Research On The Structural And Functional Changes Of The Skeletal Muscle During Ca～(2+) Release From Sarcoplasmic Reticulum By Calcium Induced Spark

The course of skeletal muscular contraction includes latency, systole and period of relaxation. During the course, the skeletal muscle must have a series of structural and functional changes. In the skeletal muscular fiber, the T-tube, sarcoplasmic reticulum(SR) and Ca2+ ion play a key role during the excitation- contraction (e-c) coupling course, but it was still not clear what was the mechanism of Ca2+ releasing and what happened structurally and functionally during SR releasing Ca2+ ion. All these concern are very hot aspects in cell biology, physiology and biophysics. The time course of skeletal muscular e-c coupling is very short (measured at millisecond), but the conventional method of chemical fixation (measured at minute) can't reserve the ultrastructure as functional changes quickly. Because of the restriction of these researching methods, conventional measure of studying on cellular ultrastructure can't obtain the skeletal muscular structural changes in millisecond level. It is still at the beginning of studying the ultrastructural and morphological changes during the course of skeletal muscular contraction and there are no influential reports in these fields. However, it is possible to study the Real-time ultra structural changes on every time spots during the course.On the point of the mechanism of skeletal muscular e-c coupling, foreign scholar had put forward two hypotheses via physiological experiments: direct contact signal transduction (DCT for short) and calcium induced calcium release (CICR for short). DCT considers when the e-c coupling takes place, SR membrane depolarized and induced electricity action, so an allosteric signal is transmitted to the calcium release channels (RyR) in the membrane of SR and then the RyR opens, which makes the skeletal muscular fibers contract. That means e-c coupling should induce changes of SR membrane in the moment of Ca + released from SR when the skeletal muscle contracts. This hypothesis had been proved by some experiments. CICR considers it is within two milliseconds which is the prophase of e-c coupling after electric stimulation, the extracellular calcium are induced to flow in through depolarized L-type channels (DHPR) in the membrane of T-tube, resulting in the calcium concentration rising just out of the SR membrane, which cause the opening of the RyR, then the calcium release gradually from SR.At the end of 1980s, foreign scholars studied the mechanism of calcium releasing from SRduring the e-c coupling through ultra-hypothermy quick cryofixation, transmission electronic microscopy(TEM) and electronic probe X-radiation micro-analysis(EPXMA), but it was very difficult to fix the skeletal muscle at the millisecond level to obtain the functional changes of the skeletal muscular e-c coupling, so they didn't acquire real-time ultrastructural changes, calcium concentration and distribution situs changes. These problems could be resolved by two-way infrared detectors (developed in our lab) with a computer to control the signal of electric stimulation and ultra-hypothermy quick freezing fixation at millisecond level technique, freezing displacement and TEM. We could study the ultrastructural time-morphological changes of SR and T-tube of the skeletal muscle in latency and systole during the e-c coupling course when the skeletal muscle is fixed on different time spots (Oms to 24ms) after electrical stimulated, then we would acquire the morphological structure, calcium concentration and distribution situs changes at the moment of calcium releasing from SR when induced. Combination with confocal microscopy, the entireness of Ca2+spark could be observed on submicroscopic level, and its mechanism would be analyzed. Colligating the two methods, the mechanism of calcium releasing from SR would be proved in morphology when the e-c coupling takes place.To study the controlling of calcium channels in sarcolemma could make clear the ultrastructural and functional changes in e-c coupling result from calcium induced calcium release. Nefidipine is a blocker of DHP...