Filopodial protrusion driven by density-dependent Ena-TOCA-1 interactions.

Filopodia are narrow actin-rich protrusions with important roles in neuronal development where membrane-binding adaptor proteins, such as I-BAR- and F-BAR-domain-containing proteins, have emerged as upstream regulators that link membrane interactions to actin regulators such as formins and proteins of the Ena/VASP family. Both the adaptors and their binding partners are part of diverse and redundant protein networks that can functionally compensate for each other. To explore the significance of the F-BAR domain-containing neuronal membrane adaptor TOCA-1 (also known as FNBP1L) in filopodia we performed a quantitative analysis of TOCA-1 and filopodial dynamics in Xenopus retinal ganglion cells, where Ena/VASP proteins have a native role in filopodial extension. Increasing the density of TOCA-1 enhances Ena/VASP protein binding in vitro, and an accumulation of TOCA-1, as well as its coincidence with Ena, correlates with filopodial protrusion in vivo. Two-colour single-molecule localisation microscopy of TOCA-1 and Ena supports their nanoscale association. TOCA-1 clusters promote filopodial protrusion and this depends on a functional TOCA-1 SH3 domain and activation of Cdc42, which we perturbed using the small-molecule inhibitor CASIN. We propose that TOCA-1 clusters act independently of membrane curvature to recruit and promote Ena activity for filopodial protrusion.

A survey of stapling methods to increase affinity, activity, and stability of ghrelin analogues.

The growth hormone secretagogue receptor (GHSR) is a G protein-coupled receptor which regulates various important physiological and pathophysiological processes in the body such as energy homeostasis, growth hormone secretion and regulation of appetite. As a result, it has been postulated as a potential therapeutic target for the treatment of cancer cachexia and other metabolic disorders, as well as a potential imaging agent target for cancers and cardiovascular diseases. Ghrelin is the primary high affinity endogenous ligand for GHSR and has limited secondary structure in solution, which makes it proteolytically unstable. This inherent instability in ghrelin can be overcome by incorporating helix-inducing staples that stabilize its structure and improve affinity and activity. We present an analysis of different stapling methods at positions 12 and 16 of ghrelin(1-20) analogues with the goal of increasing proteolytic stability and to retain or improve affinity and activity towards the GHSR. Ghrelin(1-20) analogues were modified with a wide range of chemical staples, including a lactam staple, triazole staple, hydrocarbon staple, Glaser staple, and xylene-thioether staple. Once synthesized, the receptor affinity and α-helicity were measured using competitive binding assays and circular dichroism spectroscopy, respectively. Generally, an increase in alpha-helicity using a flexible staple linker led to improved affinity towards GHSR. Ghrelin(1-20) analogues with a lactam, triazole, and hydrocarbon staple resulted in helical analogues with stronger affinity towards GHSR than unstapled ghrelin(1-20), a compound that lacks helical character. Compounds were also investigated for their agonist activity through ß-arrestin 1 & 2 recruitment BRET assays and for their metabolic stability through serum stability analysis.

Climate resilience and risks of rigidity traps in Iceland's fisheries.

Iceland's fisheries system is well-governed, data-rich, and has adapted to past ecological change. It thus provides an opportunity to identify social-ecological attributes of climate resilience and interactions among them. We elicited barriers and enabling conditions for adaptation in Iceland's fisheries from semi-structured expert interviews, using projections of fish habitat shifts by mid-century to guide discussion. Interviewees highlighted flexible management, highly connected institutions that facilitate learning, ample assets to expand adaptive options, and cultural comfort with change. However, examining how these attributes interact in reinforcing feedback loops revealed potential rigidity traps, where optimization for resilience to stock shifts may render the system more vulnerable to extreme environmental change and social backlash. This study articulates resilience attributes that Iceland and other fisheries systems might prioritize as the climate changes. It further explores circumstances in which these same attributes risk forming traps, and potential pathways to escape them.

Cognitive behavioural techniques reduce exercise anxiety and improve adherence to a resistance training program for people with anxiety-related disorders: A randomized controlled trial.

Exercise is a cost-efficient alternative to other interventions for anxiety-related disorders (ARDs; e.g., psychotherapy, pharmaceutical) that is also associated with health benefits. Several exercise modalities, including resistance training (RT), have demonstrated efficacy at reducing symptoms of ARDs; however, there are challenges associated with effectively implementing such protocols, most notably, exercise avoidance or early discontinuation. Researchers have identified exercise anxiety as a contributor to exercise avoidance for people with ARDs. Exercise-based interventions for people with ARDs may need to include strategies for helping these individuals cope with exercise anxiety to facilitate long-term exercise engagement; however, research in this area is lacking. The primary purpose of this randomized controlled trial (RCT) was to examine the effects of combining cognitive behavioural techniques (CBT) with a RT program on changes in exercise anxiety, exercise frequency, disorder-specific anxiety symptoms, and physical activity for people with ARDs. A secondary purpose was to explore group differences in exercise motivation and exercise self-efficacy across time. A total of 59 physically inactive individuals with ARDs were randomized into either RT + CBT, RT, or waitlist (WL). Primary measures were assessed at baseline, weekly during the 4-week active phase, and at 1-week, 1-month, and 3-month follow-ups. Findings indicate both RT and RT + CBT can reduce exercise anxiety; however, the addition of CBT techniques may help facilitate improvements in exercise self-efficacy, reductions in disorder-specific anxiety, and increases in long-term exercise behaviour and vigorous physical activity. These techniques may be useful for researchers and clinicians alike in supporting individuals with ARDs interested in using exercise to cope with elevated anxiety.

The Myth of "Reliable Research" in Pediatric Gender Medicine: A critical evaluation of the Dutch Studies-and research that has followed.

Two Dutch studies formed the foundation and the best available evidence for the practice of youth medical gender transition. We demonstrate that this work is methodologically flawed and should have never been used in medical settings as justification to scale this "innovative clinical practice." Three methodological biases undermine the research: (1) subject selection assured that only the most successful cases were included in the results; (2) the finding that "resolution of gender dysphoria" was due to the reversal of the questionnaire employed; (3) concomitant psychotherapy made it impossible to separate the effects of this intervention from those of hormones and surgery. We discuss the significant risk of harm that the Dutch research exposed, as well as the lack of applicability of the Dutch protocol to the currently escalating incidence of adolescent-onset, non-binary, psychiatrically challenged youth, who are preponderantly natal females. "Spin" problems-the tendency to present weak or negative results as certain and positive-continue to plague reports that originate from clinics that are actively administering hormonal and surgical interventions to youth. It is time for gender medicine to pay attention to the published objective systematic reviews and to the outcome uncertainties and definable potential harms to these vulnerable youth.

Occupational therapists' practice with complex trauma: A profile.

INTRODUCTION: Many children in Aotearoa (New Zealand) and Australia experience complex trauma and its developmental impacts. Internationally, occupational therapists work with complex trauma and use sensory-based, integrative, and functional approaches. The practices of occupational therapists in Aotearoa and Australia with children experiencing complex trauma have not previously been described. METHODS: This article reports the quantitative results of a mixed-methods study which profiled occupational therapists' practice in Aotearoa and Australia with children aged 0 to 12 years old who experienced complex trauma. Twenty-five participants completed the survey. The average age of participants was 43 years (SD = 10.65), all were female (n = 25), and most identified as New Zealand European (n = 11) or Australian European (n = 9). A survey was distributed via Occupational Therapy New Zealand - Whakaora Ngangahau Aotearoa and Occupational Therapy Australia. RESULTS: The majority of participants had a bachelor's degree (64%) and worked in community settings (76%). Fourteen participants (56%) used sensory approaches. The most common assessments used were those of sensory processing (n = 12, 48%) and observation (n = 12, 48%). The Sensory Profile was the most popular standardised assessment (n = 8, 32%). The most common interventions used with children experiencing complex trauma were sensory (n = 13, 52%) and play based (n = 13, 52%). Most participants reported not adapting their practices for Maori or Aboriginal children. Most participants felt somewhat prepared (n = 15) for working with complex trauma, with most reporting a lack of experience in this area (n = 10). Supervision was suggested by 92% (n = 23) of the participants. CONCLUSION: Sensory-based practices were most common among occupational therapists in Aotearoa and Australia who worked with children experiencing complex trauma. The participants suggested supervision, social support, and practical training when working with complex trauma. Advocacy and research are required within this subspecialty, and further professional engagement in the application of culturally safe practice.

Reconsidering Informed Consent for Trans-Identified Children, Adolescents, and Young Adults.

In less than a decade, the western world has witnessed an unprecedented rise in the numbers of children and adolescents seeking gender transition. Despite the precedent of years of gender-affirmative care, the social, medical and surgical interventions are still based on very low-quality evidence. The many risks of these interventions, including medicalizing a temporary adolescent identity, have come into a clearer focus through an awareness of detransitioners. The risks of gender-affirmative care are ethically managed through a properly conducted informed consent process. Its elements-deliberate sharing of the hoped-for benefits, known risks and long-term outcomes, and alternative treatments-must be delivered in a manner that promotes comprehension. The process is limited by: erroneous professional assumptions; poor quality of the initial evaluations; and inaccurate and incomplete information shared with patients and their parents. We discuss data on suicide and present the limitations of the Dutch studies that have been the basis for interventions. Beliefs about gender-affirmative care need to be separated from the established facts. A proper informed consent process can both prepare parents and patients for the difficult choices that they must make and can ease professionals' ethical tensions. Even when properly accomplished, however, some clinical circumstances exist that remain quite uncertain.

Sex differences in mental disorder symptoms among Canadian police officers: the mediating role of social support, stress, and sleep quality.

Police officers, and specifically women officers, report elevated mental disorder rates relative to the general population, which may be impacted by sleep quality, policing-related stress, and social support. In a sample of Canadian police officers, sex was indirectly related to post traumatic stress, depression, generalized anxiety, panic, and social anxiety symptoms through its relationships with social support and sleep quality, but not through policing-related stress. Sex was indirectly related to problematic alcohol use symptoms through sleep quality only. Differences in clinical symptom severity between both sexes may be partially accounted for by the worsened sleep quality reported by women officers relative to their men counterparts. Conversely, general social support appears to be a protective, albeit insufficient, factor influencing the mental health of women police officers. Male and women police officers did not differ in their reports of policing-related stress. The current results underscore the importance of incorporating strategies to improve sleep practices into police workplace environments. Additionally, findings that general social support and policing-related stress do not help explain the trend of increased clinical severity reported by women police suggest that more research is still needed to identify and delineate other contributing factors.

Commentary: The Signal and the Noise-questioning the benefits of puberty blockers for youth with gender dysphoria-a commentary on Rew et al. (2021).

This commentary is a critique of a recent systematic review of the evidence for the use of puberty blockers for youth with gender dysphoria (GD) by Rew et al. (2021). In our view, the review suffers from several methodological oversights including the omission of relevant studies and suboptimal analysis of the quality of the included studies. This has resulted in an incomplete and incorrect assessment of the evidence base for the use of puberty blockers. We find that Rew et al.'s conclusions and clinician recommendations are problematic, especially when discussing suicidality. A key message of the review's abstract appears to be that puberty blockers administered in childhood reduce adult suicidality. However, the study used for the basis of this conclusion (Turban et al., 2020) did not make a causal claim between puberty blockers and decreased adult suicidality. Rather, it reported a negative association between using puberty blockers and lifetime suicidal ideation. The study design did not allow for determination of causation. Our commentary concludes by demonstrating how the GD medical literature, as it moves from one publication to the next, can overstate the evidence underpinning clinical practice recommendations for youth with GD.

Stochastic combinations of actin regulatory proteins are sufficient to drive filopodia formation.

Assemblies of actin and its regulators underlie the dynamic morphology of all eukaryotic cells. To understand how actin regulatory proteins work together to generate actin-rich structures such as filopodia, we analyzed the localization of diverse actin regulators within filopodia in Drosophila embryos and in a complementary in vitro system of filopodia-like structures (FLSs). We found that the composition of the regulatory protein complex where actin is incorporated (the filopodial tip complex) is remarkably heterogeneous both in vivo and in vitro. Our data reveal that different pairs of proteins correlate with each other and with actin bundle length, suggesting the presence of functional subcomplexes. This is consistent with a theoretical framework where three or more redundant subcomplexes join the tip complex stochastically, with any two being sufficient to drive filopodia formation. We provide an explanation for the observed heterogeneity and suggest that a mechanism based on multiple components allows stereotypical filopodial dynamics to arise from diverse upstream signaling pathways.

A direct role for SNX9 in the biogenesis of filopodia.

Filopodia are finger-like actin-rich protrusions that extend from the cell surface and are important for cell-cell communication and pathogen internalization. The small size and transient nature of filopodia combined with shared usage of actin regulators within cells confounds attempts to identify filopodial proteins. Here, we used phage display phenotypic screening to isolate antibodies that alter the actin morphology of filopodia-like structures (FLS) in vitro. We found that all of the antibodies that cause shorter FLS interact with SNX9, an actin regulator that binds phosphoinositides during endocytosis and at invadopodia. In cells, we discover SNX9 at specialized filopodia in Xenopus development and that SNX9 is an endogenous component of filopodia that are hijacked by Chlamydia entry. We show the use of antibody technology to identify proteins used in filopodia-like structures, and a role for SNX9 in filopodia.

Factor Structure and Measurement Invariance of the Alcohol Use Disorders Identification Test (AUDIT) in a Sample of Military Veterans with and without PTSD.

Background: The Alcohol Use Disorders Identification Test (AUDIT) was developed as a screening tool for problematic alcohol use and an intervention framework to aid those who drink excessively. While the AUDIT is widely used with at-risk populations, such as military veterans, major gaps exist in the research literature regarding the construct validity of the AUDIT in military samples. Objectives: This study assessed the factor structure and measurement invariance of the AUDIT in a large sample of Canadian military veterans (N = 1669; 94.94% male). Methods: Exploratory factor analysis (EFA) was conducted using a random subsample (n = 825) to assess the underlying factor structure of the AUDIT. Confirmatory factor analysis (CFA), using the second subsample (n = 844), was used to cross-validate the factor structure revealed by EFA and compare it to other model variants. Finally, multigroup CFAs were conducted using the whole sample to further cross-validate the factor structure and examine measurement invariance in military veterans with and without clinical elevations in posttraumatic stress disorder (PTSD) symptoms. Results: Factor analyses revealed that a modified two-factor model provided a statistically better fit to the data compared to all other model variants; yet, the results did not confirm measurement invariance across military veterans with and without clinically significant symptoms of PTSD. Conclusions/Importance: The findings are in line with increasing evidence suggesting that two subscale scores should be calculated for the AUDIT. Results further suggest that care should be taken in interpreting AUDIT scores when PTSD symptoms are present for military veterans.

Assessing the Relative Impact of Diverse Stressors among Public Safety Personnel.

Public Safety Personnel (PSP; e.g., correctional workers and officers, firefighters, paramedics, police officers, and public safety communications officials (e.g., call center operators/dispatchers)) are regularly exposed to potentially psychologically traumatic events (PPTEs). PSP also experience other occupational stressors, including organizational (e.g., staff shortages, inconsistent leadership styles) and operational elements (e.g., shift work, public scrutiny). The current research quantified occupational stressors across PSP categories and assessed for relationships with PPTEs and mental health disorders (e.g., anxiety, depression). The participants were 4820 PSP (31.7% women) responding to established self-report measures for PPTEs, occupational stressors, and mental disorder symptoms. PPTEs and occupational stressors were associated with mental health disorder symptoms (ps < 0.001). PSP reported substantial difficulties with occupational stressors associated with mental health disorder symptoms, even after accounting for diverse PPTE exposures. PPTEs may be inevitable for PSP and are related to mental health; however, leadership style, organizational engagement, stigma, sleep, and social environment are modifiable variables that appear significantly related to mental health.

Relationships between health behaviors, posttraumatic stress disorder, and comorbid general anxiety and depression.

Poor health outcomes associated with posttraumatic stress disorder (PTSD) may reflect engagement in unhealthy behaviors that increase morbidity risk and disengagement in healthy behaviors that decrease morbidity risk. Although research supports this pattern, findings are not definitive, particularly for healthy behaviors. Many studies have not controlled for effects of concurrent generalized anxiety and depression, which might explain conflicting findings. To address this limitation, we used an online cross-sectional research design and multivariate multilevel modelling to evaluate associations between a multitude of health behaviors (i.e. sedentary behavior, sleep quality, physical activity, eating habits, alcohol use and substance use) and PTSD, while adjusting for comorbid generalized anxiety and depression, in a sample of trauma-exposed individuals (N = 246). Our results indicate that PTSD and comorbid generalized anxiety and depression symptoms were differentially associated with specific health constructs. Specifically, sedentary behavior and poor sleep quality were associated with PTSD, whereas low physical activity, poor sleep quality, and unhealthy eating habits were associated with depression. Both increased alcohol and substance use were associated with generalized anxiety. Results from our study highlight the need to conceptualize associations between health behaviors and specific psychological symptoms in a comprehensive manner as part of clinical presentations of PTSD.

A longitudinal assessment of the road to mental readiness training among municipal police.

Police agencies increasingly implement training programs to protect mental health. The Road to Mental Readiness (R2MR) program was designed by the Canadian military to increase mental health resilience. A version of R2MR was adapted for municipal police by the Mental Health Commission of Canada (MHCC). The current research was designed to assess the R2MR program, as adapted and delivered by the MHCC, in a municipal police sample. Participants were 147 Canadian police agency employees (57% women) who received a single R2MR training session. Participants completed pre- and post-training self-report questionnaires, and follow-ups at 6 and 12 months. The questionnaires assessed mental health symptoms, work engagement, resiliency, mental health knowledge, and stigma. Multilevel modeling analyses assessed for within-participant changes over time. The results were consistent with other single session interventions; specifically, there were no significant changes in mental health symptoms, resilience, or work engagement (p > .05). There were small, but significant (p < .05), reductions in stigma at post-training that may facilitate help-seeking among police; relatedly, in open-ended response fields, participants commonly described the training as helpful for changing attitudes and improving communication. More engagement with the material may produce larger, sustained gains, but more published research is critically needed.

Control of actin polymerization via the coincidence of phosphoinositides and high membrane curvature.

The conditional use of actin during clathrin-mediated endocytosis in mammalian cells suggests that the cell controls whether and how actin is used. Using a combination of biochemical reconstitution and mammalian cell culture, we elucidate a mechanism by which the coincidence of PI(4,5)P2 and PI(3)P in a curved vesicle triggers actin polymerization. At clathrin-coated pits, PI(3)P is produced by the INPP4A hydrolysis of PI(3,4)P2, and this is necessary for actin-driven endocytosis. Both Cdc42⋅guanosine triphosphate and SNX9 activate N-WASP-WIP- and Arp2/3-mediated actin nucleation. Membrane curvature, PI(4,5)P2, and PI(3)P signals are needed for SNX9 assembly via its PX-BAR domain, whereas signaling through Cdc42 is activated by PI(4,5)P2 alone. INPP4A activity is stimulated by high membrane curvature and synergizes with SNX9 BAR domain binding in a process we call curvature cascade amplification. We show that the SNX9-driven actin comets that arise on human disease-associated oculocerebrorenal syndrome of Lowe (OCRL) deficiencies are reduced by inhibiting PI(3)P production, suggesting PI(3)P kinase inhibitors as a therapeutic strategy in Lowe syndrome.

Filopodyan: An open-source pipeline for the analysis of filopodia.

Filopodia have important sensory and mechanical roles in motile cells. The recruitment of actin regulators, such as ENA/VASP proteins, to sites of protrusion underlies diverse molecular mechanisms of filopodia formation and extension. We developed Filopodyan (filopodia dynamics analysis) in Fiji and R to measure fluorescence in filopodia and at their tips and bases concurrently with their morphological and dynamic properties. Filopodyan supports high-throughput phenotype characterization as well as detailed interactive editing of filopodia reconstructions through an intuitive graphical user interface. Our highly customizable pipeline is widely applicable, capable of detecting filopodia in four different cell types in vitro and in vivo. We use Filopodyan to quantify the recruitment of ENA and VASP preceding filopodia formation in neuronal growth cones, and uncover a molecular heterogeneity whereby different filopodia display markedly different responses to changes in the accumulation of ENA and VASP fluorescence in their tips over time.