Mechanotransduction in spider slit sensilla

Mechanoreception is a vital constituent of several sensory modalities and a wide range of internal regulatory processes, but fundamental mechanisms for neural detection of mechanical stimuli have been difficult to characterize because of the morphological properties of most mechanoreceptors and the nature of the stimulus itself. An invertebrate preparation, the VS-3 lyriform slit sense organ of the spider, Cupiennius salei, has proved useful because it possesses large mechanosensory neurons, whose cell bodies are close to the sites of sensory transduction, and accessible to intracellular recording during mechanotransduction. This has made it possible to observe and experiment with all the major stages of mechanosensation. Here, we describe several important findings from this preparation, including the estimated number, conductance and ionic selectivity of the ion channels responsible for mechanotransduction, the major voltage-activated ion channels responsible for action potential encoding and control of the dynamic properties of the neurons, the location of action potential initiation following mechanical stimulation, and the efferent control of mechanoreception. While many details of mechanosensation remain to be discovered, the VS-3 system continues to offer important opportunities to advance our understanding of this crucial physiological process.     

Feeling the pressure in mammalian somatosensation

Mechanoreceptor cells of the somatosensory system initiate the perception of touch and pain. Molecules required for mechanosensation have been identified from invertebrate neurons, and recent functional studies indicate that ion channels of the transient receptor potential and degenerin/epithelial Na+ channel families are likely to be transduction channels. The expression of related channels in mammalian somatosensory neurons has fueled the notion that these channels mediate mechanotransduction in vertebrates; however, genetic disruption and heterologous expression have not yet revealed a direct role for any of these candidates in somatosensory mechanotransduction. Thus, new systems are needed to define the function of these ion channels in somatosensation and to pinpoint molecules or signaling pathways that underlie mechanotransduction in vertebrates.     

Structural correlates of mechanosensory transduction and adaptation in identified neurons of spider slit sensilla

We used isolated but functionally intact preparations of the lyriform slit-sense organ VS-3 from the leg of the spider, Cupiennius salei Keys, to examine the role of prominent fine-structural elements for mechanosensory transduction and adaptation. Slit sensilla act as strain sensors in the cuticular exoskeleton; each slit is innervated by two mechanosensitive neurons. Punctate mechanical deformation at four points along the dendrites demonstrated that mechanical excitability is confined to membrane sites at the extreme dendrite tips that are enclosed by cuticular slit structures. Depletion of microtubules in VS-3 neurons by prolonged mechanical stimulation and application of 5 mmol l(-1) colchicine did not disrupt the generation of a receptor potential. Hence, putative gating mechanisms of the mechanically activated membrane channels at the dendrite tips appear to be largely independent of microtubular structures. The discrete adaptation pattern in each of the two partner neurons, rapidly adapting versus slowly adapting, did not depend on the distinct mode of dendrite attachment to cuticular slit structures, and even persisted in isolated neurons after their dendrite tips and auxiliary structures were lost. We suggest that the two discrete adaptation patterns are based on intrinsic differences in the action potential encoding process rather than differences in stimulus transformation or mechanotransduction.     

High-threshold mechanosensitive ion channels blocked by a novel conopeptide mediate pressure-evoked pain

Little is known about the molecular basis of somatosensory mechanotransduction in mammals. We screened a library of peptide toxins for effects on mechanically activated currents in cultured dorsal root ganglion neurons. One conopeptide analogue, termed NMB-1 for noxious mechanosensation blocker 1, selectively inhibits (IC(50) 1 microM) sustained mechanically activated currents in a subset of sensory neurons. Biotinylated NMB-1 retains activity and binds selectively to peripherin-positive nociceptive sensory neurons. The selectivity of NMB-1 was confirmed by the fact that it has no inhibitory effects on voltage-gated sodium and calcium channels, or ligand-gated channels such as acid-sensing ion channels or TRPA1 channels. Conversely, the tarantula toxin, GsMTx-4, which inhibits stretch-activated ion channels, had no effects on mechanically activated currents in sensory neurons. In behavioral assays, NMB-1 inhibits responses only to high intensity, painful mechanical stimulation and has no effects on low intensity mechanical stimulation or thermosensation. Unexpectedly, NMB-1 was found to also be an inhibitor of rapid FM1-43 loading (a measure of mechanotransduction) in cochlear hair cells. These data demonstrate that pharmacologically distinct channels respond to distinct types of mechanical stimuli and suggest that mechanically activated sustained currents underlie noxious mechanosensation. NMB-1 thus provides a novel diagnostic tool for the molecular definition of channels involved in hearing and pressure-evoked pain.     

DEG/ENaC but not TRP channels are the major mechanoelectrical transduction channels in a C. elegans nociceptor

Many nociceptors detect mechanical cues, but the ion channels responsible for mechanotransduction in these sensory neurons remain obscure. Using in?vivo recordings and genetic dissection, we identified the DEG/ENaC protein, DEG-1, as the major mechanotransduction channel in ASH, a polymodal nociceptor in Caenorhabditis elegans. But DEG-1 is not the only mechanotransduction channel in ASH: loss of deg-1 revealed a minor current whose properties differ from those expected of DEG/ENaC channels. This current was independent of two TRPV channels expressed in ASH. Although loss of these TRPV channels inhibits behavioral responses to noxious stimuli, we found that both mechanoreceptor currents and potentials were essentially wild-type in TRPV mutants. We propose that ASH nociceptors rely on two genetically distinct mechanotransduction channels and that TRPV channels contribute to encoding and transmitting information. Because mammalian and insect nociceptors also coexpress DEG/ENaCs and TRPVs, the cellular functions elaborated here for these ion channels may be conserved.     

Central nervous projections of mechanoreceptors in the spider Cupiennius salei Keys.

Summary The basic organization of sensory projections in the suboesophageal central nervous system of a spider (Cupiennius salei Keys.) was analyzed with anterograde cobalt fills and a modified Golgi rapid method. The projections of three lyriform slit sense organs and of tactile hairs located proximally on the legs are described and related to central nerve tracts. There are five main longitudinal sensory tracts in the central region of the suboesophageal nervous mass arranged one above the other. Whereas the three dorsal ones contain fibers from the lyriform organs, the two ventral ones contain axons from the hair receptors. Axons from all three lyriform organs have typical shapes and widely arborizing ipsilateral intersegmental branches and a few contralateral ones. The terminal branches of the afferent projections from identical lyriform organs on each leg form characteristic longitudinal pathways, typical of each organ: U-shaped, O-shaped, or two parallel bundles. The terminations of the hair sensilla are ipsilateral and intersegmental. Two large bilaterally arranged longitudinal sensory association tracts receive inputs from all legs including the dense arborizations from tactile hairs, lyriform organs, and other sense organs. These tracts may serve as important integrating neuropils of the suboesophageal central nervous system.     

Recording of mechanosensitive currents using piezoelectrically driven mechanostimulator

Mechanotransduction constitutes the basis of a variety of physiological processes, such as the senses of touch, balance, proprioception and hearing. In vertebrates, mechanosensation is mediated by mechanosensory receptors. The aptitude of these mechanoreceptors for detecting mechanical information relies on the presence of mechanosensitive channels that transform mechanical forces into electrical signals. However, advances in understanding mechanical transduction processes have proven difficult because sensory nerve endings have historically been inaccessible to patch-clamp recording. We report here an in vitro model of mechanotransduction that allows the application of focal force on sensory neuron membrane during whole-cell patch clamping. This technique, called mechano-clamp, provides an opportunity to explore the properties and identities of mechanotransducer channels in mammalian sensory neurons. The protocol-from tissue dissociation to patch-clamp recording-can be completed in 7 h.     

Proprioceptors and sensory nerves in the legs of a spider,Cupiennius salei (Arachnida,Araneida)

Summary Retrograde CoS-impregnation was used to trace and map the course of sensory nerves and the distribution and innervation of the various proprioceptor types in all leg segments of Cupiennius salei, a Ctenid spider.1. Sensory nerve branches. In both the tibia and femur, axons of all proprioceptor types ascend in just two lateral nerves which do not merge with the main leg nerve until they reach the next proximal joint region. In the short segments — coxa, trochanter, patella, and tarsus — axons of the internal joint receptors often run separately from those of the other sensilla. Axons of the large lyriform slit sense organ at the dorsal metatarsus and of the trichobothria join with only a few hair axons and form their own nerve branches (Figs. 1, 2, 3).2. Proprioceptors. Each of the seven leg joints is supplied with at least one set of the well-known internal joint receptors, slit sensilla (single slits and lyriform organs), and long cuticular hairs. In addition, we found previously unnoticed hair plates on both sides of the coxa, near the prosoma/coxa joint; they are deflected by the articular membrane during joint movements (Fig. 4).3. Sensory cells and innervation. CoS-impregnation shows that each slit of the slit sense organs — be it a single slit or several slits in a lyriform organ — is innervated by two bipolar sensory cells (Fig. 6). We also confirm previous reports of multiple innervation in the internal joint receptors and in the long joint hairs and cuticular spines.Most of the ascending nerve branches run just beneath the cuticle for at least a short distance (Fig. 5); hence they are convenient sites for electrophysiological recordings of sensory activity even in freely walking spiders.     

Proprioceptor distribution and control of a muscle reflex in the tibia of spider legs

In spiders, retrograde cobalt staining was used to clarify the distribution and detailed innervation of the three types of proprioceptors in the tibio-metatarsal leg joint: internal joint receptors, lyriform slit sense organs, and cuticular spines and hairs. The axons of all these receptors run in just two lateral, ascending nerves, which had previously been associated only with the internal receptors. Each nerve contains several hundred axons ranging in diameter from 0.1 micron to ca. 10 micron. Each slit of the four tibial lyriform organs is innervated by two bipolar sensory neurons. The lateral nerves are entirely sensory and run just beneath the cuticle, a convenient site for electrophysiological recording. We demonstrate simultaneous nerve and muscle recordings from intact spiders; these, in combination with selective sensory ablations, show that a resistance reflex in the flexor metatarsi muscles is elicited by internal joint-receptor units.     

Neurosensory mechanotransduction through acid‐sensing ion channels

Acid‐sensing ion channels (ASICs) are voltage‐insensitive cation channels responding to extracellular acidification. ASIC proteins have two transmembrane domains and a large extracellular domain. The molecular topology of ASICs is similar to that of the mechanosensory abnormality 4‐ or 10‐proteins expressed in touch receptor neurons and involved in neurosensory mechanotransduction in nematodes. The ASIC proteins are involved in neurosensory mechanotransduction in mammals. The ASIC isoforms are expressed in Merkel cell–neurite complexes, periodontal Ruffini endings and specialized nerve terminals of skin and muscle spindles, so they might participate in mechanosensation. In knockout mouse models, lacking an ASIC isoform produces defects in neurosensory mechanotransduction of tissue such as skin, stomach, colon, aortic arch, venoatrial junction and cochlea. The ASICs are thus implicated in touch, pain, digestive function, baroreception, blood volume control and hearing. However, the role of ASICs in mechanotransduction is still controversial, because we lack evidence that the channels are mechanically sensitive when expressed in heterologous cells. Thus, ASIC channels alone are not sufficient to reconstruct the path of transducing molecules of mechanically activated channels. The mechanotransducers associated with ASICs need further elucidation. In this review, we discuss the expression of ASICs in sensory afferents of mechanoreceptors, findings of knockout studies, technical issues concerning studies of neurosensory mechanotransduction and possible missing links. Also we propose a molecular model and a new approach to disclose the molecular mechanism underlying the neurosensory mechanotransduction.     

Ein atlas der spaltsinnesorgane von Cupiennius salei keys. Chelicerata (Araneae)

To facilitate further physiological investigation, a survey was undertaken of all the slit sense organs to be found on the body of the spider Cupiennius salei. We counted and mapped more than 3 000 sensory slits in the cuticle about half of which are combined to small groups of up to 29 slits forming compound or lyriform organs.     

Mechanosensitive ion channels push cancer progression

In many cases, the mechanical properties of a tumor are different from those of the host tissue. Mechanical cues regulate cancer development by affecting both tumor cells and their microenvironment, by altering cell migration, proliferation, extracellular matrix remodeling and metastatic spread. Cancer cells sense mechanical stimuli such as tissue stiffness, shear stress, tissue pressure of the extracellular space (outside-in mechanosensation). These mechanical cues are transduced into a cellular response (e. g. cell migration and proliferation; inside-in mechanotransduction) or to a response affecting the microenvironment (e. g. inducing a fibrosis or building up growth-induced pressure; inside-out mechanotransduction). These processes heavily rely on mechanosensitive membrane proteins, prominently ion channels. Mechanosensitive ion channels are involved in the Ca²⁺-signaling of the tumor and stroma cells, both directly, by mediating Ca²⁺ influx (e. g. Piezo and TRP channels), or indirectly, by maintaining the electrochemical gradient necessary for Ca²⁺ influx (e. g. K_2P, K_Ca channels). This review aims to discuss the diverse roles of mechanosenstive ion channels in cancer progression, especially those involved in Ca²⁺-signaling, by pinpointing their functional relevance in tumor pathophysiology.     

HCN Channels Are Not Required for Mechanotransduction in Sensory Hair Cells of the Mouse Inner Ear

The molecular composition of the hair cell transduction channel has not been identified. Here we explore the novel hypothesis that hair cell transduction channels include HCN subunits. The HCN family of ion channels includes four members, HCN1-4. They were orginally identified as the molecular correlates of the hyperpolarization-activated, cyclic nucleotide gated ion channels that carry currents known as I_f, I_Q or I_h. However, based on recent evidence it has been suggested that HCN subunits may also be components of the elusive hair cell transduction channel. To investigate this hypothesis we examined expression of mRNA that encodes HCN1-4 in sensory epithelia of the mouse inner ear, immunolocalization of HCN subunits 1, 2 and 4, uptake of the transduction channel permeable dye, FM1-43 and electrophysiological measurement of mechanotransduction current. Dye uptake and transduction current were assayed in cochlear and vestibular hair cells of wildtype mice exposed to HCN channel blockers or a dominant-negative form of HCN2 that contained a pore mutation and in mutant mice that lacked HCN1, HCN2 or both. We found robust expression of HCNs 1, 2 and 4 but little evidence that localized HCN subunits in hair bundles, the site of mechanotransduction. Although high concentrations of the HCN antagonist, ZD7288, blocked 50–70% of the transduction current, we found no reduction of transduction current in either cochlear or vestibular hair cells of HCN1- or HCN2- deficient mice relative to wild-type mice. Furthermore, mice that lacked both HCN1 and HCN2 also had normal transduction currents. Lastly, we found that mice exposed to the dominant-negative mutant form of HCN2 had normal transduction currents as well. Taken together, the evidence suggests that HCN subunits are not required for mechanotransduction in hair cells of the mouse inner ear.     

Drosophila Mechanotransduction—Linking Proteins and Functions

The sensation of touch, gravity, and sound all rely on dedicated ion channels that transduce mechanical stimulus forces into electrical response signals. The functional workings and molecular identities of these mechanotransducer channels are little understood. Recent work shows that the mechanotransducers for fly and vertebrate hearing share equivalent gating mechanisms, whereby this mechanism can be probed non-invasively in the mechanics of the Drosophila ear. Here, we describe how this mechanics can be used to evaluate the roles of identified proteins in the process of mechanosensation and, specifically, their contributions to mechanotransduction.     

蜘蛛的机械感器

蜘蛛的机械感器是最灵敏的机械感受器之一,文章从蜘蛛机械感器的分布、形状和功能、竖琴状感器的结构、感器神经元的电生理特性、机械感觉转导的位点、动作电位的起始位点、受体通道的性质、输出神经对感觉神经元的调控方面对蜘蛛机械感受器进行了介绍,并提出了有待进一步解决的问题。     

Drosophila mechanotransduction--linking proteins and functions

The sensation of touch, gravity, and sound all rely on dedicated ion channels that transduce mechanical stimulus forces into electrical signals. The functional workings and molecular identities of these mechanotransducer channels are little understood. Recent work shows that the mechanotransducers for fly and vertebrate hearing share equivalent gating mechanisms, whereby this mechanism can be probed non-invasively in the mechanics of the Drosophila ear. Here, we describe how this mechanics can be used to evaluate the roles of identified proteins in the process of mechanosensation and, specifically, their contributions to mechanotransduction.     

Plasma Membrane Mechanical Stress Activates TRPC5 Channels

Mechanical forces exerted on cells impose stress on the plasma membrane. Cells sense this stress and elicit a mechanoelectric transduction cascade that initiates compensatory mechanisms. Mechanosensitive ion channels in the plasma membrane are responsible for transducing the mechanical signals to electrical signals. However, the mechanisms underlying channel activation in response to mechanical stress remain incompletely understood. Transient Receptor Potential (TRP) channels serve essential functions in several sensory modalities. These channels can also participate in mechanotransduction by either being autonomously sensitive to mechanical perturbation or by coupling to other mechanosensory components of the cell. Here, we investigated the response of a TRP family member, TRPC5, to mechanical stress. Hypoosmolarity triggers Ca²⁺ influx and cationic conductance through TRPC5. Importantly, for the first time we were able to record the stretch-activated TRPC5 current at single-channel level. The activation threshold for TRPC5 was found to be 240 mOsm for hypoosmotic stress and between −20 and −40 mmHg for pressure applied to membrane patch. In addition, we found that disruption of actin filaments suppresses TRPC5 response to hypoosmotic stress and patch pipette pressure, but does not prevent the activation of TRPC5 by stretch-independent mechanisms, indicating that actin cytoskeleton is an essential transduction component that confers mechanosensitivity to TRPC5. In summary, our findings establish that TRPC5 can be activated at the single-channel level when mechanical stress on the cell reaches a certain threshold.     

Shaping the mammalian auditory sensory organ by the planar cell polarity pathway

The human ear is capable of processing sound with a remarkable resolution over a wide range of intensity and frequency. This ability depends largely on the extraordinary feats of the hearing organ, the organ of Corti and its sensory hair cells. The organ of Corti consists of precisely patterned rows of sensory hair cells and supporting cells along the length of the snail-shaped cochlear duct. On the apical surface of each hair cell, several rows of actin-containing protrusions, known as stereocilia, form a "V"-shaped staircase. The vertices of all the "V"-shaped stereocilia point away from the center of the cochlea. The uniform orientation of stereocilia in the organ of Corti manifests a distinctive form of polarity known as planar cell polarity (PCP). Functionally, the direction of stereociliary bundle deflection controls the mechanical channels located in the stereocilia for auditory transduction. In addition, hair cells are tonotopically organized along the length of the cochlea. Thus, the uniform orientation of stereociliary bundles along the length of the cochlea is critical for effective mechanotransduction and for frequency selection. Here we summarize the morphological and molecular events that bestow the structural characteristics of the mammalian hearing organ, the growth of the snail-shaped cochlear duct and the establishment of PCP in the organ of Corti. The PCP of the sensory organs in the vestibule of the inner ear will also be described briefly.     

The sensory equipment of a spider – A morphological survey of different types of sensillum in both sexes of Argiope bruennichi (Araneae,Araneidae)

Spiders show a wide range of sensory capabilities as evidenced by behavioural observations. Accordingly, spiders possess diverse sensory structures like mechano-, hygro-, thermo- or chemoreceptive sensilla. As to chemoreceptive structures, only trichoid tip-pore sensilla were found so far that were tested for gustation. That spiders are also able to receive airborne signals is corroborated by numerous behavioural experiments but the responsible structures have not been determined yet. Here, we provide sensilla distribution maps of pedipalps and walking legs of both sexes of the wasp spider Argiope bruennichi whose biology and mating system is well explored. By means of scanning electron microscopy, we scrutinized whether there is in fact only one type of trichoid pore sensillum and if so, if there are deviations in the outer structure of the tip-pore sensilla depending on their position on the body. We also describe the external structure and distribution of slit sense organs, trichobothria and tarsal organs. Our study shows that all four sensillum types occur on pedipalps and walking legs of both sexes. As to chemosensory organs, only tip-pore sensilla were found, suggesting that this sensillum type is used for both gustation and olfaction. The highest numbers of tip-pore sensilla were observed on metatarsi and tarsi of the first two walking legs. Mechanosensitive slit sense organs occur as single slit sensilla in rows along all podomers or as lyriform organs next to the joints. The mechanosensitive trichobothria occur on the basal part of tibiae and metatarsi. Tarsal organs occur on the dorsal side of all tarsi and the male cymbium. The distribution maps of the sensilla are the starting point for further exploration of internal, morphological differences of the sensilla from different regions on the body. Cryptic anatomical differences might be linked to functional differences that can be explored in combination with electrophysiological analyses. Consequently, the maps will help to elucidate the sensory world of spiders.     

Intracellular recording from a spider vibration receptor

The present study introduces a new preparation of a spider vibration receptor that allows intracellular recording of responses to natural mechanical or electrical stimulation of the associated mechanoreceptor cells. The spider vibration receptor is a lyriform slit sense organ made up of 21 cuticular slits located on the distal end of the metatarsus of each walking leg. The organ is stimulated when the tarsus receives substrate vibrations, which it transmits to the organ’s cuticular structures, reducing the displacement to about one tenth due to geometrical reasons. Current clamp recording was used to record action potentials generated by electrical or mechanical stimuli. Square pulse stimulation identified two groups of sensory cells, the first being single-spike cells which generated only one or two action potentials and the second being multi-spike cells which produced bursts of action potentials. When the more natural mechanical sinusoidal stimulation was applied, differences in adaptation rate between the two cell types remained. In agreement with prior extracellular recordings, both cell types showed a decrease in the threshold tarsus deflection with increasing stimulus frequency. Off-responses to mechanical stimuli have also been seen in the metatarsal organ for the first time.