Young L H, Zaret B L, Barrett E J
Am J Physiol. 1987 Jun;252(6 Pt 2):H1070-6. doi: 10.1152/ajpheart.1987.252.6.H1070.
The transport of L-alanine and L-leucine into membrane vesicles isolated from mature canine ventricular myocardium was studied. Transport was assessed in purified sarcolemma and in vesicles differentially enriched either for sarcolemma or sarcoplasmic reticulum to further localize these transport systems. An imposed inward gradient of a NaNO3 stimulated uptake of L-alanine but not L-leucine by these vesicles. Amino acid uptake by these vesicles occurred into an osmotically active space. The stimulatory effect of Na+ on alanine transport was most striking in the purified sarcolemma vesicles, where Na+-stimulated alanine flux was 45 +/- 14 pmol X mg-1 X min-1. Furthermore, Na+-dependent alanine transport activity appeared to copurify with Na+-K+-ATPase activity, which served as a marker for sarcolemma membrane when these activities were compared in the three different membrane preparations. Leucine transport by sarcolemma was not altered by an imposed Na+ gradient. However, leucine uptake was a saturable function of extravesicular leucine and was inhibited by valine. In contrast, in sarcoplasmic reticulum membrane vesicles leucine uptake increased proportionately with increasing media leucine and was unaffected by valine. Our results demonstrate the feasibility of directly studying the transport of naturally occurring amino acids in membrane vesicles from mammalian heart, and the presence of Na+-dependent alanine transport system and a Na+-independent leucine transporter in the sarcolemma but not in sarcoplasmic reticulum of canine ventricular myocardium.
研究了L-丙氨酸和L-亮氨酸转运至从成年犬心室心肌分离出的膜囊泡中的过程。在纯化的肌膜以及分别富含肌膜或肌浆网的囊泡中评估转运情况，以进一步确定这些转运系统的位置。施加的NaNO₃内向梯度刺激了这些囊泡对L-丙氨酸的摄取，但未刺激对L-亮氨酸的摄取。这些囊泡对氨基酸的摄取发生在一个具有渗透活性的空间内。Na⁺对丙氨酸转运的刺激作用在纯化的肌膜囊泡中最为显著，其中Na⁺刺激的丙氨酸通量为45±14 pmol·mg⁻¹·min⁻¹。此外，当在三种不同的膜制剂中比较这些活性时，Na⁺依赖性丙氨酸转运活性似乎与Na⁺-K⁺-ATP酶活性共同纯化，而Na⁺-K⁺-ATP酶活性可作为肌膜的标志物。肌膜对亮氨酸的转运不受施加的Na⁺梯度的影响。然而，亮氨酸的摄取是囊泡外亮氨酸的饱和函数，并受到缬氨酸的抑制。相反，在肌浆网膜囊泡中，亮氨酸的摄取随培养基中亮氨酸的增加而成比例增加，且不受缬氨酸的影响。我们的结果证明了直接研究哺乳动物心脏膜囊泡中天然存在的氨基酸转运的可行性，以及犬心室心肌肌膜中存在Na⁺依赖性丙氨酸转运系统和Na⁺非依赖性亮氨酸转运体，而肌浆网中不存在。
