Spatial and dynamic changes in intracellular Ca2+ measured by confocal laser-scanning microscopy in bullfrog sympathetic ganglion cells.
Confocal laser scanning microscopy (CLSM) was used to record spatial and dynamic changes in the intracellular Ca2+ [(Ca2+]i) of bullfrog sympathetic ganglion cells in excised tissue or in culture. A CLSM utilizing Ar ion laser (488 nm) and recording fluo-3 fluorescence yielded the sliced image of ganglion cells, while conventional epifluorescence microscopy provided the cell image of a convex structure. A high K+ (50 mM) solution, caffeine (3-10 mM) and electrical stimulation (10-20 Hz, 0.5-10 s) caused a homogeneous increase in fluo-3 fluorescence with or without regional differences, possibly due to intracellular organelles and other constituents. Scanning a single horizontal line across the cytoplasm with He-Cd laser (325 nm) and recording indo-1 fluorescence demonstrated that the rate of rise in [Ca2+]i following action potentials depends on the distance from the cell membrane and on the cytoplasmic constituents, showing an inward spread of 'Ca(2+)-wave' at variable speeds of 17-219 microns/s. These results suggest that heterogeneity of the cytoplasmic structures and constituents affects dynamic and spatial changes of [Ca2+]i in response to stimuli in neurones. Such heterogenic changes in [Ca2+]i would better be studied by CLSM.
Kuba
K
K
Department of Physiology, Saga Medical School, Japan.
Hua
S Y
SY
Nohmi
M
M
D000814:Aniline Compounds; D000818:Animals; D001001:Anura; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D004558:Electric Stimulation; D005456:Fluorescent Dyes; D005728:Ganglia, Sympathetic / Q000166:cytology / Q000378:metabolism*; D007091:Image Processing, Computer-Assisted; D007425:Intracellular Membranes / Q000378:metabolism*; D007834:Lasers; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000378:metabolism*; D009994:Osmolar Concentration; D011188:Potassium / Q000494:pharmacology; D012680:Sensitivity and Specificity; D014966:Xanthenes
D000814:Aniline Compounds; D005456:Fluorescent Dyes; D014966:Xanthenes; C059715:Fluo-3; D002110:Caffeine; D011188:Potassium; D002118:Calcium