A semi-in-vivo preparation for optical recording of the insect brain

We describe a method for optically recording neuronal activity from an intact insect brain, upon natural sensory stimulation. In this preparation, the head capsule of the honeybee, Apis mellifera, is isolated from the body while leaving the entire brain undamaged. In short, a hole is first cut into the cuticule to allow optical access to the brain and to allow the removal of tracheae and glands. Then the head is cut free and placed into a dye-loaded and cooled ringer solution in a staining chamber for 1 h. Subsequently, the head is fixed in a recording chamber, covered with a cover-slip, and imaged under the microscope with a cooled CCD camera. The whole preparation leaves the antennae dry, free to move, and functional throughout the experiment, allowing for natural odour stimulation of the olfactory system. Using calcium sensitive or potential sensitive dyes (calcium-green or RH795), we could record the processing of olfactory information at the glomerular level in the antennal lobe of the bee.