The Journal of general physiology
Nourse, JL;Leung, VM;Abuwarda, H;Evans, EL;Izquierdo-Ortiz, E;Ly, AT;Truong, N;Smith, S;Bhavsar, H;Bertaccini, G;Monuki, ES;Panicker, MM;Pathak, MM;
PMID: 36069933 | DOI: 10.1085/jgp.202213084
Mechanical forces and tissue mechanics influence the morphology of the developing brain, but the underlying molecular mechanisms have been elusive. Here, we examine the role of mechanotransduction in brain development by focusing on Piezo1, a mechanically activated ion channel. We find that Piezo1 deletion results in a thinner neuroepithelial layer, disrupts pseudostratification, and reduces neurogenesis in E10.5 mouse embryos. Proliferation and differentiation of Piezo1 knockout (KO) mouse neural stem cells (NSCs) isolated from E10.5 embryos are reduced in vitro compared to littermate WT NSCs. Transcriptome analysis of E10.5 Piezo1 KO brains reveals downregulation of the cholesterol biosynthesis superpathway, in which 16 genes, including Hmgcr, the gene encoding the rate-limiting enzyme of the cholesterol biosynthesis pathway, are downregulated by 1.5-fold or more. Consistent with this finding, membrane lipid composition is altered, and the cholesterol levels are reduced in Piezo1 KO NSCs. Cholesterol supplementation of Piezo1 KO NSCs partially rescues the phenotype in vitro. These findings demonstrate a role for Piezo1 in the neurodevelopmental process that modulates the quantity, quality, and organization of cells by influencing cellular cholesterol metabolism. Our study establishes a direct link in NSCs between PIEZO1, intracellular cholesterol levels, and neural development.
Li, X;Zhang, C;Bowman, HH;Stambough, JB;Stronach, BM;Mears, SC;Barnes, LC;Ambrogini, E;Xiong, J;
PMID: 37147884 | DOI: 10.1111/acel.13846
As we age, our bones undergo a process of loss, often accompanied by muscle weakness and reduced physical activity. This is exacerbated by decreased responsiveness to mechanical stimulation in aged skeleton, leading to the hypothesis that decreased mechanical stimulation plays an important role in age-related bone loss. Piezo1, a mechanosensitive ion channel, is critical for bone homeostasis and mechanotransduction. Here, we observed a decrease in Piezo1 expression with age in both murine and human cortical bone. Furthermore, loss of Piezo1 in osteoblasts and osteocytes resulted in an increase in age-associated cortical bone loss compared to control mice. The loss of cortical bone was due to an expansion of the endosteal perimeter resulting from increased endocortical resorption. In addition, expression of Tnfrsf11b, encoding anti-osteoclastogenic protein OPG, decreases with Piezo1 in vitro and in vivo in bone cells, suggesting that Piezo1 suppresses osteoclast formation by promoting Tnfrsf11b expression. Our results highlight the importance of Piezo1-mediated mechanical signaling in protecting against age-associated cortical bone loss by inhibiting bone resorption in mice.
Ichijo, R;Maki, K;Kabata, M;Murata, T;Nagasaka, A;Ishihara, S;Haga, H;Honda, T;Adachi, T;Yamamoto, T;Toyoshima, F;
| DOI: 10.1038/s43587-022-00244-6
Stem cell loss causes tissue deterioration associated with aging. The accumulation of genomic and oxidative stress-induced DNA damage is an intrinsic cue for stem cell loss1 [/articles/s43587-022-00244-6#ref-CR1],2 [/articles/s43587-022-00244-6#ref-CR2]; however, whether there is an external microenvironmental cue that triggers stem cell loss remains unclear. Here we report that the involution of skin vasculature causes dermal stiffening that augments the differentiation and hemidesmosome fragility of interfollicular epidermal stem cells (IFESCs) in aged mouse skin. Aging-related IFESC dysregulation occurs in plantar and tail skin, and is correlated with prolonged calcium influx, which is contributed by the mechanoresponsive ion channel Piezo1 (ref. 3 [/articles/s43587-022-00244-6#ref-CR3]). Epidermal deletion of Piezo1 ameliorated IFESC dysregulation in aged skin, whereas Piezo1 activation augmented IFESC differentiation and hemidesmosome fragility in young mice. The dermis stiffened with age, which was accompanied by dermal vasculature atrophy. Conversely, induction of the dermal vasculature softened the dermis and ameliorated IFESC dysregulation in aged skin. Single-cell RNA sequencing of dermal fibroblasts identified an aging-associated anti-angiogenetic secretory molecule, pentraxin 3 (ref. 4 [/articles/s43587-022-00244-6#ref-CR4]), which caused dermal sclerotization and IFESC dysregulation in aged skin. Our findings show that the vasculature softens the microenvironment for stem cell maintenance and provide a potential mechanobiology-based therapeutic strategy against skin disorders in aging.
Hennes A, Held K, Boretto M, De Clercq K, Van den Eynde C, Vanhie A, Van Ranst N, Benoit M, Luyten C, Peeraer K, Tomassetti C, Meuleman C, Voets T, Vankelecom H, Vriens J.
PMID: 30741991 | DOI: 10.1038/s41598-018-38376-8
Successful pregnancy requires the establishment of a complex dialogue between the implanting embryo and the endometrium. Knowledge regarding molecular candidates involved in this early communication process is inadequate due to limited access to primary human endometrial epithelial cells (EEC). Since pseudo-pregnancy in rodents can be induced by mechanical scratching of an appropriately primed uterus, this study aimed to investigate the expression of mechanosensitive ion channels in EEC. Poking of EEC provoked a robust calcium influx and induced an increase in current densities, which could be blocked by an inhibitor of mechanosensitive ion channels. Interestingly, RNA expression studies showed high expression of PIEZO1 in EEC of mouse and human. Additional analysis provided further evidence for the functional expression of PIEZO1 since stimulation with Yoda1, a chemical agonist of PIEZO1, induced increases in intracellular calcium concentrations and current densities in EEC. Moreover, the ion channel profile of human endometrial organoids (EMO) was validated as a representative model for endometrial epithelial cells. Mechanical and chemical stimulation of EMO induced strong calcium responses supporting the hypothesis of mechanosensitive ion channel expression in endometrial epithelial cells. In conclusion, EEC and EMO functionally express the mechanosensitive PIEZO1 channel that could act as a potential target for the development of novel treatments to further improve successful implantation processes.
Generation of hiPSC-derived low threshold mechanoreceptors containing axonal termini resembling bulbous sensory nerve endings and expressing Piezo1 and Piezo2
Zhu, S;Stanslowsky, N;Fernández-Trillo, J;Mamo, T;Yu, P;Kalmbach, N;Ritter, B;Eggenschwiler, R;Ouwendijk, W;Mzinza, D;Tan, L;Leffler, A;Spohn, M;Brown, R;Kropp, K;Kaever, V;Ha, T;Narayanan, P;Grundhoff, A;Förster, R;Schambach, A;Verjans, G;Schmidt, M;Kispert, A;Cantz, T;Gomis, A;Wegner, F;Viejo-Borbolla, A;
| DOI: 10.1016/j.scr.2021.102535
Somatosensory low threshold mechanoreceptors (LTMRs) sense innocuous mechanical forces, largely through specialized axon termini termed sensory nerve endings, where the mechanotransduction process initiates upon activation of mechanotransducers. In humans, a subset of sensory nerve endings is enlarged, forming bulb-like expansions, termed bulbous nerve endings. There is no in vitro human model to study these neuronal endings. Piezo2 is the main mechanotransducer found in LTMRs. Recent evidence shows that Piezo1, the other mechanotransducer considered absent in dorsal root ganglia (DRG), is expressed at low level in somatosensory neurons. We established a differentiation protocol to generate, from iPSC-derived neuronal precursor cells, human LTMR recapitulating bulbous sensory nerve endings and heterogeneous expression of Piezo1 and Piezo2. The derived neurons express LTMR-specific genes, convert mechanical stimuli into electrical signals and have specialized axon termini that morphologically resemble bulbous nerve endings. Piezo2 is concentrated within these enlarged axon termini. Some derived neurons express low level Piezo1, and a subset co-express both channels. Thus, we generated a unique, iPSCs-derived human model that can be used to investigate the physiology of bulbous sensory nerve endings, and the role of Piezo1 and 2 during mechanosensation.
Functional roles for Piezo1 and Piezo2 in urothelial mechanotransduction and lower urinary tract interoception
Dalghi, MG;Ruiz, WG;Clayton, DR;Montalbetti, N;Daugherty, SL;Beckel, JM;Carattino, MD;Apodaca, G;
PMID: 34464353 | DOI: 10.1172/jci.insight.152984
The mechanisms that link visceral mechanosensation to the perception of internal organ status (i.e., interoception) remain elusive. In response to bladder filling, the urothelium releases ATP, which is hypothesized to stimulate voiding function by communicating the degree of bladder fullness to subjacent tissues including afferent nerve fibers. To determine if PIEZO channels function as mechanosensors in these events, we generated conditional urothelial Piezo1-, Piezo2-, and dual Piezo1/2-knockout (KO) mice. While functional PIEZO1 channels were expressed in all urothelial cell layers, Piezo1-KO mice had a limited phenotype. Piezo2 expression was limited to a small subset of superficial umbrella cells, yet male Piezo2-KO mice exhibited incontinence (i.e., leakage) when their voiding behavior was monitored during their active dark phase. Dual Piezo1/2-KO mice had the most significant phenotype, characterized by decreased urothelial responses to mechanical stimulation, diminished ATP release, bladder hypoactivity in anesthetized Piezo1/2-KO females, but not male ones, and urinary incontinence in both male and female Piezo1/2-KO mice during their dark phase, but not inactive light one. Our studies reveal that the urothelium functions in a sex and circadian manner to link urothelial PIEZO1/2 channel-driven mechanotransduction to normal voiding function and behavior, and in the absence of these signals, bladder dysfunction ensues.
Ali Marandi Ghoddousi, R;Magalong, VM;Kamitakahara, AK;Levitt, P;
PMID: 36313803 | DOI: 10.1016/j.crmeth.2022.100316
Spatial gene expression, achieved classically through in situ hybridization, is a fundamental tool for topographic phenotyping of cell types in the nervous system. Newly developed techniques allow for visualization of multiple mRNAs at single-cell resolution and greatly expand the ability to link gene expression to tissue topography, yet there are challenges in efficient quantification and analysis of these high-dimensional datasets. We have therefore developed the single-cell automated multiplex pipeline for RNA (SCAMPR), facilitating rapid and accurate segmentation of neuronal cell bodies using a dual immunohistochemistry-RNAscope protocol and quantification of low- and high-abundance mRNA signals using open-source image processing and automated segmentation tools. Proof of principle using SCAMPR focused on spatial mapping of gene expression by peripheral (vagal nodose) and central (visual cortex) neurons. The analytical effectiveness of SCAMPR is demonstrated by identifying the impact of early life stress on gene expression in vagal neuron subtypes.
Lee, P;Ha, T;Choi, H;Lee, S;Kim, H;Kim, C;Hong, G;
| DOI: 10.2139/ssrn.4392157
Mechanosensation begins with the sensing of pressure by mechanically activated (MA) channels in the nerve endings of dorsal root ganglion (DRG) neurons. Piezo1, a fast-inactivating MA channel, has surfaced to be involved in pruriception. However, the pressure-dependent activation mechanism and its physiological role in mechanical pain remain unidentified. Here, we report that _Piezo1_ is expressed in a small DRG subpopulation, which is largely positive for _TRPV1_ rather than _MRGPRD_, which is known for nociceptors. To investigate the molecular function of _Piezo1_ in DRG neurons, we reclassified DRG neurons based on the MA current type. The silencing of the _Piezo1_ gene resulted in two subgroups—intermediately adapting (IA) and intermediately slowly adapting (ISA) responders of DRG neurons. Silencing _Piezo1_ in mice via specific lumbar DRG-targeted ganglionic injection of shRNA virus reduced tactile pain hypersensitivity in formalin- and carrageenan-dependent inflammation. _Piezo1_ mediates mechanical pain by acting as a nociceptive MA channel.
Woo SH, Lukacs V, de Nooij JC, Zaytseva D, Criddle CR, Francisco A, Jessell TM, Wilkinson KA, Patapoutian A.
PMID: 26551544 | DOI: 10.1038/nn.4162.
Proprioception, the perception of body and limb position, is mediated by proprioceptors, specialized mechanosensory neurons that convey information about the stretch and tension experienced by muscles, tendons, skin and joints. In mammals, the molecular identity of the stretch-sensitive channel that mediates proprioception is unknown. We found that the mechanically activated nonselective cation channel Piezo2 was expressed in sensory endings of proprioceptors innervating muscle spindles and Golgi tendon organs in mice. Two independent mouse lines that lack Piezo2 in proprioceptive neurons showed severely uncoordinated body movements and abnormal limb positions. Moreover, the mechanosensitivity of parvalbumin-expressing neurons that predominantly mark proprioceptors was dependent on Piezo2 expression in vitro, and the stretch-induced firing of proprioceptors in muscle-nerve recordings was markedly reduced in Piezo2-deficient mice. Together, our results indicate that Piezo2 is the major mechanotransducer of mammalian proprioceptors.
Nonomura K, Woo SH, Chang RB, Gillich A, Qiu Z, Francisco AG, Ranade SS, Liberles SD, Patapoutian A.
PMID: 28002412 | DOI: 10.1038/nature20793
Respiratory dysfunction is a notorious cause of perinatal mortality in infants and sleep apnoea in adults, but the mechanisms of respiratory control are not clearly understood. Mechanical signals transduced by airway-innervating sensory neurons control respiration; however, the physiological significance and molecular mechanisms of these signals remain obscured. Here we show that global and sensory neuron-specific ablation of the mechanically activated ion channel Piezo2 causes respiratory distress and death in newborn mice. Optogenetic activation of Piezo2+ vagal sensory neurons causes apnoea in adult mice. Moreover, induced ablation of Piezo2 in sensory neurons of adult mice causes decreased neuronal responses to lung inflation, an impaired Hering-Breuer mechanoreflex, and increased tidal volume under normal conditions. These phenotypes are reproduced in mice lacking Piezo2 in the nodose ganglion. Our data suggest that Piezo2 is an airway stretch sensor and that Piezo2-mediated mechanotransduction within various airway-innervating sensory neurons is critical for establishing efficient respiration at birth and maintaining normal breathing in adults.
Yao, Y;Barger, Z;Saffari Doost, M;Tso, CF;Darmohray, D;Silverman, D;Liu, D;Ma, C;Cetin, A;Yao, S;Zeng, H;Dan, Y;
PMID: 36170850 | DOI: 10.1016/j.neuron.2022.08.027
Sleep disturbances are strongly associated with cardiovascular diseases. Baroreflex, a basic cardiovascular regulation mechanism, is modulated by sleep-wake states. Here, we show that neurons at key stages of baroreflex pathways also promote sleep. Using activity-dependent genetic labeling, we tagged neurons in the nucleus of the solitary tract (NST) activated by blood pressure elevation and confirmed their barosensitivity with optrode recording and calcium imaging. Chemogenetic or optogenetic activation of these neurons promoted non-REM sleep in addition to decreasing blood pressure and heart rate. GABAergic neurons in the caudal ventrolateral medulla (CVLM)-a downstream target of the NST for vasomotor baroreflex-also promote non-REM sleep, partly by inhibiting the sympathoexcitatory and wake-promoting adrenergic neurons in the rostral ventrolateral medulla (RVLM). Cholinergic neurons in the nucleus ambiguous-a target of the NST for cardiac baroreflex-promoted non-REM sleep as well. Thus, key components of the cardiovascular baroreflex circuit are also integral to sleep-wake brain-state regulation.
Nature. 2014 Dec 4;516(7529):121-5.
Ranade SS, Woo SH, Dubin AE, Moshourab RA, Wetzel C, Petrus M, Mathur J, Bégay V, Coste B, Mainquist J, Wilson AJ, Francisco AG, Reddy K, Qiu Z, Wood JN, Lewin GR, Patapoutian A.
PMID: 25471886 | DOI: 10.1038/nature13980.
The sense of touch provides critical information about our physical environment by transforming mechanical energy into electrical signals1. It is postulated that mechanically activated cation channels initiate touch sensation, but the identity of these molecules in mammals has been elusive2. Piezo2 is a rapidly adapting, mechanically activated ion channel expressed in a subset of sensory neurons of the dorsal root ganglion and in cutaneous mechanoreceptors known as Merkel-cell–neurite complexes3, 4. It has been demonstrated that Merkel cells have a role in vertebrate mechanosensation using Piezo2, particularly in shaping the type of current sent by the innervating sensory neuron4, 5, 6; however, major aspects of touch sensation remain intact without Merkel cell activity4, 7. Here we show that mice lacking Piezo2 in both adult sensory neurons and Merkel cells exhibit a profound loss of touch sensation. We precisely localize Piezo2 to the peripheral endings of a broad range of low-threshold mechanoreceptors that innervate both hairy and glabrous skin. Most rapidly adapting, mechanically activated currents in dorsal root ganglion neuronal cultures are absent in Piezo2 conditional knockout mice, and ex vivo skin nerve preparation studies show that the mechanosensitivity of low-threshold mechanoreceptors strongly depends on Piezo2. This cellular phenotype correlates with an unprecedented behavioural phenotype: an almost complete deficit in light-touch sensation in multiple behavioural assays, without affecting other somatosensory functions. Our results highlight that a single ion channel that displays rapidly adapting, mechanically activated currents in vitro is responsible for the mechanosensitivity of most low-threshold mechanoreceptor subtypes involved in innocuous touch sensation. Notably, we find that touch and pain sensation are separable, suggesting that as-yet-unknown mechanically activated ion channel(s) must account for noxious (painful) mechanosensation.
