Pit Organs Research Articles

Embryonic development and cranial ossification of the Japanese Aodaishō, Elaphe climacophora (Serpentes: Colubridae): with special reference to the prootic bone and auditory evolution in snakes.

Snakes show remarkably deviated "body plan" from other squamate reptiles. In addition to limb loss, they have accomplished enormous anatomical specialization of the skull associated with the pit organs and the reduction of the tympanic membranes and auditory canals in the outer ears. Despite being the most diverse group of snakes, our knowledge of the embryonic staging for organogenesis and cranial ossification has been minimal for Colubridae. Therefore, in the present observation, we provide the first embryonic description of the Japanese rat snake Elaphe climacophora. We based our study on the Standard Event System (SES) for external anatomical characters and on a description of the cranial ossification during post-ovipositional development. We further estimated the relative ossification timing of each cranial bony element and compared it with that of selected other snakes, lizards, turtles, and crocodilians. The present study shows that the relative ossification timing of the palatine and pterygoid bones is relatively early in squamates when compared to other reptiles, implying the developmental integration as the palate-pterygoid complex in this clade and functional demands for the unique feeding adaptation to swallow large prey with the help of their large palatine and pterygoid teeth. Furthermore, unlike in species with pit organs, the prootic bone of Ela. climacophora is expanded to provide articulation with the supratemporal, thereby contributing to the hearing system by detecting substrate vibration. We also demonstrate that the relative timing of the prootic ossification is significantly accelerated in colubrids compared to snakes with pit organs. Our finding suggests that the temporal changes of the prootic ossification underpin the evolution of the perception of the ground-bourne sound signals among snakes.

Comparative SEM Study of Sensilla and Tyloid Structures in the Antennae of Vespinae (Hymenoptera: Vespidae).

This study investigates the distribution, morphology, and potential functions of antennal sensilla in various wasp species, including Dolichovespula flora, D. intermedia, Vespula structor, Vl. vulgaris, Provespa barthelemyi, Vespa bicolor, V. ducalis, V. mocsaryana, and V. velutina var. nigothorax. The study thoroughly analyzes the antennal structure of these species, representing all four genera of the yellow-jacket and hornet subfamily Vespinae. Using scanning electron microscopy (SEM), the study identifies a total of nineteen types of sensilla, including sensilla trichodea (ST-I, ST-II, ST-III), sensilla campaniform (SCF-I, SCF-II, SCF-III), pit organs (SCO-I, SCO-II, and SA), sensilla placodea (SP-I, SP-II), sensilla chaetica (SCH-I, SCH-II), sensilla basiconica (SB-I, SB-II), sensilla agmon (SAG-I, SAG-II), and sensilla coelocapitular (SCA). Additionally, tyloids were observed in the males of seven species, except for Vl. structor and Vl. vulgaris. The study provides insights into these sensilla types' morphology, abundance, and distribution. It discusses the variations in sensilla morphology among different species and the presence of gender-specific sensilla. This study provides new data about the morphology and distribution patterns of sensilla and tyloid.

Molecular Basis of CO2 Sensing in Hyphantria cunea.

Carbon dioxide (CO2) released by plants can serve as a cue for regulating insect behaviors. Hyphantria cunea is a widely distributed forestry pest that may use CO2 as a cue for foraging and oviposition. However, the molecular mechanism underlying its ability to sense CO2 has not been elucidated. Our initial study showed that CO2 is significantly attractive to H. cunea adults. Subsequently, 44 H. cunea gustatory receptors (GRs) were identified using transcriptome data, and 3 candidate CO2 receptors that are specifically expressed in the labial palps were identified. In vivo electrophysiological assays revealed that the labial palp is the primary organ for CO2 perception in H. cunea, which is similar to findings in other lepidopteran species. By using the Xenopus oocyte expression system, we showed that the HcunGR1 and HcunGR3 co-expressions produced a robust response to CO2, but HcunGR2 had an inhibitory effect on CO2 perception. Finally, immunohistochemical staining revealed sexual dimorphism in the CO2-sensitive labial pit organ glomerulus (LPOG). Taken together, our results clarified the mechanism by which H. cunea sense CO2, laying the foundation for further investigations into the role of CO2 in the rapid spread of H. cunea.

Glomerular organization in the antennal lobe of the oriental armyworm Mythimna separata

In insects, the number and specificity of antennal lobe glomeruli often reflect the number and diversity of expressed chemosensory genes, which are linked to its ecological niche and specific olfactory needs. The oriental armyworm, Mythimna separata, is an important and common lepidopteran pest of cereal crops. Given its reliance on the olfactory system for crucial behaviors, understanding the evolutionary potential of this system requires a thorough characterization of the anatomical structure of the primary olfactory center. Here, we systematically identified all antennal lobe glomeruli of M. separata based on synaptic antibody immunostaining and mass staining of the olfactory sensory neurons. A total of 69 glomeruli were identified in females and 65 in males, and an intuitive nomenclature based on glomerular positions was applied. Our findings uncovered some sex-specific glomeruli in this species. There were ten female-specific glomeruli and three male-specific glomeruli, except for the macroglomerular complex (MGC) units, with a notable observation that the female labial pit organ glomerulus was larger than its male counterpart. Additionally, we identified four antennal-lobe tracts (ALTs) and retrograde labeling from the calyx revealed that all glomeruli were innervated by the medial ALT projection neurons. The comparison of the olfactory system structures between M. separata and sympatric moths supports their evolutionary convergence in noctuid moths. These results collectively lay the foundation for future studies on olfactory processing in M. separata.

A bifurcation integrates information from many noisy ion channels and allows for millikelvin thermal sensitivity in the snake pit organ

A bifurcation integrates information from many noisy ion channels and allows for millikelvin thermal sensitivity in the snake pit organ

A bifurcation integrates information from many noisy ion channels and allows for milli-Kelvin thermal sensitivity in the snake pit organ

In various biological systems, information from many noisy molecular receptors must be integrated into a collective response. A striking example is the thermal imaging organ of pit vipers. Single nerve fibers in the organ reliably respond to milli-Kelvin (mK) temperature increases, a thousand times more sensitive than their molecular sensors, thermo-transient receptor potential (TRP) ion channels. Here, we propose a mechanism for the integration of this molecular information. In our model, amplification arises due to proximity to a dynamical bifurcation, separating a regime with frequent and regular action potentials (APs), from a regime where APs are irregular and infrequent. Near the transition, AP frequency can have an extremely sharp dependence on temperature, naturally accounting for the thousand-fold amplification. Furthermore, close to the bifurcation, most of the information about temperature available in the TRP channels' kinetics can be read out from the times between consecutive APs even in the presence of readout noise. A key model prediction is that the coefficient of variation in the distribution of interspike times decreases with AP frequency, and quantitative comparison with experiments indeed suggests that nerve fibers of snakes are located very close to the bifurcation. While proximity to such bifurcation points typically requires fine-tuning of parameters, we propose that having feedback act from the order parameter (AP frequency) onto the control parameter robustly maintains the system in the vicinity of the bifurcation. This robustness suggests that similar feedback mechanisms might be found in other sensory systems which also need to detect tiny signals in a varying environment.

Embryonic development of the neotropical pit viper Bothrops atrox (Serpentes: Viperidae: Crotalinae), with emphasis on pit organ morphogenesis and its evolution in snakes.

Bothrops atrox is a pit viper with a loreal pit organ, and its embryological development remains undescribed. Here, we provide a comprehensive description of the embryology of B. atrox, focusing on the loreal pit organ and cephalic scales. We characterized 13 developmental stages of B. atrox based on external features consistent with the embryogenesis of previously described snake species. The loreal pit organ originates from the circumorbital region and migrates to its final position. In Crotalinae, the pit organ first becomes visible at stage 28, whereas in Pythonidae labial, pit organs appear at Stage 35. Pit organs evolved independently three times in Serpentes, encompassing Boidae, Pythonidae, and Crotalinae. Boidae lacks embryological information for pit organs. Furthermore, we observed that head scalation onset occurs at Stage 33 in B. atrox, with fusion of scales surrounding the loreal pit organ. The embryology of pit organs in Pythonidae and Boidae species remains poorly understood. Our detailed embryological descriptions are critical for proposing developmental scenarios for pit organs and guiding future research on these structures.

Microstructural comparison of pit organ between Hybrid sturgeon and Chinese sturgeon Acipenser sinensis

The present study was conducted to investigate the distribution of pit organ in the critically endangered Chinese sturgeon (Acipenser sinensis), the similarities and differences in the microstructure of pit organ between Hybrid sturgeon and Chinese sturgeon, and to provide a histological basis for studying the pit organ in sturgeon species. We used hematoxylin-eosin (HE) staining, nissl staining, multi-layer fine scanning, and scanning electron microscopy to compare and analyse the microstructure of pit organ between Hybrid sturgeon and Chinese sturgeon. The results showed that the structure of pit organ of Hybrid sturgeon and Chinese sturgeon are similar. The distribution density of pit organ in different parts of Chinese sturgeon was different, and the density of pit organ on the ventral surface of the snout was the highest. Scanning electron microscopy revealed that the pit organ of Hybrid sturgeon and Chinese sturgeon were densely packed and about 500–1000 μm wide. Nissl staining revealed that more peripheral nerves are distributed under the sensory and support cells of the pit organ. Based on previous research and our results, the increase in electrical receptivity of sturgeons with individual development is not only due to the increase in the number of inductive receptors, but may also be related to the increase in the width of the pit organ.

A genome assembly for the southern Pacific rattlesnake, Crotalus oreganus helleri, in the western rattlesnake species complex.

Rattlesnakes play important roles in their ecosystems by regulating prey populations, are involved in complex coevolutionary dynamics with their prey, and exhibit a variety of unusual adaptations, including maternal care, heat-sensing pit organs, hinged fangs, and medically-significant venoms. The western rattlesnake (Crotalus oreganus) is one of the widest ranging rattlesnake species, with a distribution from British Columbia, where it is listed as threatened, to Baja California and east across the Great Basin to western Wyoming, Colorado and New Mexico. Here, we report a new reference genome assembly for one of six currently recognized subspecies, C. oreganus helleri, as part of the California Conservation Genomics Project (CCGP). Consistent with the reference genomic sequencing strategy of the CCGP, we used Pacific Biosciences HiFi long reads and Hi-C chromatin-proximity sequencing technology to produce a de novo assembled genome. The assembly comprises a total of 698 scaffolds spanning 1,564,812,557 base pairs, has a contig N50 of 64.7 Mb, a scaffold N50 of 110.8 Mb, and BUSCO complete score of 90.5%. This reference genome will be valuable for studies on the genomic basis of venom evolution and variation within Crotalus, in resolving the taxonomy of C. oreganus and its relatives, and for the conservation and management of rattlesnakes in general.

Sensory Structures on the Antenniform Legs of Whip Spider, Phrynichus phipsoni (Arachnida, Amblypygi), from the Indian State of Goa: Scanning Electron Microscopic Elucidation

Amblypygi have a tropical and sub-tropical distribution and their first of the four pairs of legs, is modified as feelers. Their activity period and dark habitats render visual cues ineffective and they rely on non-visual sensory perception. There is agreement among researchers that the whip spiders invest in olfactory and tactile senses imparted by sensory structures concentrated on the tarsal segment of the first pair of long, well-articulated non-ambulatory legs that are constantly moved in all directions to sample the environment for detecting location of prey, as also for navigating. This is the first attempt to elucidate the sensory structures present on the antenniform legs of the whip spider, Phrynichus phipsoni (Pocock, 1894), collected from the state of Goa, India; using scanning electron micro[1]scope, and paves way for further research on sensory biology of this cryptic arachnid order. The examined specimen generally conforms to the diversity and morphology of sensory assemblage on the antenniform legs reported in other amblypygi species. Sensory structures elucidated include terminal trident tarsal claws, tarsal organ, bristles, porous sensilla, club sensilla, rod sensilla, slit sensilla, trichobothria, plate organ, pit organ and foliate leaf like hairs.

The Role of Honey Bee Derived Aliphatic Esters in the Host-Finding Behavior of Varroa destructor

Varroa destructor is an obligate ectoparasite of honey bees and shifted from its original host Apis cerana to the new host Apis mellifera in the first half of the twentieth century. The host shift has resulted in a great threat to the health and survival of A. mellifera colonies worldwide. Chemical signals play a crucial role in all aspects of the Varroa life cycle, including host finding. However, the chemical cues that affect the host finding behavior of Varroa mites are still not fully understood. In this study, we systematically profiled the headspace volatiles of both worker and drone larvae of the two honey bee species by using solid phase micro-extraction coupled to gas chromatography-mass spectrometry (SPME-GC-MS), and then used electrophysiological recording and Y-tube olfactometer bioassay to study the potential roles of the selected compounds. The chemical profiling showed that there were four aliphatic esters, ethyl myristate (EM), methyl palmitate (MP), ethyl palmitate (EP), and ethyl oleate (EO) commonly detected from all four types of larval hosts. Among them, EM was a new substance identified from honey bee headspace volatiles. Results from electrophysiological recordings indicated that all the aliphatic esters could elicit significant responses of Varroa pit organs on its forelegs. Moreover, behavioral analyses revealed that EM could significantly attract V. destructor at a medium dosage (10 µg), while MP had no observable effect on the mites and both EP and EO were able to repel the parasites. Our findings suggest an important role of host-derived aliphatic esters in Varroa host finding, and provide new chemicals for Varroa monitoring and control.

Pit viper thermography: the pit organ used by crotaline snakes to detect thermal contrast has poor spatial resolution.

Pit vipers detect infrared radiation by means of temperature contrasts created on their pit organ membranes. Signals from pit organs integrate with visual signals in the optic tectum, leading to the conjecture that the facial pits operate as an extension of the visual system. Because similar mechanisms underlie thermal imaging technology, imagery from thermal cameras is often used to infer how pit vipers perceive their environment. However, pit organs lack a focusing mechanism, and biophysical models predict that pit organs should have poor spatial resolution compared with thermal imaging cameras. Nevertheless, behavioral studies occasionally suggest pits may have better resolution than predicted by biophysical models, indicating that processing in the central nervous system may improve imaging. To estimate the spatial resolution of the neural image informing behavior, we recorded snake responses evoked by targets moving across backgrounds composed of two contrasting temperatures with an average temperature equal to the target temperature. An unresolved background would appear uniform; thus, the target would be detectable only if the background pattern were resolved. Western rattlesnakes (Crotalus oreganus) displayed no statistically significant responses to targets presented in front of patterned backgrounds, regardless of the temperature contrasts or spatial frequencies within the background, but responded strongly to targets presented in front of homogeneous backgrounds. We found no evidence that the pit organ system can resolve spatial details subtending an angle of 9deg or less. We discuss the implications of these results for understanding pit organ function in ecologically relevant habitats with thermal heterogeneity.

Uncooled self-powered hemispherical biomimetic pit organ for mid- to long-infrared imaging.

Infrared vision is highly desirable for applications in multifarious fields. Of the few species with this visual capability, snakes have exceptional infrared perception with the assistance of pit organs. Inspired by the pit organ design we present here a hemispherical biomimetic infrared imaging device. The devices use high-density ionic thermoelectric polymer nanowire arrays that serve as the sensing nerve cells. The individual nanowires exhibit notable voltage response to temperature variation in test objects. An infrared sensor array with 625 pixels on the hemispherical substrate is successfully demonstrated with an ultrawide field of view up to 135°. The device can image body temperature objects without a cooling system and external power supply. This work opens up opportunities for the design and fabrication of bioinspired infrared imaging devices based on emerging ionic thermoelectric materials.

Pit organ-based infrared discrimination sensitivity and signal transduction in the Burmese python (Python molurus bivitattus)

Burmese pythons (Python molurus bivitattus) use a unique infrared (IR) targeting system to acquire prey, avoid predators and seek thermoregulatory sites through detection of IR energy in the environment. Previous studies of sensitivity of the python IR system that relied on analysis of complex, natural behaviors lacked robust, reliable responses in animals habituated to experiments, and in vitro electrophysiological study failed to test behavioral function of the implicated protein thermoreceptor, TRPA1. The present study used conditioned discrimination procedures to analyze behavioral sensitivity and signal transduction in the python IR system. Pythons trained to behaviorally discriminate thermal stimuli averaged 70% correct choices, but failed to make correct choices when pit organs were physically occluded with IR-blocking material. The pythons exhibited greater sensitivity to thermal stimuli than previously reported, evident by correct choices that exceeded chance in response to a 14 × 10−6 W cm−2 irradiance contrast, or 0.5 °C thermal differential. Finally, in a test of the behavioral role of the putative thermoreceptor protein TRPA1, despite pit organ treatment with a TRPA1 inhibitor, python performance exceeded chance and was similar to baseline discrimination and control trials. Collectively, the results suggest that the IR system is a high sensitivity, broad-spectrum thermosensor that may operate through different and/or multiple thermoreceptive proteins with overlapping spectral response profiles. The findings reported here provide a better understanding of the relationship between the brain, behavior and environment in driving survival and ecological success of the Burmese python, especially as an invasive megapredator in the southern United States.

Fine Structure and Olfactory Reception of the Labial Palps of Spodoptera frugiperda

The olfactory system of insects is essential in many crucial behaviors, such as host seeking, mate recognition, and locating oviposition sites. Lepidopteran moths possess two main olfactory organs, including antennae and labial palps. Compared to antennae, the labial palps are relatively specific and worthy of further investigation due to the labial-palp pit organ (LPO), which contains a large number of sensilla located on the tip segment. The fall armyworm, Spodoptera frugiperda, is a worldwide lepidopteran pest, which can damage more than 350 plants and cause significant economic losses. In this study, we surveyed the structure of the labial palps and LPO of S. frugiperda using a super-high magnification lens zoom 3D microscope. Then, the distribution and fine structure of sensilla located in the LPO of S. frugiperda were investigated using scanning electron microscopy. Subsequently, the electrophysiological responses of labial palps to CO2 and 29 plant volatiles were recorded by using electrolabialpalpography. Our results showed the fine structure of labial palps, the LPO, and the sensilla located in the LPO of S. frugiperda. Moreover, we demonstrated that the labial palps are olfactory organs that respond to both CO2 and other volatile compounds. Our work established a foundation for further study of the roles of labial palps in insect olfactory related behaviors. Further investigations on the function of labial palps and their biological roles together with CO2 and volatile compound responses in S. frugiperda are necessary, as they may provide better insect behavioral regulators for controlling this pest.

Species Identification of Shed Snake Skins by Scanning Electron Microscopy, with Verification of Intraspecific Variations and Phylogenetic Comparative Analyses of Microdermatoglyphics

Shed skins of snakes have practical applications and can provide information on biodiversity. We examined shed skins or scales stripped from 53 native species of snakes from Taiwan and adjacent islands and 13 foreign species, verified the intraspecific variation of microstructures on the scales, conducted phylogenetic comparative analyses of microdermatoglyphics, and developed a guide and key to identify shed skins by scanning electron microscopy. We examined several microstructural characteristics of scales including the form and dimensions of oberhautchen cells, denticulations, microridges, nanopits, and nanochannels. Furthermore, we noted scale shape and presence or absence of rounded tubercles on the scales. The microstructures of apical (and central) regions of scales were similar to each other, but different from those on basal and lateral margins, the keel, or apical pits. The microdermatoglyphics on cranial scales (cephalic shields) differed from those on scales from other regions of the body. Those from the dorsal and dorsolateral surfaces of the body, and the dorsal surface of the tail were similar, as were those on scales from the ventral surfaces of the body and tail. For adults or juveniles, there was no significant ontogenetic variation in many microdermatoglyphic characteristics. By conducting phylogenetic mapping, we found that symmetrical (dorsal) scales without apical pit organs or rounded tubercles, keelless and rounded scales, flat microdermatoglyphic patterns (without longitudinal microridges), (sub)lamellate oberhautchen cells, cell borders without denticulations or with tiny and keelless denticulations, and relatively small nanopits at denticulation joints or covering cells, are likely to be plesiomorphic character states. We also examined character evolution models based on the microdermatoglyphics and their phylogenetic signals, conducted phylogenetic generalized least-squares regressions to examine the correlations among snake habitat type and the morphological traits on scales, and confirmed a hypothesis that the evolution of scale microstructures is dominated by both phylogenetic and functional (ecological) constraints. This study also indicates that scale microstructure can be used to identify snakeskin products, roadkill specimens and fragmented samples, scale remnants in predator feces, and wild snake sloughs.

Neuronal architecture of the second-order CO2 pathway in the brain of a noctuid moth

Many insects possess the ability to detect fine fluctuations in the environmental CO2 concentration. In herbivorous species, plant-emitted CO2, in combination with other sensory cues, affect many behaviors including foraging and oviposition. In contrast to the comprehensive knowledge obtained on the insect olfactory pathway in recent years, we still know little about the central CO2 system. By utilizing intracellular labeling and mass staining, we report the neuroanatomy of projection neurons connected with the CO2 sensitive antennal-lobe glomerulus, the labial pit organ glomerulus (LPOG), in the noctuid moth, Helicoverpa armigera. We identified 15 individual LPOG projection neurons passing along different tracts. Most of these uniglomerular neurons terminated in the lateral horn, a previously well-described target area of plant-odor projection neurons originating from the numerous ordinary antennal-lobe glomeruli. The other higher-order processing area for odor information, the calyces, on the other hand, was weakly innervated by the LPOG neurons. The overlapping LPOG terminals in the lateral horn, which is considered important for innate behavior in insects, suggests the biological importance of integrating the CO2 input with plant odor information while the weak innervation of the calyces indicates the insignificance of this ubiquitous cue for learning mechanisms.

Soft Matter Mechanics and the Mechanisms Underpinning the Infrared Vision of Snakes

Summary Pit-bearing snakes (vipers, pythons, and boas) have the extraordinary ability to “see” and accurately locate their prey and predators in total darkness. These animals use the infrared radiation emanating from objects that are warmer relative to the background environment to form a thermal image. Although enormous progress has been made to identify the key physiological features that enable the infrared vision of these snakes and a few other animals, the precise thermoelectric transduction mechanism that mediates the conversion of infrared heat to processable electrical signals has remained elusive. In this work, we quantitatively outline how cells in the snake's pit membrane organ act as apparent pyroelectric materials and convert infrared radiation into electrical signals. Despite the exceptional simplicity of our proposed mechanism and model, we are able to explain many central experimental results pertaining to the transduction process.

How Crucial is the Functional Pit Organ for the Varroa Mite?

Olfaction as well as gustation, are essential for animal survival, allowing behavioral modulation according to environmental input. We focused our study on an obligate ecto-parasitic mite of honey bees, the Varroa destructor Anderson and Trueman (Parasitiformes, Mesostigmata, Varroidae). By mechanically blocking the main olfactory organ on Varroa forelegs by varnishing with nail polish, we were able to show that other sensory organs cannot significantly compensate chemosensory abilities required for mite’s host selection, identification as well as reproduction. In fact, we found that mites with blocked forelegs had a significantly lower ability to reach a host bee than those with varnished idiosoma and unvarnished control. Furthermore, fewer foreleg blocked mites were feeding on the nurse bees and their reproduction in the brood cells was significantly impaired. The inhibition of reproduction was also reflected in altered expression levels of vitellogenin and vitellogenin receptor genes in foreleg-blocked mites.

Revisiting the Labial Pit Organ Pathway in the Noctuid Moth, Helicoverpa armigera.

Lepidopteran species detect CO2 via a specialized organ located on the peripheral segment of the labial palps, the labial palp pit organ (LPO). Based on tracing of LPO sensory neurons targeting one distinct antennal-lobe glomerulus, Kent and her colleagues described the projections originating from the LPO in the sphinx moth as “an accessory olfactory pathway in Lepidoptera” already in the 1980 ties. In spite of similar reports from studies of other lepidopteran species, however, it has been an unresolved issue whether additional termination areas of the labial nerve, such as the gnathal ganglion (GNG) and the ventral nerve cord, are actually output sites of LPO neurons. Since the previous studies have interpreted slightly differently about the projection pattern occurring from the classical mass staining, we performed selective mass staining from the inside of the pit and from the outer surface of the peripheral palp. The results demonstrated that the LPO sensory neurons project exclusively to the LPO glomerulus (LPOG), whereas the non-LPO sensory neurons target the GNG and the ventral nerve cord. Additional iontophoretic staining of individual LPO sensory neurons, performed from the LPO and the LPOG, showed three morphological neuron types: one bilateral targeting the LPOG in both antennal lobes, one unilateral targeting the ipsilateral LPOG only, and one contralateral targeting the LPOG in the other antennal lobe. Finally, to explore putative differences in the projection pattern of neurons housed by two previously reported sensillum types in the pit, i.e., hair-shaped sensilla located distally and club-shaped sensilla located proximally, we performed mass staining from two different levels of the peripheral palp. We found a projection pattern implying stronger innervation of the ipsi- than the contralateral LPOG in both staining experiments.