Near J A, Bigelow J C, Wightman R M
Department of Chemistry, Indiana University, Bloomington.
J Pharmacol Exp Ther. 1988 Jun;245(3):921-7.
Uptake of dopamine (DA) by chopped tissue prepared from rat corpus striatum has been examined to determine whether one or two kinetically distinct uptake sites exist. Two methods were used: direct measurement of accumulated [3H]DA and determination of the rate of formation of 3,4-dihydroxyphenylacetic acid (DOPAC) after exposure to DA. The rate of formation of DOPAC in the latter experiments is a direct function of the rate of DA uptake. The rates of [3H]DA uptake and DOPAC formation are both linear with time in the presence of 10 microM substrate. Studies of [3H]DA accumulation into chopped tissue reveal two apparent components with Km values of 160 nM and 3.8 microM, whereas similar experiments with striatal homogenate or synaptosomes yield a single uptake component with a Km equivalent to the lower value found in chopped tissue. Evaluation of DA uptake via the rate of DOPAC formation gave a Km value of 2.3 microM. (High substrate values were used, so a lower value for Km is not apparent in the data.) The high Km-value component was absent in animals with a lesioned striatum induced by prior nigral injections of 6-hydroxydopamine. Several pharmacologic agents (benztropine, amfonelic acid, bupropion, nomifensine and ouabain) were evaluated. All reduced the uptake of DA in chopped tissue, but with reduced potencies compared with the effect of these agents in synaptosomes. The high Km activity in chopped tissue, as well as the apparent reduced potency of uptake inhibitor, appears to arise from the diffusional barrier present inside more intact tissue. This barrier is not present in homogenates or synaptosomes, and, thus, a single uptake process is seen.
已对从大鼠纹状体制备的切碎组织对多巴胺（DA）的摄取进行了研究，以确定是否存在一个或两个动力学上不同的摄取位点。使用了两种方法：直接测量积累的[³H]DA以及在暴露于DA后测定3,4-二羟基苯乙酸（DOPAC）的形成速率。在后者的实验中，DOPAC的形成速率是DA摄取速率的直接函数。在存在10微摩尔底物的情况下，[³H]DA摄取速率和DOPAC形成速率均随时间呈线性关系。对切碎组织中[³H]DA积累的研究揭示了两个明显的组分，其Km值分别为160纳摩尔和3.8微摩尔，而对纹状体匀浆或突触体进行的类似实验产生了一个单一的摄取组分，其Km值相当于在切碎组织中发现的较低值。通过DOPAC形成速率评估DA摄取得到的Km值为2.3微摩尔。（使用了高底物值，因此在数据中较低的Km值不明显。）在先前通过黑质注射6-羟基多巴胺诱导纹状体损伤的动物中，不存在高Km值组分。评估了几种药理剂（苯海索、安非尼酸、安非他酮、诺米芬辛和哇巴因）。所有这些药剂都降低了切碎组织中DA的摄取，但与它们在突触体中的作用相比，效力有所降低。切碎组织中的高Km活性以及摄取抑制剂明显降低的效力，似乎源于更完整组织内部存在的扩散屏障。在匀浆或突触体中不存在这种屏障，因此可以看到单一的摄取过程。
