Receptor And Electrophysiological Mechanisms Of Prostaglandin E₂ In The Lateral Parabrachial Nucleus Participating In LPS-induced Fever

Research background and purpose:The prostaglandin（PG）E₂,the final mediator of lipopolysaccharide（LPS）-induced fever,elicits febrile response via EP3 receptors in the preoptic area（POA,thermoregulatory command center）.However,the lateral parabrachial nucleus（LPB）,a crucial relay for afferent signals from the skin that promote defensive thermoregulatory responses,also plays a role in the PGE₂-induced fever.Our and others’experiments showed that PGE₂can be locally synthesized in LPB during LPS-induced fever,and microinjection of PGE₂in LPB induces fever by increased BAT thermogenesis,shivering and decreased heat dissipation in the tail skin via exciting LPBel→Mn PO pathway and inhibiting LPBd→Mn PO pathway.However,there is no direct evidence that LPB is involved in LPS-induced fever,and its receptor and electrophysiological mechanisms have not been fully clarified and need to be further studied.On the basis of previous studies,adult male SD rats were used as experimental subjects in this study to further investigate the following points:（1）Whether LPB was involved in systemic fever induced by LPS?（2）What type of receptors mediated PGE₂-induced fever in LPB?（3）What was the electrophysiological mechanism underlying the fever induced by PGE₂in LPB.Materials and Methods:1.Chemical lesion,immunofluorescence,thionin staining and radiotelemetry were used to observe the effect of LPB-lesion on the fever induced by intraperitoneal injection of LPS in ratsFirstly,ibotenate（IBO）was microinjected into LPB to chemically destroy LPB neurons.One week later,immunofluorescence and thionin staining were used to observe the lesion of neurons,so as to confirm the effect and range of LPB lesion.Finally,radiotelemetry method was used to monitor the changes of core temperature(T_core)and activity of rats after intraperitoneal injection of LPS,so as to observe the effects of LPB lesion on the LPS-induced fever in rats,and analyze whether LPB participates in LPS-induced systemic fever.2.Radiotelemetry and wired physiological signal recording were used to study the type of receptors mediating the fever induced by PGE₂in LPB Firstly,the changes of T_coreand energy expenditure（EE）in conscious rats after localy injection of PGE₂receptor agonists and antagonists plus PGE₂in LPB were monitored by animal monitoring system,so as to identify the receptor type mediating LPB PGE₂-induced fever.The results were further verified in anesthetized rats.The BAT sympathetic firing activity（BAT SNA）and neck muscle electromyographic（EMG）activity of anesthetized rats were recorded by BL-420S biological function experimental system,the rectal temperature(T_rec),BAT temperature(T_BAT)and tail temperature(T_tail)of anesthetized rats were simultaneously recorded by eight-channel temperature recorder.The effects of EP3 receptor agonist and EP3 receptor antagonist plus PGE₂on the above parameter were observed.3.The electrophysiological mechanisms of PGE₂-induced fever in LPB were investigated by patch clamp technique combined with retrobeads retrograde tracingOne week after Mn PO was injected with retrobeads,the immunofluorescence labeling technique was used to detect the expression of EP3 receptors in Mn PO-projecting LPB neurons labeled with red beads.The effects of EP3 agonist and EP3 antagonist plus PGE₂on the firing activity of Mn PO-projecting LPB neurons were observed using patch-clamp technique.Finally,the effects of PGE₂,EP3 agonist and EP3 antagonist plus PGE₂on the synaptic activity and I_Acurrents of Mn PO-projecting LPBel neurons and Mn PO-projecting LPBd neurons were observed by patch clamp technique combined with retrograde tracing technique,respectively.Result:1.After LPB lesion in rats,almost no Neu N-immunopositive neurons were found in the damaged area,neuron bodies were lost,a large number of Nissl bodies disappeared,and glial cells were significantly increased.2.Febrile response was also observed after intraperitoneal injection of the LPS in rats with LPB lesion,but was significantly attenuated compared with the control.3.Locally injection of EP3 agonist sulprostone in LPB of conscious rats elicited the fever similar to that of PGE₂,resulting in increased heat production and body temperature.The fever induced by PGE₂in LPB was abolished by administration of EP3 receptor antagonist L-798106.However,EP1 and EP4 agonists and antagonists had no above effects.4.Microinjection of PGE₂into the LPB showed the increases in rectal temperature,BAT temperature,BAT sympathetic nerve discharge activity,and neck muscle EMG as well as a decrease in tail skin temperature in anesthetized rats.The above effects of PGE₂in LPB were mimicked by EP3 receptor agonist administration,whereas these effects were completely abolished by EP3 receptor antagonist administration.5.EP3 receptors were expressed on most Mn PO-projecting LPB neurons.6.The EP3 receptor agonist sulprostone mimicked the excitatory effect of PGE₂on Mn PO-projecting LPBel neurons and its inhibitory effect on Mn PO-projecting LPBd neurons.Besides,the electrophysiological effect of PGE₂was completely abolished by blocking the EP3 receptors with L-798106.7.PGE₂enhanced glutamatergic excitatory synaptic transmission through EP3receptors in the postsynaptic membrane,which in turn excited Mn PO-projecting LPBel neurons.8.PGE₂inhibited Mn PO-projecting LPBd neurons by activating I_Acurrents through EP3 receptor.Conclusions:1.Chemical lesion of LPB partially eliminates the LPS-induced fever,indicating that LPB plays a role in the LPS-induced fever.2.EP3 receptors mediate the fever induced by PGE₂in the LPB.3.The enhanced excitatory synaptic transmission in Mn PO-projecting LPBel neurons and activation of I_Acurrents in the Mn PO-projecting LPBd neurons excited LPBel→POA pathway and inhibited LPBd→POA pathway,respectively.