The Subgenual Organ Complex in Stick Insects: Functional Morphology and Mechanical Coupling of a Complex Mechanosensory Organ

Leg chordotonal organs in insects show different adaptations to detect body movements, substrate vibrations, or airborne sound. In the proximal tibia of stick insects occur two chordotonal organs: the subgenual organ, a highly sensitive vibration receptor organ, and the distal organ, of which the function is yet unknown. The distal organ consists of a linear set of scolopidial sensilla extending in the tibia in distal direction toward the tarsus. Similar organs occur in the elaborate hearing organs in crickets and bushcrickets, where the auditory sensilla are closely associated with thin tympanal membranes and auditory trachea in the leg. Here, we document the position and attachment points for the distal organ in three species of stick insects without auditory adaptations (Ramulus artemis, Sipyloidea sipylus, and Carausius morosus). The distal organ is located in the dorsal hemolymph channel and attaches at the proximal end to the dorsal and posterior leg cuticle by tissue strands. The central part of the distal organ is placed closer to the dorsal cuticle and is suspended by fine tissue strands. The anterior part is clearly separated from the tracheae, while the distal part of the organ is placed over the anterior trachea. The distal organ is not connected to a tendon or muscle, which would indicate a proprioceptive function. The sensilla in the distal organ have dendrites oriented in distal direction in the leg. This morphology does not reveal obvious auditory adaptations as in tympanal organs, while the position in the hemolymph channel and the direction of dendrites indicate responses to forces in longitudinal direction of the leg, likely vibrational stimuli transmitted in the leg’s hemolymph. The evolutionary convergence of complex chordotonal organs with linear sensilla sets between tympanal hearing organs and atympanate organs in stick insects is emphasized by the different functional morphologies and sensory specializations.

Neuronal Innervation of the Subgenual Organ Complex and the Tibial Campaniform Sensilla in the Stick Insect Midleg

Mechanosensory organs in legs play are crucial receptors in the feedback control of walking and in the detection of substrate-borne vibrations. Stick insects serve as a model for the physiological role of chordotonal organs and campaniform sensilla. This study documents, by axonal tracing, the neural innervation of the complex chordotonal organs and groups of campaniform sensilla in the proximal tibia of the midleg in Sipyloidea sipylus. In total, 6 nerve branches innervate the different sensory structures, and the innervation pattern associates different sensilla types by their position. Sensilla on the anterior and posterior tibia are innervated from distinct nerve branches. In addition, the variation in innervation is studied for five anatomical branching points. The most common variation is the innervation of the subgenual organ sensilla by two nerve branches rather than a single one. The fusion of commonly separated nerve branches also occurred. However, a common innervation pattern can be demonstrated, which is found in >75% of preparations. The variation did not include crossings of nerves between the anterior and posterior side of the leg. The study corrects the innervation of the posterior subgenual organ reported previously. The sensory neuroanatomy and innervation pattern can guide further physiological studies of mechanoreceptor organs and allow evolutionary comparisons to related insect groups.

Neuronal Regression of Internal Leg Vibroreceptor Organs in a Cave-Dwelling Insect (Orthoptera: Rhaphidophoridae: Dolichopoda araneiformis)

Animals' adaptations to cave habitats generally include elaboration of extraoptic senses, and in insects the receptor structures located on the legs are supposed to become more prominent in response to constant darkness. The receptors for detecting substrate vibrations are often highly sensitive scolopidial sensilla localized within the legs or the body. For troglobitic insects the evolutionary changes in vibroreceptor organs have not been studied. Since rock is an extremely unfavorable medium for vibration transmission, selection on vibration receptors may be weakened in caves, and these sensory organs may undergo regressive evolution. We investigated the anatomy of the most elaborate internal vibration detection system in orthopteroid insects, the scolopidial subgenual organ complex in the cave cricket Dolichopoda araneiformis (Orthoptera: Ensifera: Rhaphidophoridae). This is a suitable model species which shows high levels of adaptation to cave life in terms of both phenotypic and life cycle characteristics. We compared our data with data on the anatomy and physiology of the subgenual organ complex from the related troglophilic species Troglophilus neglectus. In D. araneiformis, the subgenual organ complex contains three scolopidial organs: the subgenual organ, the intermediate organ, and the accessory organ. The presence of individual organs and their innervation pattern are identical to those found in T. neglectus, while the subgenual organ and the accessory organ of D. araneiformis contain about 50% fewer scolopidial sensilla than in T. neglectus. This suggests neuronal regression of these organs in D. araneiformis, which may reflect a relaxed selection pressure for vibration detection in caves. At the same time, a high level of overall neuroanatomical conservation of the intermediate organ in this species suggests persistence of the selection pressure maintaining this particular organ. While regressive evolution of chordotonal organs has been documented for insect auditory organs, this study shows for the first time that internal vibroreceptors can also be affected.

The subgenual organ complex in the cave cricket Troglophilus neglectus (Orthoptera: Rhaphidophoridae): comparative innervation and sensory evolution

Comparative studies of the organization of nervous systems and sensory organs can reveal their evolution and specific adaptations. In the forelegs of some Ensifera (including crickets and tettigoniids), tympanal hearing organs are located in close proximity to the mechanosensitive subgenual organ (SGO). In the present study, the SGO complex in the non-hearing cave cricket Troglophilus neglectus (Rhaphidophoridae) is investigated for the neuronal innervation pattern and for organs homologous to the hearing organs in related taxa. We analyse the innervation pattern of the sensory organs (SGO and intermediate organ (IO)) and its variability between individuals. In T. neglectus , the IO consists of two major groups of closely associated sensilla with different positions. While the distal-most sensilla superficially resemble tettigoniid auditory sensilla in location and orientation, the sensory innervation does not show these two groups to be distinct organs. Though variability in the number of sensory nerve branches occurs, usually either organ is supplied by a single nerve branch. Hence, no sensory elements clearly homologous to the auditory organ are evident. In contrast to other non-hearing Ensifera, the cave cricket sensory structures are relatively simple, consistent with a plesiomorphic organization resembling sensory innervation in grasshoppers and stick insects.

Behaviour-locked signal analysis reveals weak 200–300 Hz comb vibrations during the honeybee waggle dance

Waggle-dancing honeybees produce vibratory movements that may facilitate communication by indicating the location of the waggle dancer. However, an important component of these vibrations has never been previously detected in the comb. We developed a method of fine-scale behavioural analysis that allowed us to analyze separately comb vibrations near a honeybee waggle dancer during the waggle and return phases of her dance. We simultaneously recorded honeybee waggle dances using digital video and laser-Doppler vibrometry, and performed a behaviour-locked Fast Fourier Transform analysis on the substratum vibrations. This analysis revealed significantly higher-amplitude 200–300 Hz vibrations during the waggle phase than during the return phase (P=0.012). We found no significant differences in the flanking frequency regions between 100–200 Hz (P=0.227) and 300–400 Hz (P=0.065). We recorded peak waggle phase vibrations from 206 to 292 Hz (244+/−28 Hz; mean +/− s. d., N=11). The maximum measured signal - noise level was +12.4 dB during the waggle phase (mean +5.8+/−2.7 dB). The maximum vibrational velocity, calculated from a filtered signal, was 128 microm s(−)(1) peak-to-peak, corresponding to a displacement of 0.09 microm peak-to-peak at 223 Hz. On average, we measured a vibrational velocity of 79+/−28 microm s(−)(1) peak-to-peak from filtered signals. These signal amplitudes overlap with the detection threshold of the honeybee subgenual organ.