前列腺素E2激活LPBel-POA通路诱导发热的作用及机制研究

Prostaglandin E2 (PGE2), a central effector of pyrogen fever, has preliminarily been shown to be locally synthetized and increase the firing rate and thermosensitivity of neurons in the lateral parabrachial nucleus (LPB), one of the brain sites mediating PGE2-induced fever. However, the neuronal mechanisms for pyrogenic effects of PGE2 in the LPB are still unknown. On the basis of our preliminary experiments and the recent findings that cooling-activated external lateral subnucleus of LPB - preoptic area (LPBel-POA) pathway evokes thermogenic responses similar to those evoked by PGE2 directly injected into the LPB, we propose that the LPBel-POA pathway should be an essential neural substrate for pyrogenic effects of PGE2. In this project, radiotelemetry, immunofluorescence histochemistry, RNA interference, whole cell patch-clamp recordings combined with Fluoro-Gold retrograde tracing or single cell RT-PCR are used to probe whether LPBel-POA pathway mediates pyrogenic effects of PGE2 in the LPB at three levels of whole animal, brain slice, and cell respectively, and further ascertain the subtype(s) of EP3 receptor mediating LPBel-POA pathway activation by PGE2 and its electrophysiological mechanisms. This research not only verifies our hypothesis that LPBel-POA pathway mediates pyrogenic effects of PGE2 in the LPB, but also prepares grounds for further investigating the central circuitries for PGE2-induced fever.

臂旁外侧核（LPB）是前列腺素E2（PGE2）致热作用部位，预实验观察到PGE2能在LPB局部合成，并增加LPB神经元放电频率和温度敏感性，但不清楚LPB内PGE2致热作用的神经机制。最近发现，皮肤冷信号经LPB外亚核-视前区（LPBel-POA）通路引起与LPB局部应用PGE2相似的产热效应，据此推测LPBel-POA通路可能是LPB内PGE2致热作用的神经基础。本课题拟采用无线遥测、免疫荧光、RNA干扰、膜片钳结合荧光金逆行示踪、单细胞RT-PCR等技术，以成年雄性SD大鼠为实验对象，在整体-离体脑片-细胞分子水平，研究PGE2能否激活LPBel-POA通路诱导发热，并明确PGE2激活LPBel-POA通路的EP3受体亚型和电生理机制，验证我们提出的PGE2激活LPBel-POA通路诱导发热的假说，为深入探索PGE2引起发热的中枢神经回路奠定理论基础。

臂旁外侧核(lateral parabrachial nucleus, LPB) 是前列腺素E2（PGE2）致热作用部位之一，但LPB内PGE2的致热机理至今尚未阐明。最近发现，LPB是前馈体温调节上传通路的重要中继站，其背亚核（LPBd）经LPBd-POA通路介导产热减少，散热增加的热防御反应。而LPB外亚核（LPBel）经LPBel-POA通路介导产热增加，散热减少的冷防御反应，这与在LPB局部应用PGE2引起的发热效应一致，我们据此推测LPBel-POA通路可能是LPB内PGE2致热作用的神经基础。本项目采用无线遥测、化学毁损、RNA干扰、RT-PCR、膜片钳结合逆行示踪、病毒顺行或逆行示踪结合免疫荧光双标、能量代谢测定、外周电生理记录等方法技术，证实LPB局部合成的PGE2参与了LPS致热过程，其致热作用与BAT产热，战栗产热增加和尾部皮肤散热减少有关；PGE2差异性调节LPB不同亚核神经元放电活动，分别通过增强LPBel→MnPO投射神经元谷氨酸能兴奋性突触传递激活LPBel→POA通路，激活LPBd→MnPO投射神经元IA电流抑制LPBd→POA通路诱导发热；EP3受体介导了LPB PGE2致热效应；PGE2对LPB不同亚核神经元的差异性调节作用由不同表型神经元介导，PGE2分别兴奋LPBelSST+神经元，抑制LPBdpdyn+神经元诱导发热。这些实验结果基本验证了我们提出的PGE2激活LPBel-POA通路诱导发热的假说，并根据实验结果，我们将假说修正补充为“PGE2激活LPBel-POA通路，抑制LPBd→POA通路诱导发热”。本项目假说的验证和补充，不仅阐明LPB内PGE2致热作用的神经电生理机制，也为发热反应的中枢机制研究提供一个新思路。