Zonal heterogeneity of calcium distribution in rat hepatocytes: an electron microscopic study with a combined glutaraldehyde-osmium-pyroantimonate technique

Potassium pyroantimonate in combination with osmium tetroxide is an excellent technique for investigation of the subcellular distribution of calcium ions in glutaraldehyde-fixed liver tissue. Under appropriate incubation conditions potassium ions are replaced by calcium ions to form an insoluble electron-dense calcium antimonate precipitate. The presence of calcium within this antimonate precipitate is confirmed by the electron spectroscopic imaging (ESI) technique. Chelator treatment with EGTA [ethylene glycol-bis-(beta-amino-ethyl ether) N',N',N'-tetraacetic acid] is usually used as a control, preventing the precipitation of calcium. In our investigation, computerized image analysis revealed a gradient of the calcium distribution from the periportal to the pericentral area. A finely dispersed precipitate was found in the mitochondrial matrix and in the euchromatin of the nuclei of the periportal hepatocytes, and in the organelles of the pericentral area a coarser deposit in lower concentration was recognized. Our findings indicate that periportal and pericentral hepatocytes retain their zonal characteristics also with respect to calcium concentration in mitochondria and nuclei.