A technique for demonstrating the nerve supply of whole larynges

At present there is a large gap in our understanding of the exact distribution of sensory and motor nerves within the larynx. Gross dissection is only accurate for large-caliber nerves, whereas microscopic sections show only isolated examples of terminal branching. To fill this gap, we modified an old technique called Sihler's stain to process five whole canine larynges. The technique requires eight steps that take approximately 3 months. The result is an entire larynx in which muscle and cartilage are rendered translucent, while nerve is counterstained. Nerve branching can be continuously traced from the large-caliber nerves to the terminal branches. We found that the nerve patterns are surprisingly complex and contain frequent anastomotic networks. This technique is a powerful tool for studying certain neurolaryngological problems.