High resolution ultrastructural mapping of total calcium: electron spectroscopic imaging/electron energy loss spectroscopy analysis of a physically/chemically processed nerve-muscle preparation
| Autor: | Jacopo Meldolesi, Roberta Pezzati, F. Grohovaz, Francesca Torri Tarelli, Mario Bossi |
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| Rok vydání: | 1996 |
| Předmět: |
Analytical chemistry
Neuromuscular Junction chemistry.chemical_element Calcium In Vitro Techniques Synaptic vesicle Microanalysis Neuromuscular junction medicine Animals Terminal cisternae Motor Neurons Organelles Multidisciplinary Histocytochemistry Endoplasmic reticulum Electron energy loss spectroscopy Histological Techniques Rana pipiens Sodium Sarcoplasmic Reticulum medicine.anatomical_structure chemistry Biophysics Ultrastructure Potassium Synaptic Vesicles Electron Probe Microanalysis Research Article |
| Zdroj: | Proceedings of the National Academy of Sciences of the United States of America. 93(10) |
| ISSN: | 0027-8424 |
| Popis: | We report on a procedure for tissue preparation that combines thoroughly controlled physical and chemical treatments: quick-freezing and freeze-drying followed by fixation with OsO4 vapors and embedding by direct resin infiltration. Specimens of frog cutaneous pectoris muscle thus prepared were analyzed for total calcium using electron spectroscopic imaging/electron energy loss spectroscopy (ESI/EELS) approach. The preservation of the ultrastructure was excellent, with positive K/Na ratios revealed in the fibers by x-ray microanalysis. Clear, high-resolution EELS/ESI calcium signals were recorded from the lumen of terminal cisternae of the sarcoplasmic reticulum but not from longitudinal cisternae, as expected from previous studies carried out with different techniques. In many mitochondria, calcium was below detection whereas in others it was appreciable although at variable level. Within the motor nerve terminals, synaptic vesicles as well as some cisternae of the smooth endoplasmic reticulum yielded positive signals at variance with mitochondria, that were most often below detection. Taken as a whole, the present study reveals the potential of our experimental approach to map with high spatial resolution the total calcium within individual intracellular organelles identified by their established ultrastructure, but only where the element is present at high levels. |
| Databáze: | OpenAIRE |
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