Rescue of myocytes and locomotion through AAV2/9-2YF intracisternal gene therapy in a rat model of creatine transporter deficiency.

Creatine deficiency syndromes (CDS), caused by mutations in GATM (AGAT), GAMT, and SLC6A8, mainly affect the central nervous system (CNS). CDS show brain creatine (Cr) deficiency, intellectual disability with severe speech delay, behavioral troubles, epilepsy, and motor dysfunction. AGAT/GAMT-deficient patients lack brain Cr synthesis but express the Cr transporter SLC6A8 at the blood-brain barrier and are thus treatable by oral supplementation of Cr. In contrast, no satisfactory treatment has been identified for Cr transporter deficiency (CTD), the most frequent of CDS. We used our Slc6a8Y389C CTD rat model to develop a new AAV2/9-2YF-driven gene therapy re-establishing the functional Slc6a8 transporter in rat CNS. We show, after intra-cisterna magna AAV2/9-2YF-Slc6a8-FLAG vector injection of postnatal day 11 pups, the transduction of Slc6a8-FLAG in cerebellum, medulla oblongata, and spinal cord as well as a partial recovery of Cr in these brain regions, together with full prevention of locomotion defaults and impairment of myocyte development observed in Slc6a8Y389 C/y male rats. While more work is needed to correct those CTD phenotypes more associated with forebrain structures, this study is the first demonstrating positive effects of an AAV-driven gene therapy on CTD and thus represents a very encouraging approach to treat the so-far untreatable CTD.

Adolescent Stress-Induced Ventral Hippocampus Redox Dysregulation Underlies Behavioral Deficits and Excitatory/Inhibitory Imbalance Related to Schizophrenia.

BACKGROUND AND HYPOTHESIS: Redox dysregulation has been proposed as a convergent point of childhood trauma and the emergence of psychiatric disorders, such as schizophrenia (SCZ). A critical region particularly vulnerable to environmental insults during adolescence is the ventral hippocampus (vHip). However, the impact of severe stress on vHip redox states and their functional consequences, including behavioral and electrophysiological changes related to SCZ, are not entirely understood. STUDY DESIGN: After exposing adolescent animals to physical stress (postnatal day, PND31-40), we explored social and cognitive behaviors (PND47-49), the basal activity of pyramidal glutamate neurons, the number of parvalbumin (PV) interneurons, and the transcriptomic signature of the vHip (PND51). We also evaluated the impact of stress on the redox system, including mitochondrial respiratory function, reactive oxygen species (ROS) production, and glutathione (GSH) levels in the vHip and serum. STUDY RESULTS: Adolescent-stressed animals exhibited loss of sociability, cognitive impairment, and vHip excitatory/inhibitory (E/I) imbalance. Genome-wide transcriptional profiling unveiled the impact of stress on redox system- and synaptic-related genes. Stress impacted mitochondrial respiratory function and changes in ROS levels in the vHip. GSH and glutathione disulfide (GSSG) levels were elevated in the serum of stressed animals, while GSSG was also increased in the vHip and negatively correlated with sociability. Additionally, PV interneuron deficits in the vHip caused by adolescent stress were associated with oxidative stress. CONCLUSIONS: Our results highlight the negative impact of adolescent stress on vHip redox regulation and mitochondrial function, which are partially associated with E/I imbalance and behavioral abnormalities related to SCZ.

The brain's go-getter circuit: Anterior cingulate cortex to nucleus accumbens and its disruption by stress.

In this issue of Neuron, Fetcho, Parekh, et al.1 show that neurons in the anterior cingulate cortex (ACC) projecting to the nucleus accumbens (NAc) are essential for integrating reward and effort evaluation in mice, and that this circuit is sensitive to exposure to stress hormones.

Blunted Glucocorticoid Responsiveness to Stress Causes Behavioral and Biological Alterations That Lead to Posttraumatic Stress Disorder Vulnerability.

BACKGROUND: Understanding why only a subset of trauma-exposed individuals develop posttraumatic stress disorder is critical for advancing clinical strategies. A few behavioral (deficits in fear extinction) and biological (blunted glucocorticoid levels, small hippocampal size, and rapid-eye-movement sleep [REMS] disturbances) traits have been identified as potential vulnerability factors. However, whether and to what extent these traits are interrelated and whether one of them could causally engender the others are not known. METHODS: In a genetically selected rat model of reduced corticosterone responsiveness to stress, we explored posttraumatic stress disorder-related biobehavioral traits using ex vivo magnetic resonance imaging, cued fear conditioning, and polysomnographic recordings combined with in vivo photometric measurements. RESULTS: We showed that genetic selection for blunted glucocorticoid responsiveness led to a correlated multitrait response, including impaired fear extinction (observed in males but not in females), small hippocampal volume, and REMS disturbances, supporting their interrelatedness. Fear extinction deficits and concomitant disruptions in REMS could be normalized through postextinction corticosterone administration, causally implicating glucocorticoid deficiency in two core posttraumatic stress disorder-related risk factors and manifestations. Furthermore, reduced REMS was accompanied by higher norepinephrine levels in the hippocampal dentate gyrus that were also reversed by postextinction corticosterone treatment. CONCLUSIONS: Our results indicate a predominant role for glucocorticoid deficiency over the contribution of reduced hippocampal volume in engendering both REMS alterations and associated deficits in fear extinction consolidation, and they causally implicate blunted glucocorticoids in sustaining neurophysiological disturbances that lead to fear extinction deficits.

Transcriptomic analysis reveals mitochondrial pathways associated with distinct adolescent behavioral phenotypes and stress response.

Adolescent individuals exhibit great variability in cortical dynamics and behavioral outcomes. The developing adolescent brain is highly sensitive to social experiences and environmental insults, influencing how personality traits emerge. A distinct pattern of mitochondrial gene expression in the prefrontal cortex (PFC) during adolescence underscores the essential role of mitochondria in brain maturation and the development of mental illnesses. Mitochondrial features in certain brain regions account for behavioral differences in adulthood. However, it remains unclear whether distinct adolescent behavioral phenotypes and the behavioral consequences of early adolescent stress exposure in rats are accompanied by changes in PFC mitochondria-related genes and mitochondria respiratory chain capacity. We performed a behavioral characterization during late adolescence (postnatal day, PND 47-50), including naïve animals and a group exposed to stress from PND 31-40 (10 days of footshock and 3 restraint sessions) by z-normalized data from three behavioral domains: anxiety (light-dark box tests), sociability (social interaction test) and cognition (novel-object recognition test). Employing principal component analysis, we identified three clusters: naïve with higher-behavioral z-score (HBZ), naïve with lower-behavioral z-score (LBZ), and stressed animals. Genome-wide transcriptional profiling unveiled differences in the expression of mitochondria-related genes in both naïve LBZ and stressed animals compared to naïve HBZ. Genes encoding subunits of oxidative phosphorylation complexes were significantly down-regulated in both naïve LBZ and stressed animals and positively correlated with behavioral z-score of phenotypes. Our network topology analysis of mitochondria-associated genes found Ndufa10 and Cox6a1 genes as central identifiers for naïve LBZ and stressed animals, respectively. Through high-resolution respirometry analysis, we found that both naïve LBZ and stressed animals exhibited a reduced prefrontal phosphorylation capacity and redox dysregulation. Our findings identify an association between mitochondrial features and distinct adolescent behavioral phenotypes while also underscoring the detrimental functional consequences of adolescent stress on the PFC.

Brain mitochondrial diversity and network organization predict anxiety-like behavior in male mice.

The brain and behavior are under energetic constraints, limited by mitochondrial energy transformation capacity. However, the mitochondria-behavior relationship has not been systematically studied at a brain-wide scale. Here we examined the association between multiple features of mitochondrial respiratory chain capacity and stress-related behaviors in male mice with diverse behavioral phenotypes. Miniaturized assays of mitochondrial respiratory chain enzyme activities and mitochondrial DNA (mtDNA) content were deployed on 571 samples across 17 brain areas, defining specific patterns of mito-behavior associations. By applying multi-slice network analysis to our brain-wide mitochondrial dataset, we identified three large-scale networks of brain areas with shared mitochondrial signatures. A major network composed of cortico-striatal areas exhibited the strongest mitochondria-behavior correlations, accounting for up to 50% of animal-to-animal behavioral differences, suggesting that this mito-based network is functionally significant. The mito-based brain networks also overlapped with regional gene expression and structural connectivity, and exhibited distinct molecular mitochondrial phenotype signatures. This work provides convergent multimodal evidence anchored in enzyme activities, gene expression, and animal behavior that distinct, behaviorally-relevant mitochondrial phenotypes exist across the male mouse brain.

Mitofusin-2 in nucleus accumbens D2-MSNs regulates social dominance and neuronal function.

The nucleus accumbens (NAc) is a brain hub regulating motivated behaviors, including social competitiveness. Mitochondrial function in the NAc links anxiety with social competitiveness, and the mitochondrial fusion protein mitofusin 2 (Mfn2) in NAc neurons regulates anxiety-related behaviors. However, it remains unexplored whether accumbal Mfn2 levels also affect social behavior and whether Mfn2 actions in the emotional and social domain are driven by distinct cell types. Here, we found that subordinate-prone highly anxious rats show decreased accumbal Mfn2 levels and that Mfn2 overexpression promotes dominant behavior. In mice, selective Mfn2 downregulation in NAc dopamine D2 receptor-expressing medium spiny neurons (D2-MSNs) induced social subordination, accompanied by decreased accumbal mitochondrial functions and decreased excitability in D2-MSNs. Instead, D1-MSN-targeted Mfn2 downregulation affected competitive ability only transiently and likely because of an increase in anxiety-like behaviors. Our results assign dissociable cell-type specific roles to Mfn2 in the NAc in modulating social dominance and anxiety.

Powering the social brain: Mitochondria in social behaviour.

A central role of brain mitochondria in regulating and influencing social behaviour is emerging. In addition to its important roles as the "powerhouses" of the cell, mitochondria possess a plethora of cellular functions, such as regulating ion homeostasis, neurotransmitter levels, and lipid metabolism. Findings in the last decade are revealing an integral role for mitochondria in the regulation of behaviours, including those from the social domain. Here, we discuss recent evidence linking mitochondrial functions and dynamics to social behaviour and deficits, including examples in which social behaviours are modulated by stress in the context of mitochondrial changes, as well as potential therapeutic strategies and outstanding questions in the field.

Glucocorticoid-based pharmacotherapies preventing PTSD.

Posttraumatic stress disorder (PTSD) is a highly disabling psychiatric condition that may arise after exposure to acute and severe trauma. It is a highly prevalent mental disorder worldwide, and the current treatment options for these patients remain limited due to low effectiveness. The time window right after traumatic events provides clinicians with a unique opportunity for preventive interventions against potential deleterious alterations in brain function that lead to PTSD. Some studies pointed out that PTSD patients present an abnormal function of the hypothalamic-pituitary-adrenal axis that may contribute to a vulnerability toward PTSD. Moreover, glucocorticoids have arisen as a promising option for preventing the disorder's development when administered in the aftermath of trauma. The present work compiles the recent findings of glucocorticoid administration for the prevention of a PTSD phenotype, from human studies to animal models of PTSD. Overall, glucocorticoid-based therapies for preventing PTSD demonstrated moderate evidence in terms of efficacy in both clinical and preclinical studies. Although clinical studies point out that glucocorticoids may not be effective for all patients' subpopulations, those with adequate traits might greatly benefit from them. Preclinical studies provide precise insight into the mechanisms mediating this preventive effect, showing glucocorticoid-based prevention to reduce long-lasting behavioral and neurobiological abnormalities caused by traumatic stress. However, further research is needed to delineate the precise mechanisms and the extent to which these interventions can translate into lower PTSD rates and morbidity. This article is part of the Special Issue on 'Fear, Anxiety and PTSD'.

Epigenome-wide DNA methylation in externalizing behaviours: A review and combined analysis.

DNA methylation (DNAm) is one of the most frequently studied epigenetic mechanisms facilitating the interplay of genomic and environmental factors, which can contribute to externalizing behaviours and related psychiatric disorders. Previous epigenome-wide association studies (EWAS) for externalizing behaviours have been limited in sample size, and, therefore, candidate genes and biomarkers with robust evidence are still lacking. We 1) performed a systematic literature review of EWAS of attention-deficit/hyperactivity disorder (ADHD)- and aggression-related behaviours conducted in peripheral tissue and cord blood and 2) combined the most strongly associated DNAm sites observed in individual studies (p < 10-3) to identify candidate genes and biological systems for ADHD and aggressive behaviours. We observed enrichment for neuronal processes and neuronal cell marker genes for ADHD. Astrocyte and granulocytes cell markers among genes annotated to DNAm sites were relevant for both ADHD and aggression-related behaviours. Only 1 % of the most significant epigenetic findings for ADHD/ADHD symptoms were likely to be directly explained by genetic factors involved in ADHD. Finally, we discuss how the field would greatly benefit from larger sample sizes and harmonization of assessment instruments.

Glutathione in the nucleus accumbens regulates motivation to exert reward-incentivized effort.

Emerging evidence is implicating mitochondrial function and metabolism in the nucleus accumbens in motivated performance. However, the brain is vulnerable to excessive oxidative insults resulting from neurometabolic processes, and whether antioxidant levels in the nucleus accumbens contribute to motivated performance is not known. Here, we identify a critical role for glutathione (GSH), the most important endogenous antioxidant in the brain, in motivation. Using proton magnetic resonance spectroscopy at ultra-high field in both male humans and rodent populations, we establish that higher accumbal GSH levels are highly predictive of better, and particularly, steady performance over time in effort-related tasks. Causality was established in in vivo experiments in rats that, first, showed that downregulating GSH levels through micro-injections of the GSH synthesis inhibitor buthionine sulfoximine in the nucleus accumbens impaired effort-based reward-incentivized performance. In addition, systemic treatment with the GSH precursor N-acetyl-cysteine increased accumbal GSH levels in rats and led to improved performance, potentially mediated by a cell-type-specific shift in glutamatergic inputs to accumbal medium spiny neurons. Our data indicate a close association between accumbal GSH levels and an individual's capacity to exert reward-incentivized effort over time. They also suggest that improvement of accumbal antioxidant function may be a feasible approach to boost motivation.

Deletion of Crtc1 leads to hippocampal neuroenergetic impairments associated with depressive-like behavior.

Mood disorders (MD) are a major burden on society as their biology remains poorly understood, challenging both diagnosis and therapy. Among many observed biological dysfunctions, homeostatic dysregulation, such as metabolic syndrome (MeS), shows considerable comorbidity with MD. Recently, CREB-regulated transcription coactivator 1 (CRTC1), a regulator of brain metabolism, was proposed as a promising factor to understand this relationship. Searching for imaging biomarkers and associating them with pathophysiological mechanisms using preclinical models can provide significant insight into these complex psychiatric diseases and help the development of personalized healthcare. Here, we used neuroimaging technologies to show that deletion of Crtc1 in mice leads to an imaging fingerprint of hippocampal metabolic impairment related to depressive-like behavior. By identifying a deficiency in hippocampal glucose metabolism as the underlying molecular/physiological origin of the markers, we could assign an energy-boosting mood-stabilizing treatment, ebselen, which rescued behavior and neuroimaging markers. Finally, our results point toward the GABAergic system as a potential therapeutic target for behavioral dysfunctions related to metabolic disorders. This study provides new insights on Crtc1's and MeS's relationship to MD and establishes depression-related markers with clinical potential.

Exploring associations between diurnal cortisol, stress, coping and psychopathology in adolescents and young adults with 22q11.2 deletion syndrome.

Background: 22q11.2 deletion syndrome (22q11DS) is a neurogenetic condition associated to a high risk for psychiatric disorders, including psychosis. Individuals with 22q11DS are thought to experience increased levels of chronic stress, which could lead to alterations in hypothalamic-pituitary-adrenocortical (HPA)-axis functioning. In the current study, we investigated for the first time diurnal salivary cortisol profiles in adolescents and young adults with 22q11DS as well as their link with stress exposure, coping strategies and psychopathology, including psychotic symptoms. Methods: Salivary cortisol was collected from adolescents and young adults with 22q11DS (n = 30, age = 19.7) and matched healthy controls (HC; n = 36, age = 18.5) six times a day for two days. Exposure to stressful life events, including peer victimization, coping strategies and general psychopathology were assessed with questionnaires. Psychotic symptoms and psychiatric comorbidities were evaluated with clinical interviews. Results: We observed similar daily levels and diurnal profiles of salivary cortisol in adolescents and young adults with 22q11DS compared to HCs. However, participants with 22q11DS reported less frequent exposure to stress than HCs. In 22q11DS, we observed a significant association between the use of non-adaptive coping strategies and the severity of psychotics symptoms. Cortisol level was not associated to severity of psychotic symptoms, but elevated cortisol awakening response (CAR) was found in participants with 22q11DS with higher levels of general psychopathology. Conclusions: Our results do not support earlier propositions of altered HPA-axis functioning in 22q11DS but highlight the need to further investigate diurnal cortisol as an indicator of HPA-axis functioning and its link with (earlier) stress exposure and psychopathology in this population. Interventions should target the development of adaptive coping skills in preventing psychosis in 22q11DS.

Pre-pandemic Psychobiological Features Predict Impact of COVID-19 Confinement on Loneliness.

The coronavirus disease 2019 (COVID-19) pandemic led to various government-imposed limitations on social interaction and strict home confinement. Such involuntary social-distancing policies can exacerbate feelings of loneliness and alter emotional well-being. Dysregulation of the hypothalamic-pituitary-adrenocortical (HPA) axis is a potential mechanism for loneliness' deleterious health effects. In this study, we explored whether pre-pandemic diurnal cortisol output (AUC G ), a measure of HPA axis function, may predict the propensity to changes in loneliness during long-term COVID-19 home confinement and if extraversion would moderate this relationship. This association has been explored by analysing the impact of COVID-19 pandemic and strict home confinement on social and emotional loneliness in 45 Spanish young adults. Diurnal cortisol levels were measured from five saliva samples obtained across a day just before the pandemic, and data about participants' perceived loneliness, empathic state, extraversion, and prospective volunteering were obtained both before and during the confinement. Participants' social and family loneliness increased during long-term strict home confinement, while prospective volunteering tendencies and extraversion decreased. Importantly, after adjusting for relevant confounders, moderation analyses revealed that in young adults with high pre-pandemic extraversion, a higher AUC G predicted a larger increase in social loneliness during confinement, while in individuals with low extraversion, AUC G was negatively related to change in loneliness. Our findings highlight the utility of pre-pandemic diurnal cortisol output in predicting the social impact of COVID-19 home confinement, presenting this hormone as a potential biomarker for a priori identification of at-risk groups during public health crises.

Acute stress affects peripersonal space representation in cortisol stress responders.

Peripersonal space is the representation of the space near the body. It is implemented by a dedicated multisensory-motor network, whose purpose is to predict and plan interactions with the environment, and which can vary depending on environmental circumstances. Here, we investigated the effect on the PPS representation of an experimentally induced stress response and compared it to a control, non-stressful, manipulation. We assessed PPS representation in healthy humans, before and after a stressful manipulation, by quantifying visuotactile interactions as a function of the distance from the body, while monitoring salivary cortisol concentration. While PPS representation was not significantly different between the control and experimental group, a relation between cortisol response and changes in PPS emerged within the experimental group. Participants who showed a cortisol stress response presented enhanced visuotactile integration for stimuli close to the body and reduced for far stimuli. Conversely, individuals with a less pronounced cortisol response showed a reduced difference in visuotactile integration between the near and the far space. In our interpretation, physiological stress resulted in a freezing-like response, where multisensory-motor resources are allocated only to the area immediately surrounding the body.

Opposite effects of stress on effortful motivation in high and low anxiety are mediated by CRHR1 in the VTA.

Individuals frequently differ in their behavioral and cognitive responses to stress. However, whether motivation is differently affected by acute stress in different individuals remains to be established. By exploiting natural variation in trait anxiety in outbred Wistar rats, we show that acute stress facilitates effort-related motivation in low anxious animals, while dampening effort in high anxious ones. This model allowed us to address the mechanisms underlying acute stress-induced differences in motivated behavior. We show that CRHR1 expression levels in dopamine neurons of the ventral tegmental area (VTA)-a neuronal type implicated in the regulation of motivation-depend on animals' anxiety, and these differences in CRHR1 expression levels explain the divergent effects of stress on both effortful behavior and the functioning of mesolimbic DA neurons. These findings highlight CRHR1 in VTA DA neurons-whose levels vary with individuals' anxiety-as a switching mechanism determining whether acute stress facilitates or dampens motivation.