Micromanipulation with stereoscopic imaging

The neuronal organization of the hippocampus is extensively studied. However, the synaptic connections between neurons of the hilus and neurons of the granule cell layer are still debate. In the present study, we utilized automated micromanipulation technique combined with visual image processing to analyze the synaptic connections between hilar mossy cells or mossy fibers, and granule cells or basket cells in the granule cell layer. The stereoscopic microscope image of the dentate gyrus of coronal slice in adult rat brain was converted into a binary image. The area of the dentate gyrus is detected by genetic algorithms. This was matched with the template image made from a microscopic photo of cresyl violet-stained coronal section of the rat brain by parallel and rotation matching. A glass microelectrode inserted into the apex of the dentate gyrus by the micromanipulation system. Lipophilic fluorescent tracer DiI or DiD was injected into the hilus by a nanoinjector.