Subcellular Stucture in Glial Networks

Office Of The Director, Nih

Investigators

Abstract

The aim of this project is to study ultrastructural features at the supramolecular scale in glial cells in the zones where mitochondria and endoplasmic reticulum adjoin. Interactions between these two types of organelle are associated with calcium signaling, through which glial cells communicate with each other during central nervous system development and synaptic activity. To preserve glial networks in their physiological state, high-pressure freezing is being used. Cultured cells are first grown on specially coated gold specimen carriers, which are then frozen at a pressure of >2000 atmospheres at liquid nitrogen temperature to avoid ice crystal damage. Specimens are then freeze-substituted with osmium fixative before plastic embedding, sectioning and staining. In the future, suitably well-preserved specimens are to be imaged in a digital energy-filtering transmission electron microscope. Tilt series will be then be recorded to obtain a three-dimensional reconstruction of the zones surrounding mitochondria in the glial cells.