SummaryIn developing neurons, phosphoinositide 3-kinases (PI3Ks) control axon growth and branching by positively regulating PI3K/PI(3,4,5)P3, but how neurons are able to generate sufficient PI(3,4,5)P3 in the presence of high levels of the antagonizing phosphatase PTEN is difficult to reconcile. We find that normal axon morphogenesis involves homeostasis of elongation and branch growth controlled by accumulation of PI(3,4,5)P3 through PTEN inhibition. We identify a plasma membrane-localized protein-protein interaction of PTEN with plasticity-related gene 2 (PRG2). PRG2 stabilizes membrane PI(3,4,5)P3 by inhibiting PTEN and localizes in nanoclusters along axon membranes when neurons initiate their complex branching behavior. We demonstrate that PRG2 is both sufficient and necessary to account for the ability of neurons to generate axon filopodia and branches in dependence on PI3K/PI(3,4,5)P3 and PTEN. Our data indicate that PRG2 is part of a neuronal growth program that induces collateral branch growth in axons by conferring local inhibition of PTEN.

Drebrin (DBN) regulates cytoskeletal functions during neuronal development, and is thought to contribute to structural and functional synaptic changes associated with aging and Alzheimer’s disease. Here we show that DBN coordinates stress signalling with cytoskeletal dynamics, via a mechanism involving kinase ataxia-telangiectasia mutated (ATM). An excess of reactive oxygen species (ROS) stimulates ATM-dependent phosphorylation of DBN at serine-647, which enhances protein stability and accounts for improved stress resilience in dendritic spines. We generated a humanized DBN Caenorhabditis elegans model and show that a phospho-DBN mutant disrupts the protective ATM effect on lifespan under sustained oxidative stress. Our data indicate a master regulatory function of ATM-DBN in integrating cytosolic stress-induced signalling with the dynamics of actin remodelling to provide protection from synapse dysfunction and ROS-triggered reduced lifespan. They further suggest that DBN protein abundance governs actin filament stability to contribute to the consequences of oxidative stress in physiological and pathological conditions.

Background: Individual vulnerability to stress manifests in the interaction of innate properties and environment. There is a growing interest in the individual variability in vulnerability to stress and how it contributes to the development of psychiatric disorders. Intake of palatable substances is often measured in animal models. We have previously demonstrated that the consumption of sucrose solution is a stable trait in rats. Aims: The present study aimed to compare the sensitivity of rats with high vs low liquid sucrose consumption to chronic variable stress and the stress effect on behavioural sensitization to amphetamine. Methods: Male Wistar rats were subjected to a chronic stress regimen and subsequent repeated treatment with amphetamine (1 mg/kg, intraperitoneally). Fifty-kHz ultrasonic vocalizations, locomotor activity and stereotypies were measured. Results: In no-stress baseline conditions, the behavioural response to acute amphetamine was similar in rats with high vs low sucrose consumption. Prior chronic stress potentiated the effect of amphetamine only in rats with high sucrose consumption. Behavioural sensitization to repeated administration of amphetamine was observed in non-stressed rats with lower sucrose preference, but not in the respective stressed group that had increased monoamine turnover in the nucleus accumbens. In contrast, in rats with high sucrose preference the amphetamine sensitization effect was prevalent in stressed rats, but not in non-stressed animals. Interpretation: Chronic stress can change the psychostimulant effect but this depends on the inherent reward sensitivity of the animal. Trait-wise, sucrose intake reflects vulnerability to chronic stress and may interact with the development of addiction.

Plasma membrane phospholipid phosphatase-related proteins as pleiotropic regulators of neuron growth and excitability.

Western blotting is a standard laboratory method used to detect proteins and assess their expression levels. Unfortunately, poor western blot image display practices and a lack of detailed methods reporting can limit a reader’s ability to evaluate or reproduce western blot results. While several groups have studied the prevalence of image manipulation or provided recommendations for improving western blotting, data on the prevalence of common publication practices are scarce. We systematically examined 551 articles published in the top 25% of journals in neurosciences (n=151) and cell biology (n=400) that contained western blot images, focusing on practices that may omit important information. Our data show that most published western blots are cropped and blot source data are not made available to readers in the supplement. Publishing blots with visible molecular weight markers is rare, and many blots additionally lack molecular weight labels. Western blot methods sections often lack information on the amount of protein loaded on the gel, blocking steps and antibody labeling protocol. Important antibody identifiers like source, catalog number or RRID were omitted frequently for primary antibodies, and regularly for secondary antibodies. We present detailed descriptions and visual examples to help scientists, peer reviewers and editors to publish more informative western blot figures and methods. Additional resources include a toolbox to help scientists produce more reproducible western blot data, teaching slides in English and Spanish and an antibody reporting template.