Effects Of Naloxone On IL-1β-induced Fever And The Content Of CAMP And HSP70 In Hypothalamus In Rats

INTRODUCTIONFever is one of simultaneous phenomenon of many clinical diseases. Cyto-kines are involved in the activation of thermogenesis and fever. IL - 1β (inter-leukin - 1β) has been widely considered one of endogenous pyrogens, and many experiments have demonstrated that direct administrating IL -1β into the lateral cerebral ventricle of rat brain induce the febrile response of animals.Naloxone is a specific opioid receptor antagonist, which is used on treating a lot of diseases of endogenous opioid peptides. Recently many researchers have found that the opioid system are involved in the febrile response caused by endogenous pyrogens. Central or peripheral pretreatment with injection of the opioid receptor antagonists can attenuate or even block the fever induced by IL -1β,tumur necrosis factor(TNF),interferon(IFN),interleukin -6 (IL - 6) and so on.Cyclic adenosine monophosphate( cAMP ) is a crucial mediator regulating cell function and synapsis transmition, and is also a major central neurogen in febrile response.Heat Shock Protein70 ( HSP70) is a kind of non - specificity cytoprotective protein , which is rapidly induced when exposing the cell and tissue to thermal stimulation in order to enhance the cell'resistance to the heat damage.In this study, we observed the effect of non - specific opioid receptor antagonist naloxone on IL - 1β - induced fever in rats after central injection, and detected the content of cAMP and HSP70 expression in hypothalamus , in order to explore the effect of opioid receptor on IL - 1β - induced fever and its mechanism.MATERIALS AND METHODS1. Animals and SurgeryHealthy male Sprague - Dawley rats, weighing between 200 and 250 g, were provided by China Medical University Animal Center. They were maintained in a room at (22 ± 2) X. and individually housed in wire - mesh cages with a 12 - h light: 12 - h dark cycle and fed with dry powder chow, with tap water available ad libitum. Under intraperitoneal anesthesia ( Ketamine hydro-chloride and xylazine) , a 1 mm of diameter cannula was implanted into the lateral cerebral ventricle by stereotaxic apparatus according to the rat brain altas, and the cannula was anchored with dental acrylic, sealing off upper end. Rats were individually housed a week before experiments. Examine body temperature: the digital thermometer probe were inserted 6 centimeters in rats'rectum. Value was recorded after stabilization. Mean of three values 1 hour before administration was taken as baseline of pre - handling. After administration, rectal temperature was recorded respectively during experiments (to measure once every other 15 min in 0 - lh, every other 30 min in 1 - 2h, every other 60 min in 2 - 8h). When the experiment was over, cannula & position were checked to discard incorrect ones.2. cAMP level assayRats were put to death immediately at the peak of IL - 1 f$ - induced fever , then the rat brain tissues were put into liquid nitrogen. 50mg hypothalamus tissue was weighted, homogenate, centurfuged and assay concentration of cAMP with radio - immunoassay.3. The expression of HSP70 in rat hypothalamus tissue assay Hypothalamus tissue was homogenated, centrifuged and the supernatantswere used to measure the expression of HSP70 by Western blot assay.4. Statistical analysisResults were presented as mean ± SD (x ± s). Change in body temperature were presented and analyzed with ATco. Data were analyzed using t - test.RESULTS1. In the febrile rats induced by i. c. v. IL - 1(3, cAMP level and HSP70 expression in hypothalamus were significantly increased.2. Pretreatment with nal could inhibit the fever induced by IL - 1(3, and meanwhile the content of cAMP and HSP70 lever were lower.DISCUSSIONIn our study, the body temperature of the rats began to rise at 30 minutes after intracerebroventricular administration of IL - 1 (3, and peaked at 2 hours. The fever lasted more than 6 hours. And the cAMP level in hypothalamus was significantly higher,which was consistent with the previous report.Recently many researchers have found that the opioid system are involved in the febrile response caused by cytokines as IL - 1 (3, TNF, IFN, IL - 6 and so on. Three opioid receptors (8,|x and *y) have been cloned and all are localized within the preoptic area of the anterior hypothalamus. Central or peripheral pretreatment with injection of the opioid receptor antagonists can attenuate or even block the fever caused by cytokines. Our research observed that IL - 1 p - induced fever was obviously inhibited by pretreatment with i. c. v. the non - specific opioid receptor antagonist Nal, and cAMP lever in hypothalamus was significantly lower than IL - 1 (3 group, which suggested that the effect of Nal on IL -1 (3 - induced fever might be caused by inhibition of cAMP synthesis in hypothalamus.Exposed to high temperature or other stress, the cells can be induced to produce a highly conserved set of polypeptides termed heat shock proteins ( HSPs). Recently many studies have revealed that these cellular stress proteins form the protection role against heat. As a non - specific cellular protective protein, HSP70 serves as molecular chaperones and enhances the ability of cells to survive thermal damage , and repairs the deformed protein caused by heat - related casualty(HRC). In this study, with the development of febrile response, theexpression of HSP70 in hypothalamus was gradually increased after i. c. v. IL -1(3. Pretreatment with Nal, in addition to inhibiting the IL - 1(3 febrile responses , significantly decreased the expression of HSP70, which further demonstrated that opioid receptor played a critical role in the course of IL -1 p - induced fever and induced the potential effect to organism by fever.These findings suggest that the activation of endogenous opioid system play a crital role in the IL - 1(3 - induced fever and epigenetic HSP70 expression. It required further experimental confirmation that whether HSP70 are involved in the febrile ceiling by inhibition of central febrile mediator and if opioid system is involved in the activation of HSP70.CONCLUSION1. cAMP is involved in IL - 1 (3 induced febrile response.2. The opioid receptor plays a critical role in the course of IL - 1(3 - induced fever.3. The opioid receptor antagonist naloxone can inhibite the IL - 1 (3 - induced fever in rats, and its mechanism may be related to the decrease of cAMP synthesis in hypothalamus;meanwhile the expression of HSP70 is reduced.