Effect And Mechanism Of Cognitive Impairment Associated With Higher DAPK1 Expression In Major Depressive Disorder

Depression is the most common psychiatric disorder plaguing humans since the21 st century.It affects more than 350 million people globally,and the World Health Organization predicts that it will become the first major disease in the world by 2030.Hence,early diagnosis and effective treatment to improve its symptoms are urgently needed.Current studies of cognitive dysfunction in depression focus mainly on major depressive disorder(MDD),a multi-factorial mood disorder in which patients often experience symptoms such as slowness of thinking,depressed mood,and reduced learning and memory ability.High prevalence,long disease duration,difficult treatment,and easy recurrence are the main features of MDD.Currently,there are many studies on cognitive dysfunction in MDD.While most of them focus on memory and executive functions,the specific mechanism of cognitive dysfunction is unclear.Clinical investigations have reported a close correlation between Alzheimer’s disease(AD)and depression,which is a high-risk predisposing factor and antecedent clinical manifestation of AD.Other studies also found that they share the same comorbidity basis,including increased neuroinflammation,decreased number of central synapses,and reduced levels of brain-derived neurotrophic factor and monoamine neurotransmitters.Cognitive dysfunction is the most obvious symptom of AD,which is mainly characterized by reduced spatial cognition,learning,and memory function,accompanied by some psychiatric symptoms.Around 20–50% of AD patients present with depressive symptoms,and antidepressants can alleviate two major pathological features of AD,namely deposition of amyloid β and hyperphosphorylation of tau protein,thereby relieving the symptoms of AD.Death-associated protein kinase 1(DAPK1)is a calcium/calmodulin-regulated serine/ threonine kinase that plays an important role in tau phosphorylation in AD.Our previous experiments found that DAPK1 was increased in the hippocampus of a mouse model of MDD.However,whether changes in DAPK1 expression are related to cognitive dysfunction and contribute to the improvement of cognitive function in patients with MDD remains unclear.In this study,C57BL/6 male mice were used to establish a mouse model of major depression through single cage feeding and chronic unpredictable stress.The depression indices,learning and memory functions,and spatial recognition ability of mice in each group were elevated by behavioral tests.Changes in the expression of DAPK1 and tau-related proteins in the hippocampus of mice were detected using western blot and immunohistochemistry.Furthermore,we investigated the effects of learning and memory training and intracerebral stereotaxic injection of a DAPK1 inhibitor on depressive behavior and cognitive dysfunction in MDD mice,thereby clarifying the effect and mechanism of DAPK1 on cognitive dysfunction in MDD and provide a theoretical basis for the clinical treatment of MDD.Methods1.The MDD mouse model was established using single-cage housing plus chronic unpredictable stress.Depressive behavior was evaluated using the sucrose preference,suspended tail,and forced swimming tests.Learning and memory function and spatial discrimination ability of mice were assessed using the Morris water maze and Y maze.Expression of DAPK1 and tau-related proteins in the hippocampus was quantified using western blotting.The in-situ analysis of DAPK1 and tau-related protein in the hippocampus was performed using immunohistochemical staining.Immuno-fluorescence staining was used to observe the colocalization of DAPK1 and Tau5 in the hippocampus of MDD mice.2.After the MDD mice underwent learning and memory training,the Morris water maze test was used to evaluate spatial learning and memory abilities.The Y-maze was used to appraise working memory.Western blotting was performed to estimate the expression of DAPK1 and phosphorylated tau protein in the hippocampus.3.DAPK1 inhibitor was administered through stereotaxic injection into the hippocampal CA3 region of MDD mice.Changes in depressive behavior and cognitive function were detected using behavioral tests.The expression of DAPK1 and tau-related proteins in the hippocampus was evaluated using western blotting.Immunofluorescence staining was used to observe the colocalization of DAPK1 and Tau5 in the hippocampus.The in-situ distribution of DAPK1 and tau proteins was observed by immunohistochemical staining.Results1.After the establishment of the MDD mouse model,the Y-maze test showed that the correct rate of spontaneous alternation was significantly decreased in the MDD group,indicating impaired working memory.The Morris water maze task showed that the average escape latency time of MDD mice was prolonged on days 4and 5,and the percentage of time that MDD mice crossed the platform in the target quadrant was decreased,demonstrating damaged spatial memory function.Western blot analysis demonstrated that the expression of DAPK1,Tau5,p T231,p S262,and p S396 was increased in the hippocampus of MDD mice.Immunohistochemistry results indicated that the expression of DAPK1 in hippocampal CA3 was elevated,and the distribution appeared unconventionally in MDD mice.Normally,DAPK1 is distributed in the neurites of neurons,with little cell membrane and cytoplasmic distribution,but a significant increase in the cytoplasmic distribution of DAPK1 in addition to the neurites in MDD mice was observed here.Immunofluorescence results showed increased cytoplasmic colocalization of DAPK1 and Tau5 in the hippocampal CA3 region of MDD mice.2.After learning and memory training in MDD mice,the spatial learning and memory abilities of MDD mice were significantly improved in the Morris water maze task.The Y-maze test showed an increased rate of correct spontaneous alternation,indicating enhanced working memory.Western blot results showed that the phosphorylation of tau protein at the Thr231 and Ser262 sites was significantly decreased.3.After injection of DAPK1 inhibitor in the hippocampal CA3 region of MDD mice,behavioral tests indicated that the working memory and spatial learning and memory abilities of MDD mice were significantly improved.Western blot results showed that tau protein phosphorylation was significantly decreased at the Thr31 and Ser262 sites.Meanwhile,the cytosolic colocalization of DAPK1 and Tau5 was decreased in the hippocampal CA3 region of MDD mice by immunofluorescence colocalization assay.Immunohistochemistry results showed that Thr231 expression was decreased in the hippocampus and cortex of MDD mice.Conclusions1.Single cage feeding plus chronic unpredictable stress can effectively establish a mouse model of MDD,and cognitive dysfunction in MDD mice may be associated with elevated DAPK1 and tau protein expression in the hippocampus.2.Learning and memory training can improve cognitive dysfunction in MDD mice,and this process may be achieved by inhibiting DAPK1 expression and tau protein phosphorylation in the hippocampus.3.DAPK1 plays an important role in the cognitive impairment of MDD mice by regulating the phosphorylation of tau protein.