Study On The Relationship Between Gap Junction Dysfunction In The Prefrontal Cortex And Depressive-like Behaviors In Rats

Major depressive disorder (MDD) is a common and disabling illness affecting a rising percentage of the world's population. Despite intensive research on the pathophysiology of depression, the neurobiological substrates underlying the disorder remain poorly characterized. Growing evidence in clinical and preclinical studies has implicated astroglial anomalies in the pathophysiology of MDD. Reduced number and altered morphology of astrocytes in several brain regions, in particular the prefrontal cortex (PFC) were described in many studies, whereas impaired cortical astroglial function was rarely reported. Gap junctional intercellular communication (GJIC) is a main determinant of astrocytic function. Alterations of GJIC in astrocytes have been reported in diverse pathological situations, such as epilepsy, ischemia, Alzheimer's disease, Parkinson's disease and Huntington's disease. However, whether or not altered astrocyte GJIC is involved in the development of major depression remains unanswered.In this study, we sought to address the above issue in the following two aspects: changes in the function of astrocyte gap junction occurring in the rat PFC after chronic unpredictable stress (CUS), a well-documented animal model of depression; the effects of pharmacological gap junction blockade in the prelimbic cortex (PLC) on depressive-like behaviors.First of all, we characterized the effects of CUS and chronic treatment with typical antidepressants including fluoxetine, a selective serotonin reuptake inhibitor (SSRI) and duloxetine, a serotonin and noradrenaline reuptake inhibitor (SNRI), on sucrose preference test (SPT) and novelty suppressed feeding test (NSFT).Secondly, in the above experimental system, dye transfer assay was applied to determine whether astrocytic gap junctions in the PFC are functional. Transmission electron microscopy (TEM) was applied to examine the detailed ultrastructure of gap junctions. Western blotting (WB), RT-PCR and immunohistochemistry were used to detect the expression of connexin43(Cx43), a major component of astrocyte gap junction.Thirdly, considering that CUS model is characterized by abnormalities in the hypothalamic-pituitary-adrenal (HPA) axis activity such as elevated corticosterone (CORT) concentrations, which may support the validity of the CUS-induced rat model of psychotic depression, we analyzed the effects of mifepristone, proved to lead to the rapid amelioration of psychotic depression as glucocorticoid receptor (GR) antagonist, on CUS-induced depressive-like behaviors and alterations of astrocyte gap junction using methods similar to the above.Then, we infused three doses of gap junction blocker carbenoxolone (CBX) into the PLC and measured sucrose preference known to assess anhedonia as well as the latency to feed in a novel environment, an indication of anxiety levels. We also examined the influence of CBX on the density of astrocytes visualized by glial fibrillary acidic protein (GFAP) immunoreactivity, and investigated the effect on Cx43protein levels using WB analysis.Finally, we infused more specific gap junction blockers Cx43mimetic peptides Gap27and Gap26into the PLC and measured sucrose preference known to assess anhedonia, a core symptom of depression.The results showed that animals exposed to CUS and showing behavioral deficits in SPT and NSFT exhibited significant decreases in diffusion of gap junction channel-permeable dye and abnormal gap junctional ultrastructure, as well as reductions in Cx43protein and mRNA levels and Cx43puncta density in the PFC. The behavioral and cellular alterations induced by CUS were reversed or blocked by chronic treatment with the typical antidepressant fluoxetine or duloxetine or by subchronic treatment with the GR antagonist mifepristone. The results also demonstrate that CBX infusions induced anhedonia in SPT, and anxiety in NSFT, and CBX infusions do not decrease the density of astrocytes in the PFC, nor provoke a reduction of the length and complexity of the processes of GFAP-positive cells, although there were signs of gap junction impairment, including the obvious decreases of Cx43protein level. In accordance with the dose-dependent alterations of Cx43protein level, the effects of CBX on SPT and NSFT are also dose-dependent. Furthermore, Cx43mimetic peptides Gap27and Gap26infusions also induced anhedonia, a core symptom of depression.In summary, animals exposed to CUS exhibited behavioral deficits in tests measuring anhedonia and anxiety, as well as functional and ultrastructural deficits of astrocytic gap junction and reductions in Cx43expression and Cx43puncta density in the PFC. The cellular and behavioral alterations induced by CUS were reversed and/or blocked by treatment with typical antidepressants or mifepristone, indicating the mechanism of their antidepressant action may involve the amelioration of gap junction dysfunction and the cellular changes may be related to GR activation. Pharmacological blockade of gap junction in the PLC with CBX or Cx43mimetic peptides produced depressive-like behaviors without glial ablation. Together, this study supports the hypothesis that gap junction dysfunction contributes to the pathophysiology of depression.