percussion-induced brain injury (FPI) or sham operation. Of these, 264 were assigned to magnetic resonance imaging (MRI cohort, 43 sham, 221 traumatic brain injury [TBI]) and 261 to electrophysiological follow-up (EEG cohort, 41 sham, 220 TBI). A major effort was made to harmonize the rats, materials, equipment, procedures, and monitoring systems. On the 7th post-TBI month, rats were video-EEG monitored for epilepsy diagnosis. RESULTS: A total of 245 rats were video-EEG phenotyped for epilepsy on the 7th postinjury month (121 in MRI cohort, 124 in EEG cohort). In the whole cohort (n = 245), the prevalence of PTE in rats with TBI was 22%, being 27% in the MRI and 18% in the EEG cohort (p > .05). Prevalence of PTE did not differ between the three study sites (p > .05). The average seizure frequency was .317 ± .725 seizures/day at University of Eastern Finland (UEF; Finland), .085 ± .067 at Monash University (Monash; Australia), and .299 ± .266 at University of California, Los Angeles (UCLA; USA; p < .01 as compared to Monash). The average seizure duration did not differ between UEF (104 ± 48 s), Monash (90 ± 33 s), and UCLA (105 ± 473 s; p > .05). Of the 219 seizures, 53% occurred as part of a seizure cluster (≥3 seizures/24 h; p >.05 between the study sites). Of the 209 seizures, 56% occurred during lights-on period and 44% during lights-off period (p > .05 between the study sites). SIGNIFICANCE: The PTE phenotype induced by lateral FPI is reproducible in a multicenter design. Our study supports the feasibility of performing preclinical multicenter trials in PTE to increase statistical power and experimental rigor to produce clinically translatable data to combat epileptogenesis after TBI.

Levetiracetam Pharmacokinetics and Brain Uptake in a Lateral Fluid Percussion Injury Rat Model.

Post-traumatic epilepsy (PTE) occurs in some patients after moderate/severe traumatic brain injury (TBI). Although there are no approved therapies to prevent epileptogenesis, levetiracetam (LEV) is commonly given for seizure prophylaxis due to its good safety profile. This led us to study LEV as part of the Epilepsy Bioinformatics Study for Antiepileptogenic Therapy (EpiBioS4Rx) Project. The objective of this work is to characterize the pharmacokinetics (PK) and brain uptake of LEV in naïve control rats and in the lateral fluid percussion injury (LFPI) rat model of TBI after either single intraperitoneal doses or a loading dose followed by a 7-day subcutaneous infusion. Sprague-Dawley rats were used as controls and for the LFPI model induced at the left parietal region using injury parameters optimized for moderate/severe TBI. Naïve and LFPI rats received either a bolus injection (intraperitoneal) or a bolus injection followed by subcutaneous infusion over 7 days. Blood and parietal cortical samples were collected at specified time points throughout the study. LEV concentrations in plasma and brain were measured using validated high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) methods. Noncompartmental analysis and a naive-pooled compartmental PK modeling approach were used. Brain-to-plasma ratios ranged from 0.54 to 1.4 to 1. LEV concentrations were well fit by one-compartment, first-order absorption PK models with a clearance of 112 ml/h per kg and volume of distribution of 293 ml/kg. The single-dose pharmacokinetic data were used to guide dose selection for the longer-term studies, and target drug exposures were confirmed. Obtaining LEV PK information early in the screening phase allowed us to guide optimal treatment protocols in EpiBioS4Rx. SIGNIFICANCE STATEMENT: The characterization of levetiracetam pharmacokinetics and brain uptake in an animal model of post-traumatic epilepsy is essential to identify target concentrations and guide optimal treatment for future studies.

Hepatitis C drug prescriptions and Medicaid policies--four states, Indian health care system, USA 2018.

Medicaid, the state-level public insurance in the United States, has widely differing criteria treatment for hepatitis C virus (HCV) such as stage of liver fibrosis, documented sobriety, and specialist consultation. In a rural health network, facilities located in two less restrictive states prescribed HCV drugs at a significantly higher rate than two more restrictive states (rate ratio 4.7, CI 2.6-8.5). Prescription rates per population were highly associated with HCV treatment policies.

Genes involved in convergent evolution of eusociality in bees.

Eusociality has arisen independently at least 11 times in insects. Despite this convergence, there are striking differences among eusocial lifestyles, ranging from species living in small colonies with overt conflict over reproduction to species in which colonies contain hundreds of thousands of highly specialized sterile workers produced by one or a few queens. Although the evolution of eusociality has been intensively studied, the genetic changes involved in the evolution of eusociality are relatively unknown. We examined patterns of molecular evolution across three independent origins of eusociality by sequencing transcriptomes of nine socially diverse bee species and combining these data with genome sequence from the honey bee Apis mellifera to generate orthologous sequence alignments for 3,647 genes. We found a shared set of 212 genes with a molecular signature of accelerated evolution across all eusocial lineages studied, as well as unique sets of 173 and 218 genes with a signature of accelerated evolution specific to either highly or primitively eusocial lineages, respectively. These results demonstrate that convergent evolution can involve a mosaic pattern of molecular changes in both shared and lineage-specific sets of genes. Genes involved in signal transduction, gland development, and carbohydrate metabolism are among the most prominent rapidly evolving genes in eusocial lineages. These findings provide a starting point for linking specific genetic changes to the evolution of eusociality.

Folie et Société: eroding the body-mind relationship via dysfunctional paternalistic systems.

This theoretical perspective examines the proposition of shared complex trauma between a parent and child, arising from blurred relational boundaries and societal oppression, leading to inequality both at home and within the larger paternalistic system of society. Specifically, the focus is on living within a paternalistic, authoritarian system where rules are unjust, demanding obedience and compliance without questioning the behaviors of the authority. Individuals growing up in these circumstances are subject to adverse and emotionally overwhelming experiences, which lead to the creation of emotional memory images (EMIs). The delusion in which the child is caught up becomes a reality for the child as time passes. This phenomenon is recognized in psychiatry as "Folie à deux" (the madness of two or more) at the micro level, and "Folie et Société" (the madness of society) on the macro level. Complex trauma, derived from a child's exposure to multiple adverse events, can erode the mind-body relationship, impacting both mental and physical health. These traumatic experiences in early childhood can manifest as body-focused disorders in adolescents, prevailing throughout adulthood if left unattended. This article provides a theoretical perspective on dealing with the dissociation and chronic stress related to oppressive and authoritarian family systems. The broader implications of this article include highlighting the psychophysiological underpinnings of complex trauma, the relationship of a highly oppressive paternalistic authoritarian system imposed on children and adolescents, and the role of Split-Second Unlearning as a therapeutic intervention to clear EMIs and improve overall health outcomes.

Tau Phosphorylation Patterns in the Rat Cerebral Cortex After Traumatic Brain Injury and Sodium Selenate Effects: An Epibios4rx Project 2 Study.

Sodium selenate (SS) activates protein phosphatase 2 (PP2A) and reduces phosphorylated tau (pTAU) and late post-traumatic seizures after lateral fluid percussion injury (LFPI). In EpiBioS4Rx Project 2, a multi-center international study for post-traumatic targets, biomarkers, and treatments, we tested the target relevance and modification by SS of pTAU forms and PP2A and in the LFPI model, at two sites: Einstein and Melbourne. In Experiment 1, adult male rats were assigned to LFPI and sham (both sites) and naïve controls (Einstein). Motor function was monitored by neuroscores. Brains were studied with immunohistochemistry (IHC), Western blots (WBs), or PP2A activity assay, from 2 days to 8 weeks post-operatively. In Experiment 2, LFPI rats received SS for 7 days (SS0.33: 0.33 mg/kg/day; SS1: 1 mg/kg/day, subcutaneously) or vehicle (Veh) post-LFPI and pTAU, PR55 expression, or PP2A activity were studied at 2 days and 1 week (on treatment), or 2 weeks (1 week off treatment). Plasma selenium and SS levels were measured. In Experiment 1 IHC, LFPI rats had higher cortical pTAU-Ser202/Thr205-immunoreactivity (AT8-ir) and pTAU-Ser199/202-ir at 2 days, and pTAU-Thr231-ir (AT180-ir) at 2 days, 2 weeks, and 8 weeks, ipsilaterally to LFPI, than controls. LFPI-2d rats also had higher AT8/total-TAU5-ir in cortical extracts ipsilateral to the lesion (WB). PP2A (PR55-ir) showed time- and region-dependent changes in IHC, but not in WB. PP2A activity was lower in LFPI-1wk than in sham rats. In Experiment 2, SS did not affect neuroscores or cellular AT8-ir, AT180-ir, or PR55-ir in IHC. In WB, total cortical AT8/total-TAU-ir was lower in SS0.33 and SS1 LFPI rats than in Veh rats (2 days, 1 week); total cortical PR55-ir (WB) and PP2A activity were higher in SS1 than Veh rats (2 days). SS dose dependently increased plasma selenium and SS levels. Concordant across-sites data confirm time and pTAU form-specific cortical increases ipsilateral to LFPI. The discordant SS effects may either suggest SS-induced reduction in the numbers of cells with increased pTAU-ir, need for longer treatment, or the involvement of other mechanisms of action.

Perspectives on emotional memory images and the persistence of pain.

Multiple influences prevent recovery from pain. Our viewpoint is that non-conscious emotional memory images (EMIs) triggers outdated stress responses contributing to the intractability of pain. In this perspectives article we explore the concept that EMIs contribute to the persistence of pain. We contend that psychophysiological "stress" responses, resulting from first-time, novel and unprecedented pernicious or adverse events form EMIs within very short time frames (split-second learning). Subsequently, these EMIs are re-triggered in daily living, "re-playing" stress responses. We postulate that EMIs continually "raise the alarm" to socio-ecological stimuli by re-triggering the HPA-axis and amplifying neural input associated with threat, fear, anxiety, and pain, creating a debilitating state of psychophysiological dis-ease. We position the EMI within a philosophical debate on the nature and locus of memory and explain how the EMI, irrespective of whether it is a "thing" or a metaphor, can create a basis of understanding for the client to grasp. We describe a therapeutic approach (Split-Second Unlearning) to "clear" EMIs and the "stickiness" of pain and help people embark on a healing journey. This involves surveillance of clients for micro-expression(s) signifying an in-the-moment stress response, representative of the presence of an EMI, and encouraging the client to become a curious observer within/of their own experience. This helps the client detach their EMI from its stress response. We contend that this occurs rapidly without the need to get bogged down in a whole-life narrative. We advocate further exploration of our EMI model of dis-ease in the context of intractable pain.

Past Adversity Influencing Now (PAIN): perspectives on the impact of temporal language on the persistence of pain.

Persistent pain is a significant healthcare issue, often unresponsive to traditional treatments. We argue for incorporating non-biomedical perspectives in understanding pain, promoting more comprehensive solutions. This article explores how language, specifically time-related terms, may affect the persistence (stickiness) of pain. We delve into how language influences one's experience of the world, especially in understanding pain through spatial metaphors. Notably, time perceptions differ across languages and cultures and there is no absolute construct of temporal pain experience. In English, time is viewed linearly as past, present, and future. We introduce a framework called Past Adversity Influencing Now (PAIN) which includes various temporal phases of pain; Past Perfect, Past Imperfect, Present, Future Imperfect, and Future Perfect. We suggest that past negative memories (emotional memory images) can "trap" individuals in a "sticky" pain state. We speculate that the process of diagnosing pain as "chronic" may solidify this "stickiness", drawing from the ancient Greek idea of "logos", where pain communicates a message across time and space needing recognition. Our PAIN framework encourages examining pain through a temporal lens, guiding individuals towards a more positive future.

Hidden family rules: perspective on a dysfunctional paternalistic system and the persistence of pain.

This article explores how paternalistic control and power reside within the family system and how this may influence pain and its persistence. Drawing upon clinical case studies and existing literature, this exploration emphasises the role of paternal dysfunction in creating emotional memory images and delves into how this may influence the chronification and treatment resistance of pain (i.e., making pain "sticky"). We argue that a dysfunctional paternalistic family system, often characterised by authoritarian dynamics, emotional neglect, and abuse, results in adverse experiences and emotional memory images that create a fertile ground for the entrenchment and propagation of psychosomatic symptoms, including pain. Further, the paper emphasizes the potential intergenerational effects of such a scenario, where inherited "Family Rules" drive maladaptive coping mechanisms, which contribute to the persistence of psychological and physiological distress across generations. Understanding these complexities offers new perspectives on treating psychological disorders and their physiological ramifications. It also highlights the urgency of addressing dysfunctional familial dynamics in psychotherapeutic interventions for both immediate and long-term psychophysiological health outcomes.

Perspectives on the insidious nature of pain metaphor: we literally need to change our metaphors.

Metaphorical language is used to convey one thing as representative or symbolic of something else. Metaphor is used in figurative language but is much more than a means of delivering "poetic imagination". A metaphor is a conceptual tool for categorising, organizing, thinking about, and ultimately shaping reality. Thus, metaphor underpins the way humans think. Our viewpoint is that metaphorical thought and communication contribute to "painogenicity", the tendency of socio-ecological environments (settings) to promote the persistence of pain. In this perspectives article, we explore the insidious nature of metaphor used in pain language and conceptual models of pain. We explain how metaphor shapes mental organisation to govern the way humans perceive, navigate and gain insight into the nature of the world, i.e., creating experience. We explain how people use metaphors to "project" their private sensations, feelings, and thoughts onto objects and events in the external world. This helps people to understand their pain and promotes sharing of pain experience with others, including health care professionals. We explore the insidious nature of "warmongering" and damage-based metaphors in daily parlance and demonstrate how this is detrimental to health and wellbeing. We explore how metaphors shape the development and communication of complex, abstract ideas, theories, and models and how scientific understanding of pain is metaphorical in nature. We argue that overly simplistic neuro-mechanistic metaphors of pain contribute to fallacies and misnomers and an unhealthy focus on biomedical research, in the hope of developing medical interventions that "prevent pain transmission [sic]". We advocate reconfiguring pain language towards constructive metaphors that foster a salutogenic view of pain, focusing on health and well-being. We advocate reconfiguring metaphors to align with contemporary pain science, to encourage acceptance of non-medicalised strategies to aid health and well-being. We explore the role of enactive metaphors to facilitate reconfiguration. We conclude that being cognisant of the pervasive nature of metaphors will assist progress toward a more coherent conceptual understanding of pain and the use of healthier pain language. We hope our article catalyses debate and reflection.

Definition and attributes of the emotional memory images underlying psychophysiological dis-ease.

Background: Previously, we proposed a "Split-second Unlearning" model to explain how emotional memories could be preventing clients from adapting to the stressors of daily living, thus forming a barrier to learning, health and well-being. We suggested that these emotional memories were mental images stored inside the mind as 'emotional memory images' (EMIs). Objective: To elaborate on the nature of these emotional memory images within the context of split-second learning and unlearning and the broader field of psychoanalysis, to initiate a conversation among scholars concerning the path that future healthcare research, practice, and policy should take. Method: A narrative review of the attributes of EMIs utilizing relevant and contentious research and/or scholarly publications on the topic, facilitated by observations and approaches used in clinical practice. Results: We propose a refined definition of EMIs as Trauma induced, non-conscious, contiguously formed multimodal mental imagery, which triggers an amnesic, anachronistic, stress response within a split-second. The systematic appraisal of each attribute of an EMI supports the idea that the EMI is distinct from similar entities described in literature, enabling further sophistication of our Split-second Unlearning model of psychophysiological dis-ease. Conclusion: Exploration of the concept of EMIs provides further insight on mechanisms associated with psychophysiological dis-ease and opportunities for therapeutic approaches.

Postnatal developmental trajectory of sex-biased gene expression in the mouse pituitary gland.

BACKGROUND: The pituitary gland regulates essential physiological processes such as growth, pubertal onset, stress response, metabolism, reproduction, and lactation. While sex biases in these functions and hormone production have been described, the underlying identity, temporal deployment, and cell-type specificity of sex-biased pituitary gene regulatory networks are not fully understood. METHODS: To capture sex differences in pituitary gene regulation dynamics during postnatal development, we performed 3' untranslated region sequencing and small RNA sequencing to ascertain gene and microRNA expression, respectively, across five postnatal ages (postnatal days 12, 22, 27, 32, 37) that span the pubertal transition in female and male C57BL/6J mouse pituitaries (n = 5-6 biological replicates for each sex at each age). RESULTS: We observed over 900 instances of sex-biased gene expression and 17 sex-biased microRNAs, with the majority of sex differences occurring with puberty. Using miRNA-gene target interaction databases, we identified 18 sex-biased genes that were putative targets of 5 sex-biased microRNAs. In addition, by combining our bulk RNA-seq with publicly available male and female mouse pituitary single-nuclei RNA-seq data, we obtained evidence that cell-type proportion sex differences exist prior to puberty and persist post-puberty for three major hormone-producing cell types: somatotropes, lactotropes, and gonadotropes. Finally, we identified sex-biased genes in these three pituitary cell types after accounting for cell-type proportion differences between sexes. CONCLUSION: Our study reveals the identity and postnatal developmental trajectory of sex-biased gene expression in the mouse pituitary. This work also highlights the importance of considering sex biases in cell-type composition when understanding sex differences in the processes regulated by the pituitary gland.

Learning associations between action and perception: effects of incompatible training on body part and spatial priming.

Observation of another person executing an action primes the same action in the observer's motor system. Recent evidence has shown that these priming effects are flexible, where training of new associations, such as making a foot response when viewing a moving hand, can reduce standard action priming effects (Gillmeister, Catmur, Liepelt, Brass, & Heyes, 2008). Previously, these effects were obtained after explicit learning tasks in which the trained action was cued by the content of a visual stimulus. Here we report similar learning processes in an implicit task in which the participant's action is self-selected, and subsequent visual effects are determined by the nature of that action. Importantly, we show that these learning processes are specific to associations between actions and viewed body parts, in that incompatible spatial training did not influence body part or spatial priming effects. Our results are consistent with models of visuomotor learning that place particular emphasis on the repeated experience of watching oneself perform an action.

Split-Second Unlearning: Developing a Theory of Psychophysiological Dis-ease.

Psychophysiological "stress" underpins many conditions including anxiety, depression, phobias, chronic fatigue syndrome and non-specific musculoskeletal pain such as fibromyalgia. In this article we develop an understanding of chronic psychophysiological stress from a psychological educational perspective, by drawing on supporting evidence that significant emotional events in early life (traumatic and benign) can influence health and well-being later in life. We suggest that traumatic events instigate psychophysiological "stress" responses and the formation of emotional memory images (EMIs) within very short time frames, i.e., "split-second learning." Once formed these emotional memories are triggered in daily living "re-playing" psychophysiological stress responses, resulting in chronic psychophysiological "dis-ease." We describe a novel therapeutic approach to scan clients for mannerisms signifying a subconscious "freeze-like" stress response that involves the client as a curious observer within their own experience, feeding back the non-verbal cues as they arrive in the moment. By breaking down the observable fragments of their split-second Pavlovian response to the trigger, clients can detach their EMI from the psychophysiology stress response, i.e., "split-second unlearning." Our split-second unlearning model recognizes the EMI as a barrier to moving forward and needs to be unlearned before the client can become naturally adaptive again. We argue that this approach places the client at the center of the work without the need of getting bogged down in a life-long narrative.

Apocynin attenuates diaphragm oxidative stress and protease activation during prolonged mechanical ventilation.

OBJECTIVE: To investigate whether apocynin protects the diaphragm from wasting and oxidative stress during mechanical ventilation (MV). DESIGN: Prospective, randomized, controlled study. SETTING: Research laboratory. SUBJECTS: Adult female Sprague-Dawley rats. INTERVENTIONS: Rats were randomly assigned to one of five experimental groups: 1) acutely anesthetized control, 2) spontaneous breathing control, 3) spontaneously breathing control with administration of the nicotinamide adenine dinucleotide phosphate oxidase inhibitor, apocynin, 4) mechanically ventilated, and 5) mechanically ventilated with apocynin. MEASUREMENTS AND MAIN RESULTS: Apocynin attenuated MV-induced diaphragmatic oxidative stress, contractile dysfunction, and type I, type IIa, and type IIb/IIx myofiber atrophy. The apocynin-induced attenuation of MV-induced diaphragmatic atrophy and contractile dysfunction occurred in conjunction with a reduction in the small increase in nicotinamide adenine dinucleotide phosphate oxidase activity as well as the preservation of total glutathione levels, glutathione peroxidase protein abundance, and a decrease in the activation of the cysteine proteases, calpain-1 and caspase-3. Interestingly, independent of MV, apocynin increased diaphragmatic levels of calpastatin, an endogenous calpain inhibitor. Furthermore, treatment of skeletal muscle cells in culture (C2C12 myotubes) with apocynin resulted in an increase in both calpastatin mRNA levels and protein abundance. CONCLUSIONS: Our results suggest that the protective effects of apocynin on the diaphragm during prolonged MV seem to be linked to both its functions as an antioxidant and role in cellular signaling regulating the cysteine protease inhibitor calpastatin.

Harmonization of pipeline for preclinical multicenter plasma protein and miRNA biomarker discovery in a rat model of post-traumatic epileptogenesis.

The Epilepsy Bioinformatics Study for Antiepileptogenic Therapy (EpiBioS4Rx) is an international, multicenter, multidisciplinary study aimed at preventing epileptogenesis (EpiBioS4Rx: https://epibios.loni.usc.edu/). One of the study's major objectives is the discovery of diagnostic, prognostic, and predictive plasma protein and microRNA (miRNA) biomarkers that are sensitive, specific, and translatable to the human condition. Epilepsy due to structural brain abnormalities, secondary to neurological insults such as traumatic brain injury (TBI), currently represents ∼50% of all epilepsy cases. In the preclinical EpiBioS4Rx study, TBI was induced in adult male Sprague Dawley rats using a standardized protocol for lateral fluid-percussion injury. Whole blood was collected from the tail vein at baseline and 2, 9 and 30 days post-injury and processed for plasma separation. Biomaterial properties, sample preparation and integrity, and choice of analysis platform can significantly impact measured marker levels and, in turn, interpretation with respect to injury and/or other variables. We present here the results of procedural harmonization for the first 320 rats included in the EpiBioS4Rx study study, from three international research centers, and preliminary proteomic and miRNA analyses. We also discuss experimental considerations for establishing rigorous quality controls with the goal of harmonizing operating procedures across study sites, and delivering high-quality specimens for preclinical biomarker discovery in a rat model of post-traumatic epilepsy (PTE).

Unintended consequences of invasive predator control in an Australian forest: overabundant wallabies and vegetation change.

Over-abundance of native herbivores is a problem in many forests worldwide. The abundance of native macropod wallabies is extremely high at Booderee National Park (BNP) in south-eastern Australia. This has occurred because of the reduction of exotic predators through an intensive baiting program, coupled with the absence of other predators. The high density of wallabies at BNP may be inhibiting the recruitment of many plant species following fire-induced recruitment events. We experimentally examined the post-fire response of a range of plant species to browsing by wallabies in a forest heavily infested with the invasive species, bitou bush Chrysanthemoides monilifera. We recorded the abundance and size of a range of plant species in 18 unfenced (browsed) and 16 fenced (unbrowsed) plots. We found the abundance and size of bitou bush was suppressed in browsed plots compared to unbrowsed plots. Regenerating seedlings of the canopy or middle storey tree species Eucalyptus pilularis, Acacia implexa, Allocasuarina littoralis, Breynia oblongifolia and Banksia integrifolia were either smaller or fewer in number in grazed plots than treatment plots as were the vines Kennedia rubicunda, Glycine tabacina and Glycine clandestina. In contrast, the understorey fern, Pteridium esculentum increased in abundance in the browsed plots relative to unbrowsed plots probably because of reduced competition with more palatable angiosperms. Twelve months after plots were installed the community structure of the browsed and unbrowsed plots was significantly different (P = 0.023, Global R = 0.091). The relative abundance of C. monilifera and P. esculentum contributed most to the differences. We discuss the possible development of a low diversity bracken fern parkland in Booderee National Park through a trophic cascade, similar to that caused by overabundant deer in the northern hemisphere. We also discuss its implications for broad scale fox control in southern Australian forests.