On-off regulation of hypothalamic GRF secretion by somatostatin

生长抑素(SRIF)是一种十四肽，最初是作为下丘脑促垂体素生长激素(GH)抑制因子分离出来的。1988年，我们在体内证明了内源性SRIF的突然停用可诱导GH释放因子(GRF)的分泌，并提示SRIF在下丘脑中起GRF抑制因子的作用。我们的研究项目得到了1990-1991年助学金的支持，目的是扩展我们自己的这个新概念。为此，我们建立了大鼠下丘脑孵育的方法。我们还建立了大鼠下丘脑正中隆起和灌流系统的分离神经末梢培养体系。GRF和SRIF用高度特异的RIA测定。为了去除SRIF，我们使用了硫醇试剂半胱胺和用蛋白A-琼脂糖从兔SRIF抗血清中分离出的丙种球蛋白部分。半胱胺以浓度和钙离子依赖的方式刺激孵育的下丘脑GRF基础分泌和K~(2+)诱导的GRF分泌。用Sep-Pak C18和反相高效液相色谱鉴定GRF分泌到培养基中。我们还证明，SRIF对GRF分泌的抑制在新生早期是不存在的，大约在出生后7-10天发展，这与大鼠脑垂体门静脉建立的时间相对应。这一发现支持SRIF对GRF分泌的开关调节的生理学意义。此外，我们还观察到，在培养液中加入抗SRIF丙种球蛋白可以增加下丘脑突触体内GRF的释放，并且周期性地注入半胱胺可以刺激灌流的大鼠下丘脑GRF的分泌。总而言之，这些发现支持SRIF在下丘脑水平上作为GRF抑制因子的概念。

Somatostatin (SRIF) is a tetradecapeptide originally isolated as a hypothalamic hypophysiotropic growth hormone (GH)-inhibiting factor. In 1988, we demonstrated that sudden withdrawal of endogenous SRIF induced secretion of GH-releasing factor (GRF) in vivo and suggested that SRIF acts as a GRF-inhibiting factor within the hypothalamus. Our research project, supported by a Grant-in-Aid 1990-1991, was to extend this novel concept of our own. For this porupose, we have-established the method of rat hypothalamic incubation. We have also developed culture system of isolated nerve endings of the stalk-median eminence and the perifusion system of rat hypothalami. GRF and SRIF were measured by highly specific RIAs. To deplete SRIF, we used a thiol agent cysteamine and a gamma-globulin fraction separated from rabbit SRIF antiserum by Protein A-Sepharose. An addition of cysteamine to the medium stimulated basal GRF secretion and enhanced K^+-evoked GRF secretion from incubated hypothalami in a concentration- and Ca^<2+>-dependent manner. GRF secreted into the medium was characterized by Sep-Pak C18 and reverse-phase HPLC. We have also demonstrated that SRIF inhibition of GRF secretion is absent in an early neonatal period and develops approximately 7-10 days after birth, which correspond the time when the hypophysial portal vessels are established in the rat. This finding supports physiological significance of the on off regulation of GRF secretion by SRIF. Furthermore, we have observed that an addition of anti-SRIF gamma-globulin to the medium augments GRF release from hypothalamic synaptosomes and that a periodicinfusion of cysteamine can stimulate GRF secretion from perifused rat hypothalami. Collectively, these findings support the concept that SRIF acts as a GRF-inhibiting factor at the level of the hypothalamus.