| Chronic stress is the main cause or inducement of many diseases,which can cause the organism production performance to decline,the elimination rate and the mortality to rise,which adversely affects the healthy development of our country’s livestock industry.In the process of modern intensive livestock production,animals will inevitably be stimulated by a variety of stressors,such as abrupt climate change,long-distance transportation,high population density,non-standard veterinary treatment,and so on.Chronic stress can cause tissue damage in the Prefrontal cortex(PFC)and induce depression-like behavior in animals,resulting in a series of adverse factors such as weight loss,decreased animal product quality,and increased mortality,resulting in certain economic losses.Therefore,it has become a research hotspot in the fields of veterinary medicine and medicine to explore effective drugs and elucidate the key pharmacodynamic mechanisms.Related studies have shown that chronic stress can reduce the content of 5-Hydroxytryptamine(5-HT)in the brain,and subsequently inhibit the cAMP/CREB/BDNF pathway,damaging the structure and function of neurons.Chlorogenic acid(CGA),a natural drug,has strong neuroprotective effects,such as improving neuron loss.However,whether CGA has a protective effect on chronic stress-induced PFC injury and its mechanism of action remain unclear.Therefore,in this study,the CGA intervention was conducted by establishing a rat model of Chronic restraint stress(CRS).Based on the cAMP/CREB/BDNF pathway,the protective effects of 5-HT on PFC injury in rats induced by chronic stress and its mechanism were explored.A total of 70 healthy male Wistar rats were randomly divided into 7 groups(n=10): control group(CON group),chronic restraint stress group(CS group),50 mg/kg CGA intervention group(CS+CAL group),100 mg/kg CGA intervention group(CS+CAM group),150 mg/kg CGA intervention group(CS+CAH group),100 mg/kg CGA control group(CON+CAM group),and 150mg/kg CGA control group(CON+CAH group).In the CRS model,rats were fixed in the restraint apparatus from 9:00～15:00 every day for 21 days of restraint stress.At the beginning of self-modeling,CGA was dissolved in water 2 hours before CRS every day,and rats in each group were given intragastric administration according to different doses,while rats in the CON group were placed in rat cages without being disturbed.The body weight of each group was recorded every three days.Behavioral tests such as the sugar water preference test,the open field test,the tail suspension test,and the forced swimming test were conducted after the stress procedure.After behavioral testing,all rats were anesthetized with isoflurane,and blood samples were taken quickly and sacrificed.Corticosterone(CORT)content was detected in the serum.PFC tissue structure,5-HT content,cAMP/CREB/BDNF signaling pathway,IDO and SERT changes were detected.Furthermore,the regulatory mechanism of CGA on 5-HT and its protective effect on PFC under chronic stress were clarified.Results: The body weight measurement results of rats showed that compared with the CON group,the body weight of rats in the CS group decreased significantly(p<0.001),and compared with the CS group,the body weight of rats in the CS+CAL,CS+CAM and CS+CAH groups had a trend of increasing,but the body weight of rats in the CS+CAM group increased significantly(p<0.01).ELISA detection of serum CORT content in rats showed that the serum CORT content of rats in the CS group was significantly increased compared with the CON group(p<0.01),while CORT content in the CS+CAM and the CS+CAH groups was significantly decreased(p<0.01,p<0.05).The results of behavioral tests showed that compared with the CON,the preference rate of rats in the CS group for sugar water decreased significantly(p<0.001),the total distance,the number of cross cells,the number of standing times and the central area activity time decreased significantly,and the stationary time increased significantly in tail suspension and forced swimming(p<0.01).However,100 mg/kg of CGA can improve the depression-like behavior of the above rats.The histopathological results showed that the PFC nerve cells in the CON,CON+CAH and CS+CAM groups were complete in structure and abundant in Nissl bodies.In the CS group,CS+CAL group and CS+CAH group,large area hyperemia,blurred neuronal structure,disappearance of nucleoli,glial cell nucleus pyknotic,and disappearance of Nissl bodies were observed.ELISA showed that compared with CON,5-HT content in the PFC of rats in the CS group was significantly decreased(p<0.01),and 5-HT content in rats in the CS+CAM group was significantly increased(p<0.05).Therefore,the dose of 100 mg/kg CGA will be used for subsequent experimental exploration.In addition,there was no damage to the body after the 150mg/kg CGA blank control intervention,indicating that CGA has a high degree of safety and reliability.The ultramicrostructure of PFC showed that the synaptic structure of the CON group and the CS+CAM group was complete and clear,and the gap was regular and tight.The number of CS synapses decreased significantly,the gap boundaries blurred,and the anterior and posterior membranes fused.The results of ELISA,Western blot,immunohistochemistry and Real-time PCR detection of cAMP/CREB/BDNF path-related proteins and genes showed that the expression levels of cAMP,PKA,p-CREB and BDNF in the CS group were significantly lower than those in the CON group and the CS+CAM group.Western blot,immunohistochemical and Real-time PCR detection of IDO and SERT showed that the levels of IDO and SERT in the CS group were significantly higher than those in the CON group and the CS+CAM group.Molecular docking techniques and CETSA results showed that IDO and SERT were closely bound to CGA,suggesting that they could be used as potential utility targets.Comprehensive results of various parts of the experiment showed that CRS caused a decrease in the content of neurotransmitter 5-HT in rat PFC,thereby inhibiting the cAMP/CREB/BDNF signal pathway,reducing the nutritional and regenerative function of nerve cells,causing damage to the PFC and producing depressive behavior in it.However,CGA can directly or indirectly increase the content of 5-HT by acting on the potential targets IDO and SERT,promote the cAMP/CREB/BDNF signal pathway to play a protective role in the PFC injury of rats caused by CRS,and thus improve the depressive behavior of rats. |
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