Imperato A, Di Chiara G
J Neurosci. 1984 Apr;4(4):966-77. doi: 10.1523/JNEUROSCI.04-04-00966.1984.
A method for the estimation in rats of the in vivo release and metabolism of dopamine (DA) is described. The method is based on the dialysis principle and consists of inserting transversally in the striatum a thin (0.2 mm) dialysis tube (Amicon Vitafiber) which is then perfused with Ringer. The Ringer, flowing at a constant rate of 2 microliters/min in the dialysis tube, extracts low molecular weight substances from the surrounding tissue by way of simple diffusion along a concentration gradient. At the distal end of the dialysis tube, the Ringer is collected every 10 to 20 min and directly injected into a high performance liquid chromatographer (HPLC) equipped with reverse phase octadecyl sulfate columns which separate DA and its metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). These substances are then quantitatively estimated by oxidative electrochemical detection. The basal output of DA is 0.3 pmol/20 min, whereas the outputs of DOPAC and HVA are 60 and 20 pmol/20 min, respectively. In basal conditions the output of DA, DOPAC, and HVA is stable over at least 10 hr. Histological examination of the track left by the dialysis probe in rats after 10 hr of continuous dialysis reveals very little damage and normal neuronal morphology in the vicinity of the dialysis tube. Increase of the K+ concentration in the Ringer to 30 mM produced a sharp, reversible increase of DA output. Both the basal and K+-stimulated release were Ca++ dependent, because omission of Ca++ abolished basal and K+-stimulated DA release. Electrical stimulation of the nigrostriatal DA neurons in the medial forebrain bundle sharply increased DA output. Amphetamine sulfate in low doses (1.0 mg/kg, i.v.) produced a 9-fold increase in DA release and decreased DOPAC and HVA output. alpha-Methyl tyrosine (150 mg/kg, i.v.) reduced within 2 hr DA release to 15% of basal values and in parallel also decreased the output of DOPAC and HVA. Reserpine (5 mg/kg, i.p.) reduced DA release but in a slower fashion than alpha-methyl tyrosine and increased DOPAC and HVA. Pargyline (75 mg/kg, i.p.) produced a 4-fold increase of DA release, while it rapidly brought to zero DOPAC and HVA output. gamma-Butyrolactone (700 mg/kg, i.p.) rapidly and lastingly reduced DA, DOPAC, and HVA output. The biochemical and histological results obtained indicate that the method is suitable to estimate in the rat the changes in the release o f endogenous DA and its metabolites which take place in vivo under administration of centrally acting drug.
本文描述了一种在大鼠体内估算多巴胺（DA）释放及代谢情况的方法。该方法基于透析原理，具体操作是将一根细的（0.2毫米）透析管（密理博Vitafiber）横向插入纹状体，然后用林格氏液进行灌注。林格氏液以2微升/分钟的恒定流速在透析管中流动，通过沿着浓度梯度的简单扩散从周围组织中提取低分子量物质。在透析管的远端，每10至20分钟收集一次林格氏液，并直接注入配备有反相十八烷基硫酸盐柱的高效液相色谱仪（HPLC）中，该柱可分离DA及其代谢产物二羟基苯乙酸（DOPAC）和高香草酸（HVA）。然后通过氧化电化学检测对这些物质进行定量估算。DA的基础输出量为0.3皮摩尔/20分钟，而DOPAC和HVA的输出量分别为60皮摩尔/20分钟和20皮摩尔/20分钟。在基础条件下，DA、DOPAC和HVA的输出在至少10小时内保持稳定。对大鼠进行10小时连续透析后，透析探针留下的轨迹进行组织学检查发现，透析管附近损伤极小，神经元形态正常。将林格氏液中的K⁺浓度提高到30毫摩尔/升会使DA输出量急剧且可逆地增加。基础释放和K⁺刺激释放均依赖于Ca²⁺，因为去除Ca²⁺会消除基础释放和K⁺刺激的DA释放。电刺激内侧前脑束中的黑质纹状体DA神经元会使DA输出量急剧增加。低剂量（1.0毫克/千克，静脉注射）的硫酸苯丙胺会使DA释放增加9倍，并降低DOPAC和HVA的输出量。α-甲基酪氨酸（150毫克/千克，静脉注射）在2小时内将DA释放量降至基础值的15%，同时也降低了DOPAC和HVA的输出量。利血平（5毫克/千克，腹腔注射）会降低DA释放，但速度比α-甲基酪氨酸慢，并增加DOPAC和HVA的输出量。帕吉林（75毫克/千克，腹腔注射）使DA释放量增加4倍，同时迅速将DOPAC和HVA的输出量降至零。γ-丁内酯（700毫克/千克，腹腔注射）迅速且持久地降低DA、DOPAC和HVA的输出量。所获得的生化和组织学结果表明，该方法适用于估算在给予中枢作用药物的情况下，大鼠体内内源性DA及其代谢产物释放的变化。
