TO DEVELOP A NEW SYSTEM TO CONTIMUOUSLY MONITOR THE FUNCTIONS AT THE CELLULAR AND MOLECULAR LEVELS OF THE VASCULAR ENDOTHELIAL AND SMOOTH MUSCLE CELLS IN VIVO.

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1. THE DEVELOPMENT OF A NEW SYSTEM FOR MONITORING THE FUNCTIONS OF THE VASCULAR ENDOTHELIAL AND SMOOTH MUSCLE CELLS : The system we have developed has following characteristics ; (1) Using a grating and a chopper mirror driven by a stepping motor, it became possible to alternately (10 Hz) illuminate the tissue sample with 340 and 380 nm excitation lights for the measurement of intracellular calcium concentration, [Ca] i. (2) the chopper system is controlled through a personal computer by investigator designed software. (3) Fluorescence images are obtained using a chopper mirror and a SIT camera. A display flame is 640x400 dots and a 8 bit flame. (4) Fluorescence ratio (F340/F380) images can be obtained at 5 Hz. (5) It is possible to simultaneously record changes in the [Ca] i levels at 30 points in an image on the TV monitor. (6) Low cost for making the system.2. NEW FINDINGS OBTAINED USING THIS SYSTEM,FRONT-SURFACE FLUORIMETRY AND TECHNIQUES IN MOLECULAR BIOLOGY : (1) Using the newly developed technique, the simultaneous fluorimetry of fura-2 and diaminonaphthalene, we found the [Ca] i-NO production relationships in the endothelial cells in situ. (2) Using front-sueface fluorimetry, reverse transcription polymerase chain reaction, and the renal and coronary arteries, we found that both adrenomedullin and endothelin might act as autocrine/paracrione signal transmitters in these tissues.3. HUMANA PRESS (NJ : U.S.A) asked us to write a chapter "Measurement of [Ca] i in muscle strips using front-surface fluorimetry" of a book on "Calcium signaling protocols (edited by DG Lambert)", a part of the Methods in Molecular Biology series. It is in press.