Molecular mechanisms of pain signal transmission and evaluation of analgesias inn vitro

1) 5-羟色胺（5-HT）增强了表达5-HT_<2A>和5-HT_7受体的大鼠背根神经节（DRG）细胞对辣椒素的Ca^<2+>反应。蛋白激酶C或蛋白激酶ea的阻滞剂抑制了增加的反应。2)在离体新生大鼠脊髓中，电刺激L3前侧根引起快速单突触反射电位（MSR），随后在同侧L3前侧根引起极慢电位变化（sVRP）。α2肾上腺素受体激动剂、右美托咪定（DEX）、噻嗪（XY）和可乐定（CLO）或阿片受体激动剂、吗啡（MOR）和DPDPE均可抑制sVRP。效价顺序为：DEX>DPDPE=MOR>CLO>XY。3)用含20% CO_2 （pH6.7）的碳酸氢盐缓冲生理溶液灌注离体脊髓，可抑制MSR和sVRP。腺苷A1受体拮抗剂可减弱高碳酸血症的抑制作用。腺苷激酶抑制剂对MSR和sVRP也有类似的抑制作用。4)锌选择性增强sVRP，使其受到谷氨酸NMDA受体拮抗剂、氯胺酮和AP-5的抑制。5)DEC、XY和MOR对福尔马林诱导的挣扎体运动有抑制作用。DEC和MOR抑制辣椒素诱导的体运动，但不抑制XY，表明XY作用部位的差异。6)克隆了猪香草样蛋白受体蛋白编码cDNA(2484个碱基，猪TRPV1)，与大鼠或人TRPV1同源性为84%。7)TRPV1在HEK293细胞中表达。在这些细胞中，辣椒素引起细胞内Ca^<2+>浓度和非选择性阳离子电流的增加。降低细胞外pH值或升高灌注液温度（> - 42℃）可激活猪TRPV1，表明其在Ca2+g多模态受体中的功能。8)尽管猪TRPV1对香草样受体激动剂的反应性与大鼠TRPV1非常相似，但降低细胞外pH在猪中比在大鼠TRPV1中更有效。

1)Serotonin (5-HT) potentiated Ca^<2+> response to capsaicin in cultured rat dorsal root ganglion (DRG) cells expressing 5-HT_<2A> and 5-HT_7 receptors. The increased response was inhibited by the blocker of protein kinase C or protein kinaseA. 2)In isolated neonatal rat spinal cord, electrical stimulation of the L3 ventral root elicited a fast monosynaptic reflex potential (MSR) followed by very slow potential change (sVRP) in the ipsilateral L3 ventral root. The sVRP was inhibited by α2 adrenoceptor agonists, dexmedetomidine (DEX), xylazine (XY) and clonidine (CLO) or opiate receptor agonists, morphine (MOR) and DPDPE. The order of potency was DEX>DPDPE=MOR>CLO>XY. 3)MSR and sVRP were inhibited by perfusion of the isolated spinal cord with bicarbonate-buffeted physiological solution bubbled with 20% CO_2 (pH6.7). The inhibitory effect of hypercapnia was attenuated by adenosine A1 receptor antagonists. An adenosine kinase inhibitor evoked a similar inhibitory effect on MSR and sVRP. 4)Zinc selectively potentiated sVRP, which inhibited by glutamate NMDA receptor antagonists, ketamine and AP-5. 5)DEC, XY and MOR inhibited struggling body movement induced by formalin. DEC and MOR inhibited capsaicin-induced body movement but not XY, indicative of the difference in the site of action of XY. 6)We cloned cDNA coding porcine vanilloid receptor protein (2484 bases, porcine TRPV1), which was 84% homology to rat or human TRPV1. 7)TRPV1 were expressed in HEK293 cells. In these cells, capsaicin caused increases in intracellular Ca^<2+> concentration and non-selective cation currents. Porcine TRPV1 was activated by reducing extracellular pH or increasing temperature of the perfusing solution (>42℃), indicative of fiunctionin Ca2+g polymodal receptors. 8)Although responsiveness to vanilloid agonists in porcine TRPV1 was quite similar to that in rat TRPV1, the reduction of extracellular pH was more effective in porcine than rat TRPV1.