Study On Mechanism Of Secretion Regulation In Gonadotrope Endocrine Cells And DRG Neuron Cells

Secretion forms the basic for Signal transmission between tne neurons and hormone release from endocrine cells,which are important in maintaining normal physiological function of organisms. Deficiency in hormone secretion from gonadotropes will impact development and mature of sex. The study on the mechanism of secretion and its regulation is important to further understand the essence of secretion and to treat secretion-relevant diseases. In this study, we applied advanced biophysical techniques,such as microfluoremetric technique, membrane capacitance measurement, modeling and statistic. Systemic investigation on the mechanism of secretion and its regulation have been carried out in the rat gonadotropes. We have previously reported that protein kinase C (PKC) activation sensitizes the Ca²⁺ sensor for exocytosis in pituitary gonadotropes. We propose that PKC sensitizes exocytosis by reducing the number of calcium binding sites on the Ca sensor (from three to two) without significantly altering the Ca²⁺-binding kinetics. The reduction in the umber of Ca²⁺-binding steps lowers the threshold for release and up-regulates release of fusion-competent vesicles distant from Ca²⁺ channels. Thus, our results seem to point to a novel target of action, distinct from Munc13. Main result of the Study are as follows: (1) It has been widely report that PKC activation sensitizes the Ca²⁺ sensor for exocytosis. Recently, we have shown that PKC activation sensitizes the Ca²⁺ sensor for exocytosis in pituitary gonadotropes. We proved the Ca²⁺ sensor of gonadotropes contains some isoform of synaptotagmin whose Ca²⁺ sensitivity can be modulated by phosphorylation. It is not known whether the number of binding sites on the Ca²⁺ sensor can be modulated or not. With two or three Ca²⁺-binding sites models (Scheme 1 and 2), we simulated the exocytotic burst recorded from control cells and PMA treated cells. Closer inspection of the onset of the response immediately after the flash shows that Scheme 1 reproduces the sigmoidicity better than Scheme 2. We therefore fitted all the experimental responses to both models. This difference is also clear from the cumulative histogram plot. To further confirm the modulatory mechanism of PMA on Ca2+ sensing, we employed the so-called "Ca2+ ramp"technique. The time courses of the Cm response to a Ca2+ ramp also fitted well to Scheme 1 for control and Scheme 2 for PMA-treated cells. We think the reduction in the number of Ca2+-binding steps lowers the threshold for release and up-regulates release of fusion-competent vesicles distant from Ca channels. (2) A controversy surrounds the issue of whether an additional mechanism exists to account for the initiation of transmitter release. To understand the characteristic of the voltage-dependent Cm change, a suitable model is highly desirable. One aim is first to derive a model that can be used to describe the experimental data of the voltage-dependent Cm signals, and to predict the underlying mechanism of the Ca2+-independent exocytosis and endocytosis in the somata of DRG neurons. The current study will help us to further validate the other types of cells with CIVDS and to understand its physiological significance. The model may also provide more valuable information for other modeling work. (3) In order to develop voltage-dependent secretion, a softare based on MATLAB was designed mainly by me. Now this softeare was supported by HUST fund and we will try hard to perfect it and applied it the area of teaching.