New approaches for studying the functional anatomy of the phallus in Lepidoptera

Abstract

Existing techniques for examining the everted vesica (endophallus) of Lepidoptera are based primarily on cuticular preparations macerated with a caustic solution for taxonomic study. These techniques destroy muscles and other soft tissue, thus studies of the functional anatomy of the skeletomuscular apparatus of the phallus are not possible. Injection of formaldehyde solution into the phallus of fresh specimens is proposed as a new approach for studying the intact anatomy of this structure. The new technique results in simultaneously everting and fixing the vesica, resulting in a better approximation of its functional shape. This method produces properly fixed tissues and the whole structure can be processed for various further studies, including histology sectioning, scanning electron microscopy, and confocal laser scanning microscopy. The commonly used stubs for scanning electron microscopes do not allow observation of the sample from all aspects. This problem was solved by the modification of a commercially available stub. The device allows 360° rotation of the phallus, and the concept can be applied for observation of other objects as well.