Yulia Golub

Reduced hippocampus volume in the mouse model of Posttraumatic Stress Disorder

Some, but not all studies in patients with posttraumatic stress disorder (PTSD), report reduced hippocampus (HPC) volume. In particular it is unclear, whether smaller hippocampal volume represents a susceptibility factor for PTSD rather than a consequence of the trauma. To gain insight into the relationship of brain morphology and trauma exposure, we investigated volumetric and molecular changes of the HPC in a mouse model of PTSD by means of in vivo Manganese Enhanced Magnetic Resonance Imaging (MEMRI) and ex vivo ultramicroscopic measurements. Exposure to a brief inescapable foot shock led to a volume reduction in both left HPC and right central amygdala two months later. This volume loss was mirrored by a down-regulation of growth-associated protein-43 (GAP43) in the HPC. Enriched housing decreased the intensity of trauma-associated contextual fear, independently of whether it was provided before or after the shock. Beyond that, enriched housing led to an increase in intracranial volume, including the lateral ventricles and the hippocampus, and to an up-regulation of GAP43 as revealed by MEMRI and Western blot analysis, thus partially compensating for trauma-related HPC volume loss and down-regulation of GAP43 expression. Together these data demonstrate that traumatic experience in mice causes a reduction in HPC and central amygdala volume possibly due to a shrinkage of axonal protrusions. Enriched housing might induce trophic changes, which may contribute to the amelioration of trauma-associated PTSD-like symptoms at behavioural, morphological and molecular levels.

Consequences of extinction training on associative and non-associative fear in a mouse model of Posttraumatic Stress Disorder (PTSD)

A common approach to the clinical treatment of Posttraumatic Stress Disorder (PTSD) has focused on the facilitation of fear extinction through cognitive behavioural therapy that involves both safe exposure to the trauma-related cues and subsequent changes in conditioned stimulus-unconditioned stimulus (CS-US) contingency expectations. PTSD symptoms can be tracked back to pathologically modified associative fear, hyperarousal and a time-dependent fear generalization. We have used a mouse model of PTSD that is based on a brief exposure to an inescapable foot shock in order to investigate the influence of early (starting 1 day after the shock) and late (starting 1 month after the shock) extinction training. Both early and late extinction training led to a long-lasting reduction of contextual and generalized fear, but only early extinction caused an amelioration of hyperarousal. Consequently, our results suggest early post-shock intervention as a successful strategy for reducing hyperarousal in the aftermath of a trauma.

aCaMKII Autophosphorylation Controls the Establishment of Alcohol Drinking Behavior.

The α-Ca2+/calmodulin-dependent protein kinase II (αCaMKII) is a crucial enzyme controlling plasticity in the brain. The autophosphorylation of αCaMKII works as a ‘molecular memory’ for a transient calcium activation, thereby accelerating learning. We investigated the role of αCaMKII autophosphorylation in the establishment of alcohol drinking as an addiction-related behavior in mice. We found that alcohol drinking was initially diminished in αCaMKII autophosphorylation-deficient αCaMKIIT286A mice, but could be established at wild-type level after repeated withdrawals. The locomotor activating effects of a low-dose alcohol (2 g/kg) were absent in αCaMKIIT286A mice, whereas the sedating effects of high-dose (3.5 g/kg) were preserved after acute and subchronic administration. The in vivo microdialysis revealed that αCaMKIIT286A mice showed no dopamine (DA) response in the nucleus accumbens to acute or subchronic alcohol administration, but enhanced serotonin (5-HT) responses in the prefrontal cortex. The attenuated DA response in αCaMKIIT286A mice was in line with altered c-Fos activation in the ventral tegmental area after acute and subchronic alcohol administration. In order to compare findings in mice with the human condition, we tested 23 single-nucleotide polymorphisms (SNPs) in the CAMK2A gene for their association with alcohol dependence in a population of 1333 male patients with severe alcohol dependence and 939 controls. We found seven significant associations between CAMK2A SNPs and alcohol dependence, one of which in an autophosphorylation-related area of the gene. Together, our data suggest αCaMKII autophosphorylation as a facilitating mechanism in the establishment of alcohol drinking behavior with changing the DA–5-HT balance as a putative mechanism.

AKT/GSK-3β/β-catenin signalling within hippocampus and amygdala reflects genetically determined differences in posttraumatic stress disorder like symptoms

Only a small percentage of individuals develop posttraumatic stress disorder (PTSD) in the aftermath of a trauma. It is still largely unknown to what extent gene-environment interactions contribute to the inter-individual differences in PTSD susceptibility and resilience and what cellular processes may underlie long-term maintenance of the disorder. Here we employed a mouse model of PTSD to unravel the contribution of genetic background and maternal influences on long-lasting changes in kinase and transcription factor activities in PTSD-susceptible C57BL/6NCrl (B6N) and resilient C57BL/6JOlaHsd (B6JOla) mice. Mice received an inescapable foot shock and were tested for activity changes in the AKT/GSK-3beta/beta-catenin-pathway in specific brain structures 42 days later. To control for prenatal and postnatal environmental (i.e. maternal) factors part of the experiments were performed with animals originating from within-strain and between-strain embryo transfers. In PTSD-susceptible B6N mice, long-term maintenance of contextual and sensitized fear was accompanied by (i) increased levels of phosphorylated AKT within the dorsal hippocampus and (ii) higher levels of phosphorylated AKT and GSK-3beta and increased beta-catenin levels within the basolateral amygdala. In animals originating from embryo transfers, levels of phosphorylated GSK-3beta and of beta-catenin were decreased in the dorsal hippocampus, but increased in the basolateral amygdala of shocked B6N mice compared to shocked B6JOla mice. This was independent of the genotype of the recipient mothers. At the behavioural level, these differences coincided with sustained sensitized and more pronounced contextual fear of B6N compared to B6JOla mice. Taken together our study identifies lasting changes in the AKT/GSK-3beta/beta-catenin cascade within the hippocampus and amygdala as molecular correlates of genetically determined differences in the severity of PTSD-like symptoms.

Long-Lasting Hippocampal Synaptic Protein Loss in a Mouse Model of Posttraumatic Stress Disorder

Despite intensive research efforts, the molecular pathogenesis of posttraumatic stress disorder (PTSD) and especially of the hippocampal volume loss found in the majority of patients suffering from this anxiety disease still remains elusive. We demonstrated before that trauma-induced hippocampal shrinkage can also be observed in mice exhibiting a PTSD-like syndrome. Aiming to decipher the molecular correlates of these trans-species posttraumatic hippocampal alterations, we compared the expression levels of a set of neurostructural marker proteins between traumatized and control mice at different time points after their subjection to either an electric footshock or mock treatment which was followed by stressful re-exposure in several experimental groups. To our knowledge, this is the first systematic in vivo study analyzing the long-term neuromolecular sequelae of acute traumatic stress combined with re-exposure. We show here that a PTSD-like syndrome in mice is accompanied by a long-lasting reduction of hippocampal synaptic proteins which interestingly correlates with the strength of the generalized and conditioned fear response but not with the intensity of hyperarousal symptoms. Furthermore, we demonstrate that treatment with the serotonin reuptake inhibitor (SSRI) fluoxetine is able to counteract both the PTSD-like syndrome and the posttraumatic synaptic protein loss. Taken together, this study demonstrates for the first time that a loss of hippocampal synaptic proteins is associated with a PTSD-like syndrome in mice. Further studies will have to reveal whether these findings are transferable to PTSD patients.

αCaMKII controls the establishment of cocaine's reinforcing effects in mice and humans

Although addiction develops in a considerable number of regular cocaine users, molecular risk factors for cocaine dependence are still unknown. It was proposed that establishing drug use and memory formation might share molecular and anatomical pathways. Alpha-Ca2+/calmodulin-dependent protein kinase-II (αCaMKII) is a key mediator of learning and memory also involved in drug-related plasticity. The autophosphorylation of αCaMKII was shown to accelerate learning. Thus, we investigated the role of αCaMKII autophosphorylation in the time course of establishing cocaine use-related behavior in mice. We found that αCaMKII autophosphorylation-deficient αCaMKIIT286A mice show delayed establishment of conditioned place preference, but no changes in acute behavioral activation, sensitization or conditioned hyperlocomotion to cocaine (20 mg kg−1, intraperitoneal). In vivo microdialysis revealed that αCaMKIIT286A mice have blunted dopamine (DA) and blocked serotonin (5-HT) responses in the nucleus accumbens (NAcc) and prefrontal cortex after acute cocaine administration (20 mg kg−1, intraperitoneal), whereas noradrenaline responses were preserved. Under cocaine, the attenuated DA and 5-HT activation in αCaMKIIT286A mice was followed by impaired c-Fos activation in the NAcc. To translate the rodent findings to human conditions, several CAMK2A gene polymorphisms were tested regarding their risk for a fast establishment of cocaine dependence in two independent samples of regular cocaine users from Brazil (n=688) and Switzerland (n=141). A meta-analysis across both samples confirmed that CAMK2A rs3776823 TT-allele carriers display a faster transition to severe cocaine use than C-allele carriers. Together, these data suggest that αCaMKII controls the speed for the establishment of cocaine’s reinforcing effects.

Inverse effects of lipopolysaccharides on anxiety in pregnant mice and their offspring

This study aimed to evaluate the effects of the bacterial lipopolysaccharide (LPS) exposure during early pregnancy on anxiety-related behaviour of both pregnant female mice and their male offspring. Pregnant NMRI mice were treated with subcutaneous injections of LPS (30, 60, 120, 240 and 480 μg/kg) on the tenth gestational day of pregnancy. Pro-inflammatory cytokines, such as TNF-α, IL-1β, IL-6 and corticosterone levels, were measured in maternal serum 1.5h following the LPS injections. Baseline anxiety levels of pregnant mice (1.5h after LPS administration) and their male offspring (at postnatal days 60-70) were investigated with the elevated plus maze (EPM) test. In addition, anxiety levels in the offspring were measured after 2h restraint stress or TNF-α (10 μg/kg) administration. Our results demonstrate that LPS administration induces anxiety-like behaviour and a significant increase in cytokines and corticosterone levels in maternal serum. However, in male offspring, prenatal LPS administration has no significant effects on serum cytokines and corticosterone secretion with an exception of the lowest LPS dose that slightly reduced corticosterone levels. Interestingly, prenatal LPS treatment seemed to decrease the baseline anxiety levels, while pretreatment with restraint stress or TNF-α abolished this anxiolytic effects. In summary, our results suggest that prenatal exposure to LPS during early pregnancy may result in reduced baseline anxiety in adult male offspring.

αCaMKII autophosphorylation mediates neuronal activation in the hippocampal dentate gyrus after alcohol and cocaine in mice

Psychoactive drug-induced cellular activation is a key mechanism to promote neuronal plasticity and addiction. Alpha Ca(2+)/calmodulin-dependent protein kinase II (αCaMKII) and its autophosphorylation play a key role in the development of drug use associated behaviours. It has been suggested that αCaMKII autophosphorylation is necessary for drug-induced neuronal activation in the mesolimbic system. Here, we show an alcohol- and cocaine-induced increase in c-fos expression in the hippocampal dentate gyrus, which is absent in αCaMKII(T286A) autophosphorylation deficient mice. These findings may suggest a role in hippocampal αCaMKII autophosphorylation in the acute neuroplastic effects of alcohol and cocaine.

Activation of GABAA receptors in the medial prefrontal cortex produces an anxiolytic-like response.

Objectives There has been increasing evidence that the γ-aminobutyric acid (GABA)ergic system is involved in the neurobiology of anxiety. The present study aimed to investigate the role of GABAergic systems in the modulation of anxiety in the medial prefrontal cortex (mPFC) of rats using the elevated plus maze test. Methods Rats were anaesthetised with a mixture of ketamine and xylazine, and then special cannulae were inserted stereotaxically into the mPFC. After 5–7 days of recovery, the effects of intra-mPFC administration of GABAergic agents were studied. Results Bilateral injection of the GABAA receptor agonist muscimol (0.25, 0.5 and 1 μg/rat) produces an anxiolytic-like effect, shown by significant increases in the percentage of open-arm time (%OAT) and percentage of open-arm entries (%OAE). Intra-mPFC administration of the GABAA receptor antagonist bicuculline (0.25, 0.5 and 1 μg/rat) produces significant anxiogenic-like behaviour. However, intra-mPFC injection of the GABAB receptor agonist baclofen (0.05, 0.1 and 0.2 μg/rat) and the GABAB receptor antagonist CGP35348 (5, 10 and 15 μg/rat) did not alter %OAT and %OAE significantly. Conclusion The results of the present study demonstrate that the GABAergic system of the mPFC modulates anxiety-related behaviours of rats through GABAA receptors.

Corrigendum: ACaMKII autophosphorylation controls the establishment of alcohol drinking behavior (Neuropsychopharmacology (2013) 38 (2735))

Correction to: Neuropsychopharmacology (2013) 38, 1636–1647; doi:10.1038/npp.2013.60 In this article, the following collaborators of The GESGA Consortium are being added to the byline: Jens Treutlein, Sven Cichon, Monika Ridinger, Manuel Mattheisen, Stefan Herms, Norbert Wodarz, Peter Zill, Wolfgang Maier, Rainald Mössner, Wolfgang Gaebel, Norbert Dahmen, Norbert Scherbaum, Christine Schmäl, Michael Steffens, Susanne Lucae, Marcus Ising, Bertram Müller-Myhsok, Markus M Nöthen, Karl Mann, and Marcella Rietschel Department of Genetic Epidemiology in Psychiatry, Central Institute of Mental Health, Medical Faculty Mannheim/Heidelberg University, Heidelberg, Germany; Department of Genomics, Life and Brain Center, University of Bonn, Bonn, Germany; Institute of Human Genetics, University of Bonn, Bonn, Germany; Institute of Neuroscience and Medicine (INM-1), Research Center Juelich, Juelich, Germany; Division of Medical Genetics, Department of Biomedicine, University Hospital Basel, University of Basel, Basel, Germany; Department of Psychiatry, University Medical Center Regensburg, University of Regensburg, Regensburg, Germany; Institute for Medical Biometry, Informatics, and Epidemiology, University of Bonn, Bonn, Germany; Department of Biostatistics, Harvard School of Public Health, Boston, MA, USA; Department of Psychiatry, University of Munich, Munich, Germany; Department of Psychiatry, University of Bonn, Bonn, Germany; Department of Psychiatry and Psychotherapy, University of Düsseldorf, Düsseldorf, Germany; Department of Psychiatry, University of Mainz, Mainz, Germany; Addiction Research Group at the Department of Psychiatry and Psychotherapy, LVR Hospital Essen, University of Duisburg-Essen, Essen, Germany; Department of Psychiatric Pharmacogenetics, Max-Planck-Institute of Psychiatry, Munich, Germany; Department of Molecular Psychology, Max-Planck-Institute of Psychiatry, Munich, Germany; Department of Statistical Genetics, Max-PlanckInstitute of Psychiatry, Munich, Germany; German Center for Neurodegenerative Diseases (DZNE), Bonn, Germany; Department of Addictive Behavior and Addiction Medicine, Central Institute of Mental Health, Medical Faculty Mannheim/Heidelberg University, Heidelberg, Germany.