Differential innervation within a transverse plane of spinal gray matter by sensorimotor cortices, with special reference to the somatosensory cortices.

The corticospinal (CS) neurons projecting to the cervical cord distribute not only in motor-related cortical areas, but also in somatosensory areas, including the primary somatosensory cortex (S1). The exact functions of these widely distributed CS neurons are largely unknown, however. In this study, we injected mice with adeno-associated virus encoding membrane-binding fluorescent proteins to investigate the distribution of axons from CS neurons in different regions within a broad cortical area. We found that CS axons from the primary motor cortex (M1), the rostral part of S1 (S1r), and the caudal part of S1 (S1c) differentially project to specific compartments within the spinal gray matter of the seventh cervical cord segment: (a) M1 projects mainly to intermediate and ventral areas, (b) S1r to the mediodorsal area, and (c) S1c to the dorsolateral area. We also found that the projection from S1r, which corresponds to the forelimb area, largely overlaps the cutaneous afferent terminals from the forepaw (hand) in the dorsal horn, and we detected a similar relation between S1c and the trunk. Our findings suggest the existence of considerably fine somatotopic compartments within the dorsal horn that process somatosensation and descending information, which is provided mainly by S1 CS neurons and contribute to delicate control of sensory information in generation of movement.

Connectomics of the zebrafish's lateral-line neuromast reveals wiring and miswiring in a simple microcircuit.

The lateral-line neuromast of the zebrafish displays a restricted, consistent pattern of innervation that facilitates the comparison of microcircuits across individuals, developmental stages, and genotypes. We used serial blockface scanning electron microscopy to determine from multiple specimens the neuromast connectome, a comprehensive set of connections between hair cells and afferent and efferent nerve fibers. This analysis delineated a complex but consistent wiring pattern with three striking characteristics: each nerve terminal is highly specific in receiving innervation from hair cells of a single directional sensitivity; the innervation is redundant; and the terminals manifest a hierarchy of dominance. Mutation of the canonical planar-cell-polarity gene vangl2, which decouples the asymmetric phenotypes of sibling hair-cell pairs, results in randomly positioned, randomly oriented sibling cells that nonetheless retain specific wiring. Because larvae that overexpress Notch exhibit uniformly oriented, uniformly innervating hair-cell siblings, wiring specificity is mediated by the Notch signaling pathway.

Serotonergic paraneurones in the female mouse urethral epithelium and their potential role in peripheral sensory information processing.

AIM: The mechanisms underlying detection and transmission of sensory signals arising from visceral organs, such as the urethra, are poorly understood. Recently, specialized ACh-expressing cells embedded in the urethral epithelium have been proposed as chemosensory sentinels for detection of bacterial infection. Here, we examined the morphology and potential role in sensory signalling of a different class of specialized cells that express serotonin (5-HT), termed paraneurones. METHODS: Urethrae, dorsal root ganglia neurones and spinal cords were isolated from adult female mice and used for immunohistochemistry and calcium imaging. Visceromotor reflexes (VMRs) were recorded in vivo. RESULTS: We identified two morphologically distinct groups of 5-HT+ cells with distinct regional locations: bipolar-like cells predominant in the mid-urethra and multipolar-like cells predominant in the proximal and distal urethra. Sensory nerve fibres positive for calcitonin gene-related peptide, substance P, and TRPV1 were found in close proximity to 5-HT+ paraneurones. In vitro 5-HT (1 µm) stimulation of urethral primary afferent neurones, mimicking 5-HT release from paraneurones, elicited changes in the intracellular calcium concentration ([Ca2+ ]i ) mediated by 5-HT2 and 5-HT3 receptors. Approximately 50% of 5-HT responding cells also responded to capsaicin with changes in the [Ca2+ ]i . In vivo intra-urethral 5-HT application increased VMRs induced by urethral distention and activated pERK in lumbosacral spinal cord neurones. CONCLUSION: These morphological and functional findings provide insights into a putative paraneurone-neural network within the urethra that utilizes 5-HT signalling, presumably from paraneurones, to modulate primary sensory pathways carrying nociceptive and non-nociceptive (mechano-sensitive) information to the central nervous system.

Afferent synaptogenesis between ectopic hair-cell-like cells and neurites of spiral ganglion induced by Atoh1 in mammals in vitro.

Newly formed ectopic hair-cell-like cells (EHCLCs) induced by overexpression of atonal homolog 1 (Atoh1) in vitro were found to possess features of endogenous hair cells (HCs) in previous reports and in the present study. However, limited information is available regarding whether EHCLCs and native spiral ganglion neurons (SGNs) form afferent synapses, which are important for the restoration of hearing. In the current study, we focused on the afferent synaptogenesis between EHCLCs and SGN-derived dendrites. Cochlear explants of auditory epithelia with native SGNs retained were cultured in vitro, and human adenovirus serotype 5 (Ad5) vectors encoding Atoh1 were used to overexpress Atoh1 and induce EHCLCs. We observed that the neurites of the original SGNs extended toward the lesser epithelial ridge (LER) and innervated the EHCLCs. Immunohistochemical analyses revealed the expression of presynaptic ribbon C-terminal-binding protein 2 (CtBP2) and postsynaptic density protein (PSD)-95 in the nerve endings of SGN-derived neurons adjacent to EHCLCs. PSD-95 was located directly opposite CtBP2-positive puncta in the terminals of branches of SGNs, demonstrating that the neurites of SGNs formed afferent-like synaptic connections with EHCLCs. However, the expression of glutamate receptor type 2 (GluR2) could not be detected in the terminals of branches of SGNs surrounding EHCLCs. In addition, we found that the presynaptic ribbon (CtBP2) formation in EHCLCs preceded neural innervation. Furthermore, CtBP2-positive puncta increased and then decreased in EHCLCs, similar to the changes observed in endogenous HCs in terms of their number and distribution. Our finding of the generation of cochlear afferent synapses between EHCLCs and original SGNs will lay the foundation for regenerative approaches to restoring hearing after hair cell loss.

Stimulation of renal afferent fibers leads to activation of catecholaminergic and non-catecholaminergic neurons in the medulla oblongata.

Presympathetic neurons in the rostral ventrolateral medulla (RVLM) including the adrenergic cell groups play a major role in the modulation of several reflexes required for the control of sympathetic vasomotor tone and blood pressure (BP). Moreover, sympathetic vasomotor drive to the kidneys influence natriuresis and diuresis by inhibiting the cAMP/PKA pathway and redistributing the Na+/H+ exchanger isoform 3 (NHE3) to the body of the microvilli in the proximal tubules. In this study we aimed to evaluate the effects of renal afferents stimulation on (1) the neurochemical phenotype of Fos expressing neurons in the medulla oblongata and (2) the level of abundance and phosphorylation of NHE3 in the renal cortex. We found that electrical stimulation of renal afferents increased heart rate and BP transiently and caused activation of tyrosine hydroxylase (TH)-containing neurons in the RVLM and non-TH neurons in the NTS. Additionally, activation of the inhibitory renorenal reflex over a 30-min period resulted in increased natriuresis and diuresis associated with increased phosphorylation of NHE3 at serine 552, a surrogate for reduced activity of this exchanger, in the contralateral kidney. This effect was not dependent of BP changes considering that no effects on natriuresis or diuresis were found in the ipsilateral-stimulated kidney. Therefore, our data show that renal afferents leads to activation of catecholaminergic and non-catecholaminergic neurons in the medulla oblongata. When renorenal reflex is induced, NHE3 exchanger activity appears to be decreased, resulting in decreased sodium and water reabsorption in the contralateral kidney.

CNS sites activated by renal pelvic epithelial sodium channels (ENaCs) in response to hypertonic saline in awake rats.

In some patients, renal nerve denervation has been reported to be an effective treatment for essential hypertension. Considerable evidence suggests that afferent renal nerves (ARN) and sodium balance play important roles in the development and maintenance of high blood pressure. ARN are sensitive to sodium concentrations in the renal pelvis. To better understand the role of ARN, we infused isotonic or hypertonic NaCl (308 or 500mOsm) into the left renal pelvis of conscious rats for two 2hours while recording arterial pressure and heart rate. Subsequently, brain tissue was analyzed for immunohistochemical detection of the protein Fos, a marker for neuronal activation. Fos-immunoreactive neurons were identified in numerous sites in the forebrain and brainstem. These areas included the nucleus tractus solitarius (NTS), the lateral parabrachial nucleus, the paraventricular nucleus of the hypothalamus (PVH) and the supraoptic nucleus (SON). The most effective stimulus was 500mOsm NaCl. Activation of these sites was attenuated or prevented by administration of benzamil (1µM) or amiloride (10µM) into the renal pelvis concomitantly with hypertonic saline. In anesthetized rats, infusion of hypertonic saline but not isotonic saline into the renal pelvis elevated ARN activity and this increase was attenuated by simultaneous infusion of benzamil or amiloride. We propose that renal pelvic epithelial sodium channels (ENaCs) play a role in activation of ARN and, via central visceral afferent circuits, this system modulates fluid volume and peripheral blood pressure. These pathways may contribute to the development of hypertension.

Different types of spinal afferent nerve endings in stomach and esophagus identified by anterograde tracing from dorsal root ganglia.

In visceral organs of mammals, most noxious (painful) stimuli as well as innocuous stimuli are detected by spinal afferent neurons, whose cell bodies lie in dorsal root ganglia (DRGs). One of the major unresolved questions is the location, morphology, and neurochemistry of the nerve endings of spinal afferents that actually detect these stimuli in the viscera. In the upper gastrointestinal (GI) tract, there have been many anterograde tracing studies of vagal afferent endings, but none on spinal afferent endings. Recently, we developed a technique that now provides selective labeling of only spinal afferents. We used this approach to identify spinal afferent nerve endings in the upper GI tract of mice. Animals were anesthetized, and injections of dextran-amine were made into thoracic DRGs (T8-T12). Seven days post surgery, mice were euthanized, and the stomach and esophagus were removed, fixed, and stained for calcitonin gene-related peptide (CGRP). Spinal afferent axons were identified that ramified extensively through many rows of myenteric ganglia and formed nerve endings in discrete anatomical layers. Most commonly, intraganglionic varicose endings (IGVEs) were identified in myenteric ganglia of the stomach and varicose simple-type endings in the circular muscle and mucosa. Less commonly, nerve endings were identified in internodal strands, blood vessels, submucosal ganglia, and longitudinal muscle. In the esophagus, only IGVEs were identified in myenteric ganglia. No intraganglionic lamellar endings (IGLEs) were identified in the stomach or esophagus. We present the first identification of spinal afferent endings in the upper GI tract. Eight distinct types of spinal afferent endings were identified in the stomach, and most of them were CGRP immunoreactive. J. Comp. Neurol. 524:3064-3083, 2016. © 2016 Wiley Periodicals, Inc.

Optogenetic inhibition of cortical afferents in the nucleus accumbens simultaneously prevents cue-induced transient synaptic potentiation and cocaine-seeking behavior.

Animal models of relapse reveal that the motivation to seek drug is regulated by enduring morphological and physiological changes in the nucleus accumbens, as well as transient synaptic potentiation in the accumbens core (NAcore) that parallels drug-seeking behavior. The current study sought to examine the link between the behavioral and synaptic consequences of cue-induced cocaine seeking by optically silencing glutamatergic afferents to the NAcore from the prelimbic cortex (PL). Adeno-associated virus coding for the inhibitory opsin archaerhodopsin was microinjected into PL, and optical fibers were targeted to NAcore. Animals were trained to self-administer cocaine followed by extinction training, and then underwent cue-induced reinstatement in the presence or absence of 15 min of optically induced inhibition of PL fibers in NAcore. Inhibiting the PL-to-NAcore projection blocked reinstated behavior and was paralleled by decreased dendritic spine head diameter and AMPA/NMDA ratio relative to sham-laser control rats. Interestingly, while spine density was elevated after extinction training, no further effects were observed by cued reinstatement or optical inhibition. These findings validate the critical role for PL afferents to the NAcore in simultaneously regulating both reinstated behavior and the associated transient synaptic potentiation.

Probing diversity within subpopulations of locomotor-related V0 interneurons.

The V0 interneuronal population is derived from Dbx1 expressing progenitors. Initial studies on these interneurons in the mouse spinal cord demonstrated that they project commissural axons and are involved in coordinating left-right alternation during locomotion. Subsequent work has indicated that the V0 population can be divided into genetically distinct ventral (V0V) and dorsal (V0D) subpopulations, and experimental evidence suggests that each is responsible for left-right alternation at different locomotor speeds. In this study, we perform a series of experiments to probe the location and connectivity of these subpopulations in neonatal mice and demonstrate that they are more diverse than previously predicted. While the distribution of either subpopulation remains consistent along the extent of the lumbar spinal cord, a cluster of V0D cells lateral to the central canal receive substantial input from primary afferents. Retrograde tracing and activity dependent labeling experiments demonstrate that a group of V0 interneurons located in this same region preferentially project axons towards contralateral motoneurons via an oligosynaptic pathway, and are active during fictive locomotion. Our results suggest that this subset of V0 interneurons may be primarily responsible for coordination of left-right alternation during locomotion. Furthermore these experiments indicate that while genetic identity is one determinant of the function of a neuron during locomotion, the specific position in which the cell is located may also play a key role.

Coordinated Recruitment of Cortical-Subcortical Circuits and Ascending Dopamine and Serotonin Neurons During Inhibitory Control of Cocaine Seeking in Rats.

People with cocaine addiction retain some degree of prefrontal cortex (PFC) inhibitory control of cocaine craving, a brain capacity that may underlie the efficacy of cognitive behavioral therapy for addiction. Similar findings were recently found in rats after extended access to and escalation of cocaine self-administration. Rats' inhibitory control of cocaine seeking was flexible, sufficiently strong to suppress cocaine-primed reinstatement and depended, at least in part, on neuronal activity within the prelimbic (PL) PFC. Here, we used a large-scale and high-resolution Fos mapping approach to identify, beyond the PL PFC, how top-down and/or bottom-up PFC-subcortical circuits are recruited during inhibition of cocaine seeking. Overall, we found that effective inhibitory control of cocaine seeking is associated with the coordinated recruitment of different top-down cortical-striatal circuits originating from different PFC territories, and of different bottom-up dopamine (DA) and serotonin (5-HT) midbrain subsystems that normally modulate activity in these circuits. This integrated brain response suggests that rats concomitantly engage and experience intricate cognitive and affective processes when they have to inhibit intense cocaine seeking. Thus, even after extended drug use, rats can be successfully trained to engage whole-brain inhibitory control mechanisms to suppress cocaine seeking.

Projection neurons of the vestibulo-sympathetic reflex pathway.

Changes in head position and posture are detected by the vestibular system and are normally followed by rapid modifications in blood pressure. These compensatory adjustments, which allow humans to stand up without fainting, are mediated by integration of vestibular system pathways with blood pressure control centers in the ventrolateral medulla. Orthostatic hypotension can reflect altered activity of this neural circuitry. Vestibular sensory input to the vestibulo-sympathetic pathway terminates on cells in the vestibular nuclear complex, which in turn project to brainstem sites involved in the regulation of cardiovascular activity, including the rostral and caudal ventrolateral medullary regions (RVLM and CVLM, respectively). In the present study, sinusoidal galvanic vestibular stimulation was used to activate this pathway, and activated neurons were identified through detection of c-Fos protein. The retrograde tracer Fluoro-Gold was injected into the RVLM or CVLM of these animals, and immunofluorescence studies of vestibular neurons were conducted to visualize c-Fos protein and Fluoro-Gold concomitantly. We observed activated projection neurons of the vestibulo-sympathetic reflex pathway in the caudal half of the spinal, medial, and parvocellular medial vestibular nuclei. Approximately two-thirds of the cells were ipsilateral to Fluoro-Gold injection sites in both the RVLM and CVLM, and the remainder were contralateral. As a group, cells projecting to the RVLM were located slightly rostral to those with terminals in the CVLM. Individual activated projection neurons were multipolar, globular, or fusiform in shape. This study provides the first direct demonstration of the central vestibular neurons that mediate the vestibulo-sympathetic reflex.

Thalamic synaptic transmission of sensory information modulated by synergistic interaction of adenosine and serotonin.

The thalamic synapses relay peripheral sensory information to the cortex, and constitute an important part of the thalamocortical network that generates oscillatory activities responsible for different vigilance (sleep and wakefulness) states. However, the modulation of thalamic synaptic transmission by potential sleep regulators, especially by combination of regulators in physiological scenarios, is not fully characterized. We found that somnogen adenosine itself acts similar to wake-promoting serotonin, both decreasing synaptic strength as well as short-term depression, at the retinothalamic synapse. We then combined the two modulators considering the coexistence of them in the hypnagogic (sleep-onset) state. Adenosine plus serotonin results in robust synergistic inhibition of synaptic strength and dramatic transformation of short-term synaptic depression to facilitation. These synaptic effects are not achievable with a single modulator, and are consistent with a high signal-to-noise ratio but a low level of signal transmission through the thalamus appropriate for slow-wave sleep. This study for the first time demonstrates that the sleep-regulatory modulators may work differently when present in combination than present singly in terms of shaping information flow in the thalamocortical network. The major synaptic characters such as the strength and short-term plasticity can be profoundly altered by combination of modulators based on physiological considerations.

Nociceptive afferents to the premotor neurons that send axons simultaneously to the facial and hypoglossal motoneurons by means of axon collaterals.

It is well known that the brainstem premotor neurons of the facial nucleus and hypoglossal nucleus coordinate orofacial nociceptive reflex (ONR) responses. However, whether the brainstem PNs receive the nociceptive projection directly from the caudal spinal trigeminal nucleus is still kept unclear. Our present study focuses on the distribution of premotor neurons in the ONR pathways of rats and the collateral projection of the premotor neurons which are involved in the brainstem local pathways of the orofacial nociceptive reflexes of rat. Retrograde tracer Fluoro-gold (FG) or FG/tetramethylrhodamine-dextran amine (TMR-DA) were injected into the VII or/and XII, and anterograde tracer biotinylated dextran amine (BDA) was injected into the caudal spinal trigeminal nucleus (Vc). The tracing studies indicated that FG-labeled neurons receiving BDA-labeled fibers from the Vc were mainly distributed bilaterally in the parvicellular reticular formation (PCRt), dorsal and ventral medullary reticular formation (MdD, MdV), supratrigeminal nucleus (Vsup) and parabrachial nucleus (PBN) with an ipsilateral dominance. Some FG/TMR-DA double-labeled premotor neurons, which were observed bilaterally in the PCRt, MdD, dorsal part of the MdV, peri-motor nucleus regions, contacted with BDA-labeled axonal terminals and expressed c-fos protein-like immunoreactivity which induced by subcutaneous injection of formalin into the lip. After retrograde tracer wheat germ agglutinated horseradish peroxidase (WGA-HRP) was injected into VII or XII and BDA into Vc, electron microscopic study revealed that some BDA-labeled axonal terminals made mainly asymmetric synapses on the dendritic and somatic profiles of WGA-HRP-labeled premotor neurons. These data indicate that some premotor neurons could integrate the orofacial nociceptive input from the Vc and transfer these signals simultaneously to different brainstem motonuclei by axonal collaterals.

Feline immunodeficiency virus as a gene transfer vector in the rat nucleus tractus solitarii.

Gene transfer has been used to examine the role of putative neurotransmitters in the nucleus tractus solitarii (NTS). Most such studies used adenovirus vector-mediated gene transfer although adenovirus vector transfects both neuronal and non-neuronal cells. Successful transfection in the NTS has also been reported with lentivirus as the vector. Feline immunodeficiency virus (FIV), a lentivirus, may preferentially transfect neurons and could be a powerful tool to delineate physiological effects produced by altered synthesis of transmitters in neurons. However, it has not been studied in NTS. Therefore, we sought to determine whether FIV transfects rat NTS cells and to define the type of cell transfected. We found that injection of FIV encoding LacZ gene (FIVLacZ) into the NTS led to transfection of numerous NTS cells. Injection of FIVLacZ did not alter immunoreactivity (IR) for neuronal nitric oxide synthase, which we have shown resides in NTS neurons. A majority (91.7 +/- 3.9%) of transfected cells contained IR for neuronal nuclear antigen, a neuronal marker; 2.1 +/- 3.8% of transfected cells contained IR for glial fibrillary acidic protein, a glial marker. No transfected neurons or fibers were observed in the nodose ganglion, which sends afferents to the NTS. We conclude that FIV almost exclusively transfects neurons in the rat NTS from which it is not retrogradely transported. The cell-type specificity of FIV in the NTS may provide a molecular method to study local physiological functions mediated by potential neurotransmitters in the NTS.

The postsynaptic function of type II cochlear afferents.

The mammalian cochlea is innervated by two classes of sensory neurons. Type I neurons make up 90-95% of the cochlear nerve and contact single inner hair cells to provide acoustic analysis as we know it. In contrast, the far less numerous type II neurons arborize extensively among outer hair cells (OHCs) and supporting cells. Their scarcity and smaller calibre axons have made them the subject of much speculation, but little experimental progress for the past 50 years. Here we record from type II fibres near their terminal arbors under OHCs to show that they receive excitatory glutamatergic synaptic input. The type II peripheral arbor conducts action potentials, but the small and infrequent glutamatergic excitation indicates a requirement for strong acoustic stimulation. Furthermore, we show that type II neurons are excited by ATP. Exogenous ATP depolarized type II neurons, both directly and by evoking glutamatergic synaptic input. These results prove that type II neurons function as cochlear afferents, and can be modulated by ATP. The lesser magnitude of synaptic drive dictates a fundamentally different role in auditory signalling from that of type I afferents.

Dorsal horn neurons presynaptic to lamina I spinoparabrachial neurons revealed by transynaptic labeling.

Sensory input to supraspinally projecting lamina I (LI) neurons arises both directly from primary afferents and via neurons intrinsic to the spinal dorsal horn. The types of neurons presynaptic to those projection neurons remain poorly known. To address this question we used retrogradely transported adenoviral vectors encoding green fluorescent protein (GFP) and a GFP-TTC (fragment C of the tetanus toxin) fusion protein, labeling respectively spinoparabrachial projection neurons and neurons presynaptic to them. The expression of GFP by infected neurons labeled the entire dendritic tree, enabling a more complete and quantitative morphological description of spinoparabrachial neurons than previous methods. These neurons were located in spinal LI, with dendritic arbors oriented extensively in the rostrocaudal axis (1,089.8 +/- 91.5 microm) and displaying low spine density. In contrast, their dendrites did not extend significantly ventrally (29.2 +/- 3.5 microm). The use of transynaptic tracer GFP-TTC revealed a population of local circuit LI neurons presynaptic to LI projection neurons. These local circuit LI neurons had distinct morphological properties, in particular significantly longer ventrally oriented dendrites (80.1 +/- 10.1 microm). The transynaptic tracer also revealed a population of stalked cells, some being highly spiny, directly in contact with spinal projection neurons. However, stalked cells were not the only lamina II cells in direct contact with projection neurons. Intracellular injections with Lucifer yellow in parasagittal slices of fixed tissue confirmed the above observations. Overall, these experiments demonstrated that neurons projecting to the parabrachial nucleus had their dendritic branching almost exclusively in LI and had sparse dendritic spines, in contrast with local circuit neurons that often extended ventrally and could be very spiny.

Research progress in transient receptor potential vanilloid 1 of sensory nervous system.

The transient receptor potential vanilloid subfamily member 1 (TRPV1) is a protein mainly expressed in sensory neurons and fibers, such as in trigeminal ganglion and dorsal root ganglion, and has been indicated to be involved in several physiological and pathological processes. Studies on thermal activation have revealed that phosphorylation is involved in TRPV1 activation and 2 putative phosphorylation sites, Ser residues 502 (Ser-502) and Ser residues 800 (Ser-800), have been recently confirmed to possess the capability of resensitizing TRPV1. In addition to acidification, alkalization has also been proved to be a highly effective stimulator for TRPV1. TRPV1 could be regulated by various physical and chemical modulators, as well as the chronic pain. TRPV1 plays a crucial role in the transmission of pain signals, especially under inflammation and the neoplasm conditions, and it can also modulate nociceptive afferents by reinforcing morphine tolerance. The present review mainly focused on the structural and functional complexities of TRPV1, together with its activation and modulation by a wide variety of physical and chemical stimuli. Its pharmacological manipulation (sensitization/desensitization) and therapeutical targets were also discussed.

RETouching upon mechanoreceptors.

The rapidly adapting (RA) low-threshold mechanoreceptors respond to movement of the skin and vibration and are critical for the perception of texture and shape. In this issue of Neuron, two papers (Bourane et al. and Luo et al.) demonstrate that early-born Ret+ sensory neurons are RA mechanoreceptors, whose peripheral nerve terminals are associated with Meissner corpuscles, longitudinal lanceolate endings, and Pacinian corpuscles. The studies further show that Ret signaling is essential for the development of these mechanoreceptors.

Molecular identification of rapidly adapting mechanoreceptors and their developmental dependence on ret signaling.

In mammals, the first step in the perception of form and texture is the activation of trigeminal or dorsal root ganglion (DRG) mechanosensory neurons, which are classified as either rapidly (RA) or slowly adapting (SA) according to their rates of adaptation to sustained stimuli. The molecular identities and mechanisms of development of RA and SA mechanoreceptors are largely unknown. We found that the "early Ret(+)" DRG neurons are RA mechanoreceptors, which form Meissner corpuscles, Pacinian corpuscles, and longitudinal lanceolate endings. The central projections of these RA mechanoreceptors innervate layers III through V of the spinal cord and terminate within discrete subdomains of the dorsal column nuclei. Moreover, mice lacking Ret signaling components are devoid of Pacinian corpuscles and exhibit a dramatic disruption of RA mechanoreceptor projections to both the spinal cord and medulla. Thus, the early Ret(+) neurons are RA mechanoreceptors and Ret signaling is required for the assembly of neural circuits underlying touch perception.

Low-threshold mechanoreceptor subtypes selectively express MafA and are specified by Ret signaling.

Low-threshold mechanoreceptor neurons (LTMs) of the dorsal root ganglia (DRG) are essential for touch sensation. They form highly specialized terminations in the skin and display stereotyped projections in the spinal cord. Functionally defined LTMs depend on neurotrophin signaling for their postnatal survival and functioning, but how these neurons arise during development is unknown. Here, we show that specific types of LTMs can be identified shortly after DRG genesis by unique expression of the MafA transcription factor, the Ret receptor and coreceptor GFRalpha2, and find that their specification is Ngn2 dependent. In mice lacking Ret, these LTMs display early differentiation defects, as revealed by reduced MafA expression, and at later stages their central and peripheral projections are compromised. Moreover, in MafA mutants, a discrete subset of LTMs display altered expression of neurotrophic factor receptors. Our results provide evidence that genetic interactions involving Ret and MafA progressively promote the differentiation and diversification of LTMs.