Metabolic Chronopathology in Depression: Functional Astrocyte-Neuron Interactions in the Cellular Model of Glucocorticoid Resistance

Description

Major depressive disorder (MDD) is a leading cause of global disease burden, potentially lethal, due to the high risk of suicide. Current therapies of depression are not optimal, therapies are long-lasting and often ineffective. These shortcomings are mostly due to insufficient knowledge of molecular mechanisms of the disease. To improve the situation, several global initiatives exist to resolve biological basics of distinct symptoms of depression at the cellular and molecular level. These joined efforts provided evidence that glial cells, significantly contribute to pathophysiology of depression. In this project, we focus on one subtype of glia, namely astrocytes. These cells support neurons in several ways, e.g. they regulate levels of main excitatory neurotransmitter, glutamate, and they link this process to brain energy metabolism. Importantly, it has been reported that both, glutamate and energy homeostasis, are defective in the brain of patients suffering from depression, pointing to astrocytes as the possible cellular site of these disease. Strikingly, beneficial effects of recently developed antidepressant, ketamine, encompass restoration of glutamatergic synapse plasticity and normalization of glucose metabolism, suggesting that astrocytes may be a target of the drug. In this project, we aim to understand metabolic dysfunction of astrocytes in the context of depression to identify novel and more efficient therapeutic strategies.

Summary of project results

Major depressive disorder (MDD) is a chronic, recurrent and heterogeneous disorder, currently affecting >300M people world-wide. Majority of current MDD therapies not optimal: they are prescribed largely on a trial-and-error basis, therapies are long-lasting, and approx. 33% of patients fail to achieve full remission. The complexity of MDD makes delineation of its pathophysiology extremely troublesome. It is now widely accepted that the susceptibility or resilience to develop MDD stems from both genetic predispositions which alter physiological stress response, and this maladaptation leads to synaptic deficits in brain regions controlling behavior. To decipher the exact mechanisms of the dysfunction, several global initiatives highlighted the need of investigating individual biological phenotypes related to psychiatric disorders which can be resolved at the level of defined circuits, cell types, pathways or genes. Crucially, these units can be studied in animal and cellular models recapitulating biological phenomena related to MDD.

The project MiChroFan “Metabolic chronopathology in depression: functional astrocyte-neuron interactions in cellular model of glucocorticoid resistance.” focused on establishing a new cellular model for investigating the interplay between genetic risk factors and one of the most frequent physiological symptoms in depression, i.e. dysfunctional circadian clock. In particular, since

a) disrupted circadian rhythmicity and impaired metabolism are physiological symptoms of depression;

b) biological effects of stress are mediated by glucocorticoids;

c) glucocorticoids act as a molecular effector of a circadian clock; and

d) we previously discovered that astrocytes mediate glucocorticoid signaling in the brain;

in MiChroFan, we hypothesized that aberrant GR signalling in astrocytes contributes to dysregulation of circadian rhythmicity.

We therefore set out to establish models enabling the investigation of molecular components of the GR pathway in neural cells and its role in the regulation of circadian rhythmicity and metabolic performance. In particular, we investigated how genetic predisposition exerted by a key regulator of the glucocorticoid receptor pathway, a protein called FKBP51, impacts the GR-induced changes in astrocytes.

Transcriptomic profiling of astrocytes or neurons generated from human induced pluripotent stem cells (hiPSCs) derived from individuals carrying distinct variants of the FKBP5 gene revealed variant-dependent differences. Interestingly, the molecular profile overlapped with pathways differentiating mouse strains carrying the human version of the FKBP5 gene with corresponding ‘high’ or ‘low’ risk FKBP5 variants. In particular, genes engaged in circadian entrainment constituted a shared subset between mouse and human carriers of distinct FKBP5 variants. This discovery suggests a novel function of FKBP51 protein in shaping cell-specific GR control of circadian rhythmicity in the brain. These results were published in a leading journal in the field, Molecular Psychiatry [1].

To monitor the GR-induced oscillations of clock genes and metabolites in cellular models we opted for direct monitoring of fluorescent indicators. To visualize those changes in a real time, we delivered to astrocytes modified viral vectors expressing the fluorescent protein (mVenus) under the control of core clock gene promoters (either Cry1 or Bmal1), an approach which previously was successfully used. We monitored fluorescent changes with an automated imaging system, consisting of the fluorescent microscope, equipped with an incubation chamber for long-term cell culture. We observed reliable oscillations of both reporters with a phase of approx. 26h, with Bmal1 reporter showing higher amplitude than Cry1. We also obtained a pilot data on GR-induced clock oscillations from hIPSC-derived astrocytes, a system to be used in future applications.

After successful establishing of a robust monitoring system, we investigated whether FKBP51 is required for GR-induced oscillation of clock genes. To this end, we used primary astrocyte cultures from WT mice, as well as from mice lacking both copies of Fkbp5 (knockout, KO) gene. We observed reliable induction of oscillations of our reporters, however, the 2nd and 3rd circadian peak of Cry1-mVenus were diminished in Fkbp5 KO astrocytes. This data revealed for a first time that a genetic risk factor of depression is required for GR control of clock gene oscillations in astrocytes.

In summary, we discovered a new mechanism through which a genetic risk factor of depression, FKBP51, affects glucocorticoid regulation of circadian rhythmicity in astrocytes. Future studies will identify pharmacological means of counteracting this deficit.  

Scientific deliverables:

1. High-impact factor, open-access publication summarizing a discovery of a moderating role of a genetic risk factor for depression in neuro-glia interactions.
2. Establishing cutting-edge tools enabling long-term top notch research in the host institution.
3. Development of a new model to investigate biological phenomenon related to psychiatric disorders: chronopathology.
4. Several oral and poster presentations in international conferences.
5. A review article on the contribution of glucocorticoid signalling in astrocytes to biological phenotypes associated with depression.

Socioeconomic deliverables

1. Sustainable high performance and increased competitiveness of a new research group in Łukasiewicz – PORT Polish Center for Technology Development, Wrocław, Poland.
2. Increased potential of the new research group ‘Biology of Astrocytes’ led by Dr Michał Ślęzak, as manifested by several grants obtained in international collaborations and a major HE grant for hosting institute as a coordinator of an international consortium engaging EU leaders of neuropsychiatric research.
3. Entering a PhD program and winning a Ministry of Science and Higher Education fellowship for Industry PhD program by Ms Tansu Gover, young investigator in the project
4. Enhanced internationalization of the research arena, manifested directly (facilitation of returning PI accommodation, attracting 1 PhD student from abroad) and indirectly (facilitation of attracting 2 postdoctoral researchers from abroad).
5. Improved technological readiness of a newly emerged center for applied biomedical research in Poland.
6. Multiplier effect through dissemination of data in international conferences, as well as active participation in cross-sectorial events (e.g. match-making), to ensure increased visibility of the research group and the institution.