Trendelenburg A U, Klebroff W, Hein L, Starke K
Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, Freiburg, Germany.
Naunyn Schmiedebergs Arch Pharmacol. 2001 Aug;364(2):117-30. doi: 10.1007/s002100100423.
The function of presynaptic alpha2-autoreceptors was studied in the hippocampus, occipito-parietal cortex, atria and vas deferens of NMRI mice, mice in which the alpha2A/D-, the alpha2B- or alpha2c-adrenoceptor gene had been disrupted (alpha2A/DKO, alpha2BKO and alpha2CKO, respectively), and the wildtype mice from which the knockout animals had been generated. Tissue pieces were preincubated with 3H-noradrenaline and then superfused and stimulated electrically. The alpha2-adrenoceptor agonist medetomidine reduced the electrically evoked overflow of tritium in all tissues from all mouse strains (stimulation with single pulses or single high-frequency pulse trains, called POPs, i.e. pulse patterns leading to minimal autoinhibition). The effects of medetomidine did not differ in NMRI, wildtype, alpha2BKO and alpha2CKO mice but were greatly reduced in alpha2A/DKO brain preparations and to a lesser extent in alpha2A/DKO atria and vasa deferentia. Six drugs were tested as antagonists against medetomidine. Their pKd values indicated that the hippocampal and occipito-parietal alpha2-autoreceptors in NMRI and wildtype mice were alpha2D (the rodent variant of the alpha2A/D-adrenoceptor) whereas the atrial and vas deferens alpha2-autoreceptors in NMRI and wildtype mice could not be identified with a single alpha2 subtype. Deletion of the alpha2A/D gene changed the pKd values in all tissues so that they now reflected alpha2C properties, whereas deletion of the alpha2C gene changed the pKd values in atria and vasa deferentia so that they now had alpha2D properties (as they had in NMRI and wildtype brain preparations). Autoinhibition by released noradrenaline was created using trains of up to 64 pulses or up to 4 POPs, and the overflow-enhancing effect of the alpha2 antagonist rauwolscine was determined. Results did not differ, irrespective of whether preparations were obtained from NMRI, wildtype, alpha2BKO or alpha2CKO mice: the overflow of tritium elicited by p pulses or POPs was much smaller than p times the overflow elicited by a single pulse or POP, and rauwolscine greatly increased the evoked overflow. Results differed, however, in tissues taken from alpha2A/DKO mice: in these tissues, the overflow of tritium elicited by p pulses or POPs was close to p times the overflow elicited by a single pulse or POP, and rauwolscine did not increase the evoked overflow of tritiumor increased it only marginally. When a greater degree of autoinhibition was produced in atria and vasa deferentia by stimulation with 120 pulses, both disruption of the alpha2A/D gene and disruption of the alpha2C gene but not disruption of the alpha2B gene attenuated the overflow-enhancing effects of phentolamine and rauwolscine. In NMRI and wildtype atria and vasa deferentia, the relative potencies of phentolamine and rauwolscine at enhancing the evoked overflow were not easily compatible with a single alpha2 subtype. In alpha2A/DKO atria and vasa deferentia, the relative potencies of phentolamine and rauwolscine indicated that the autoinhibition-mediating receptors were alpha2C, whereas in alpha2CKO atria and vasa deferentia the relative potencies indicated that the autoinhibition-mediating receptors were alpha2D. It is concluded that alpha2-autoreceptors function identically in NMRI mice and the wildtype mice from which the receptor-deficient animals had been generated. There is no evidence from the experiments for any contribution of alpha2B-adrenoceptors to autoreceptor function. The main presynaptic alpha2-autoreceptors are alpha2A/D, both as sites of action of exogenous agonists and as sites of action of previously released noradrenaline. However, there are in addition non-alpha2A/D-, probably alpha2C-autoreceptors. They are less prominent in mediating the inhibitory effects of exogenous agonists and the negative feedback effect of released noradrenaline. They operate not only after deletion of the alpha2A/D-adrenoceptors but also in normal (NMRI, wildtype) mice without gene deletion.
在NMRI小鼠、α2A/D -、α2B -或α2C -肾上腺素能受体基因已被破坏的小鼠（分别为α2A/DKO、α2BKO和α2CKO）以及产生基因敲除动物的野生型小鼠的海马体、枕顶叶皮质、心房和输精管中研究了突触前α2 -自身受体的功能。将组织块用3H -去甲肾上腺素预孵育，然后进行灌流并电刺激。α2 -肾上腺素能受体激动剂美托咪定降低了所有小鼠品系所有组织中电诱发的氚溢出（用单个脉冲或单个高频脉冲串刺激，即导致最小自抑制的脉冲模式）。美托咪定的作用在NMRI、野生型、α2BKO和α2CKO小鼠中没有差异，但在α2A/DKO脑制备物中大大降低，在α2A/DKO心房和输精管中降低程度较小。测试了六种药物作为美托咪定的拮抗剂。它们的pKd值表明，NMRI和野生型小鼠海马体和枕顶叶的α2 -自身受体是α2D（α2A/D -肾上腺素能受体的啮齿动物变体），而NMRI和野生型小鼠心房和输精管的α2 -自身受体不能用单一的α2亚型来识别。α2A/D基因的缺失改变了所有组织中的pKd值，使其现在反映α2C特性，而α2C基因的缺失改变了心房和输精管中的pKd值，使其现在具有α2D特性（如同在NMRI和野生型脑制备物中一样）。使用多达64个脉冲或多达4个脉冲串产生由释放的去甲肾上腺素引起的自抑制，并测定α2拮抗剂萝芙木碱的溢出增强作用。无论制备物是从NMRI、野生型、α2BKO还是α2CKO小鼠获得，结果都没有差异：由p个脉冲或脉冲串引起的氚溢出远小于p乘以由单个脉冲或脉冲串引起的溢出，并且萝芙木碱大大增加了诱发的溢出。然而，从α2A/DKO小鼠获取的组织结果不同：在这些组织中，由p个脉冲或脉冲串引起的氚溢出接近p乘以由单个脉冲或脉冲串引起的溢出，并且萝芙木碱没有增加诱发的氚溢出或仅略微增加。当通过120个脉冲刺激在心房和输精管中产生更大程度的自抑制时，α2A/D基因的破坏和α2C基因的破坏而非α2B基因的破坏减弱了酚妥拉明和萝芙木碱的溢出增强作用。在NMRI和野生型心房和输精管中，酚妥拉明和萝芙木碱增强诱发溢出的相对效力不容易与单一的α2亚型相匹配。在α2A/DKO心房和输精管中，酚妥拉明和萝芙木碱的相对效力表明介导自抑制的受体是α2C，而在α2CKO心房和输精管中相对效力表明介导自抑制的受体是α2D。得出的结论是，α2 -自身受体在NMRI小鼠和产生受体缺陷动物的野生型小鼠中功能相同。实验没有证据表明α2B -肾上腺素能受体对自身受体功能有任何贡献。主要的突触前α2 -自身受体是α2A/D，既是外源性激动剂的作用位点，也是先前释放的去甲肾上腺素的作用位点。然而，此外还有非α2A/D -的，可能是α2C -自身受体。它们在介导外源性激动剂的抑制作用和释放的去甲肾上腺素的负反馈作用方面不太突出。它们不仅在α2A/D -肾上腺素能受体缺失后起作用，而且在没有基因缺失的正常（NMRI、野生型）小鼠中也起作用。
