Title Identifying the mechanisms of action in mice lacking brain

Name and affiliation of supervisors Charité, Berlin MDC-Berlin Prof. Dr. Golo Kronenberg Prof. Dr. Michael Bader Supervisor Co-Supervisor Principle Investigator Principle Investigator [email protected] [email protected]

Prof. Rainer Hellweg Friederike Klempin, PhD Collaborator Natalia Alenina, PhD

Short description of the PhD project 1. State of the art and preliminary work Major depression is a serious mental disorder, which affects up to 15% of the population. Pharmacological manipulation of serotonin (5-HT) leads to clinical improvement associated with a delayed increase in hippocampal neurogenesis [1, 2]. “State of the art” therapy consists of selective serotonin inhibitors (SSRIs), however, not all patients respond to this treatment. Intriguingly, , which acts as a selective serotonin reuptake enhancer (SSRE) [3, 4] is also a potent antidepressant and has therefore been called a “hypothesis killer”. Here we propose that the lack of a clear mechanism for SSRI function significantly limits our ability to effectively manage depression. In this proposal we a) take advantage of our recent insights into the role of serotonin in activity-dependent neurogenesis, b) apply a powerful genetic model that lacks brain serotonin to test a variety of pro-, and antidepressive stimuli that will directly translate to the clinical situation, and c) explore the interplay of brain-derived neurotrophic factor (BDNF) and serotonin. At the center of the proposal is a powerful genetic tool. Tryptophan hydroxylase 2 deficient mice are selectively depleted in brain serotonin [5, 6] and exhibit increased aggression and a depression-like state [7]. This combination of symptoms is highly reminiscent of the clinical presentation of depressed patients with reduced central serotonin function (i.e., low 5-hydroxyindoleacetic acid [5-HIAA] levels in cerebrospinal fluid) [8]. Surprisingly, Tph2-/- mice have no measurable change in cell proliferation in the dentate gyrus at baseline despite serotonin’s established role as neuromodulator. However, we recently established that Tph2 is required for activity-induced neurogenesis [9]. We further pursue the question of how neurogenic factors collaborate with, and potentially compensate for the lack of each another. One aim of this proposal is to determine BDNF levels in wild type animals and Tph2-/- mice at baseline and following antidepressant treatment. Altered BDNF signaling could be relevant to mechanisms of serotonin therapy, and preliminary data achieved in the collaboration of both research teams from Charité and MDC already revealed changes in hippocampal BDNF protein levels in Tph2-/- mice following running. Our results indicate that the serotonin system crucially shapes the brain’s response to physical exercise. Here, we will test whether the serotonin system also modulates the effects of stress, SSRIs/tianeptine and of electroconvulsive therapy. Insights from these studies will add to our conceptual framework of risk factors and therapeutic targets for depression.

Figure 1. Lack of TPH2 prevents the pro-proliferative effect of running [9].

2. Work program Project during the 1st year – Aim1. Comparison of the physiological effects mediated by SSRI and SSRE treatment. This aim will establish the role of a novel downstream mediator of serotonin action, BDNF. We will specifically test whether the known citalopram (SSRI) and tianeptine (SSRE) differentially affect adult hippocampal neurogenesis in wild type animals (i.p. injections over 21 days) (1). Neurotrophic signaling might be involved in their action, and BDNF protein (ELISA), and mRNA (RT-PCR) levels will be determined (2) as well as changes in 5-HT, and 5-HIAA levels, and further monoamine and , e.g. dopamine and glutamate (HPLC) (3). Further analysis comprises mRNA expression of glucocorticoid , and SERT in depression-related brain regions (4). Project during the 2nd year – Aim2. Elucidating resilience in a chronic stress model and the response to SSRI treatment in life-long serotonin depletion. This aim will specifically address the effects of long-term 5-HT deficiency on novel adaptive mechanisms. We will utilize Tph2-/- mice to test whether the SSRI citalopram also acts in the absence of serotonin, and to investigate and compare the phenotypes of KO and wild type in response to chronic stress. The chronic stress paradigm is well established by the research team at Charité [10-12]. Experiments will comprise analysis of the HPA- system and the assessment of the above Aim1 (1-4). We will in addition perform a variety of behavioral tests to define the degree of psychological stress when serotonin is absent. Project during the 3rd year – Aim3. Test electroconvulsive therapy (ECT) in a model with serotonin depletion. ECT is a highly effective treatment for major depression. In this aim we want to test whether serotonin signaling is essential for the well-established effects of ECT, namely the induction of hippocampal neurogenesis and BDNF. We will study how Tph2-/- mice react to a series of 5 daily electroconvulsive seizure (ECS) sessions. Experimental readout will be again the determination of (1-4) of the above Aims.