An optogenetic method for the controlled release of single molecules.

We developed a system for optogenetic release of single molecules in cells. We confined soluble and transmembrane proteins to the Golgi apparatus via a photocleavable protein and released them by short pulses of light. Our method allows for a light dose-dependent delivery of functional proteins to the cytosol and plasma membrane in amounts compatible with single-molecule imaging, greatly simplifying access to single-molecule microscopy of any protein in live cells. We were able to reconstitute ion conductance by delivering BK and LRRC8/volume-regulated anion channels to the plasma membrane. Finally we were able to induce NF-kB signaling in T lymphoblasts stimulated by interleukin-1 by controlled release of a signaling protein that had been knocked out. We observed light-induced formation of functional inflammatory signaling complexes that triggered phosphorylation of the inhibitor of nuclear factor kappa-B kinase only in activated cells. We thus developed an optogenetic method for the reconstitution and investigation of cellular function at the single-molecule level.

Unique variants in CLCN3, encoding an endosomal anion/proton exchanger, underlie a spectrum of neurodevelopmental disorders.

The genetic causes of global developmental delay (GDD) and intellectual disability (ID) are diverse and include variants in numerous ion channels and transporters. Loss-of-function variants in all five endosomal/lysosomal members of the CLC family of Cl- channels and Cl-/H+ exchangers lead to pathology in mice, humans, or both. We have identified nine variants in CLCN3, the gene encoding CIC-3, in 11 individuals with GDD/ID and neurodevelopmental disorders of varying severity. In addition to a homozygous frameshift variant in two siblings, we identified eight different heterozygous de novo missense variants. All have GDD/ID, mood or behavioral disorders, and dysmorphic features; 9/11 have structural brain abnormalities; and 6/11 have seizures. The homozygous variants are predicted to cause loss of ClC-3 function, resulting in severe neurological disease similar to the phenotype observed in Clcn3-/- mice. Their MRIs show possible neurodegeneration with thin corpora callosa and decreased white matter volumes. Individuals with heterozygous variants had a range of neurodevelopmental anomalies including agenesis of the corpus callosum, pons hypoplasia, and increased gyral folding. To characterize the altered function of the exchanger, electrophysiological analyses were performed in Xenopus oocytes and mammalian cells. Two variants, p.Ile607Thr and p.Thr570Ile, had increased currents at negative cytoplasmic voltages and loss of inhibition by luminal acidic pH. In contrast, two other variants showed no significant difference in the current properties. Overall, our work establishes a role for CLCN3 in human neurodevelopment and shows that both homozygous loss of ClC-3 and heterozygous variants can lead to GDD/ID and neuroanatomical abnormalities.

Eye movement desensitisation and reprocessing as add-on treatment to enhanced cognitive behaviour therapy for patients with anorexia nervosa reporting childhood maltreatment: A quasi-experimental multicenter study.

OBJECTIVE: This quasi-experimental study aimed to compare the outcome of patients with Anorexia Nervosa (AN) reporting moderate/severe childhood maltreatment (CM) treated exclusively with Enhanced Cognitive Behaviour Therapy (CBT-E) or with CBT-E plus Eye Movement Desensitisation and Reprocessing (EMDR). METHOD: A total of 75 patients with AN reporting moderate/severe CM were initially assessed regarding body mass index (BMI), general and eating disorder (ED)-specific psychopathology, and dissociative symptoms, and re-evaluated after 40 CBT-E sessions (T1). Then, 18 patients received EMDR, whereas the others were placed on a waiting list and continued CBT-E. T2 assessment was performed after 20-25 sessions of EMDR or CBT-E. A control group of 67 patients without CM was also enroled and treated with CBT-E. RESULTS: Contrary to patients without CM, neither of the traumatised groups improved in BMI, general and ED psychopathology, or dissociation at T1. However, at T2, both traumatised groups improved in BMI and ED-specific psychopathology, with the CBT + EMDR group demonstrating greater improvements. Moreover, only the CBT + EMDR group improved in general psychopathology and dissociative symptoms. The reduction of ED symptoms in traumatised patients was mediated by the amelioration of dissociation. DISCUSSION: The addition of EMDR to CBT-E may benefit patients with AN reporting moderate/severe CM.

Molecular determinants underlying volume-regulated anion channel subunit-dependent oxidation sensitivity.

The volume-regulated anion channel (VRAC) is formed by LRRC8 subunits. Besides their role in the maintenance of cell homeostasis, VRACs are critically involved in oxidative stress mechanisms: reactive oxygen species directly modulate VRACs in a subunit-dependent manner. It was reported that LRRC8A-LRRC8E heteromeric channels are activated by oxidation, whereas LRRC8A-LRRC8C heteromers are inhibited. Here we adopted chimeric- as well as concatemeric-based strategies to identify residues responsible for the divergent effect of oxidants. We identified two cysteines in the first two leucine rich repeats of LRRC8E, C424 and C448, as the targets of oxidation. Oxidation likely results in the formation of a disulfide bond between the two cysteines, which in turn induces a conformational change leading to channel activation. Additionally, we found that LRRC8C inhibition is caused by oxidation of the first methionine. We thus identified crucial molecular elements involved in channel activation, which are conceivably relevant in determining physiological ROS effects. KEY POINTS: Volume-regulated anion channels (VRACs) are heterohexameric complexes composed of an essential LRRC8A subunit and a variable number of LRRC8B-E subunits. VRACs are directly regulated by oxidation, with LRRC8A-LRRC8E heteromers being potentiated and LRRC8A-LRRC8C heteromers being inhibited by oxidation. We identified two LRRC8E specific intracellular cysteines that form a disulfide bond upon oxidation leading to LRRC8A-LRRC8E potentiation. Inhibition of LRRC8A-LRRC8C heteromers is mediated by the oxidation of the start methionine, being additionally dependent on the identity of the LRR domain. Besides providing physiological insights concerning the outcome of reactive oxygen species modulation, the results point to key structural elements involved in VRAC activation.

Eating disturbances in eating disorders and in high-functioning autism spectrum disorders: a preliminary study.

PURPOSE: The relationship between autism spectrum disorders (ASDs) and eating disorders (EDs) has been widely studied in the last decades. We aimed to directly compare patients with EDs, individuals with high-functioning ASDs (HF-ASDs) and healthy controls (HC) at measures detecting: (1) symptoms of eating disorders, (2) eating disturbances known to be characteristic of autism. METHODS: Thirty-four patients with EDs, 34 individuals with HF-ASDs and 35 HC, all females, completed the eating attitude test (EAT-26) and the Swedish eating assessment for autism spectrum disorders (SWEAA), two self-report questionnaires assessing, respectively, symptoms and concerns characteristic of eating disorders and ASD-related eating disturbances. RESULTS: At the EAT-26, patients with EDs scored significantly higher than individuals with HF-ASDs, and both of them scored higher than HC (p < 0.05, ηp2 = 0.283). Conversely, at the SWEAA, no differences between individuals with HF-ASDs and patients with EDs emerged (p = 901), but they both scored higher than HC (p < 0.05, ηp2 = 0.247). CONCLUSION: Individuals with HF-ASDs did not seem to reach the same level of EDs symptomatology as patients with EDs. Patients with EDs did not seem to present a different amount of autistic-eating behaviours than subjects with HF-ASDs. Patients with EDs and individuals with HF-ASDs scored higher than HC at both scales. Our results give further preliminary evidence of the overlap between autistic traits and EDs symptomatology, and should be taken into account in the definition of a shared model between EDs and ASDs. LEVEL OF EVIDENCE: Level II; Evidence obtained from controlled trial without randomization.

Mechanisms of Activation of LRRC8 Volume Regulated Anion Channels.

Volume regulated anion channels (VRACs) are ubiquitously expressed in all vertebrate cells. Despite many years of research, the fundamental mechanisms underlying VRAC activation are not understood. The recent molecular identification of the LRRC8 genes underlying VRAC revealed that VRACs are formed by a hexameric assembly of members of the LRRC8 gene family. Knowing the genes underlying VRACs allowed the discovery of novel VRAC functions into cell volume regulation, and first structure function studies revealed important insight in channel activation mechanisms. The determination of cryo-EM structures of homomeric LRRC8A and LRRC8D complexes provide a framework for a rational approach to investigate biophysical mechanisms. We discuss several recent advances within the structural framework, and we critically review the literature on the main mechanisms proposed to be involved in VRAC activation, including low intracellular ionic strength, membrane unfolding, oxidation, phosphorylation and G-protein coupling.

TRPM2 Oxidation Activates Two Distinct Potassium Channels in Melanoma Cells through Intracellular Calcium Increase.

Tumor microenvironments are often characterized by an increase in oxidative stress levels. We studied the response to oxidative stimulation in human primary (IGR39) or metastatic (IGR37) cell lines obtained from the same patient, performing patch-clamp recordings, intracellular calcium ([Ca2+]i) imaging, and RT-qPCR gene expression analysis. In IGR39 cells, chloramine-T (Chl-T) activated large K+ currents (KROS) that were partially sensitive to tetraethylammonium (TEA). A large fraction of KROS was inhibited by paxilline-a specific inhibitor of large-conductance Ca2+-activated BK channels. The TEA-insensitive component was inhibited by senicapoc-a specific inhibitor of the Ca2+-activated KCa3.1 channel. Both BK and KCa3.1 activation were mediated by an increase in [Ca2+]i induced by Chl-T. Both KROS and [Ca2+]i increase were inhibited by ACA and clotrimazole-two different inhibitors of the calcium-permeable TRPM2 channel. Surprisingly, IGR37 cells did not exhibit current increase upon the application of Chl-T. Expression analysis confirmed that the genes encoding BK, KCa3.1, and TRPM2 are much more expressed in IGR39 than in IGR37. The potassium currents and [Ca2+]i increase observed in response to the oxidizing agent strongly suggest that these three molecular entities play a major role in the progression of melanoma. Pharmacological targeting of either of these ion channels could be a new strategy to reduce the metastatic potential of melanoma cells, and could complement classical radio- or chemotherapeutic treatments.

The psychological impact of COVID-19-related lockdown measures among a sample of Italian patients with eating disorders: a preliminary longitudinal study.

PURPOSE: To explore the prevalence of symptoms of anxiety and depression, along with PTSD- and ED-related symptoms, across a sample of patients with Eating Disorders (EDs) compared to a group of healthy controls (HC) during the lockdown period in Italy; to assess whether patients' reported aforementioned psychiatric symptoms improved, remained stable or worsened with the easing of the lockdown measures. METHODS: t0 assessment (during lockdown): 59 ED patients and 43 HC completed an online survey, including the Depression, Anxiety and Stress Scale-21 items (DASS-21), the Impact of Event Scale-Revised (IES-R), the Perceived Stress Scale (PSS), and specific ad-hoc questions extracted from the Eating Disorder Examination-Questionnaire; t1 assessment (post-lockdown): 40 EDs patients, a subset of the t0 sample, completed the same assessment 2 months after t0. RESULTS: EDs patients scored higher than HC at the DASS-21, IES-R and PSS. At t1, levels of stress, anxiety and depression were not different than at t0, but symptoms related to post-traumatic stress disorder (PTSD), patients' reported level of psychological wellbeing and specific EDs symptomatology improved. DISCUSSION: During the lockdown, EDs patients presented significantly higher levels of stress, anxiety, depression, PTSD- and ED-related symptoms than HC. With the easing of the lockdown, PTSD- and ED-related symptoms improved, but high levels of stress, anxiety and depression persisted. LEVEL OF EVIDENCE: Level I, experimental study.

The effect of a single yoga class on interoceptive accuracy in patients affected by anorexia nervosa and in healthy controls: a pilot study.

PURPOSE: To evaluate interoceptive accuracy (Iac) before and after a single yoga class in a population of patients with anorexia nervosa (AN) and in a population of healthy controls (HC). METHODS: Fifteen patients with AN and twenty HC were included in the study. All individuals participated in a single yoga class. Before (T0) and after (T1) the yoga class, they underwent the heartbeat detection task for the evaluation of Iac. At T0, all participants also underwent a psychological assessment, including evaluation of depression, anxiety, body awareness, alexithymia, self-objectification and eating disorders psychopathology. RESULTS: Patients with AN had lower Iac than HC at T0. A significant improvement of Iac at T1 was found in the HC group but not in the group of patients with AN. CONCLUSION: We infer that our findings might be linked to the fact that patients with AN, differently from HC, did not properly attend to their bodies, despite the yoga class. This hypothesis is consistent with previous studies showing that patients with AN have decreased Iac during self-focused behavior because of body-related avoidance. Moreover, we surmise that HC might be keener to improve their perception of internal body signals even after a single yoga class because their emotional awareness system is not impaired. Patients with AN, on the contrary, may have an intrinsic impairment of their emotional awareness, making it harder for them to modulate their Iac. LEVEL OF EVIDENCE: Level III, evidence obtained from well-designed cohort or case-control analytical studies.

Risk and resilience factors for specific and general psychopathology worsening in people with Eating Disorders during COVID-19 pandemic: a retrospective Italian multicentre study.

PURPOSE: The COVID-19 pandemic restrictions had negative impact on the psychopathology of people with Eating Disorders (EDs). Factors involved in the vulnerability to stressful events have been under-investigated in this population. We aimed to assess which factors contributed to COVID-19-induced worsening in both general and specific psychopathology. METHODS: Three-hundred and twelve people with a clinically defined diagnosis of an ED and undergoing a specialist ED treatment in different Italian ED services before the spreading of COVID-19 pandemic filled in an online survey. ED specific and general psychopathology changes after COVID-19 quarantine were retrospectively evaluated. Factors related to COVID-19 concerns (financial condition, fear of contagion, perceived social isolation/support, satisfaction in peer, family or sentimental relationships), illness duration and treatment-related variables (type of treatment provided, type of access to care, satisfaction with therapeutic relationships) were included as predicting factors in a structural equational model, which included latent variables consisting of general and ED psychopathology items as outcomes. RESULTS: A perceived low quality of therapeutic relationships, fear of contagion and increased isolation were positively associated with psychopathology worsening. Reduced satisfaction with family and with friends' relationships and reduced perceived social support were associated with ED and general symptoms deterioration, respectively. No significant effect emerged for intimate relationships, illness duration, economic condition and type of treatment. CONCLUSIONS: This study provides a comprehensive evaluation of clinical variables associated with psychopathological changes during the COVID-19 lockdown period highlighting potential risk and resilience factors and, possibly, informing treatment as well as prevention strategies for EDs. LEVEL OF EVIDENCE IV: Evidence obtained from multiple time series analysis such as case studies.

Epilepsy in isolated parenchymal neurocutaneous melanosis: A systematic review.

PURPOSE: Neurocutaneous melanosis (NCM) is a rare congenital syndrome characterized by giant melanocytic cutaneous nevi and melanosis within the central nervous system (CNS), often sparing leptomeninges and concentrated in the brain parenchyma. Epilepsy and neurodevelopmental abnormalities are the only complications reported in children with isolated parenchymal melanosis. A minority of patients experience drug-resistant epilepsy, and up to now, no predictors of epilepsy prognosis have been identified. METHODS: In this systematic review, according to preferred reporting items for systematic review and meta-analysis (PRISMA) guidelines, we aggregated clinical cases of patients with isolated parenchymal melanosis affected by epilepsy, in order to recognize predictors of clinical outcome and to clarify indications of available therapeutic approaches. RESULTS: Sixteen articles (19 patients) were included in the final analysis from initial database research; 4 articles (4 patients) were selected from reference lists and 1 from conference abstracts (1 patient). In our series, distribution of parenchymal melanosis was the best predictor of epilepsy outcome: frequencies of seizure-free patients were different between cases of isolated/bilateral amygdale melanosis and those of multiple localizations (p = 0.037). Failure of antiepileptic drugs (AEDs) and/or surgical epilepsy therapy were associated with poor cognitive outcome (p = 0.03). CONCLUSION: Antiepileptic drugs were effective in the majority of patients with epilepsy with parenchymal melanosis. In case of multifocal distribution, more than one-third of patients presented a drug-resistant epilepsy. Epilepsy surgery is the best choice in patients with isolated amygdala localization. We propose the recognition of a multifactorial nature of cognitive impairment in neuromelanosis, emphasizing the role of drug-resistant epilepsy.

Gain of function of sporadic/familial hemiplegic migraine-causing SCN1A mutations: Use of an optimized cDNA.

INTRODUCTION: Familial hemiplegic migraine 3 is an autosomal dominant headache disorder associated with aura and transient hemiparesis, caused by mutations of the neuronal voltage-gated sodium channel Nav1.1. While a gain-of function phenotype is generally assumed to underlie familial hemiplegic migraine, this has not been fully explored. Indeed, a major obstacle in studying in vitro neuronal sodium channels is the difficulty in propagating and mutagenizing expression plasmids containing their cDNAs. The aim of this work was to study the functional effect of two previously uncharacterized hemiplegic migraine causing mutations, Leu1670Trp (L1670W) and Phe1774Ser (F1774S). METHODS: A novel SCN1A containing-plasmid was designed in silico and synthesized, and migraine mutations were inserted in this background. Whole-cell patch clamp was performed to investigate the functional properties of mutant Nav1.1 transiently expressed in Human Embryonic Kidney 293 cells. RESULTS AND CONCLUSIONS: We generated an optimized Nav1.1 expression plasmid that was extremely simple to handle and used the novel plasmid to study the functional effects of two migraine mutations. We observed that L1670W, but not F1774S, reduced current density and that both mutations led to a dramatic increase in persistent sodium currents, a depolarizing shift of the steady state-inactivation voltage-dependence, and a faster recovery from inactivation. The results are consistent with a major gain-of function effect underlying familial hemiplegic migraine 3. Our optimization strategy will help to characterize in an efficient manner the effect in vitro of mutations of neuronal voltage-gated sodium channels.

Transcranial direct current stimulation modulates implicit attitudes towards food in eating disorders.

OBJECTIVE: Neuromodulation of regions involved in food processing is increasingly used in studies on eating behaviors, but results are controversial. We assessed the effects of anodal transcranial direct current stimulation (a-tDCS) on food and body implicit preferences in patients with eating disorders (EDs). METHOD: Thirty-six ED patients and 36 healthy females completed three sessions with a-tDCS applied to the medial-prefrontal cortex (mPFC), the right extrastriate body area (rEBA) or in sham mode. Each participant then completed three Implicit Association Tests (IATs) on tasty/tasteless food, underweight/overweight body images, flowers versus insects as control. Differences in latency between incongruent and congruent blocks were calculated (D score). RESULTS: The tDCS by group interaction was significant for the IAT-food D score, with patients showing weaker preference for tasty food than controls in sham, but not a-tDCS sessions. In particular, rEBA stimulation significantly increased patients' D score compared to sham. Moreover, a-tDCS over mPFC and rEBA selectively increased patients' reaction times in the incongruent blocks of the IAT-food. DISCUSSION: A-tDCS on frontal and occipito-temporal cortices modulated food preferences in ED patients. The effect was specific for food images and selective in patients, but not in healthy participants. These findings suggest that neuromodulation of these regions could affect implicit food attitudes.

Advances on tinamou phylogeny: an assembled cladistic study of the volant palaeognathous birds.

Tinamous are volant terrestrial birds, endemic to the Neotropics. Here, an inclusive phenotype-based phylogenetic study of the interrelationships among all extinct and living species of tinamous is conducted. In this cladistic analysis, results are compared between main character subsets and with previous molecular studies. Special attention is paid to character definition and scoring of integumentary and behavioural characters: transformation costs are applied to analyse egg coloration and plumage characters-on the basis of pigment composition and overlap of pigmentation patterns respectively-in the context of generalized (Sankoff) parsimony. Cladistic analysis recovers the traditional subdivision between those tinamous specialized for open areas (Nothurinae) and those inhabiting forested environments (Tinaminae) and support the monophyly of recognized genera. The present study demonstrates that morphological analysis yields highly congruent results when compared with previous molecular studies; thus, it provides morphological synapomorphies for clades that have been proposed by these molecular analyses. The placement of the fossil species within the open-area (Nothurinae) and the forest-dwelling (Tinaminae) tinamous is also consistent with the palaeoenvironmental conditions inferred from the associated flora and fauna.

Anorexia Nervosa and Functional Motor Symptoms: Two Faces of the Same Coin?

The aim of this study was to compare anorexia nervosa (AN) patients and functional motor symptoms (FMS) patients by assessing their variability in demographic and clinical characteristics, risk factors, precipitators, and family history. The authors assessed levels of depression and anxiety, alexithymia, dissociation, body awareness, self-objectification, and interoception in the two groups, using 20 healthy controls (HCs) as a control group for psychometric assessment and for interoception. Unexpectedly, no differences in the three groups were observed for the measures related to awareness of physical state, including body awareness, self-objectification, and interoceptive ability via the heartbeat task. However, the AN and FMS groups were not different from each other but were different from the HC group with regard to anxiety, depression, alexithymia, and dissociation. In light of the similarities found, these data support the hypothesis of a common etiology involving emotional dysfunction in both disorders. These findings suggest that AN and FMS may be disorders belonging to the same spectrum (where emotional dysregulation is a key feature) and that there exist potential opportunities for collaborative, integrated investigations of etiology, diagnosis, and management of these disorders.

Early identification of risk for eating disorders in Italian secondary school students: a cross-sectional study.

AIM: Eating disorders are major illnesses that primarily affect adolescents and young adults and seriously threaten public health. Early identification of at-risk individuals and timely initiation of treatment is crucial to improve outcomes. The Inside Out Institute Screener (IOI-S) is a rapid self-administration screening tool for high-risk and early-stage eating disorders. This study aimed to investigate the risk of having an eating disorder in a sample of Italian students by testing the Italian version of the IOI-S. METHODS: A multicentre cross-sectional study was conducted in a population of students aged 12-19 years; validity and reliability of the IOI-Sita were investigated. RESULTS: Four-hundred and ninety-one (81.97%) students were enrolled, 24.85% of whom were found to be at "very high risk" of an eating disorder, according to IOI-Sita. Younger (p<0.001) and female (p<0.001) students had higher risk scores. The EFA confirmed the original monodimensional structure of the tool, S-CVI=0.95%. The Content Validity Index of the scale (S-CVI) was 0.95, ω coefficient was 0.927. DISCUSSION AND CONCLUSIONS: This research confirms the need to screen for eating disorders in Italian youth adequately; the psychometric properties of the IOI-Sita confirm it as a valid and reliable tool for screening high-risk and early-stage eating disorders.

Interactomic exploration of LRRC8A in volume-regulated anion channels.

Ion channels are critical in enabling ion movement into and within cells and are important targets for pharmacological interventions in different human diseases. In addition to their ion transport abilities, ion channels interact with signalling and scaffolding proteins, which affects their function, cellular positioning, and links to intracellular signalling pathways. The study of "channelosomes" within cells has the potential to uncover their involvement in human diseases, although this field of research is still emerging. LRRC8A is the gene that encodes a crucial protein involved in the formation of volume-regulated anion channels (VRACs). Some studies suggest that LRRC8A could be a valuable prognostic tool in different types of cancer, serving as a biomarker for predicting patients' outcomes. LRRC8A expression levels might be linked to tumour progression, metastasis, and treatment response, although its implications in different cancer types can be varied. Here, publicly accessible databases of cancer patients were systematically analysed to determine if a correlation between VRAC channel expression and survival rate exists across distinct cancer types. Moreover, we re-evaluated the impact of LRRC8A on cellular proliferation and migration in colon cancer via HCT116 LRRC8A-KO cells, which is a current topic of debate in the literature. In addition, to investigate the role of LRRC8A in cellular signalling, we conducted biotin proximity-dependent identification (BioID) analysis, revealing a correlation between VRAC channels and cell-cell junctions, mechanisms that govern cellular calcium homeostasis, kinases, and GTPase signalling. Overall, this dataset improves our understanding of LRRC8A/VRAC and explores new research avenues while identifying promising therapeutic targets and promoting inventive methods for disease treatment.

Updating Prospective Self-Efficacy Beliefs About Cardiac Interoception in Anorexia Nervosa: An Experimental and Computational Study.

Patients with anorexia nervosa (AN) typically hold altered beliefs about their body that they struggle to update, including global, prospective beliefs about their ability to know and regulate their body and particularly their interoceptive states. While clinical questionnaire studies have provided ample evidence on the role of such beliefs in the onset, maintenance, and treatment of AN, psychophysical studies have typically focused on perceptual and 'local' beliefs. Across two experiments, we examined how women at the acute AN (N = 86) and post-acute AN state (N = 87), compared to matched healthy controls (N = 180) formed and updated their self-efficacy beliefs retrospectively (Experiment 1) and prospectively (Experiment 2) about their heartbeat counting abilities in an adapted heartbeat counting task. As preregistered, while AN patients did not differ from controls in interoceptive accuracy per se, they hold and maintain 'pessimistic' interoceptive, metacognitive self-efficacy beliefs after performance. Modelling using a simplified computational Bayesian learning framework showed that neither local evidence from performance, nor retrospective beliefs following that performance (that themselves were suboptimally updated) seem to be sufficient to counter and update pessimistic, self-efficacy beliefs in AN. AN patients showed lower learning rates than controls, revealing a tendency to base their posterior beliefs more on prior beliefs rather than prediction errors in both retrospective and prospective belief updating. Further explorations showed that while these differences in both explicit beliefs, and the latent mechanisms of belief updating, were not explained by general cognitive flexibility differences, they were explained by negative mood comorbidity, even after the acute stage of illness.

Adaptation and validation of the Caregiver Burden Inventory in eating disorders.

BACKGROUND: Living with people diagnosed with a mental disorder is known to increase the risk of developing high levels of so-called "caregiver burden" in informal caregivers. In-depth analysis of this phenomenon and specific assessment tools for caregivers of patients diagnosed with Eating Disorders (EDs) are lacking. In this study, we aimed to evaluate the psychometric properties of the Caregiver Burden Inventory in EDs and employ this adapted tool in this category of caregivers. METHODS: A cross-sectional study was conducted in the Eating Disorders outpatient unit of an Italian University hospital. Face and content validity were investigated by calculating standard Content Validity Indices (CVI-I and CVI-S) after administering the Inventory to 6 expert nurses with at least 5 years of experience in mental health services assisting people diagnosed with Eating Disorders. Internal consistency was evaluated with Cronbach's α coefficient for the overall scale and subscales. An exploratory factor analysis (EFA) was performed to explore latent constructs. The adapted CBI was then administered to 62 informal caregivers of ED patients. RESULTS: The EFA yielded a 5-factor structure. The CVI-S was 97.2%; the Cronbach α coefficient was 0,90 (> 0.74 in each subscale). The median burden level in the experimental population was 40.0 [range = 21 to 54], in a theoretical range from 0 (no burden) to 96 (highest level of burden). CONCLUSION: The Caregiver Burden Inventory appears to be a valid and reliable instrument to assess caregiver burden in individuals diagnosed with Eating Disorders. Further research is needed to evaluate this tool's efficiency in improving individually tailored interventions on families.

Eating disorders (EDs) are characterized by the development of abnormal eating habits, dysregulation of body weight and/or body image distortion. The pervasiveness of EDs may significantly affect the life of patients' caregivers in terms of emotional load, termed Caregiver Burden (CB). This may impact the quality of life of the entire family and promote conflicts which may in turn exacerbate ED behaviours. Among several international assessment tools to measure CB, the Caregiver Burden Inventory (CBI) is perhaps the most widely employed. However, this tool has never been specifically tested in caregivers of ED patients. Here we show that CBI can adequately quantify burden in this category of caregivers. Systematic use of this instrument can support healthcare workers who aim to address the carer's discomfort and promote a focused monitoring of subjects with increased risk, in order to adequately plan targeted intervention programs.

The VRAC blocker DCPIB directly gates the BK channels and increases intracellular Ca2+ in melanoma and pancreatic duct adenocarcinoma cell lines.

BACKGROUND AND PURPOSE: The volume regulated anion channel (VRAC) is known to be involved in different aspects of cancer cell behaviour and response to therapies. For this reason, we investigated the effect of DCPIB, a presumably specific blocker of VRAC, in two types of cancer: pancreatic duct adenocarcinoma (PDAC) and melanoma. EXPERIMENTAL APPROACH: We used patch-clamp electrophysiology, supported by Ca2+ imaging, gene expression analysis, docking simulation and mutagenesis. We employed two PDAC lines (Panc-1 and MiaPaCa-2), as well as a primary (IGR39) and a metastatic (IGR37) melanoma line. KEY RESULTS: DCPIB markedly increased whole-cell currents in Panc-1, MiaPaca2 and IGR39, but not in IGR37 cells. The currents were mostly mediated by KCa 1.1 channels, commonly known as BK channels. We confirmed DCPIB activation of BK channels also in HEK293 cells transfected with α subunits of this channel. Further experiments showed that in IGR39, and to a smaller degree also in Panc-1 cells, DCPIB induced a rapid Ca2+ influx. This, in turn, indirectly potentiated BK channels and, in IGR39 cells, additionally activated other Ca2+ -dependent channels. However, Ca2+ influx was not required for activation of BK channels by DCPIB, as such activation involved the extracellular part of the protein and we have identified a residue crucial for binding. CONCLUSION AND IMPLICATIONS: DCPIB directly targeted BK channels and, also, acutely increased intracellular Ca2+ . Our findings extend the list of DCPIB effects that should be taken into consideration for future development of DCPIB-based modulators of ion channels and other membrane proteins.