Anat Milman

Rapid antidepressive-like activity of specific glycogen synthase kinase-3 inhibitor and its effect on β-catenin in mouse hippocampus

BACKGROUND Inhibition of glycogen synthase kinase-3 (GSK-3) is thought to be a key feature in the therapeutic mechanism of several mood stabilizers; however, the role of GSK-3 in depressive behavior has not been determined. In these studies, we evaluated the antidepressive effect of L803-mts, a novel GSK-3 peptide inhibitor, in an animal model of depression, the mouse forced swimming test (FST). METHODS Animals were intracerebroventricularly injected with L803-mts or with respective control peptide (cp) 1 hour, 3 hours, or 12 hours before their subjection to FST. RESULTS Animals administered L803-mts showed reduced duration of immobility at all three time points tested, as compared with cp-treated animals. Expression levels of beta-catenin, the endogenous substrate of GSK-3, increased in the hippocampus of L803-mts-treated animals by 20%-50%, as compared with cp-treated animals. CONCLUSIONS Our studies show, for the first time, that in-vivo inhibition of GSK-3 produces antidepressive-like behavior and suggest the potential of GSK-3 inhibitors as antidepressants.

Role of glycogen synthase kinase-3β in early depressive behavior induced by mild traumatic brain injury

Traumatic brain injury (TBI) is a triggering event for a set of pathophysiological changes and concomitant depressive behavior. Glycogen synthase kinase-3 (GSK-3) is a potent in vivo regulator of cell apoptosis and, in addition, is implicated in depressive behavior. In this study, we investigated the role of GSK-3 in the physiological model of mild TBI (mTBI) at both the cellular and behavior levels. mTBI resulted in increased phosphorylation of inhibitory site serine(9) of GSK-3beta, which coincided with increased serine(473) phosphorylation of its upstream kinase PKB and accumulation of its downstream target beta-catenin in the hippocampus. mTBI induced a depressive behavior which was evident as early as 24 h post-injury. Pretreatment with GSK-3 inhibitors, lithium, or L803-mts prevented mTBI-induced depression. We suggest that mTBI elicits a pro-survival cascade of PKB/GSK-3beta/beta-catenin as part of a rehabilitation program. Furthermore, the use of selective GSK-3 inhibitors may have therapeutic benefits in treatment conditions associated with brain injury.

Apoptotic and behavioral sequelae of mild brain trauma in mice

Mild traumatic brain injury (mTBI) is a not uncommon event in adolescents and young adults. Although it does not result in clear morphological brain defects, it is associated with long‐term cognitive, emotional, and behavioral problems. Herein, we characterized the biochemical and behavioral changes associated with experimental mTBI in mice that may act as either targets or surrogate markers for interventional therapy. Specifically, mTBI was induced by 30‐g and 50‐g weight drop, and at 8 and 72 hr thereafter markers of cellular apoptosis—caspase‐3, Bax, apoptosis‐inducing factor (AIF), and cytochrome‐c (Cyt‐c)—were quantified by Western blot analysis in hippocampus ipsilateral to the impact. Levels of amyloid‐β precursor protein (APP) were also measured, and specific behavioral tests—passive avoidance, open field, and forced swimming (Porsolt) paradigms—were undertaken to assess learning, emotionality, and emotional memory. In the absence of hemorrhage or infarcts, as assessed by triphenyltetrazolium chloride staining, procaspase‐3 and Bax levels were markedly altered following mTBI at both times. No cleaved caspase‐3 was detected, and levels of AIF and Cyt‐c, but not APP, were significantly changed at 72 hr. Mice subjected to mTBI were indistinguishable from controls by neurological examination at 1 and 24 hr, and by passive avoidance/open field at 72 hr, but could be differentiated in the forced swimming paradigm. In general, this model mimics the diffuse effects of mTBI on brain function associated with the human condition and highlights specific apoptotic proteins and a behavioral paradigm as potential markers for prospective interventional strategies.

Age-dependent differential expression of BACE splice variants in brain regions of tg2576 mice.

Plaques found in the brains of patients suffering from Alzheimers disease (AD) mainly consist of beta-amyloid (Abeta), which is produced by sequential cleaving of amyloid precursor protein (APP) by two proteolytic enzymes, beta- and gamma-secretases. Any change in the fine balance between these enzymes and their substrate may contribute to the etio-pathogenesis of AD. Indeed, the protein level and enzymatic activity of beta-secretase (BACE), but not its mRNA level, were found elevated in brain areas of AD patients who suffer a high load of Abeta plaque formation. Similarly, increased BACE activity but no mRNA change was observed in a transgenic mouse model of AD, tg2576, in which over expression of the Swedish mutated human APP leads to Abeta plaque formation and learning deficits. Based on the recent demonstration of four BACE splice variants with different enzymatic activity, the discrepancy between BACE activity and mRNA expression may be explained by the altered BACE alternative splicing. To test this hypothesis, we studied the expression of all BACE splice variants in different brain areas of tg2576 mice at age of 4 months and 1 year old. We found developmental and regional differences between wild-type and tg2576 mice. Our results indicate that over expression of APP in tg2576 mice leads to the altered alternative splicing of BACE and the increase of its enzymatically more active splice variant (I-501).

Behavioral effects of opioid subtypes compared with benzodiazepines in the staircase paradigm

Opioid abuse is frequently associated with the abuse of benzodiazepines (BZ). Despite the fact that benzodiazepines and opioids act at totally separate receptor sites and through different biochemical and pharmacological mechanisms, they obviously interact with each other at some level. The present study was designed to investigate the behavioral effects of agents with activity on the opioid system and with the benzodiazepine receptor complex in the staircase paradigm. The benzodiazepine agonist lorazepam was used as a reference drug and showed a benzodiazepine effect: it reduced the rearing activity in mice in a dose-dependent manner, at doses that did not suppress climbing. Dissociation between the rearing and climbing behavior was not obtained with the administration of the opioid benzodiazepine, tifluadome, although tifluadome was antagonized by the BZ antagonist, flumazenil and not by naloxone, an opioid antagonist. An administration of opioids that act on the different subtype receptors morphine, U50 and SNC80, induced a dose dependent decrease in both the rearing and climbing activity, meaning no behavioral dissociation. Naloxone was found to have an antagonistic effect on morphine and U50-488H (micro and kappa subtype receptors, respectively), but no antagonistic effect on the delta subtype receptor agonist, SNC80. The administration of nalorphine to mice had no effect on both the rearing and climbing, in the staircase paradigm. When combining naloxone with the mentioned nalorphine, results showed the same effect, which was achieved with all the other opiates. This data, once again, supports the validity of the mouse staircase test for the identification of anxiolytic agents with a GABA(A) receptor complex-mediated activity, and for the ability to differentiate between diverse effects of the different opioids.

Age of First Arrhythmic Event in Brugada Syndrome: Data From the SABRUS (Survey on Arrhythmic Events in Brugada Syndrome) in 678 Patients

Background Data on the age at first arrhythmic event (AE) in Brugada syndrome are from limited patient cohorts. The aim of this study is 2-fold: (1) to define the age at first AE in a large cohort of patients with Brugada syndrome, and (2) to assess the influence of the mode of AE documentation, sex, and ethnicity on the age at first AE. Methods and Results A survey of 23 centers from 10 Western and 4 Asian countries gathered data from 678 patients with Brugada syndrome (91.3% men) with first AE documented at time of aborted cardiac arrest (group A, n=426) or after prophylactic implantable cardioverter–defibrillator implantation (group B, n=252). The vast majority (94.2%) of the patients were 16 to 70 years old at the time of AE, whereas pediatric (<16 years) and elderly patients (>70 years) comprised 4.3% and 1.5%, respectively. Peak AE rate occurred between 38 and 48 years (mean, 41.9±14.8; range, 0.27–84 years). Group A patients were younger than in Group B by a mean of 6.7 years (46.1±13.2 versus 39.4±15.0 years; P<0.001). In adult patients (≥16 years), women experienced AE 6.5 years later than men (P=0.003). Whites and Asians exhibited their AE at the same median age (43 years). Conclusions SABRUS (Survey on Arrhythmic Events in Brugada Syndrome) presents the first analysis on the age distribution of AE in Brugada syndrome, suggesting 2 age cutoffs (16 and 70 years) that might be important for decision-making. It also allows gaining insights on the influence of mode of arrhythmia documentation, patient sex, and ethnic origin on the age at AE.

Gender Differences in Patients with Brugada Syndrome and Arrhythmic Events: Data from a Survey on Arrhythmic Events in 678 Patients

BACKGROUND There is limited information on gender differences in patients with Brugada syndrome (BrS) who experienced arrhythmic events (AEs). OBJECTIVE The purpose of this study was to compare clinical, electrocardiographic (ECG), electrophysiological, and genetic characteristics between males and females in patients with BrS with their first AE. METHODS The multicenter Survey on Arrhythmic Events in BRUgada Syndrome collected data on the first AE in 678 patients with BrS including 619 males (91.3%) and 59 females (8.7%) aged 0.27-84 years (mean age 42.5 ± 14.1 years) at the time of AE occurrence. RESULTS After excluding pediatric patients, it was found that females were older than males (49.5 ± 14.4 years vs 43 ± 12.7 years, respectively; P = .001). Higher proportions of females were observed in the pediatric and elderly populations. In Asians, the male to female ratio for AEs was ≈9-fold higher than that in White. Spontaneous type 1 BrS ECG was associated with an earlier onset of AEs in pediatric females. A similar prevalence (≈65%) of spontaneous type 1 BrS ECG was present in males and females above the age of 60 years. Females less frequently showed spontaneous type 1 BrS ECG (41% vs 69%; P < .001) or arrhythmia inducibility at electrophysiology study (36% vs 66%; P < .001). An SCN5A mutation was more frequently found in females (48% vs 28% in males; P = .007). CONCLUSION This study confirms that female patients with BrS are much rarer, display less type 1 Brugada ECG, and exhibit lower inducibility rates than do males. It shows for the first time that female patients with BrS with AE have higher SCN5A mutation rates as well as the relationship between gender vs age at the onset of AEs and ethnicity.

Fever-related arrhythmic events in the multicenter Survey on Arrhythmic Events in Brugada Syndrome

BACKGROUND The literature on fever-related arrhythmic events (AEs) in Brugada syndrome (BrS) is currently limited to few case reports and small series. OBJECTIVE The present study aimed to describe the characteristics of fever-related AE in a large cohort of patients with BrS. METHODS The Survey on Arrhythmic Events in Brugada Syndrome is a multicenter study on 678 patients with BrS with first AE documented at the time of aborted cardiac arrest (n = 426) or after prophylactic implantable cardioverter-defibrillator implantation (n = 252). RESULTS In 35 of 588 patients (6%) with available information, the AE occurred during a febrile illness. Most of the 35 patients were male (80%), Caucasian (83%), and proband (70%). The mean age at the time of AE was 29 ± 24 years (range 0.3-76 years). Most patients (80%) presented with aborted cardiac arrest and 6 (17%) with arrhythmic storm. Family history of sudden death, history of syncope, and spontaneous type 1 Brugada electrocardiogram were noted in 17%, 40%, and 71% of patients, respectively. Ventricular fibrillation was induced at electrophysiology study in 9 of 19 patients (47%). An SCN5A mutation was found in 14 of 28 patients (50%). The highest proportion of fever-related AE was observed in the pediatric population (age <16 years), with a disproportionally higher event rate in the very young (age 0-5 years) (65%). Males were involved in all age groups and females only in the pediatric and elderly groups. Fever-related AE affected 17 Caucasians aged <24 years, but no Asians aged <24 years. CONCLUSION The risk of fever-related AE in BrS markedly varies according to age group, sex, and ethnicity. Taking these factors into account could help the clinical management of patients with BrS with fever.