A mesoscopic technique for the study of the development of the peripheral system in rat foetuses

In order to study the morphogenesis of the nervous system in the rat an acetylcholinesterase in toto method for staining nervous tissue in rat foetuses was developed.

Rat foetuses of 14-22 days are fixed "en bloc" for 24 hours in a cold sucrose-formol solution. Fixed specimens are rinsed for 2 days in cold 0.22 M sucrose in a sodiumcacodylate buffer (pH 7.2). The specimens are cut (mid-)sagittally with the aid of a razor-blade, and incubated in a medium of acetylthiocholine iodide in acetate buffer (pH 5.0). Then, dehydration in glycerine/water mixtures of increasing glycerine content follows. The specimens may be stored in pure glycerine or embedded in epoxy-resin blocks and can be studied under a binocular dissecting microscope. In using this in toto staining method both the continuity of the central and peripheral parts of the nervous system as well as details up to the level of individual perikarya and motor endplates are preserved. With this mesoscopic method the three-dimensional architecture of the peripheral nervous system and its topological relations to other structures can be studies in one specimen. The exact procedure and the results as well as a method for embedding specimens in epoxy-resin blocks for teaching purposes are described. The advantages of this mesoscopic staining method for foetuses are discussed.