Collateral Projections From The Caudal Subnucleus Of Spinal Trigeminal Nucleus To The Ventral Posteromedial Thalamic Nucleus And The Parafascicular Thalamic Nucleus In The Rat

Pain is an annoying sensation triggered by definite or possible tissue damage,which is one of the most commonly seen clinical syndromes.It is characterized with multiple dimensions including sensory-discriminatory and affective-cognitive evaluative components.Physiological pain can act as warning and generate a series of defensive behaviors,thus helping people to avoid noxious stimulations.Pathological pain,however,can be intractable and dramatically interfere with patients' life quality.To date,numerous researches have been made on the mechanism of orofacial pain signaling transmission.However,whether orofacial pain signaling is transmitted to the lateral and medial thalamus by divergent axon collaterals remains illusive.Previous researches have demonstrated that projection neurons within the superficial laminae of the spinal dorsal horn project to the lateral thalamus nuclei like ventral posterior lateral(VPL)and then to the primary and secondary somatosensory cortex concerning pain sensation,which forms the lateral pain system;meanwhile,neurons in deeper laminae of the spinal dorsal horn project to the medial thalamic nuclei like midline/intralaminar nuclei(MINT)and then to key limbic nuclei including medial prefrontal cortex(mPFC),amygdala and insular cortex(IC)to process information about pain related emotions,which is termed as the medial pain system.Researches have also demonstrated that similar organization also exists in the caudal subnucleus of spinal trigeminal nucleus(Vc),which is also known as the medullary dorsal horn(MDH).However,no previous research has displayed the existence of Vc neurons which send collateral projections to the lateral and medial thalamus,respectively.The results of our earlier researches demonstrated that projection neurons in the Vc transmit orofacial pain signaling to the ventral posteriormedial thalamic nucleus(VPM).In addition,neurons in the parafascicular thalamic nucleus(Pf),an essential component of the MINT,expressed Fos protein in orofacial pain model,which implied its participation in orofacial pain transmission process.Under such circumstances,whether there exist collateral projection neurons in the Vc that respectively send projecting fibers and terminals to the VPM and Pf remains elusive.What's more,what is the neurochemical properties of these neurons and do these collateral projection neurons modulate pain transmission? All of these are questions remain unsolved.Given these scientific problems,we predominantly utilized traditional retrograde tracing methods and were attempted to provide morphological evidence of a possible collateral projection from the Vc to the VPM and Pf.Furthermore,orofacial pain model induced by formalin was applied in rats to examine whether these collateral projection neurons participate in pain transmission.The contents of the present work are divided into three parts:1.Vc sent collateral projections to the VPM and Pf in the ratsRats were stereotactically injected with fluoro-gold(FG)into the VPM and tetramethylrhodamine-dextran(TMR)into the Pf.Rats were sacrificed 7 d after injections and immunofluorescent double staining of FG and TMR was applied.The distribution of FG-/TMR-labeled neurons and FG/TMR double-labeled neurons were observed and counted.The results showed that FG/TMR double-labeled neurons were observed mainly in laminae of the Vc with contralateral predominance.2.Immunofluorescent triple staining of FG,TMR and SPR/SPSerial sections from part one were used for immunofluorescent triple staining of FG,TMR and substance P receptor(SPR)and the distribution of FG-/TMR-labeled neurons,FG/TMR double-labeled neurons and SPR-expressing neurons were observed and counted.The results exhibited that some of the collateral projection neurons in the Vc expressed SPR.Additionally,immunofluorescent triple staining of FG,TMR and substance P(SP)was also applied,which demonstrated that some collateral projection neurons in superficial laminae of the Vc were in close contacts with SP-containing fibers and terminals.3.Collateral projection neurons expressed Fos induced by orofacial formalin pain stimulationRats were intradermally injected with 4% formalin to induce an orofacial pain model.Results demonstrated that significant more Fos-immunoreactivity(Fos-ir)neurons were observed in superficial laminae of the Vc than in the control group,which implied that Vc was activated by orofacial pain stimulation.Meanwhile,serial sections of the Vc were also performed with immunofluorescent triple staining of FG,TMR and Fos protein.The results showed that collateral projection neurons in laminae ? and ? send orofacial pain signals to the VPM and Pf,respectively.In summary,the present results indicate that:(1)Vc neurons send collateral projections to the VPM and Pf through axonal branches,respectively;(2)Collateral projection neurons in the Vc expressed SPR and closely contacted with SP-immunoreactive fibers and terminals;(3)Collateral projection neurons in the Vc might participate in orofacial nociceptive information transmission.