Acid taste, evoked mainly by protons (H+), is a core taste modality for many organisms. The hedonic valence of acid taste is bidirectional: animals prefer slightly but avoid highly acidic foods. However, how animals discriminate low from high acidity remains poorly understood. To explore the taste perception of acid, we use the fruit fly as a model organism. We find that flies employ two competing taste sensory pathways to detect low and high acidity, and the relative degree of activation of each determines either attractive or aversive responses. Moreover, we establish one member of the fly Otopetrin family, Otopetrin-like a (OtopLa), as a proton channel dedicated to the gustatory detection of acid. OtopLa defines a unique subset of gustatory receptor neurons and is selectively required for attractive rather than aversive taste responses. Loss of otopla causes flies to reject normally attractive low-acid foods. Therefore, the identification of OtopLa as a low-acid sensor firmly supports our competition model of acid taste sensation. Altogether, we have discovered a binary acid-sensing mechanism that may be evolutionarily conserved between insects and mammals.

Touch and itch sensations are crucial for evoking defensive and emotional responses, and light tactile touch may induce unpleasant itch sensations (mechanical itch or alloknesis). The neural substrate for touch-to-itch conversion in the spinal cord remains elusive. We report that spinal interneurons expressing Tachykinin 2-Cre (Tac2Cre) receive direct Aβ low threshold mechanoreceptor (LTMR) input and form monosynaptic connections with GRPR neurons. Ablation or inhibition markedly reduces mechanical but not acute chemical itch nor noxious touch information. Chemogenetic inhibition of Tac2Cre neurons also displays pronounced deficit in chronic dry skin itch, a type of chemical itch in mice. Consistently, ablation of gastrin-releasing peptide receptor (GRPR) neurons, which are essential for transmitting chemical itch, also abolishes mechanical itch. Together, these results suggest that innocuous touch and chemical itch information converge on GRPR neurons and thus map an exquisite spinal circuitry hard-wired for converting innocuous touch to irritating itch.

The medial prefrontal cortex (mPFC) is implicated in processing sensory-discriminative and affective pain. Nonetheless, the underlying mechanisms are poorly understood. Here we demonstrate a role for excitatory neurons in the prelimbic cortex (PL), a sub-region of mPFC, in the regulation of pain sensation and anxiety-like behaviours. Using a chronic inflammatory pain model, we show that lesion of the PL contralateral but not ipsilateral to the inflamed paw attenuates hyperalgesia and anxiety-like behaviours in rats. Optogenetic activation of contralateral PL excitatory neurons exerts analgesic and anxiolytic effects in mice subjected to chronic pain, whereas inhibition is anxiogenic in naive mice. The intrinsic excitability of contralateral PL excitatory neurons is decreased in chronic pain rats; knocking down cyclin-dependent kinase 5 reverses this deactivation and alleviates behavioural impairments. Together, our findings provide novel insights into the role of PL excitatory neurons in the regulation of sensory and affective pain.

Many cellular programs of neural development are under combinatorial regulation by different chemoattractive or chemorepulsive factors. Here, we describe a microfluidic platform that utilizes well-controlled three-dimensional (3D) diffusion to generate molecular gradients of varied steepness in a large array of hydrogel cylinders, allowing high-throughput 3D chemotactic assays for mechanistic dissection of steepness-dependent neuronal chemotaxis. Using this platform, we examine neuronal sensitivity to the steepness of gradient composed of netrin-1, nerve growth factor, or semaphorin3A (Sema3A) proteins, and reveal dramatic diversity and complexity in the associated chemotactic regulation of neuronal development. Particularly for Sema3A, we find that serine/threonine kinase-11 and glycogen synthase kinase-3 signaling pathways are differentially involved in steepness-dependent chemotactic regulation of coordinated neurite repellence and neuronal migration. These results provide insights to the critical role of gradient steepness in neuronal chemotaxis, and also prove the technique as an expandable platform for studying other chemoresponsive cellular systems.

Steviol glycosides (SGs), such as stevioside and rebaudioside A, are natural, non-caloric sweet-tasting organic molecules, present in extracts of the scrub plant Stevia rebaudiana, which are widely used as sweeteners in consumer foods and beverages. TRPM5 is a Ca2+-activated cation channel expressed in type II taste receptor cells and pancreatic β-cells. Here we show that stevioside, rebaudioside A and their aglycon steviol potentiate the activity of TRPM5. We find that SGs potentiate perception of bitter, sweet and umami taste, and enhance glucose-induced insulin secretion in a Trpm5-dependent manner. Daily consumption of stevioside prevents development of high-fat-diet-induced diabetic hyperglycaemia in wild-type mice, but not in Trpm5-/- mice. These results elucidate a molecular mechanism of action of SGs and identify TRPM5 as a potential target to prevent and treat type 2 diabetes.

Itch is an annoying sensation consisting of both sensory and emotional components. It is known to involve the parabrachial nucleus (PBN), but the following transmission nodes remain elusive. The present study identified that the PBN-central medial thalamic nucleus (CM)-medial prefrontal cortex (mPFC) pathway is essential for itch signal transmission at the supraspinal level in male mice. Chemogenetic inhibition of the CM-mPFC pathway attenuates scratching behavior or chronic itch-related affective responses. CM input to mPFC pyramidal neurons is enhanced in acute and chronic itch models. Specifically chronic itch stimuli also alter mPFC interneuron involvement, resulting in enhanced feedforward inhibition and a distorted excitatory/inhibitory balance in mPFC pyramidal neurons. The present work underscores CM as a transmit node of the itch signal in the thalamus, which is dynamically engaged in both the sensory and affective dimensions of itch with different stimulus salience.

Fatty acids (FAs) are not only essential components of cellular energy storage and structure, but play crucial roles in signalling. Here we present a toolkit of photoswitchable FA analogues (FAAzos) that incorporate an azobenzene photoswitch along the FA chain. By modifying the FAAzos to resemble capsaicin, we prepare a series of photolipids targeting the Vanilloid Receptor 1 (TRPV1), a non-selective cation channel known for its role in nociception. Several azo-capsaicin derivatives (AzCAs) emerge as photoswitchable agonists of TRPV1 that are relatively inactive in the dark and become active on irradiation with ultraviolet-A light. This effect can be rapidly reversed by irradiation with blue light and permits the robust optical control of dorsal root ganglion neurons and C-fibre nociceptors with precision timing and kinetics not available with any other technique. More generally, we expect that photolipids will find many applications in controlling biological pathways that rely on protein-lipid interactions.

Previous work identified nociceptive Schwann cells that can initiate pain. Consistent with the existence of inherently mechanosensitive sensory Schwann cells, we found that in mice, the mechanosensory function of almost all nociceptors, including those signaling fast pain, were dependent on sensory Schwann cells. In polymodal nociceptors, sensory Schwann cells signal mechanical, but not cold or heat pain. Terminal Schwann cells also surround mechanoreceptor nerve-endings within the Meissner's corpuscle and in hair follicle lanceolate endings that both signal vibrotactile touch. Within Meissner´s corpuscles, two molecularly and functionally distinct sensory Schwann cells positive for Sox10 and Sox2 differentially modulate rapidly adapting mechanoreceptor function. Using optogenetics we show that Meissner's corpuscle Schwann cells are necessary for the perception of low threshold vibrotactile stimuli. These results show that sensory Schwann cells within diverse glio-neural mechanosensory end-organs are sensors for mechanical pain as well as necessary for touch perception.

Dietary composition affects food preference in animals. High sugar intake suppresses sweet sensation from insects to humans, but the molecular basis of this suppression is largely unknown. Here, we reveal that sugar intake in Drosophila induces the gut to express and secrete Hedgehog (Hh) into the circulation. We show that the midgut secreted Hh localize to taste sensilla and suppresses sweet sensation, perception, and preference. We further find that the midgut Hh inhibits Hh signalling in the sweet taste neurons. Our electrophysiology studies demonstrate that the midgut Hh signal also suppresses bitter taste and some odour responses, affecting overall food perception and preference. We further show that the level of sugar intake during a critical window early in life, sets the adult gut Hh expression and sugar perception. Our results together reveal a bottom-up feedback mechanism involving a "gut-taste neuron axis" that regulates food sensation and preference.

Although alkaline sensation is critical for survival, alkali-activated receptors are yet to be identified in vertebrates. Here, we showed that the OTOP1 channel can be directly activated by extracellular alkali. Notably, OTOP1 biphasically mediated proton influx and efflux with extracellular acid and base stimulation, respectively. Mutations of K221 and R554 at the S5-S6 and S11-S12 linkers significantly reduced alkali affinity without affecting acid activation, suggesting that different domains are responsible for acid- and alkali-activation of OTOP1. The selectivity for H+ was significantly higher in OTOP1 activated by alkali than that by acid, further suggesting that the two activations might be independent gating processes. Given that the alkali-activation of OTOP1 and the required key residues were conserved in the six representative vertebrates, we cautiously propose that OTOP1 participates in alkaline sensation in vertebrates. Thus, our study identified OTOP1 as an alkali-activated channel.

The mechanosensitive ion channel Piezo2 in mucosa and primary afferents transduces colonic mechanical sensation. Here we show that chemogenetic activation or nociceptor-targeted deletion of Piezo2 is sufficient to regulate colonic mechanical sensitivity in a sex dependent manner. Clozapine N-oxide-induced activation of Piezo2;hM3Dq-expressing sensory neurons evokes colonic hypersensitivity in male mice, and causes dyspnea in female mice likely due to effects on lung sensory neurons. Activation of Piezo2-expressing colonic afferent neurons also induces colonic hypersensitivity in male but not female mice. Piezo2 levels in nociceptive neurons are higher in female than in male mice. We also show that Piezo2 conditional deletion from nociceptive neurons increases body weight growth, slows colonic transits, and reduces colonic mechanosensing in female but not male mice. Piezo2 deletion blocks colonic hypersensitivity in male but not female mice. These results suggest that Piezo2 in nociceptive neurons mediates innocuous colonic mechanosensing in female mice and painful sensation in male mice, suggesting a sexual dimorphism of Piezo2 function in the colonic sensory system.

Reptiles are the most morphologically and physiologically diverse tetrapods, and have undergone 300 million years of adaptive evolution. Within the reptilian tetrapods, geckos possess several interesting features, including the ability to regenerate autotomized tails and to climb on smooth surfaces. Here we sequence the genome of Gekko japonicus (Schlegel's Japanese Gecko) and investigate genetic elements related to its physiology. We obtain a draft G. japonicus genome sequence of 2.55 Gb and annotated 22,487 genes. Comparative genomic analysis reveals specific gene family expansions or reductions that are associated with the formation of adhesive setae, nocturnal vision and tail regeneration, as well as the diversification of olfactory sensation. The obtained genomic data provide robust genetic evidence of adaptive evolution in reptiles.

Temperature is a critical environmental stimulus that has a strong impact on an organism's biochemistry. Animals can respond to changes in ambient temperature through behaviour or altered physiology. However, how animals habituate to temperature is poorly understood. The nematode C. elegans stores temperature experiences and can induce temperature habituation-linked cold tolerance. Here we show that light and pheromone-sensing neurons (ASJ) regulate cold habituation through insulin signalling. Calcium imaging reveals that ASJ neurons respond to temperature. Cold habituation is abnormal in a mutant with impaired cGMP signalling in ASJ neurons. Insulin released from ASJ neurons is received by the intestine and neurons regulating gene expression for cold habituation. Thus, temperature sensation in a light and pheromone-sensing neuron produces a robust effect on insulin signalling that controls experience-dependent temperature habituation.

Members of the transient receptor potential (TRP) ion channels conduct cations into cells. They mediate functions ranging from neuronally mediated hot and cold sensation to intracellular organellar and primary ciliary signaling. Here we report a cryo-electron microscopy (cryo-EM) structure of TRPC4 in its unliganded (apo) state to an overall resolution of 3.3 Å. The structure reveals a unique architecture with a long pore loop stabilized by a disulfide bond. Beyond the shared tetrameric six-transmembrane fold, the TRPC4 structure deviates from other TRP channels with a unique cytosolic domain. This unique cytosolic N-terminal domain forms extensive aromatic contacts with the TRP and the C-terminal domains. The comparison of our structure with other known TRP structures provides molecular insights into TRPC4 ion selectivity and extends our knowledge of the diversity and evolution of the TRP channels.

Norepinephrine is a biogenic amine neurotransmitter that has widespread effects on alertness, arousal and pain sensation. Consequently, blockers of norepinephrine uptake have served as vital tools to treat depression and chronic pain. Here, we employ the Drosophila melanogaster dopamine transporter as a surrogate for the norepinephrine transporter and determine X-ray structures of the transporter in its substrate-free and norepinephrine-bound forms. We also report structures of the transporter in complex with inhibitors of chronic pain including duloxetine, milnacipran and a synthetic opioid, tramadol. When compared to dopamine, we observe that norepinephrine binds in a different pose, in the vicinity of subsite C within the primary binding site. Our experiments reveal that this region is the binding site for chronic pain inhibitors and a determinant for norepinephrine-specific reuptake inhibition, thereby providing a paradigm for the design of specific inhibitors for catecholamine neurotransmitter transporters.

Although TRPV1 channels represent a key player of noxious heat sensation, the precise mechanisms for thermal hyperalgesia remain unknown. We report here that conditional knockout of deSUMOylation enzyme, SENP1, in mouse dorsal root ganglion (DRG) neurons exacerbated thermal hyperalgesia in both carrageenan- and Complete Freund's adjuvant-induced inflammation models. TRPV1 is SUMOylated at a C-terminal Lys residue (K822), which specifically enhances the channel sensitivity to stimulation by heat, but not capsaicin, protons or voltage. TRPV1 SUMOylation is decreased by SENP1 but upregulated upon peripheral inflammation. More importantly, the reduced ability of TRPV1 knockout mice to develop inflammatory thermal hyperalgesia was rescued by viral infection of lumbar 3/4 DRG neurons of wild-type TRPV1, but not its SUMOylation-deficient mutant, K822R. These data suggest that TRPV1 SUMOylation is essential for the development of inflammatory thermal hyperalgesia, through a mechanism that involves sensitization of the channel response specifically to thermal stimulation.

PKD2L1, also termed TRPP3 from the TRPP subfamily (polycystic TRP channels), is involved in the sour sensation and other pH-dependent processes. PKD2L1 is believed to be a nonselective cation channel that can be regulated by voltage, protons, and calcium. Despite its considerable importance, the molecular mechanisms underlying PKD2L1 regulations are largely unknown. Here, we determine the PKD2L1 atomic structure at 3.38 Å resolution by cryo-electron microscopy, whereby side chains of nearly all residues are assigned. Unlike its ortholog PKD2, the pore helix (PH) and transmembrane segment 6 (S6) of PKD2L1, which are involved in upper and lower-gate opening, adopt an open conformation. Structural comparisons of PKD2L1 with a PKD2-based homologous model indicate that the pore domain dilation is coupled to conformational changes of voltage-sensing domains (VSDs) via a series of π-π interactions, suggesting a potential PKD2L1 gating mechanism.

Somatic sensation is defined by the existence of a diversity of primary sensory neurons with unique biological features and response profiles to external and internal stimuli. However, there is no coherent picture about how this diversity of cell states is transcriptionally generated. Here, we use deep single cell analysis to resolve fate splits and molecular biasing processes during sensory neurogenesis in mice. Our results identify a complex series of successive and specific transcriptional changes in post-mitotic neurons that delineate hierarchical regulatory states leading to the generation of the main sensory neuron classes. In addition, our analysis identifies previously undetected early gene modules expressed long before fate determination although being clearly associated with defined sensory subtypes. Overall, the early diversity of sensory neurons is generated through successive bi-potential intermediates in which synchronization of relevant gene modules and concurrent repression of competing fate programs precede cell fate stabilization and final commitment.

The sensation of movement and position of the limbs is critical for normal behaviours in tetrapods. In the bony fishes it is unclear what proprioceptive feedback is provided from the paired fins, the piscine homologues of the tetrapod limbs. Here we test mechanosensory abilities of afferent nerves in the pectoral fin rays, limb structures used by many fish species in propulsion and manoeuvreing. We examine the bluegill sunfish, a fish that uses its pectoral fins extensively in locomotion. We find that the activity of fin ray nerve fibres reflects the amplitude and velocity of fin ray bending. Spike sorting analyses demonstrate the presence of both slowly and rapidly adapting afferent nerve fibres. The fin sensory abilities we describe substantially expand the diversity of known vertebrate proprioceptive capabilities, and suggest that the pectoral fins need to be considered as possible proprioceptive sensors in studies of their functional morphology, movement and evolution.

Itch is a distinct aversive sensation that elicits a strong urge to scratch. Despite recent advances in our understanding of the peripheral basis of itch, we know very little regarding how central neural circuits modulate acute and chronic itch processing. Here we establish the causal contributions of defined periaqueductal gray (PAG) neuronal populations in itch modulation in mice. Chemogenetic manipulations demonstrate bidirectional modulation of scratching by neurons in the PAG. Fiber photometry studies show that activity of GABAergic and glutamatergic neurons in the PAG is modulated in an opposing manner during chloroquine-evoked scratching. Furthermore, activation of PAG GABAergic neurons or inhibition of glutamatergic neurons resulted in attenuation of scratching in both acute and chronic pruritis. Surprisingly, PAG GABAergic neurons, but not glutamatergic neurons, may encode the aversive component of itch. Thus, the PAG represents a neuromodulatory hub that regulates both the sensory and affective aspects of acute and chronic itch.