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Gangliosides are particularly abundant in the central nervous system, where they are mainly associated with the synaptic membranes. Their structure underlies a specific role in determining several cell physiological processes of the nervous system. The high number of different gangliosides available in nature suggests that their structure, related to both the hydrophobic and hydrophilic portion of the molecule, defines a code, although not completely understood, that through hydrophobic interactions and hydrogen bonds allows the transduction of signals starting at the plasma membranes. In this short review, we describe some structural aspects responsible for the role played by gangliosides in maintaining and determining neuronal functions., The biosynthetic pathway of central nervous system sphingolipids: the scheme reports the two main pathways to obtain sphingolipids containing C18‐ or C20‐sphingosine.

The mammalian cochlea relies on the active forcing of sensory outer hair cells (OHCs) to amplify traveling wave responses along the basilar membrane. These forces are the result of electromotility, wherein current through the OHCs leads to conformational changes in the cells that provide stresses on surrounding structures. OHC transducer current can be detected via the voltage in the scala tympani (the cochlear microphonic, CM), and the CM can be used as an indicator of healthy cochlear operation. The CM represents a summation of OHC currents (the inner hair cell contribution is known to be small) and to use CM to probe the properties of OHC transduction requires a model that simulates that summation. We developed a finite element model for that purpose. The pattern of current generators (the model input) was initially based on basilar membrane displacement, with the current size based on in vitro data. The model was able to reproduce the amplitude of experimental CM results reasonably well when the input tuning was enhanced slightly (peak increased by ∼6 dB), which can be regarded as additional hair bundle tuning, and with a current/input value of 200–260 pA/nm, which is ∼4 times greater than the largest in vitro measures.

The responses of primary sensory neurons to subnanomolar and nanomolar ouabain concentrations, corresponding to the endogenous ouabain (EO) concentration, were studied. Atomic force microscopy (AFM) studies showed that exposure to EO led to an increase in neuron stiffness. Studies using local voltage clamping showed that ligand-receptor binding of EO with the Na,K-ATPase/Src complex decreased the effective charge of the activatory gate system of NaV1.8 channels. Furthermore, the EO-activated intracellular cascade in which the Na,K-ATPase/Src complex operates as a signal transducer was found to lead to a decrease in the fluorescence intensity of antibodies to NaV1.8 channels, as demonstrated by confocal laser scanning microscopy. These results suggest that EO, triggering the transduction function of the Na,K-ATPase/Src complex and the corresponding intracellular signal cascade, is able to modulate the expression of the SCN10A gene, which produces NaV1.8 channels, which are responsible for encoding nociceptive signals.

The beating heart possesses the intrinsic ability to adapt cardiac output to changes in mechanical load. The century-old Frank–Starling law and Anrep effect have documented that stretching the heart during diastolic filling increases its contractile force. However, the molecular mechanotransduction mechanism and its impact on cardiac health and disease remain elusive. Here we show that the mechanically activated Piezo1 channel converts mechanical stretch of cardiomyocytes into Ca2+ and reactive oxygen species (ROS) signaling, which critically determines the mechanical activity of the heart. Either cardiac-specific knockout or overexpression of Piezo1 in mice results in defective Ca2+ and ROS signaling and the development of cardiomyopathy, demonstrating a homeostatic role of Piezo1. Piezo1 is pathologically upregulated in both mouse and human diseased hearts via an autonomic response of cardiomyocytes. Thus, Piezo1 serves as a key cardiac mechanotransducer for initiating mechano-chemo transduction and consequently maintaining normal heart function, and might represent a novel therapeutic target for treating human heart diseases., The beating heart adapts cardiac output to changes in mechanical load via incompletely understood mechanotransduction mechanisms. Here the authors show that the mechanosensitive Piezo1 channel serves as a mechanotransducer for directly converting mechanical stretch of cardiomyocytes into Ca2+ and ROS signaling and consequently maintaining normal heart function.

The contraction-relaxation cycle of the heart is one of the most robust mechanical systems in the body that adapts rapidly to the body’s needs by changing mechanical parameters. In many respects, we can consider the cardiac system as a complex machine and can use engineering approaches to describe its function. The classical physiology of the heart also focused on understanding function but the new molecular level tools in light microscopy and nanoengineering now enable a deeper understanding of the physiology. The field of mechanobiology has emerged with a focus on how mechanical activity alters biological systems at the molecular level and how those systems in turn control mechanical parameters. In the case of mechanical activity, there are clearly benefits of exercise for the heart, for cancer patients, and for aging but we do not understand the links at a molecular level. Why does regular exercise benefit the heart? We have some preliminary clues at a molecular level about the benefits of physical activity in the cases of cancer and aging; however, there is less known about how exercise affects cardiovascular performance. Unlike the omics approaches which generally link proteins to processes, a mechanobiological understanding of a process explains how forces and mechanical activity will regulate the process through modifications of protein activities. In other words, mechanical activity is an essential component of most biological systems that is transduced into biochemical changes in protein activity. Further, it follows logically that if a mechanical parameter of the cardiac system is typically controlled, then cellular mechanosensing systems must be able to directly or indirectly measure that parameter. The challenge is to understand how changes in activity of the heart are controlled in the short term and then how the system adapts to the integrated level of activity over the longer term. By way of introduction to molecular mechanobiology, I will present examples of mechanosensing from the molecular to the cellular scale and how they may be integrated at the cell and tissue levels. An important element of Mechanobiology at the system level is the physiological state of the cell: i.e., the cell in a senescent state, a cancer state, or a normal cell state (Sheetz 2019). The background for the mechanobiological approach is discussed in “The Cell as a Machine” (Sheetz and Yu, Cambridge Univ Press, 2018), which considers cell states and the molecular systems underlying the important cellular functions. A major challenge in mechanobiology is the understanding of the transduction of mechanical activity into changes in cell function. Of particular relevance here is the benefit of exercise to cardiac performance. This has been seen in many cases and there are a variety of factors that contribute. Further, exercise will benefit cancer patients and will reverse some of the adverse effects of aging. Exercise will cause increased cardiac activity that will be sensed by many mechanosensory systems from a molecular to a cellular level both in the heart and in the vasculature. At a molecular level in cardiac systems, proteins are able to measure stress and strain and to generate appropriate signals of the magnitude of stress and strain that can regulate the cellular contractility and other parameters. The protein sensors are generally passive systems that give a transient measure of local parameters such as the stress at cell-cell junctions during contraction and the strain of the sarcomeres during relaxation. Large stresses at the junctions can activate signaling systems that can reduce contractility or over time activate remodeling of the junctions to better support larger stresses. The proteins involved and their sensory mechanisms are not known currently; however, the mechanosensitive channel, Piezo1, has been implicated in the transduction process in the vasculature (Beech 2018). In the case of strain sensors, large stretches of titin during relaxation can unfold more titin domains that can send signals to the cell. Two different mechanisms of strain sensing are likely in titin. The titin kinase domain is activated by strain but the substrates of the kinase are not know in vivo (Linke 2018). In the backbone of titin are many Ig domains that unfold at different forces and unfolding could cause the binding of proteins that would then activate enzymatic pathways to alter the contractile cycle to give the proper level of strain (Ait-Mou et al. 2017; Granzier et al. 2014; Granzier et al. 2009). The cell-matrix adhesion protein, talin, has eleven cryptic binding sites for another adhesion protein, vinculin, that are revealed by the unfolding of domains in the talin molecule (Yao et al. 2016). Since some domains unfold at lower forces than others, small strains will preferentially unfold those domains, making the system an excellent sensor of the extent of stretch as expected for titin. Because there is an ordered array of many titin molecules, the sensing of strain can be very sensitive to small changes in sarcomere length. Needless to say, titin is only one part of the regulatory system that controls sarcomere length. As one goes more deeply into the working of the system, it is evident that many additional mechanosensory elements are involved in maintaining a functioning cardiac system.

Significance Each of the sensory receptors responsible for hearing or balance—a hair cell—has a mechanosensitive hair bundle. Mechanical stimuli pull upon molecular filaments—the tip links—that open ionic channels in the hair bundle. Loud sounds can damage hearing by breaking the tip links; recovery by replacement of the constituent proteins then requires several hours. We disrupted the tip links in vitro by removing the calcium ions that stabilize them, and then monitored the electrical response or stiffness of hair bundles to determine whether the links could recover. We found that tip links recovered within seconds if their ends were brought back into contact. This form of repair might occur in normal ears to restore sensitivity after damage., Hearing and balance rely on the capacity of mechanically sensitive hair bundles to transduce vibrations into electrical signals that are forwarded to the brain. Hair bundles possess tip links that interconnect the mechanosensitive stereocilia and convey force to the transduction channels. A dimer of dimers, each of these links comprises two molecules of protocadherin 15 (PCDH15) joined to two of cadherin 23 (CDH23). The “handshake” that conjoins the four molecules can be disrupted in vivo by intense stimulation and in vitro by exposure to Ca2+ chelators. Using hair bundles from the rat’s cochlea and the bullfrog’s sacculus, we observed that extensive recovery of mechanoelectrical transduction, hair bundle stiffness, and spontaneous bundle oscillation can occur within seconds after Ca2+ chelation, especially if hair bundles are deflected toward their short edges. Investigating the phenomenon in a two-compartment ionic environment that mimics natural conditions, we combined iontophoretic application of a Ca2+ chelator to selectively disrupt the tip links of individual frog hair bundles with displacement clamping to control hair bundle motion and measure forces. Our observations suggest that, after the normal Ca2+ concentration has been restored, mechanical stimulation facilitates the reconstitution of functional tip links.

Significance When sound vibrations reach the inner ear, fine protein filaments called “tip links” stretch and open cochlear hair-cell mechanosensitive channels that trigger sensory perception. Similarly, vestibular hair cells use tip links to sense mechanical stimuli produced by head motions. Tip links are formed by cadherin-23 and protocadherin-15, two large proteins involved in hearing loss and balance disorders. Here we present multiple structures, models, and simulations that depict the lower end of the tip link, including the complete protocadherin-15 ectodomain. These models show an essential connection between cadherin-23 and protocadherin-15 with dual molecular “handshakes” and various protein sites that are mutated in inherited deafness. The simulations also reveal how the tip link responds to force to mediate hearing and balance sensing., The vertebrate inner ear, responsible for hearing and balance, is able to sense minute mechanical stimuli originating from an extraordinarily broad range of sound frequencies and intensities or from head movements. Integral to these processes is the tip-link protein complex, which conveys force to open the inner-ear transduction channels that mediate sensory perception. Protocadherin-15 and cadherin-23, two atypically large cadherins with 11 and 27 extracellular cadherin (EC) repeats, are involved in deafness and balance disorders and assemble as parallel homodimers that interact to form the tip link. Here we report the X-ray crystal structure of a protocadherin-15 + cadherin-23 heterotetrameric complex at 2.9-Å resolution, depicting a parallel homodimer of protocadherin-15 EC1-3 molecules forming an antiparallel complex with two cadherin-23 EC1-2 molecules. In addition, we report structures for 10 protocadherin-15 fragments used to build complete high-resolution models of the monomeric protocadherin-15 ectodomain. Molecular dynamics simulations and validated crystal contacts are used to propose models for the complete extracellular protocadherin-15 parallel homodimer and the tip-link bond. Steered molecular dynamics simulations of these models suggest conditions in which a structurally diverse and multimodal protocadherin-15 ectodomain can act as a stiff or soft gating spring. These results reveal the structural determinants of tip-link–mediated inner-ear sensory perception and elucidate protocadherin-15’s structural and adhesive properties relevant in disease.

Rod photoreceptors are composed of a soma and an inner segment (IS) connected to an outer segment (OS) by a thin cilium. OSs are composed of a stack of similar to 800 lipid discs surrounded by the plasma membrane where phototransduction takes place. Intracellular calcium plays a major role in phototransduction and is more concentrated in the discs, where it can be incorporated and released. To study calcium dynamics in rods, we used the fluorescent calcium dye CaSiR-1 AM working in the near-infrared (NIR) (excitation at 650 and emission at 664 nm), an advantage over previously used dyes. In this way, we investigated calcium dynamics with an unprecedented accuracy and most importantly in semidark-adapted conditions. We observed light-induced drops in [Ca2+](i) with kinetics similar to that of photoresponses recorded electrophysiologically. We show three properties of the rods. First, intracellular calcium and key proteins have concentrations that vary from the OS base to tip. At the OS base, [Ca2+](i) is similar to 80 nM and increases up to similar to 200 nM at the OS tip. Second, there are spontaneous calcium flares in healthy and functional rod OSs; these flares are highly localized and are more pronounced at the OS tip. Third, a bright flash of light at 488 nm induces a drop in [Ca2+](i) at the OS base but often a flare at the OS tip. Therefore, rod OSs are not homogenous structures but have a structural and functional gradient, which is a fundamental aspect of transduction in vertebrate photoreceptors.

Cardiomyocytes, the cells generating contractile force in the heart, are connected to each other through a highly specialised structure, the intercalated disc (ID), which ensures force transmission and transduction between neighbouring cells and allows the myocardium to function in synchrony. In addition, cardiomyocytes possess an intrinsic ability to sense mechanical changes and to regulate their own contractile output accordingly. To achieve this, some of the components responsible for force transmission have evolved to sense changes in tension and to trigger a biochemical response that results in molecular and cellular changes in cardiomyocytes. This becomes of particular importance in cardiomyopathies, where the heart is exposed to increased mechanical load and needs to adapt to sustain its contractile function. In this review, we will discuss key mechanosensing elements present at the intercalated disc and provide an overview of the signalling molecules involved in mediating the responses to changes in mechanical force.

Iatrogenic external root resorption can become a serious pathological condition with clinical tooth movement. Little is known regarding how cementum responds to mechanical loading in contrast to bone, especially under compressive stress. In the field of bone biology, several studies have established the contribution of sphingosine-1-phosphate (S1P) signaling in bone remodeling, mechanical transduction and homeostasis. As osteocytes and cementocytes share similar morphological and functional characteristics, this study aimed to investigate the mechanotransduction ability of cementocytes and to explore the contribution of S1P signaling under compressive stress induced mechanotransduction. We found that compressive stress inhibited major S1P signaling and promoted the expression of anabolic factors in IDG-CM6 cells, a novel immortalized murine cementocyte cell line. By inhibiting S1P signaling, we verified that S1P signaling played a vital role in regulating the expression of the mechanotransduction factors prostaglandin E2 (PGE2) and β-catenin, as well as factors responsible for cementogenesis and cementoclastogenesis in IDG-CM6 cells. These results support the hypothesis that cementocytes act as key mechanically responsive cells in cementum, responding to compressive stress and directing local cementum metabolism.

The invention of optical tweezers more than three decades ago has opened new avenues in the study of the mechanical properties of biological molecules and cells. Quantitative force measurements still represent a challenging task in living cells due to the complexity of the cellular environment. Here, we review different methodologies to quantitatively measure the mechanical properties of living cells, the strength of adhesion/receptor bonds, and the active force produced during intracellular transport, cell adhesion, and migration. We discuss experimental strategies to attain proper calibration of optical tweezers and molecular resolution in living cells. Finally, we show recent studies on the transduction of mechanical stimuli into biomolecular and genetic signals that play a critical role in cell health and disease.

Background Centipedes are terrestrial, predatory arthropods with specialized sensory organs. However, many aspects of their sensory biology are still unknown. This also concerns hygroreception, which is especially important for centipedes, as their epicuticle is thin and they lose water rapidly at low humidity. Thus, the detection of humid places is vital but to date no definite hygroreceptor was found in centipedes. House centipedes (Scutigeromorpha) possess a peculiar opening at the base of their antenna, termed ‘scape organ’, that houses up to 15 cone-shaped sensilla in a cavity. Lacking wall and tip-pores, these socket-less sensilla may be hypothesized to function as hygroreceptors similar to those found in hexapods. Results The cone-shaped sensilla in the scape organ as well as nearby peg-shaped sensilla are composed of three biciliated receptor cells and three sheath cells. A tip-pore is present but plugged by a highly electron-dense secretion, which also overlays the entire inner surface of the cavity. Several solitary recto-canal epidermal glands produce the secretion. Receptor cell type 1 (two cells in cone-shaped sensilla, one cell in peg-shaped sensilla) possesses two long dendritic outer segments that project to the terminal pore. Receptor cell type 2 (one cell in both sensilla) possesses two shorter dendritic outer segments connected to the first (proximal) sheath cell that establishes a scolopale-like structure, documented for the first time in detail in a myriapod sensillum. Conclusions The nearly identical configuration of receptor cells 1 with their long dendritic outer segments in both sensilla is similar to hexapod hygroreceptors. In Scutigera coleoptrata, however, the mechanism of stimulus transduction is different. Water vapor may lead to swelling and subsequent elongation of the plug pin that enters the terminal pore, thus causing stimulation of the elongated dendritic outer segments. The interconnection of receptor cell 2 with short outer dendritic segments to a scolopale-like structure potentially suits both sensilla for vibration or strain detection. Thus, both sensilla located at the antennal base of scutigeromorph centipedes fulfill a dual function.

Olfactory receptors (ORs) comprise the largest multigene family in the vertebrates. They belong to the class A (rhodopsin-like) family of G protein-coupled receptors (GPCRs), which are the most abundant membrane proteins, having widespread, significant roles in signal transduction in cells, and therefore, they are a major pharmacological target. Moreover, ORs displayed high selectivity and sensitivity towards odorant detection, a characteristic that raised the interest for developing biohybrid sensors based on ORs for the detection of volatile compounds. The transduction of odorant binding into cellular signaling by ORs is not well understood and knowing its mechanism would enable developing new pharmacology and high performance biohybrid electronic sensors. Recent findings suggest that ligand recognition by ORs is determined by the nanoscale alterations of charge distribution in the receptor structure (Ref). However, the electrical characterization of ORs and their response towards ligand binding in bulk experiments is subjected to microscopic models and assumptions [2]. Here, we have directly determined the nanoscale electrical properties of ORs with unprecedented control over the receptor orientation, and their change upon odorant binding, using electrochemical scanning tunneling microscopy (EC-STM) in near-physiological conditions. Recordings of current versus time, distance, and electrochemical potential allows determining the OR impedance parameters and their dependence with odorant binding. The simultaneous measurement of RC equivalent by means of the open-circuit voltage (VOC) allows increasing the electrical sensitivity at the single receptor level for biosensing applications. Our results allow validating OR structural-electrostatic models and their functional activation processes.

In recent years, the importance of mechanical signaling and the cellular mechanical microenvironment in affecting cellular behavior has been widely accepted. Cells in epithelial monolayers are mechanically connected to each other and the underlying extracellular matrix (ECM), forming a highly connected mechanical system subjected to various mechanical cues from their environment, such as the ECM stiffness. Changes in the ECM stiffness have been linked to many pathologies, including tumor formation. However, our understanding of how ECM stiffness and its heterogeneities affect the transduction of mechanical forces in epithelial monolayers is lacking. To investigate this, we used a combination of experimental and computational methods. The experiments were conducted using epithelial cells cultured on an elastic substrate and applying a mechanical stimulus by moving a single cell by micromanipulation. To replicate our experiments computationally and quantify the forces transduced in the epithelium, we developed a new model that described the mechanics of both the cells and the substrate. Our model further enabled the simulations with local stiffness heterogeneities. We found the substrate stiffness to distinctly affect the force transduction as well as the cellular movement and deformation following an external force. Also, we found that local changes in the stiffness can alter the cells’ response to external forces over long distances. Our results suggest that this long-range signaling of the substrate stiffness depends on the cells’ ability to resist deformation. Furthermore, we found that the cell’s elasticity in the apico-basal direction provides a level of detachment between the apical cell-cell junctions and the basal focal adhesions. Our simulation results show potential for increased ECM stiffness, e.g. due to a tumor, to modulate mechanical signaling between cells also outside the stiff region. Furthermore, the developed model provides a good platform for future studies on the interactions between epithelial monolayers and elastic substrates.Author summaryCells can communicate using mechanical forces, which is especially important in epithelial tissues where the cells are highly connected. Also, the stiffness of the material under the cells, called the extracellular matrix, is known to affect cell behavior, and an increase in this stiffness is related to many diseases, including cancers. However, it remains unclear how the stiffness affects intercellular mechanical signaling. We studied this effect using epithelial cells cultured on synthetic deformable substrates and developed a computational model to quantify the results better. In our experiments and simulations, we moved one cell to observe how the substrate stiffness impacts the deformation of the neighboring cells and thus the force transduction between the cells. Our model also enabled us to study the effect of local stiffness changes on the force transduction. Our results showed that substrate stiffness has an apparent impact on the force transduction within the epithelial tissues. Furthermore, we found that the cells can communicate information on the local stiffness changes over long distances. Therefore, our results indicate that the cellular mechanical signaling could be affected by changes in the substrate stiffness which may have a role in the progression of diseases such as cancer.

Hair cell mechanosensitivity resides in the sensory hair bundle, an apical protrusion of actin-filled stereocilia arranged in a staircase pattern. Hair bundle deflection activates mechano-electric transduction (MET) ion channels located near the tops of the shorter rows of stereocilia. The elicited macroscopic current is shaped by the hair bundle motion so that the mode of stimulation greatly influences the cell’s output. We present data quantifying the displacement of the whole outer hair cell bundle using high-speed imaging when stimulated with a fluid jet. We find a spatially non-uniform stimulation that results in splaying, where the hair bundle expands apart. Based on modeling, the splaying is predominantly due to fluid dynamics with a small contribution from hair bundle architecture. Additionally, in response to stimulation, the hair bundle exhibited a rapid motion followed by a slower motion in the same direction (creep) that is described by a double exponential process. The creep is consistent with originating from a linear passive system that can be modeled using two viscoelastic processes. These viscoelastic mechanisms are integral to describing the mechanics of the mammalian hair bundle.

In vertebrate hearing organs, mechanical vibrations are converted to ionic currents through mechanoelectrical-transduction (MET) channels. Concerted stereocilia motion produces an ensemble MET current driving the hair-cell receptor potential. Mammalian cochleae are unique in that the tuning of sensory cells is determined by their mechanical environment and the mode of hair-bundle stimulation that their environment creates. However, little is known about the in situ intra-hair-bundle motions of stereocilia relative to one another, or to their environment. In this study, high-speed imaging allowed the stereocilium and cell-body motions of inner hair cells to be monitored in an ex vivo organ of Corti (OoC) mouse preparation. We have found that the OoC rotates about the base of the inner pillar cell, the hair bundle rotates about its base and lags behind the motion of the apical surface of the cell, and the individual stereocilia move semi-independently within a given hair bundle., Wang et al. use high-speed imaging to examine the in situ movement of inner hair-cell stereocilia in the mouse ear. They report that these individual cilia move semi-independently within their given bundle, which forms part of a larger complex within the inner ear, providing unique filtering opportunity for hearing in mammals.

Internal human body normal temperature fluctuates between 36.5 and 37.5°C and it is generally measured in the oral cavity. Interestingly, most electrophysiological studies on the functioning of ion channels and their role in neuronal behavior are carried out at room temperature, which usually oscillates between 22 and 24°C, even when thermosensitive channels are studied. We very often forget that if the core of the body reached that temperature, the probability of death from cardiorespiratory arrest would be extremely high. Does this mean that we are studying ion channels in dying neurons? Thousands of electrophysiological experiments carried out at these low temperatures suggest that most neurons tolerate this aggression quite well, at least for the duration of the experiments. This also seems to happen with ion channels, although studies at different temperatures indicate large changes in both, neuron and channel behavior. It is known that many chemical, physical and therefore physiological processes, depend to a great extent on body temperature. Temperature clearly affects the kinetics of numerous events such as chemical reactions or conformational changes in proteins but, what if these proteins constitute ion channels and these channels are specifically designed to detect changes in temperature? In this review, we discuss the importance of the potassium channels of the TREK subfamily, belonging to the recently discovered family of two-pore domain channels, in the transduction of thermal sensitivity in different cell types.

In the cochlea, mechano-electrical transduction is preceded by dynamic range compression. Outer hair cells (OHCs) and their voltage dependent length changes, known as electromotility, play a central role in this compression process, but the exact mechanisms are poorly understood. Here we review old and new experimental findings and show that (1) just audible high-frequency tones evoke an ∼1-microvolt AC receptor potential in basal OHCs; (2) any mechanical amplification of soft high-frequency tones by OHC motility would have an adverse effect on their audibility; (3) having a higher basolateral K+ conductance, while increasing the OHC corner frequency, does not boost the magnitude of the high-frequency AC receptor potential; (4) OHC receptor currents display a substantial rectified (DC) component; (5) mechanical DC responses (baseline shifts) to acoustic stimuli, while insignificant on the basilar membrane, can be comparable in magnitude to AC responses when recorded in the organ of Corti, both in the apex and the base. In the basal turn, the DC component may even exceed the AC component, lending support to Dallos’ suggestion that both apical and basal OHCs display a significant degree of rectification. We further show that (6) low-intensity cochlear traveling waves, by virtue of their abrupt transition from fast to slow propagation, are well suited to transport high-frequency energy with minimal losses (∼2-dB loss for 16-kHz tones in the gerbil); (7) a 90-dB, 16-kHz tone, if transmitted without loss to its tonotopic place, would evoke a destructive displacement amplitude of 564 nm. We interpret these findings in a framework in which local dissipation is regulated by OHC motility. This article is part of the Special Issue Outer hair cell Edited by Joseph Santos-Sacchi and Kumar Navaratnam.

This paper describes the first extensive study of voltage-clamp recordings from WT and mutant mouse cone photoreceptors made from dark-adapted retinas without fluorescent markers. Cones can continue to adapt rapidly in bright light, even without known mechanisms of transduction modulation., We describe the first extensive study of voltage-clamp current responses of cone photoreceptors in unlabeled, dark-adapted mouse retina using only the position and appearance of cone somata as a guide. Identification was confirmed from morphology after dye filling. Photocurrents recorded from wild-type mouse cones were biphasic with a fast cone component and a slower rod component. The rod component could be eliminated with dim background light and was not present in mouse lines lacking the rod transducin-α subunit (Gnat1−/−) or connexin 36 (Cx36−/−). Cones from Gnat1−/− or Cx36−/− mice had resting membrane potentials between −45 and −55 mV, peak photocurrents of 20–25 picoamps (pA) at a membrane potential Vm = −50 mV, sensitivities 60–70 times smaller than rods, and a total membrane capacitance two to four times greater than rods. The rate of activation (amplification constant) was largely independent of the brightness of the flash and was 1–2 s−2, less than half that of rods. The role of Ca2+-dependent transduction modulation was investigated by recording from cones in mice lacking rod transducin (Gnat1), recoverin, and/or the guanylyl-cyclase–activating proteins (GCAPs). In confirmation of previous results, responses of Gnat1−/−;Gcaps−/− cones and triple-mutant Gnat1−/−;Gcaps−/−;Rv−/− cones recovered more slowly both to light flashes and steps and were more sensitive than cones expressing the GCAPs. Cones from all four mouse lines showed significant recovery and escaped saturation even in bright background light. This recovery occurred too rapidly to be caused by pigment bleaching or metaII decay and appears to reflect some modulation of response inactivation in addition to those produced by recoverin and the GCAPs. Our experiments now make possible a more detailed understanding of the cellular physiology of mammalian cone photoreceptors and the role of conductances in the inner and outer segment in producing cone light responses.

Cells feel the forces exerted on them by the surrounding extracellular matrix (ECM) environment and respond to them. While many cell fate processes are dictated by these forces, which are highly synchronized in space and time, abnormal force transduction is implicated in the progression of many diseases (muscular dystrophy, cancer). However, material platforms that enable transient, cyclic forces in vitro to recreate an in vivo-like scenario remain a challenge. Here, we report a hydrogel system that rapidly beats (actuates) with spatio-temporal control using a near infra-red light trigger. Small, user-defined mechanical forces (~nN) are exerted on cells growing on the hydrogel surface at frequencies up to 10 Hz, revealing insights into the effect of actuation on cell migration and the kinetics of reversible nuclear translocation of the mechanosensor protein myocardin related transcription factor A, depending on the actuation amplitude, duration and frequency., Mechanotransduction of cells is of interest for a number of reasons but model in vitro systems remain a challenge. Here, the authors report on a hydrogel which changes properties upon near infrared irradiation to create cyclic forces and demonstrate the application of these gels to study mechanotransduction.

Facilely constructed metal nanoparticles with highly active sites, high economic impact, and stable electrocatalysts are crucial to developed sensory protocol for oxygen reactive species (ORS) released from cancer cells. In this study, we control the fabrication of uniformly arranged non-metal sphere catalysts that interfacially dressed by Ru nanoparticles for ultrasensitive ORS-sensing electrode patterns from cancer cells. The actively dressing of spherical ˜10 nm-Ru nano-buds/bumps that markedly interlinked entire micrometric surfaces of well-developed sulfur-doped carbon (S–C) spheres led to create catalytic surface defects through formation anisotropic wrinkling skins due to homogenous dispersion of bump mapping networks. These anisotropic protrusions create interfacial Ru@S–C surfaces with highly active catalytic sites, caves and grooves, roughness along the exterior S–C sphere–skin matrices, enabling a well-thought-out sensory signaling of ORS released from cancer cells. Specifically, our designed Ru@S–C modified electrode offers well-defined reading out transduction and end-point stimulus-responsive signaling of ORS molecules such as H2O2. The design of wrinkled Ru@S–C sphere sensory electrode provided H2O2-sensing assay with high sensitivity (314 μA μM−1 cm−2) and selectivity, low detection limit (25 nM based on 3σ), and wide linear ranges of detection reached up to 2 mM in physiological systems. Our findings indicated that highly active electrocatalysts based on wrinkling Ru@S–C can be employed for the in vitro detection of ORS released from cancer cells (HeLa cells) with high sensitivity and selectivity, low cytotoxicity, high biocompatibility, and fast response time, as well as detection of oxidative stress effects on living cells.

Transmembrane channel like protein 1 (TMC1) is likely to be a pore-forming subunit of the transduction channel of cochlear hair cells that is mechanically gated by tension on tip links in the stereocilia bundle. To localise TMC1 precisely, we labelled mice cochleae of different ages using custom-made polyclonal antibodies to TMC1 for light and transmission electron microscopy (TEM). Immunofluorescence revealed stereocilia labelling at P9 but not at P3 in apical hair cells. Immunogold labelling for TEM confirmed that labelling was absent at P3, and showed weak labelling at P6 with no stereocilia tip labelling, increasing at P9, with specific tip labelling on shorter stereocilia and some throughout the bundle. At P12 and P21, labelling was refined mostly to stereocilia tips. Quantification showed that labelling overall reached maximum by P12, labelling per tip was relatively constant from P9 to P21, but percent tips labelled was reduced from 16% to 8%. Tmc1−/− showed no labelling. Thus TMC1 occurs at the lower end of the tip link, supporting its presence in the MET complex and likely the channel. Tip localisation from P9 onwards coincides with lipoma HMGIC fusion partner-like 5 (LHFPL5), a protein that may be involved in acquiring/maintaining TMC1 localisation.

Mammals possess a remarkable ability to sense subtle temperature deviations from the thermoneutral skin temperature of ~33 °C, which ensures precise warm sensation. However, the underlying mechanisms remain unclear. Here we show that STIM1, an endoplasmic reticulum (ER) resident transmembrane protein that responds to both ER Ca2+ depletion and heat, mediates temperature-induced Ca2+ influx in skin keratinocytes via coupling to Orai Ca2+ channels in plasma membrane. Behaviorally, the keratinocyte-specific knockout of STIM1 shifts the optimal preference temperature (OPT) of mice from ~32 °C to ~34 °C, resulting in a strikingly reversed preference between 32 °C and 34 °C. Importantly, the thermally inactive STIM1-ΔK knock-in mice show altered OPT and warm preference behaviors as well, demonstrating the requirement of STIM1 thermosensitivity for warm sensation. Furthermore, the wild-type and mutant mice prefer temperatures closer to their respective OPTs, but poorly distinguish temperatures that are equally but oppositely deviated from their OPTs. Mechanistically, keratinocyte STIM1 affects the in vivo warm responses of sensory neurons by likely involving TRPA1 as a downstream transduction channel. Collectively, our data suggest that STIM1 serves as a novel in vivo thermosensor in keratinocytes to define the OPT, which might be utilized as a peripheral reference temperature for precise warm sensation.

Calcium plays important roles in the function and survival of rod and cone photoreceptor cells. Rapid regulation of calcium in the outer segments of photoreceptors is required for the modulation of phototransduction that drives the termination of the flash response as well as light adaptation in rods and cones. On a slower time scale, maintaining proper calcium homeostasis is critical for the health and survival of photoreceptors. Decades of work have established that the level of calcium in the outer segments of rods and cones is regulated by a dynamic equilibrium between influx via the transduction cGMP-gated channels and extrusion via rod- and cone-specific Na+/Ca2+, K+ exchangers (NCKXs). It had been widely accepted that the only mechanism for extrusion of calcium from rod outer segments is via the rod-specific NCKX1, while extrusion from cone outer segments is driven exclusively by the cone-specific NCKX2. However, recent evidence from mice lacking NCKX1 and NCKX2 have challenged that notion and have revealed a more complex picture, including a NCKX-independent mechanism in rods and two separate NCKX-dependent mechanisms in cones. This review will focus on recent findings on the molecular mechanisms of extrusion of calcium from the outer segments of rod and cone photoreceptors, and the functional and structural changes in photoreceptors when normal extrusion is disrupted.

The force-induced unfolding and refolding of proteins is speculated to be a key mechanism in the sensing and transduction of mechanical signals in the living cell. Yet, little evidence has been gathered for its existencein vivo. Prominently, stretch-induced unfolding is postulated to be the activation mechanism of the twitchin/titin family of autoinhibited sarcomeric kinases linked to the mechanical stress response of muscle. To test the occurrence of mechanical kinase activation in living working muscle, we generated transgenicC. elegansexpressing twitchin containing FRET moieties flanking the kinase domain and developed a quantitative technique for extracting FRET signals in freely movingC. elegans, using tracking and simultaneous imaging of animals in three channels (donor fluorescence, acceptor fluorescence, and transmitted light). Computer vision algorithms were used to extract fluorescence signals and muscle contraction states in each frame, in order to obtain fluorescence and body curvature measurements with spatial and temporal precisionin vivo. The data revealed statistically significant periodic changes in FRET signals during muscle activity, consistent with a periodic change in the conformation of twitchin kinase. We conclude that stretch-unfolding of twitchin kinase occurs in the active muscle, whereby mechanical activity titrates the signalling pathway of this cytoskeletal kinase. We anticipate that the methods we have developed here could be applied to obtainingin vivoevidence for force-induced conformational changes or elastic behavior of other proteins not only inC. elegansbut in other animals in which there is optical transparency (e.g zebrafish).

Encouraged by recent advances of biophysical and biochemical assays we introduce a 3D finite element model of an osteoblast, seeking an analogue between exogenous forces and intracellularly activated sensory mechanisms. The cell was reverse engineered and the dimensions of the internal cellular structures were based on literature data. The model was verified and validated against atomic force microscopy experiments and four loading scenarios were considered. The stress distributions developing on the main cellular components were calculated along with their corresponding strain values. The nucleus and mitochondria exhibited similar loading trends, with the mitochondria being stressed by an order of magnitude higher than the nucleus (e.g. 1.4 vs. 0.16 MPa). Equivalent stiffness was determined to increase by almost 50%, from the apex to the cell's periphery, as was the cell's elasticity, which was lowest when the load was exerted directly above the nucleus. The assessment of how extrinsic loads are propagated to a cell's internal structures is inherently a problem of high complexity. The findings presented in this study can provide important insight into biophysical and biochemical responses elicited in cells through mechanical stimulus. This was evident in both the nuclear and mitochondrial loading and would stipulate the important contribution of even more accurate models in the interpretation of cellular events. One Sentence Summary: The results of this numerical biomechanical study demonstrated that even minor extrinsic loads irrespective of the application site, are transduced by a fraction of the cytoskeleton to its internal structure (primarily to its mitochondria and secondary to the cell's nucleus), indicating mechanical stimulus as the dominant pathway to cell expression.

The radial spoke (RS) heads of motile cilia and flagella contact projections of the central pair (CP) apparatus to coordinate motility, but the morphology is distinct for protozoa and metazoa. Here we show the murine RS head is compositionally distinct from that of Chlamydomonas. Our reconstituted murine RS head core complex consists of Rsph1, Rsph3b, Rsph4a, and Rsph9, lacking Rsph6a and Rsph10b, whose orthologs exist in the protozoan RS head. We resolve its cryo-electron microscopy (cryo-EM) structure at 3.2-A resolution. Our atomic model further reveals a twofold symmetric brake pad-shaped structure, in which Rsph4a and Rsph9 form a compact body extended laterally with two long arms of twisted Rsph1 β-sheets and potentially connected dorsally via Rsph3b to the RS stalk. Furthermore, our modeling suggests that the core complex contacts the periodic CP projections either rigidly through its tooth-shaped Rsph4a regions or elastically through both arms for optimized RS–CP interactions and mechanosignal transduction.

Inner ear hair cells detect sound-induced displacements and transduce these stimuli into electrical signals in a hair bundle that consists of stereocilia that are arranged in rows of increasing height. When stereocilia are deflected, they tug on tiny (~5 nm in diameter) extracellular tip links interconnecting stereocilia, which convey forces to the mechanosensitive transduction channels. Although mechanotransduction has been studied in live hair cells for decades, the functionally important ultrastructural details of the mechanotransduction machinery at the tips of stereocilia (such as tip link dynamics or transduction-dependent stereocilia remodeling) can still be studied only in dead cells with electron microscopy. Theoretically, scanning probe techniques, such as atomic force microscopy, have enough resolution to visualize the surface of stereocilia. However, independent of imaging mode, even the slightest contact of the atomic force microscopy probe with the stereocilia bundle usually damages the bundle. Here we present a detailed protocol for the hopping probe ion conductance microscopy (HPICM) imaging of live rodent auditory hair cells. This non-contact scanning probe technique allows time lapse imaging of the surface of live cells with a complex topography, like hair cells, with single nanometers resolution and without making physical contact with the sample. The HPICM uses an electrical current passing through the glass nanopipette to detect the cell surface in close vicinity to the pipette, while a 3D-positioning piezoelectric system scans the surface and generates its image. With HPICM, we were able to image stereocilia bundles and the links interconnecting stereocilia in live auditory hair cells for several hours without noticeable damage. We anticipate that the use of HPICM will allow direct exploration of ultrastructural changes in the stereocilia of live hair cells for better understanding of their function.

Ion channels activated by mechanical inputs are important force sensing molecules in a wide array of mammalian cells and tissues. The transient receptor potential channel, TRPV4, is a polymodal, nonselective cation channel that can be activated by mechanical inputs but only if stimuli are applied directly at the interface between cells and their substrate, making this molecule a context-dependent force sensor. However, it remains unclear how TRPV4 is activated by mechanical inputs at the cell-substrate interface, which cell intrinsic and cell extrinsic parameters might modulate the mechanical activation of the channel and how mechanical activation differs from TRPV4 gating in response to other stimuli. Here we investigated the impact of substrate mechanics and cytoskeletal components on mechanically evoked TRPV4 currents and addressed how point mutations associated with TRPV4 phosphorylation and arthropathy influence mechanical activation of the channel. Our findings reveal distinct regulatory modulation of TRPV4 from the mechanically activated ion channel PIEZO1, suggesting the mechanosensitivity of these two channels is tuned in response to different parameters. Moreover, our data demonstrate that the effect of point mutations in TRPV4 on channel activation are profoundly dependent on the gating stimulus.

Thermal transient receptor potential channels play a key role in thermal sensation. Although predictive models exist for temperature-dependent transduction through these channels and for the associated sensations of pain, the ability to predict irreversible desensitization has been lacking. We explored the role of irreversible ion channel desensitization in pain sensation and hypothesized that desensitization of ion channels would follow the kinetics similar to the denaturation of catalytic enzymes. We therefore proposed a three-state model to simulate the kinetic of temperature-sensitive ion channels from the closed state through opening and irreversible thermal desensitization. We tested the model against data obtained in vivo from a feline model. The theoretical model predicts all experimental data with reasonable accuracy, and represents an important step toward the ability for understanding of the molecular basis of nociceptor signaling providing the possibility to design local anesthesia regimens and to the prediction of postoperative pain.

The integrative study that included experimentation and mathematical modeling was carried out to analyze dynamic aspects of transient Ca(2+) signaling induced by brief pulses of GPCR agonists in mesenchymal stromal cells from the human adipose tissue (AD-MSCs). The experimental findings argued for IP(3)/Ca(2+)-regulated Ca(2+) release via IP(3) receptors (IP(3)Rs) as a key mechanism mediating agonist-dependent Ca(2+) transients. The consistent signaling circuit was proposed to formalize coupling of agonist binding to Ca(2+) mobilization for mathematical modeling. The model properly simulated the basic phenomenology of agonist transduction in AD-MSCs, which mostly produced single Ca(2+) spikes upon brief stimulation. The spike-like responses were almost invariantly shaped at different agonist doses above a threshold, while response lag markedly decreased with stimulus strength. In AD-MSCs, agonists and IP(3) uncaging elicited similar Ca(2+) transients but IP(3) pulses released Ca(2+) without pronounced delay. This suggested that IP(3) production was rate-limiting in agonist transduction. In a subpopulation of AD-MSCs, brief agonist pulses elicited Ca(2+) bursts crowned by damped oscillations. With properly adjusted parameters of IP(3)R inhibition by cytosolic Ca(2+), the model reproduced such oscillatory Ca(2+) responses as well. GEM-GECO1 and R-CEPIA1er, the genetically encoded sensors of cytosolic and reticular Ca(2+), respectively, were co-expressed in HEK-293 cells that also responded to agonists in an “all-or-nothing” manner. The experimentally observed Ca(2+) signals triggered by ACh in both compartments were properly simulated with the suggested signaling circuit. Thus, the performed modeling of the transduction process provides sufficient theoretical basis for deeper interpretation of experimental findings on agonist-induced Ca(2+) signaling in AD-MSCs.

The lingual surface potential (LSP), which hyperpolarizes in response to salt and bitter stimuli, is thought to be a bioelectrical signal associated with taste transduction in humans. In contrast, a recent study reported sweet and sour stimuli to evoke a depolarization of the LSP. We questioned the origin of such a depolarization because liquid junction potentials (JPs), which arise at the interfaces of recording electrode and taste solutions, are neglected in the report. We recorded the LSPs to sucrose and NaCl solutions on the human tongue using an Ag/AgCl electrode. To estimate JPs generated by each taste solution, we made an agar model to simulate the human tongue. The lingual surface was rinsed with a 10 mM NaCl solution that mimics the sodium content of the lingual fluid. In the human tongue, sucrose dissolved in distilled water evoked a depolarizing LSP that could be attributed to JPs, resulting from the change in electrolyte concentration of the taste solution. Sucrose dissolved in 10 mM NaCl solution evoked a hyperpolarizing LSP which became more negative in a concentration-dependent manner (300–1500 mM). Lactisole (3.75 mM), an inhibitor of sweet taste, significantly reduced the LSPs and decreased perceived intensity of sweetness by human subjects. The negative JPs generated by 100 mM NaCl in the agar model were not different from the LSPs to 100 mM NaCl. When the electrolyte environment on the lingual surface is controlled for JPs, the bioelectrical signal associated with sweet taste transduction is a hyperpolarizing potential.

International audience; Plants spend most of their life oscillating around 1–3 Hz due to the effect of the wind. Therefore, stems and foliage experience repetitive mechanical stresses through these passive movements. However, the mechanism of the cellular perception and transduction of such recurring mechanical signals remains an open question. Multimeric protein complexes forming mechanosensitive (MS) channels embedded in the membrane provide an efficient system to rapidly convert mechanical tension into an electrical signal. So far, studies have mostly focused on nonoscillatory stretching of these channels. Here, we show that the plasma-membrane MS channel MscS-LIKE 10 (MSL10) from the model plant Arabidopsis thaliana responds to pulsed membrane stretching with rapid activation and relaxation kinetics in the range of 1 s. Under sinusoidal membrane stretching MSL10 presents a greater activity than under static stimulation. We observed this amplification mostly in the range of 0.3–3 Hz. Above these frequencies the channel activity is very close to that under static conditions. With a localization in aerial organs naturally submitted to wind-driven oscillations, our results suggest that the MS channel MSL10, and by extension MS channels sharing similar properties, represents a molecular component allowing the perception of oscillatory mechanical stimulations by plants.

In cell-based therapies for cartilage lesions, the main problem is still the formation of fibrous cartilage, caused by underlying de-differentiation processes ex vivo. Biophysical stimulation is a promising approach to optimize cell-based procedures and to adapt them more closely to physiological conditions. The occurrence of mechano-electrical transduction phenomena within cartilage tissue is physiological and based on streaming and diffusion potentials. The application of exogenous electric fields can be used to mimic endogenous fields and, thus, support the differentiation of chondrocytes in vitro. For this purpose, we have developed a new device for electrical stimulation of chondrocytes, which operates on the basis of capacitive coupling of alternating electric fields. The reusable and sterilizable stimulation device allows the simultaneous use of 12 cavities with independently applicable fields using only one main supply. The first parameter settings for the stimulation of human non-degenerative chondrocytes, seeded on collagen type I elastin-based scaffolds, were derived from numerical electric field simulations. Our first results suggest that applied alternating electric fields induce chondrogenic re-differentiation at the gene and especially at the protein level of human de-differentiated chondrocytes in a frequency-dependent manner. In future studies, further parameter optimizations will be performed to improve the differentiation capacity of human cartilage cells.

Rod photoreceptors can be saturated by exposure to bright background light, so that no flash superimposed on the background can elicit a detectable response. This phenomenon, called increment saturation, was first demonstrated psychophysically by Aguilar and Stiles and has since been shown in many studies to occur in single rods. Recent experiments indicate, however, that rods may be able to avoid saturation under some conditions of illumination. We now show inex vivoelectroretinogram and single-cell recordings that in continuous and prolonged exposure even to very bright light, the rods of mice from both sexes recover as much as 15% of their dark current and that responses can persist for hours. In parallel to recovery of outer segment current is an ∼10-fold increase in the sensitivity of rod photoresponses. This recovery is decreased in transgenic mice with reduced light-dependent translocation of the G protein transducin. The reduction in outer-segment transducin together with a novel mechanism of visual-pigment regeneration within the rod itself enable rods to remain responsive over the whole of the physiological range of vision. In this way, rods are able to avoid an extended period of transduction channel closure, which is known to cause photoreceptor degeneration.SIGNIFICANCE STATEMENTRods are initially saturated in bright light so that no flash superimposed on the background can elicit a detectable response. Frederiksen and colleagues show in whole retina and single-cell recordings that, if the background light is prolonged, rods slowly recover and can continue to produce significant responses over the entire physiological range of vision. Response recovery occurs by translocation of the G protein transducin from the rod outer to the inner segment, together with a novel mechanism of visual-pigment regeneration within the rod itself. Avoidance of saturation in bright light may be one of the principal mechanisms the retina uses to keep rod outer-segment channels from ever closing for too long a time, which is known to produce photoreceptor degeneration.

Understanding the mechanism by which plants respond to cold stress and strengthen their tolerance to low temperatures is an important and challenging task in plant sciences. Experiments have established that the first step in the perception and transduction of the cold stress signal consists of a transient influx of Ca2+. This Ca2+ influx triggers the activation of a cascade of phosphorylation-dephosphorylation reactions that eventually affects the expression of C-repeat-binding factors (CBFs, notably CBF3), which were shown in many plants to control resistance to cold stress by regulating the expression of cold-regulated (COR) genes. Based on experimental observations mostly made on Arabidopsis thaliana, we build a computational model for the cold response pathway in plants, from the transduction of the cold signal via the transient influx of Ca2+ to the activation of the phosphorylation cascade leading to CBF3 expression. We explore the dynamics of this regulatory network by means of numerical simulations and compare the results with experimental observations on the dynamics of the cold response, both for the wild type and for mutants. The simulations show how, in response to cold stress, a brief Ca2+ influx, which is over in minutes, is transduced along the successive steps of the network to trigger the expression of cold response genes such as CBF3 within hours. Sometimes, instead of a single Ca2+ spike the decrease in temperature brings about a train of high-frequency Ca2+ oscillations. The model is applied to both types of Ca2+ signaling. We determine the dynamics of the network in response to a series of identical cold stresses, to account for the observation of desensitization and resensitization. The analysis of the model predicts the possibility of an oscillatory expression of CBF3 originating from the negative feedback exerted by ZAT12, a factor itself controlled by CBF3. Finally, we extend the model to incorporate the circadian control of CBF3 expression, to account for the gating of the response to cold stress by the plant circadian clock.

Touch, hearing, and blood pressure regulation require mechanically gated ion channels that convert mechanical stimuli into electrical currents. One such channel is Piezo1, which plays a key role in the transduction of mechanical stimuli in humans and is implicated in diseases, such as xerocytosis and lymphatic dysplasia. There is building evidence that suggests Piezo1 can be regulated by the membrane environment, with the activity of the channel determined by the local concentration of lipids, such as cholesterol and phosphoinositides. To better understand the interaction of Piezo1 with its environment, we conduct simulations of the protein in a complex mammalian bilayer containing more than 60 different lipid types together with electrophysiology and mutagenesis experiments. We find that the protein alters its local membrane composition, enriching specific lipids and forming essential binding sites for phosphoinositides and cholesterol that are functionally relevant and often related to Piezo1-mediated pathologies. We also identify a number of key structural connections between the propeller and pore domains located close to lipid-binding sites.

Significant cellular morphology changes in renal tubules were observed in diabetes patients and animal models. However, the interaction between insulin and tubular epithelial cells microvillar structure remains obscure. To understand microvillar dynamics, we used Scanning Ion Conductance Microscope to visualize microvillar in the living cell. Here, we found two layers of microvilli on the tubular epithelial cell surface: short compact microvilli and netlike long microvilli. Insulin treatment could increase microvilli length and density. This process was mediated by the PI3K/PLCγ signaling pathway, other than the PI3K/Arp2/3 signal pathway. In conclusion, our findings present a novel insulin signaling transduction mechanism, which contributes to understanding renal tubular epithelial cell microvilli dynamic regulation.

The detection of ambient cold is critical for mammals, who use this information to avoid tissue damage by cold and to maintain stable body temperature. The transduction of information about the environmental cold is mediated by cold-sensitive ion channels expressed in peripheral sensory nerve endings in the skin. Most transduction mechanisms for detecting temperature changes identified to date depend on transient receptor potential (TRP) ion channels. Mild cooling is detected by the menthol-sensitive TRPM8 ion channel, but how painful cold is detected remains unclear. The TRPA1 ion channel, which is activated by cold in expression systems, seemed to provide an answer to this question, but whether TRPA1 is activated by cold in neurons and contributes to the sensation of cold pain continues to be a matter of debate. Recent advances have been made in this area of investigation with the identification of several potential cold-sensitive ion channels in thermosensory neurons, including two-pore domain potassium channels (K2P), GluK2 glutamate receptors, and CNGA3 cyclic nucleotide-gated ion channels. This mini-review gives a brief overview of the way by which ion channels contribute to cold sensation, discusses the controversy around the cold-sensitivity of TRPA1, and provides an assessment of some recently-proposed novel cold-transduction mechanisms. Evidence for another unidentified cold-transduction mechanism is also presented.

Decades-old model of slow adaptation in sensory hair cells of the auditory and vestibular systems requires revamp., Hair cells detect sound and motion through a mechano-electric transduction (MET) process mediated by tip links connecting shorter stereocilia to adjacent taller stereocilia. Adaptation is a key feature of MET that regulates a cell’s dynamic range and frequency selectivity. A decades-old hypothesis proposes that slow adaptation requires myosin motors to modulate the tip-link position on taller stereocilia. This “motor model” depended on data suggesting that the receptor current decay had a time course similar to that of hair-bundle creep (a continued movement in the direction of a step-like force stimulus). Using cochlear and vestibular hair cells of mice, rats, and gerbils, we assessed how modulating adaptation affected hair-bundle creep. Our results are consistent with slow adaptation requiring myosin motors. However, the hair-bundle creep and slow adaptation were uncorrelated, challenging a critical piece of evidence upholding the motor model. Considering these data, we propose a revised model of hair cell adaptation.

Over the past two decades, substantial amount of work has been conducted to characterize different odorant receptors, neuroanatomy and odorant response properties of the early olfactory system of Drosophila melanogaster. Yet many odorant receptors remain only partially characterized, and the odorant transduction process and the axon hillock spiking mechanism of the olfactory sensory neurons (OSNs) have yet to be fully determined. Identity and concentration, two key characteristics of the space of odorants, are encoded by the odorant transduction process. Detailed molecular models of the odorant transduction process are, however, scarce for fruit flies. To address these challenges we advance a comprehensive model of fruit fly OSNs as a cascade consisting of an odorant transduction process (OTP) and a biophysical spike generator (BSG). We model odorant identity and concentration using an odorant-receptor binding rate tensor, modulated by the odorant concentration profile, and an odorant-receptor dissociation rate tensor, and quantitatively describe the mechanics of the molecular ligand binding/dissociation of the OTP. We model the BSG as a Connor-Stevens point neuron. The resulting spatio-temporal encoding model of the Drosophila antenna provides a theoretical foundation for understanding the neural code of both odorant identity and odorant concentration and advances the state-of-the-art in a number of ways. First, it quantifies on the molecular level the spatio-temporal level of complexity of the transformation taking place in the antennae. The concentration-dependent spatio-temporal code at the output of the antenna circuits determines the level of complexity of olfactory processing in the downstream neuropils, such as odorant recognition and olfactory associative learning. Second, the model is biologically validated using multiple electrophysiological recordings. Third, the model demonstrates that the currently available data for odorant-receptor responses only enable the estimation of the affinity of the odorant-receptor pairs. The odorant-dissociation rate is only available for a few odorant-receptor pairs. Finally, our model calls for new experiments for massively identifying the odorant-receptor dissociation rates of relevance to flies., Author summary Identity and concentration, intrinsically embedded in the odorant space, are two key characteristics of olfactory coding that define the level of complexity of neural processing throughout the olfactory system in the fruit fly. In this paper we advance a theoretical foundation for understanding these two characteristics by quantifying mathematically the odorant space and devising a biophysical model of the olfactory sensory neurons (OSNs). To validate our modeling approach, we propose and apply an algorithm to estimate the affinity value and the dissociation rate, the two characteristics that define odorant identity, of multiple odorant-receptor pairs. We then evaluate our model with a multitude of odorant waveforms and demonstrate that the model output reproduces the temporal responses of OSNs obtained from in vivo electrophysiology recordings. Furthermore, we evaluate the model at the OSN population level and quantify on the molecular level the spatio-temporal level of complexity of the transformation taking place between the odorant space and the OSNs. The resulting concentration-dependent spatio-temporal code determines the level of complexity of the input space driving olfactory processing in the downstream neuropils. Lastly, our model demonstrates that the currently available data for OSN responses only enables estimation of affinity value. This calls for new experiments for massively identifying the odorant-receptor dissociation rates of relevance to flies.

The cochlea of the inner ear transduces sound energy into electrical signals that are essential for audition. This transduction is processed in nano-scale vibration of the cochlear sensory epithelia. In mammals, the epithelia contain various cells and structures: inner hair cells, outer hair cells (OHCs), Deiters’ cells, basilar membrane, reticular lamina, etc. The sound elicits vibration in all these constituents. Among them, only OHCs cell body actively and periodically changes in length in association with the vibration. The unique mechanical activity of OHCs modifies the sound elicited vibration in the epithelia with a feedback mechanism. Although the modification is considered to critically contribute to the high sensitivity and sharp tuning in hearing through sensory IHCs, the real motion of OHCs remains uncertain. Vibrometrical studies of cochlear mechanics has revealed important vibration of the cell bodies involving the epithelia. However, difference in vibration pattern of the apical and basal ends of the cell has remain uncertain due to low spatial resolution of the system and low reflectivity of the cells. We performed a spectral domain OCT (SD-OCT) vibrometry by using the modified commercial SD-OCT system. Because the broad spectral bandwidth and strong power of the light source improve a performance of OCT systems in both of imaging and vibrometry, we introduced a supercontinuum light source into the commercial system. Our system achieved cellular-level tomographic imaging and subnano-scale vibration measurement in the transparent epithelia with the recording time of 100 ms in in vivo animal.

Retinal rod and cone photoreceptors mediate vision in dim and bright light, respectively, by transducing absorbed photons into electrical signals. Their phototransduction mechanisms are essentially identical. However, one distinct difference between them is that, whereas a rod visual pigment remains stable in darkness, a cone pigment is liable to spontaneously dissociate into apo-opsin and retinal (the chromophore) without isomerization. This cone-pigment property has long been reported, but it has often been overlooked in the field. Importantly, because apo-opsin has constitutive activity, it triggers transduction signaling to produce electrical noise even in darkness. Currently, the precise apo-opsin content in dark-adapted cones is unknown, as is its associated dark activity. We report here a systematic study of red (L), green (M) and blue (S) cones, finding different percentages of apo-opsin across them in darkness. Surprisingly, we also found that apo-opsin signaling in L and M cones in darkness is actually much higher than the well-known holo-pigment thermal isomerization activity. As such, given the very low signal amplification at the pigment-to-transducin/phosphodiesterase step in cone transduction, apo-opsin noise may not be easily distinguishable from light-triggered responses and may affect cone vision near threshold.

Sensory hair cells of the inner ear rely on the hair bundle, a cluster of actin-filled stereocilia, to transduce auditory and vestibular stimuli into electrical impulses. Because they are long and thin projections, stereocilia are most prone to damage at the point where they insert into the hair cell’s soma. Moreover, this is the site of stereocilia pivoting, the mechanical movement that induces transduction, which additionally weakens this area mechanically. To bolster this fragile area, hair cells construct a dense core called the rootlet at the base of each stereocilium, which extends down into the actin meshwork of the cuticular plate and firmly anchors the stereocilium. Rootlets are constructed with tightly packed actin filaments that extend from stereocilia actin filaments which are wrapped with TRIOBP; in addition, many other proteins contribute to the rootlet and its associated structures. Rootlets allow stereocilia to sustain innumerable deflections over their lifetimes and exemplify the unique manner in which sensory hair cells exploit actin and its associated proteins to carry out the function of mechanotransduction.

The basolateral membrane of the mammalian inner hair cell (IHC) expresses large voltage and Ca2+ gated outward K+ currents. To quantify how the voltage-dependent activation of the K+ channels affects the functionality of the auditory nerve innervating the IHC, this study adopts a model of mechanical-to-neural transduction in which the basolateral K+ conductances of the IHC can be made voltage-dependent or not. The model shows that the voltage-dependent activation of the K+ channels (i) enhances the phase-locking properties of the auditory fiber (AF) responses; (ii) enables the auditory nerve to encode a large dynamic range of sound levels; (iii) enables the AF responses to synchronize precisely with the envelope of amplitude modulated stimuli; and (iv), is responsible for the steep offset responses of the AFs. These results suggest that the basolateral K+ channels play a major role in determining the well-known response properties of the AFs and challenge the classical view that describes the IHC membrane as an electrical low-pass filter. In contrast to previous models of the IHC-AF complex, this study ascribes many of the AF response properties to fairly basic mechanisms in the IHC membrane rather than to complex mechanisms in the synapse.

Hearing relies on the conversion of mechanical stimuli into electrical signals. In vertebrates, this process of mechano-electrical transduction (MET) is performed by specialized receptors of the inner ear, the hair cells. Each hair cell is crowned by a hair bundle, a cluster of microvilli that pivot in response to sound vibrations, causing the opening and closing of mechanosensitive ion channels. Mechanical forces are projected onto the channels by molecular springs called tip links. Each tip link is thought to connect to a small number of MET channels that gate cooperatively and operate as a single transduction unit. Pushing the hair bundle in the excitatory direction opens the channels, after which they rapidly reclose in a process called fast adaptation. It has been experimentally observed that the hair cell’s biophysical properties mature gradually during postnatal development: the maximal transduction current increases, sensitivity sharpens, transduction occurs at smaller hair-bundle displacements, and adaptation becomes faster. Similar observations have been reported during tip-link regeneration after acoustic damage. Moreover, when measured at intermediate developmental stages, the kinetics of fast adaptation varies in a given cell depending on the magnitude of the imposed displacement. The mechanisms underlying these seemingly disparate observations have so far remained elusive. Here, we show that these phenomena can all be explained by the progressive addition of MET channels of constant properties, which populate the hair bundle first as isolated entities, then progressively as clusters of more sensitive, cooperative MET channels. As the proposed mechanism relies on the difference in biophysical properties between isolated and clustered channels, this work highlights the importance of cooperative interactions between mechanosensitive ion channels for hearing.SIGNIFICANCEHair cells are the sensory receptors of the inner ear that convert mechanical stimuli into electrical signals transmitted to the brain. Sensitivity to mechanical stimuli and the kinetics of mechanotransduction currents change during hair-cell development. The same trend, albeit on a shorter timescale, is also observed during hair-cell recovery from acoustic trauma. Furthermore, the current kinetics in a given hair cell depends on the stimulus magnitude, and the degree of that dependence varies with development. These phenomena have so far remained unexplained. Here, we show that they can all be reproduced using a single unifying mechanism: the progressive formation of channel pairs, in which individual channels interact through the lipid bilayer and gate cooperatively.

The mechanosensitivity of cells depends on the lipid-protein interactions of the plasma membrane. Affectations in the lipid region of the plasma membrane affect the transduction of mechanical forces, and any molecule that modifies the biophysical integrity of the lipid bilayer can alter the mechanical activity of the proteins inside the membrane. To understand whether inhibitors of mechanically activated ion channels affect the mechanical properties of the plasma membrane, we evaluated the rigidity of the membrane of sensory neurons of the DRG of mice using a variant of the scanning ion conductance microscopy method, which allows us to calculate the Young's modulus of individual cells before and after the perfusion of different doses of Gd3+, ruthenium red and GsMTx-4. Our results suggest that these molecules compromise the membrane by increasing the Young's modulus value, which indicates that the membrane becomes more rigid; these compounds act through different mechanisms and by a non-specific manner, each one shows a certain preference for specific cell subpopulations, depending on their cell size and their reactivity to isolectin B4. Our results support the idea that the biophysical properties that result from the interactions that arise in the membranes are part of the mechanotransduction process.

Entorhinal cortex (EC) is the initial brain region that suffers abnormal tau in Alzheimer's disease (AD). Whether overexpression of human tau (htau40) in EC disrupts cognitive function and synaptic plasticity in AD has not been fully elucidated. To investigate the effects of htau40 on the pathology and associated mechanisms of early stage of AD in mice, an adeno-associated virus-based htau40 transduced in medial EC (mEC) mouse model was established. The results showed that htau40 restrictedly expressed in mEC after transduction. The memory function and long-term potentiation (LTP) of dentate gyrus (DG) were significantly impaired by overexpression of htau40 in mEC after transduction at 3 and 6 months. However, the abnormities of neurons and neurotransmitters in mEC started at just 1 month after transduction. The resting membrane potential was increased and paired pulse facilitates was depressed, but the action potential amplitude, threshold, and half width did not alter after htau40 transduction at 1 month. The levels of inhibitory neurotransmitters were up regulated whereas level of lactate was decreased. Our study demonstrated that htau40 in mEC impaired cognition and synaptic plasticity of perforant path (PP)-DG, which simulated early stage of AD and elucidated the mechanism of that htau40 overexpression in mEC may be associated with the development of AD.

Microtubules, the most rigid components of the cytoskeleton, can be key transduction elements between external forces and the cellular environment. Mechanical forces induce microtubule deformation, which is presumed to be critical for the mechanoregulation of cellular events. However, concrete evidence is lacking. In this work, with high-speed atomic force microscopy, we unravel how microtubule deformation regulates the translocation of the microtubule-associated motor protein kinesin-1, responsible for intracellular transport. Our results show that the microtubule deformation by bending impedes the translocation dynamics of kinesins along them. Molecular dynamics simulation shows that the hindered translocation of kinesins can be attributed to an enhanced affinity of kinesins to the microtubule structural units in microtubules deformed by bending. This study advances our understanding of the role of cytoskeletal components in mechanotransduction.

Certain eukaryotic cells are believed to sense and respond to vibrational stimuli that are too small even to be transduced by mechanosensitive ion channels. One possible mechanism of signal amplification and transduction is torsional and translational resonance of the nucleus, the stiffest and densest organelle in a eukaryotic cell. To explore this possibility, we developed a theoretical model that analyzes the natural frequencies of torsional and translational vibrations of the nucleus. The model predicts that the natural frequency for torsional vibration is dependent upon cytoskeletal contractility, while that for translational vibration is dependent upon cytoskeletal stiffness. Further analysis across many species and cell types suggests that, for most eukaryotic cells, torsional vibration is the dominant form of nuclear response to higher frequency stimuli, providing a new potential mechanism for frequency-based mechanotransduction.