Wakade A R, Wakade T D, Malhotra R K
Department of Pharmacology, State University of New York, Brooklyn.
J Neurochem. 1988 Sep;51(3):820-9. doi: 10.1111/j.1471-4159.1988.tb01817.x.
The functional integrity of adrenal chromaffin storage vesicles was studied in the perfused rat adrenal gland subjected to intense exocytosis. Continuous perfusion with 55 mM K+-Krebs solution produced a large and uninterrupted secretion of catecholamines. Total amounts secreted within 45 min were 4.66 micrograms and represented almost 30% of the total tissue catecholamine content. If perfusion with excess K+ was extended to 90 min, the secretion increased further to 5.76 micrograms. Despite such a large secretory response, the catecholamine content of the K+-stimulated adrenal medulla was comparable to that of unstimulated control, suggesting an enhanced resynthesis to maintain the normal levels. Pretreatment of rats with alpha-methyl-p-tyrosine, and including this agent in the perfusion medium during stimulation with K+, caused a marked reduction in catecholamine content. The degree of depletion depended on the extent of stimulation with K+ (45% in 45 min and 60% in 90 min). Although depleted catecholamine stores did not show spontaneous recovery in 2 h, inclusion of tyrosine, L-3,4-dihydroxyphenylalanine or dopamine (but not epinephrine or norepinephrine) completely restored the catecholamine content of previously depleted adrenal medulla. Repletion achieved by tyrosine was time dependent (evident in 30 min and maximum in 2 h) and blocked by alpha-methyl-p-tyrosine but not by calcium deprivation. The ratio of epinephrine to norepinephrine remained constant during various stages of the experiment, suggesting both types of vesicles were equally affected by different treatments. The secretory response (10 Hz for 30 s) was unaffected even though tissue catecholamine stores were significantly depleted (50%). In summary, we have demonstrated that catecholamine content of the isolated perfused adrenal gland can be reduced by stimulation of exocytotic secretion in the presence of tyrosine hydroxylase inhibitor. Since the depleted stores can be fully refilled by synthesis of catecholamines from its precursors, it is suggested that chromaffin vesicles may be reutilized for the purpose of synthesis, storage, and secretion of adrenal medullary hormones.
在经历强烈胞吐作用的灌注大鼠肾上腺中，研究了肾上腺嗜铬储存囊泡的功能完整性。用55 mM K⁺ - Krebs溶液持续灌注可产生大量且不间断的儿茶酚胺分泌。45分钟内分泌的总量为4.66微克，几乎占组织儿茶酚胺总含量的30%。如果用过量K⁺灌注延长至90分钟，分泌量进一步增加至5.76微克。尽管有如此大的分泌反应，但K⁺刺激的肾上腺髓质中的儿茶酚胺含量与未刺激的对照相当，表明再合成增强以维持正常水平。用α - 甲基 - p - 酪氨酸预处理大鼠，并在K⁺刺激期间将该试剂包含在灌注培养基中，导致儿茶酚胺含量显著降低。消耗程度取决于K⁺刺激的程度（45分钟内为45%，90分钟内为60%）。尽管耗尽的儿茶酚胺储存库在2小时内未显示出自发恢复，但包含酪氨酸、L - 3,4 - 二羟基苯丙氨酸或多巴胺（但不是肾上腺素或去甲肾上腺素）可完全恢复先前耗尽的肾上腺髓质中的儿茶酚胺含量。酪氨酸实现的补充是时间依赖性的（30分钟时明显，2小时时达到最大值），并被α - 甲基 - p - 酪氨酸阻断，但不被钙剥夺阻断。在实验的各个阶段，肾上腺素与去甲肾上腺素的比率保持恒定，表明两种类型的囊泡受到不同处理的影响相同。即使组织儿茶酚胺储存库显著耗尽（50%），分泌反应（10 Hz持续30秒）也不受影响。总之，我们已经证明，在酪氨酸羟化酶抑制剂存在的情况下，通过刺激胞吐分泌可以降低分离的灌注肾上腺中的儿茶酚胺含量。由于耗尽的储存库可以通过从其前体合成儿茶酚胺而完全重新填充，因此建议嗜铬囊泡可重新用于肾上腺髓质激素的合成、储存和分泌。
