The Astrocytes In The Hypothalamus And Brainstem Involved In Water-Immersion Restraint Stress In Rats

Restraint water-immersion stress (RWIS) is considered to be a compound stressmodel, including psychological and physical stimulation. RWIS can result inhypothermia and disorded gastrointestinal function within a few hours, such ashypersecretion of gastric acid and acute gastric erosions. RWIS can imitate clinicalgastric ulcer caused by trauma, operation or sepsis, therefore it is widely used to studythe pathogenesis of stress-induced gastric lesion. Our previous work found that gastricerosions was caused by overaction of neurons in brain. Bilateral subdiaphragmaticvagotomy or pretreated with atropine significantly relieved gastric lesion, indicatinggastric function is primarily controlled by the parasympathetic nervous system, whichmainly arises from the dorsal motor nucleus of the vagus (DMV). Nucleus of solitarytract (NTS) and DMV, the primary parasympathetic center innervating gastric motilityand acid secretion, are part of the vagal complex. Sensory information from thegastrointestinal tract and other viscera can be perceived by NTS via the afferent vagusnerve fibers. Then after integrating the information, NTS secondary neurons futherlyactivate the the vagus preganglionic neurons located in the DMV, throughneurotransmitters such as glutamate. The vagal efferent fibers arising from DMV inturn projected to the gastrointestinal tract, enabling the regulation of gastrointestinalfunction. Paraventricular nucleus(PVN) and supraoptic nucleus (SON) are importantnuclei of the hypothalamus, the superior controller of gastrointestinal function.Current studies demonstrated that these two nuclei are involved in the regulation ofstress response, in addition to keeping water balance and stabilizing blood pressure orosmotic pressure. Previous studies indicated that NTS, DMV in the brainstem alongwith PVN, SON in the hypothalamus are involved in the regulation of visceralfunction. Then, due to the intimate connections between ventrolateral medulla (VLM) and neuclei such as NTS, DMV, PVN and SON, it is reasoned to infer that theVLM is actually a body-viscus integration center which can integrate somatic andvisceral afferent information and regulate visceral activities to maintain homeostasis.In previous experiments, using Fos as the marker for the activation of neurons,wefound the degree of Fos expression in NTS、DMV、PVN and SON correlated with thedurations(30min、1h、2h、3h) of RWIS. These results indicated that these nucleus takepart in RWIS. Astrocytes are the most numerous and widely distributed subtypes ofglial cells in the central nervous system(CNS). Not only compose of the CNSskeleton, astrocytes also participated in the transmission and processing ofinformation in CNS. If so:1. Whether astrocytes participate in RWIS and regulate gastric ulcer induced bythis stress model?2. After5h of RWIS, What’s going to happen to Fos expression?3. If astrocytes participate in RWIS, then which one are actived firstly as forneurons and astrocytes?4. What is the relationship between astrocytes and neurons in Restraint water-immersion stress?Thus, in the present study:1. The rats were divided into control group and experimental group(RWIS0.5、1、2、3、5h). Immunohistochemistry of GFAP and Fos was utilized to to observe theastrocytic.and neural activity in NTS, DMV, SON, PVN and Rostral ventrolateralmedulla(RVLM).2. The rats were divided into saline group、L-AA (Inhibiting the expression ofGFAP)group and ASO (Inhibiting the expression of Fos)group.To detect Fosexpression after inhibiting expression of GFAP and GFAP expression after inhibitingthe expression of Fos,and the possible “crosstalk” between astrocytes and neurons was investigated in RWIS.The present data demonstrated that:1. Astrocytes participated in RWIS.Compared with the control group, the expression of GFAP in the DMV, NTS,VLM, PVN and SON are increased with different degrees. After RWIS0.5h,expression of GFAP was increased significantly in PVN and some sections of NTSand DMV.2. After RWIS5h, the Fos expression showed a rising trend.Similar to the previous results, compared with the control group, Fos-IR neuronsin the DMV, NTS, PVN and SON were increased significantly, especially for DMVand NTS, which confirmed the role of DMV and NTS in the control ofgastrointestinal function.The peak of Fos expression occurred at RWIS1h, then theFos expression decreased gradually. But after5h of stress, Fos expression showed asignificant increase in most nuclei.3. In the RWIS model, astrocytes are activated earlier than neurons.The experimental results showd that, GFAP could be more easily activated thanFos. Compared to the control group, at30min of sress, the expression of GFAPincreased significantly whereas the expression of Fos did’t exhibit significant changein PVN and in some sections of DMV. In addition, the expression of GFAP in therosal and middle NTS increased significantly, while the expression of Fos were variedslightly. Therefore, under RWIS conditions, astrocytes may be first activated, thustransmiting stimulation information to neurons by some neurotransmitters.4. Astrocytes and neurons are cooperative relatedIn this experiment, using the oligonucleotide of c-fos(ASO) to inhibit Fosexpression, a significant reduction of GFAP expression was obtained. Moreover, using astrocytes inhibitor(L-AA) to inhibit GFAP expression, a significant reduction in Fosprotein expression was observed in addition to the decrease of GFAP expression. Allthe results suggested that the activation of astrocytes and neurons are intimatelyinteracted with each other.and collectively participate in the RWIS.