Cheer J F, Marsden C A, Kendall D A, Mason R
School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, NG7 2UH, Nottingham, UK.
Neuroscience. 2000;99(4):661-7. doi: 10.1016/s0306-4522(00)00241-4.
Cannabinoid compounds have been reported to excite ventral tegmental neurons through activation of cannabinoid CB1 receptors. More recently, biochemical and whole-cell voltage-clamp studies carried out on CB1-transfected AtT20 cells have shown a rapid desensitization of these receptors following activation of protein kinase C by 4-alpha-phorbol. To investigate the possible physiological correlates of this phenomenon, we have studied the effects of repeated cannabinoid treatment on ventral tegmental area dopaminergic neuronal firing in vitro. Rat brain slices containing the ventral tegmental area were used for single-unit extracellular recordings. Only neurons meeting established electrophysiological and pharmacological criteria for dopaminergic neurons were used in the study (firing neurons were detected either using tungsten or glass microelectrodes). The high-affinity cannabinoid agonist HU210 produced a concentration-dependent increase in firing (1-15 microM; EC(50) approximately 7 microM). Initial HU210 exposure produced a significant increase in cell firing rate in the ventral tegmental area, with a maximum approximately 3.5-fold increase over pre-drug basal firing; a subsequent exposure to HU210 produced an approximately threefold increase over basal firing. Nevertheless, the duration and onset of excitation produced by the cannabinoid differed significantly between the first and second exposures; the first excitation lasted significantly longer than the second and required less time to reach a comparable change in firing rate. The increases in firing rate and the time to return to basal firing were not significantly different between exposures. Furthermore, the cannabinoid antagonist SR141716A completely prevented the HU210-induced excitation whilst having no effect on its own, thus indicating a CB1-receptor mediated mechanism for the observed increase in firing. Ventral tegmental area neurons are also excited by the GABA(A) receptor antagonist bicuculline. To assess the role of GABA in cannabinoid-mediated excitation, HU210 was added in the presence of bicuculline. HU210 did not affect the initial bicuculline-induced increase in firing, suggesting different sites of action for the two compounds. Our data fail to support previously reported findings using repeated cannabinoid administration and cell preparations. The maintained increase in DA drive elicited by the potent cannabinoid agonist HU210 in the in vitro ventral tegmental circuit could explain some of the behavioural properties of cannabinoids, such as the lack of tolerance for the psychotropic effects of marijuana seen in human users.
据报道，大麻素化合物可通过激活大麻素CB1受体来兴奋腹侧被盖区神经元。最近，对转染CB1的AtT20细胞进行的生化和全细胞膜片钳研究表明，在用4-α-佛波醇激活蛋白激酶C后，这些受体迅速脱敏。为了研究这一现象可能的生理相关性，我们在体外研究了重复给予大麻素对腹侧被盖区多巴胺能神经元放电的影响。含有腹侧被盖区的大鼠脑片用于单细胞细胞外记录。本研究仅使用符合既定的多巴胺能神经元电生理和药理学标准的神经元（使用钨电极或玻璃微电极检测放电神经元）。高亲和力大麻素激动剂HU210产生了浓度依赖性的放电增加（1 - 15微摩尔；半数有效浓度约为7微摩尔）。最初暴露于HU210使腹侧被盖区的细胞放电率显著增加，最大增加幅度比给药前的基础放电增加约3.5倍；随后再次暴露于HU210使放电比基础放电增加约三倍。然而，大麻素引起的兴奋持续时间和起始时间在第一次和第二次暴露之间有显著差异；第一次兴奋持续时间明显长于第二次，且达到可比放电率变化所需时间更短。两次暴露之间放电率增加幅度和恢复到基础放电的时间没有显著差异。此外，大麻素拮抗剂SR141716A完全阻止了HU210诱导的兴奋，而其自身无作用，因此表明观察到的放电增加是由CB1受体介导的机制。腹侧被盖区神经元也会被GABA(A)受体拮抗剂荷包牡丹碱兴奋。为了评估GABA在大麻素介导的兴奋中的作用，在荷包牡丹碱存在的情况下加入HU210。HU210不影响最初荷包牡丹碱诱导的放电增加，表明这两种化合物的作用位点不同。我们的数据不支持先前使用重复给予大麻素和细胞制剂所报道的结果。强效大麻素激动剂HU210在体外腹侧被盖区回路中引起的多巴胺驱动的持续增加可以解释大麻素的一些行为特性，比如人类使用者对大麻精神效应缺乏耐受性。
