EMBRYONIC DEVELOPMENT OF SMOOTH MUSCLE CA2+ REGULATION

人们对平滑肌肉的发育知之甚少。 兴奋收缩耦合和松弛仍然不完全。 即使在成熟光滑的肌肉中也能理解。通过研究以下机制 他们发展出新的洞察力，可能会出现。透彻了解 正常发育将是解释异常原因所必需的 发育和病理情况。很多活动都是顺畅的 肌肉细胞(收缩或新陈代谢)受 胞浆游离钙浓度([Ca++]i)。[Ca++]i的变化可能是 通过成熟肌肉中张力的变化间接监测。处于胚胎阶段 细胞，收缩是很难或不可能监测的。最近的 引入一种无中断的方法来直接测量 小细胞中的[Ca++]i现在使这项研究变得可行。这个设计得很好的 方法使用荧光螯合剂(Quin-2)的预意酯(Quin-2/AM) 对钙具有很高的亲和力和选择性(Kd=115 nm)。蜂窝 酶使Quin-2/AM脱酯，从而释放羧基， 结合钙，使分子过于亲水而无法逃逸。案例的装订 会影响分子的荧光强度。的发展。 神经递质、神经肽和激素的受体 内脏或血管通畅的收缩或松弛信号 肌肉和钙的运输机制将被研究。这项提案将重点放在 鸡胆胚胎胆碱能受体发育的研究 平滑的肌肉。这种组织很容易拿到，也是最具特征的 到目前为止的胚胎平滑肌。将研究其发展过程。 在卵子和组织培养中。有利于组织培养的条件 适当的区分和适合Quin-2方法的将是 发展起来的。

Little is known about the development of smooth muscle. Excitation-contraction coupling and relaxation are still not fully understood even in mature smooth muscles. By studying the mechanisms as they develop new insights may emerge. A thorough understanding of the normal development will be necessary to interpret causes of abnormal development and pathological conditions. Much of the activity of smooth muscle cells (contractile or metabolic) is affected by changes in the cytoplasmic free Ca concentration ([Ca++]i). Changes in the [Ca++]i can be indirectly monitored by changes in tension in mature muscle. In embryonic cells, contractions are difficult or impossible to monitor. The recent introduction of a nondisruptive method to directly measure changes in [Ca++]i in small cells now makes this study feasible. This well-designed method uses a permeant ester (Quin-2/AM) of a fluorescent chelator (Quin-2) with a high affinity and selectivity for calcium (Kd=115 nM). Cellular enzymes de-esterify Quin-2/AM thereby freeing the carboxyl groups which bind Ca and make the molecule too hydrophilic to escape. The binding of Ca affects the fluorescence intensity of the molecule. The development of receptors for neurotransmitters, neuropeptides and hormones that initiate signals for contraction or relaxation of visceral or vascular smooth muscles and Ca transport mechanisms will be studied. This proposal focuses on the development of cholinergic receptors in embryonic chicken gizzard smooth muscle. This tissue is easily accessible and the best characterized embryonic smooth muscle to date. The developmental process will be studied in ovo and in tissue cultures. Tissue culture conditions favoring appropriate differentiation and suitable for the Quin-2 method will be developed.