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Light-controllable tools provide powerful means to manipulate and interrogate brain function with relatively low invasiveness and high spatiotemporal precision. Although optogenetic approaches permit neuronal excitation or inhibition at the network level, other technologies, such as optopharmacology (also known as photopharmacology) have emerged that provide molecular-level control by endowing light sensitivity to endogenous biomolecules. In this Review, we discuss the challenges and opportunities of photocontrolling native neuronal signalling pathways, focusing on ion channels and neurotransmitter receptors. We describe existing strategies for rendering receptors and channels light sensitive and provide an overview of the neuroscientific insights gained from such approaches. At the crossroads of chemistry, protein engineering and neuroscience, optopharmacology offers great potential for understanding the molecular basis of brain function and behaviour, with promises for future therapeutics.

Merely touching the pancreas can lead to premature zymogen activation and pancreatitis but the mechanism is not completely understood. Here we demonstrate that pancreatic acinar cells express the mechanoreceptor Piezo1 and application of pressure within the gland produces pancreatitis. To determine if this effect is through Piezo1 activation, we induce pancreatitis by intrapancreatic duct instillation of the Piezo1 agonist Yoda1. Pancreatitis induced by pressure within the gland is prevented by a Piezo1 antagonist. In pancreatic acinar cells, Yoda1 stimulates calcium influx and induces calcium-dependent pancreatic injury. Finally, selective acinar cell-specific genetic deletion of Piezo1 protects mice against pressure-induced pancreatitis. Thus, activation of Piezo1 in pancreatic acinar cells is a mechanism for pancreatitis and may explain why pancreatitis develops following pressure on the gland as in abdominal trauma, pancreatic duct obstruction, pancreatography, or pancreatic surgery. Piezo1 blockade may prevent pancreatitis when manipulation of the gland is anticipated. Manipulation of the pancreas during surgery can induce acute pancreatitis due to zymogen activation. Here the authors show that the mechanoreceptor Piezo1 is activated by pressure and its activation leads to calcium dependent pancreatic injury whereas its inhibition is protective against pancreatitis.

Increased pulmonary microvessel pressure experienced in left heart failure, head trauma, or high altitude can lead to endothelial barrier disruption referred to as capillary “stress failure” that causes leakage of protein-rich plasma and pulmonary edema. However, little is known about vascular endothelial sensing and transduction of mechanical stimuli inducing endothelial barrier disruption. Piezo1, a mechanosensing ion channel expressed in endothelial cells (ECs), is activated by elevated pressure and other mechanical stimuli. Here, we demonstrate the involvement of Piezo1 in sensing increased lung microvessel pressure and mediating endothelial barrier disruption. Studies were made in mice in which Piezo1 was deleted conditionally in ECs (Piezo1iΔEC), and lung microvessel pressure was increased either by raising left atrial pressure or by aortic constriction. We observed that lung endothelial barrier leakiness and edema induced by raising pulmonary microvessel pressure were abrogated in Piezo1iΔEC mice. Piezo1 signaled lung vascular hyperpermeability by promoting the internalization and degradation of the endothelial adherens junction (AJ) protein VE-cadherin. Breakdown of AJs was the result of activation of the calcium-dependent protease calpain and degradation of the AJ proteins VE-cadherin, β-catenin, and p120-catenin. Deletion of Piezo1 in ECs or inhibition of calpain similarly prevented reduction in the AJ proteins. Thus, Piezo1 activation in ECs induced by elevated lung microvessel pressure mediates capillary stress failure and edema formation secondary to calpain-induced disruption of VE-cadherin adhesion. Inhibiting Piezo1 signaling may be a useful strategy to limit lung capillary stress failure injury in response to elevated vascular pressures.

Hypertrophic scar (HS) formation is a skin fibroproliferative disease that occurs following a cutaneous injury, leading to functional and cosmetic impairment. To date, few therapeutic treatments exhibit satisfactory outcomes. The mechanical force has been shown to be a key regulator of HS formation, but the underlying mechanism is not completely understood. The Piezo1 channel has been identified as a novel mechanically activated cation channel (MAC) and is reportedly capable of regulating force-mediated cellular biological behaviors. However, the mechanotransduction role of Piezo1 in HS formation has not been investigated. In this work, we found that Piezo1 was overexpressed in myofibroblasts of human and rat HS tissues. In vitro, cyclic mechanical stretch (CMS) increased Piezo1 expression and Piezo1-mediated calcium influx in human dermal fibroblasts (HDFs). In addition, Piezo1 activity promoted HDFs proliferation, motility, and differentiation in response to CMS. More importantly, intradermal injection of GsMTx4, a Piezo1-blocking peptide, protected rats from stretch-induced HS formation. Together, Piezo1 was shown to participate in HS formation and could be a novel target for the development of promising therapies for HS formation.

Targeting receptor proteins, such as ligand-gated ion channels and G protein-coupled receptors, has directly enabled the discovery of most drugs developed to modulate receptor signalling. However, as the search for novel and improved drugs continues, an innovative approach — targeting receptor complexes — is emerging. Receptor complexes are composed of core receptor proteins and receptor-associated proteins, which have profound effects on the overall receptor structure, function and localization. Hence, targeting key protein–protein interactions within receptor complexes provides an opportunity to develop more selective drugs with fewer side effects. In this Review, we discuss our current understanding of ligand-gated ion channel and G protein-coupled receptor complexes and discuss strategies for their pharmacological modulation. Although such strategies are still in preclinical development for most receptor complexes, they exemplify how receptor complexes can be drugged, and lay the groundwork for this nascent area of research.Targeting protein complexes, including those containing G protein-coupled receptors or ligand-gated ion channels, could provide opportunities to increase the target and functional selectivity of novel drugs compared with existing therapies, which only target the receptors. This Review discusses the landscape of ligand-gated ion channel and G protein-coupled receptor complexes as therapeutic targets, as well as strategies for their pharmacological modulation.

P2X receptors are cation-selective ion channels gated by extracellular ATP, and are implicated in diverse physiological processes, from synaptic transmission to inflammation to the sensing of taste and pain. Because P2X receptors are not related to other ion channel proteins of known structure, there is at present no molecular foundation for mechanisms of ligand-gating, allosteric modulation and ion permeation. Here we present crystal structures of the zebrafish P2X(4) receptor in its closed, resting state. The chalice-shaped, trimeric receptor is knit together by subunit-subunit contacts implicated in ion channel gating and receptor assembly. Extracellular domains, rich in beta-strands, have large acidic patches that may attract cations, through fenestrations, to vestibules near the ion channel. In the transmembrane pore, the 'gate' is defined by an approximately 8 A slab of protein. We define the location of three non-canonical, intersubunit ATP-binding sites, and suggest that ATP binding promotes subunit rearrangement and ion channel opening.

Proton conductance across cell membranes serves many biological functions, ranging from the regulation of intracellular and extracellular pH to the generation of electrical signals that lead to sour taste perception. Otopetrins (OTOPs) are a conserved, eukaryotic family of proton-selective ion channels, one of which (OTOP1) serves as a gustatory sensor for sour tastes and ammonium chloride. As the functional properties and structures of OTOP channels were only recently described, there are presently few tools available to modulate their activity. Here, we perform subsequent rounds of molecular docking-based virtual screening against the structure of zebrafish OTOP1, followed by functional testing using whole-cell patch-clamp electrophysiology, and identify several small molecule inhibitors that are effective in the low-to-mid µM range. Cryo-electron microscopy structures reveal inhibitor binding sites in the intrasubunit interface that are validated by functional testing of mutant channels. Our findings reveal pockets that can be targeted for small molecule discovery to develop modulators for Otopetrins. Such modulators can serve as useful toolkit molecules for future investigations of structure-function relationships or physiological roles of Otopetrins. Otopetrins form proton channels in animals ranging from nematodes to humans. Here, authors identify small molecule inhibitors and characterize their binding in the intrasubunit interface region, thus highlighting the area for pharmacological targeting for channel modulation.

Ion transport across the cell membrane mediated by channels and carriers participate in the regulation of tumour cell survival, death and motility. Moreover, the altered regulation of channels and carriers is part of neoplastic transformation. Experimental modification of channel and transporter activity impacts tumour cell survival, proliferation, malignant progression, invasive behaviour or therapy resistance of tumour cells. A wide variety of distinct Ca2+ permeable channels, K+ channels, Na+ channels and anion channels have been implicated in tumour growth and metastasis. Further experimental information is, however, needed to define the specific role of individual channel isoforms critically important for malignancy. Compelling experimental evidence supports the assumption that the pharmacological inhibition of ion channels or their regulators may be attractive targets to counteract tumour growth, prevent metastasis and overcome therapy resistance of tumour cells. This short review discusses the role of Ca2+ permeable channels, K+ channels, Na+ channels and anion channels in tumour growth and metastasis and the therapeutic potential of respective inhibitors.

Targeted protein degradation/downregulation (TPD/TPDR) is a disruptive paradigm for developing therapeutics. <2% of ~600 E3 ligases have been exploited for this modality, and efficacy for multi-subunit ion channels has not been demonstrated. NEDD4-2 E3 ligase regulates myriad ion channels, but its utility for TPD/TPDR is uncertain due to complex regulatory mechanisms. Here, we identify a nanobody that binds NEDD4-2 HECT domain without disrupting catalysis sites as revealed by cryo-electron microscopy and in vitro ubiquitination assays. Recruiting NEDD4-2 to diverse ion channels (Ca V 2.2; KCNQ1; and epithelial Na + channel, ENaC, with a Liddle syndrome mutation) using divalent nanobodies (DiVas) strongly suppresses their surface density and function. Global proteomics indicates DiVa recruitment of endogenous NEDD4-2 to KCNQ1-YFP yields dramatically lower off-target effects compared to NEDD4-2 overexpression. The results establish utility of NEDD4-2 recruitment for TPD/TPDR, validate ion channels as susceptible to this modality, and introduce a general method to generate ion channel inhibitors. Researchers develop a new way to selectively remove ion channel proteins by recruiting the body’s own NEDD4-2 enzyme using custom nanobodies, offering a precise and general strategy for future drug development.

The rate of action potential firing in nociceptors is a major determinant of the intensity of pain. Possible modulators of action potential firing include the HCN ion channels, which generate an inward current, I h , after hyperpolarization of the membrane. We found that genetic deletion of HCN2 removed the cyclic adenosine monophosphate (cAMP)—sensitive component of I h and abolished action potential firing caused by an elevation of cAMP in nociceptors. Mice in which HCN2 was specifically deleted in nociceptors expressing Na v 1.8 had normal pain thresholds, but inflammation did not cause hyperalgesia to heat stimuli. After a nerve lesion, these mice showed no neuropathic pain in response to thermal or mechanical stimuli. Neuropathic pain is therefore initiated by HCN2-driven action potential firing in Na v 1.8-expressing nociceptors.