Cutaneous Sensory Neurons Research Articles

Identity of myelinated cutaneous sensory neurons projecting to nocireceptive laminae following nerve injury in adult mice

It is widely thought that, after peripheral injury, some low-threshold mechanoreceptive (LTMR) afferents "sprout" into pain-specific laminae (I-II) of the dorsal horn and are responsible for chronic pain states such as mechanical allodynia. Although recent studies have questioned this hypothesis, they fail to account for a series of compelling results from single-fiber analyses showing extensive projections from large-diameter myelinated afferents into nocireceptive layers after nerve injury. Here we show that, in the thoracic spinal cord of naïve adult mouse, all myelinated nociceptors gave rise to terminal projections throughout the superficial dorsal horn laminae (I-II). Most (70%) of these fibers had large-diameter axons with recurving flame-shaped central arbors that projected throughout the dorsal horn laminae I-V. This morphology was reminiscent of that attributed to sprouted LTMRs described in previous studies. After peripheral nerve axotomy, we found that LTMR afferents with narrow, uninflected somal action potentials did not sprout into superficial laminae of the dorsal horn. Only myelinated noiceptive afferents with broad, inflected somal action potentials were found to give rise to recurving collaterals and project into superficial "pain-specific" laminae after axotomy. We conclude that the previously undocumented central morphology of large, myelinated cutaneous nociceptors may very well account for the morphological findings previously thought to require sprouting of LTMRs.

Fine-grained fate maps for the ophthalmic and maxillomandibular trigeminal placodes in the chick embryo

Vertebrate cranial ectodermal placodes are transient, paired thickenings of embryonic head ectoderm that are crucial for the formation of the peripheral sensory nervous system: they give rise to the paired peripheral sense organs (olfactory organs, inner ears and anamniote lateral line system), as well as the eye lenses, and most cranial sensory neurons. Here, we present the first detailed spatiotemporal fate-maps in any vertebrate for the ophthalmic trigeminal (opV) and maxillomandibular trigeminal (mmV) placodes, which give rise to cutaneous sensory neurons in the ophthalmic and maxillomandibular lobes of the trigeminal ganglion. We used focal DiI and DiO labelling to produce eight detailed fate-maps of chick embryonic head ectoderm over approximately 24 h of development, from 0–16 somites. OpV and mmV placode precursors arise from a partially overlapping territory; indeed, some individual dyespots labelled both opV and mmV placode-derived cells. OpV and mmV placode precursors are initially scattered within a relatively large region of ectoderm adjacent to the neural folds, intermingled both with each other and with future epidermal cells, and with geniculate and otic placode precursors. Although the degree of segregation increases with time, there is no clear border between the opV and mmV placodes even at the 16-somite stage, long after neurogenesis has begun in the opV placode, and when neurogenesis is just beginning in the mmV placode. Finally, we find that occasional cells in the border region between the opV placode and mmV placode express both Pax3 (an opV placode specific marker) and Neurogenin1 (an mmV placode specific marker), suggesting that a few cells are responding to both opV and mmV placode-inducing signals. Overall, our results fill a large gap in our knowledge of the early stages of development of both the opV and mmV placodes, providing an essential framework for subsequent studies of the molecular control of their development.

TRPV1 unlike TRPV2 is restricted to a subset of mechanically insensitive cutaneous nociceptors responding to heat.

In the present study, a murine ex vivo somatosensory system preparation was used to determine the response characteristics of cutaneous sensory neurons staining positively for TRPV1 or TRPV2. TRPV1 immunostaining was found exclusively (11/11) in a specific set of mechanically insensitive unmyelinated (C) nociceptors that responded to heating of their receptive fields. No cutaneous C-fibers that responded to both mechanical and heat stimuli stained positively for TRPV1 (0/62). The relationship between TRPV2 and heat transduction characteristics was not as clear, as few unmyelinated or myelinated fibers that responded to heat contained TRPV2. TRPV2 was found most frequently in mechanically sensitive myelinated fibers, including both low threshold and high threshold mechanoreceptors (nociceptors). Although TRPV2 was found in only 1 of 6 myelinated polymodal nociceptors, it was found in a majority (10/16) of myelinated mechanical nociceptors. Thus, whereas the in vivo role of TRPV1 as a heat-sensitive channel in cutaneous sensory neurons is clearly defined, the role of TRPV2 in cutaneous neurons remains unknown. These results also suggest that TRPV1 may be essential for heat transduction in a specific subset of mechanically insensitive cutaneous nociceptors and that this subset may constitute a discrete heat input pathway for inflammation-induced thermal pain. The distinct subset of murine cutaneous nociceptors containing TRPV1 has many attributes in common with mechanically insensitive C-fibers in humans that are believed to play a role in pathological pain states. Therefore, these murine fibers provide a clinically relevant animal model for further study of this group of cutaneous nociceptors.

Survival and regeneration of cutaneous and muscular afferent neurons after peripheral nerve injury in adult rats

Peripheral nerve injury induces the retrograde degeneration of dorsal root ganglion (DRG) cells, which affects predominantly the small-diameter cutaneous afferent neurons. This study compares the time-course of retrograde cell death in cutaneous and muscular DRG cells after peripheral nerve transection as well as neuronal survival and axonal regeneration after primary repair or nerve grafting. For comparison, spinal motoneurons were also included in the study. Sural and medial gastrocnemius DRG neurons were retrogradely labeled with the fluorescent tracers Fast Blue (FB) or Fluoro-Gold (FG) from the homonymous transected nerves. Survival of labeled sural and gastrocnemius DRG cells was assessed at 3 days and 1-24 weeks after axotomy. To evaluate axonal regeneration, the sciatic nerve was transected proximally at 1 week after FB-labeling of the sural and medial gastrocnemius nerves and immediately reconstructed using primary repair or autologous nerve grafting. Twelve weeks later, the fluorescent tracer Fluoro-Ruby (FR) was applied 10 mm distal to the sciatic lesion in order to double-label sural and gastrocnemius neurons that had regenerated across the repair site. Counts of labeled gastrocnemius DRG neurons did not reveal any significant retrograde cell death after nerve transection. In contrast, sural axotomy induced a delayed loss of sural DRG cells, which amounted to 22% at 4 weeks and 43-48% at 8-24 weeks postoperatively. Proximal transection of the sciatic nerve at 1 week after injury to the sural or gastrocnemius nerves neither further increased retrograde DRG degeneration, nor did it affect survival of sural or gastrocnemius motoneurons. Primary repair or peripheral nerve grafting supported regeneration of 53-60% of the spinal motoneurons and 47-49% of the muscular DRG neurons at 13 weeks postoperatively. In the cutaneous DRG neurons, primary repair or peripheral nerve grafting increased survival by 19-30% and promoted regeneration of 46-66% of the cells. The present results suggest that cutaneous DRG neurons are more sensitive to peripheral nerve injury than muscular DRG cells, but that their regenerative capacity does not differ from that of the latter cells. However, the retrograde loss of cutaneous DRG cells taking place despite immediate nerve repair would still limit the recovery of cutaneous sensory functions.

Overexpression of neurotrophin‐3 enhances the mechanical response properties of slowly adapting type 1 afferents and myelinated nociceptors

Constitutive overexpression of neurotrophin-3 (NT3) in murine skin results in an increased number of sensory neurons within the dorsal root ganglia, an increase of myelinated axons in cutaneous nerves, hyperinnervation of the skin, and an increased number of Merkel cells found in flank skin. Here we used a saphenous skin/nerve preparation to determine if these anatomical changes affect the functional response characteristics of cutaneous sensory neurons. Overexpression of NT3 significantly increased the responses of slowly adapting type 1 (SA1) low-threshold mechanoreceptors and Adelta high-threshold mechanoreceptors to suprathreshold mechanical stimulation. It also resulted in significantly faster conduction velocities of SA1 fibers. In contrast to earlier findings in flank skin, no differences were noted in the numbers of Merkel cells in the touch domes in hindlimb skin of NT3-overexpressing mice. In addition, the number of dermal Merkel cells, located around hair follicles on the dorsum of the foot, was reduced by 55%. The increase in mechanical sensitivity was found to correlate with significant increases in the expression of acid-sensing ion channels (ASIC) 1 and 3. Additional experiments using intracellular recordings and staining procedures confirmed that at least some cutaneous myelinated nociceptors and SA1 mechanoreceptors stained positively for both trkC and ASIC3. These results indicate that cutaneous NT3 overexpression alters the response properties of specific cutaneous sensory neurons, and that these changes may be due to the modulation of putative mechanosensitive ion channels.

Molecular genetic visualization of a rare subset of unmyelinated sensory neurons that may detect gentle touch

C-fiber tactile afferents are a subpopulation of unmyelinated cutaneous sensory neurons activated by gentle stroking. Using a genetically encoded tracer, we found that Mas-related G protein-coupled receptor B4 marks a rare subpopulation of unmyelinated, nonpeptidergic sensory fibers that exclusively innervate hairy skin. These fibers terminate in large arborizations similar in size and distribution to C-fiber tactile afferent receptive fields, suggesting that MrgprB4 may provide genetic access to these elusive neurons in mice.

Artemin Overexpression in Skin Enhances Expression of TRPV1 and TRPA1 in Cutaneous Sensory Neurons and Leads to Behavioral Sensitivity to Heat and Cold

Artemin, a neuronal survival factor in the glial cell line-derived neurotrophic factor family, binds the glycosylphosphatidylinositol-anchored protein GFRalpha3 and the receptor tyrosine kinase Ret. Expression of the GFRalpha3 receptor is primarily restricted to the peripheral nervous system and is found in a subpopulation of nociceptive sensory neurons of the dorsal root ganglia (DRGs) that coexpress the Ret and TrkA receptor tyrosine kinases and the thermosensitive channel TRPV1. To determine how artemin affects sensory neuron properties, transgenic mice that overexpress artemin in skin keratinocytes (ART-OE mice) were analyzed. Expression of artemin caused a 20.5% increase in DRG neuron number and increased the level of mRNA encoding GFRalpha3, TrkA, TRPV1, and the putative noxious cold-detecting channel TRPA1. Nearly all GFRalpha3-positive neurons expressed TRPV1 immunoreactivity, and most of these neurons were also positive for TRPA1. Interestingly, acid-sensing ion channel (ASIC) 1, 2a, 2b, and 3 mRNAs were decreased in the DRG, and this reduction was strongest in females. Analysis of sensory neuron physiological properties using an ex vivo preparation showed that cutaneous C-fiber nociceptors of ART-OE mice had reduced heat thresholds and increased firing rates in response to a heat ramp. No change in mechanical threshold was detected. Behavioral testing of ART-OE mice showed that they had increased sensitivity to both heat and noxious cold. These results indicate that the level of artemin in the skin modulates gene expression and response properties of afferents that project to the skin and that these changes lead to behavioral sensitivity to both hot and cold stimuli.

Graded Activity of Transcription Factor Runx3 Specifies the Laminar Termination Pattern of Sensory Axons in the Developing Spinal Cord

Different functional classes of dorsal root ganglion sensory neurons project their axons to distinct target zones within the developing spinal cord. To explore the mechanisms that link sensory neuron subtype identity and axonal projection pattern, we analyzed the roles of Runx and ETS transcription factors in the laminar targeting of sensory afferents. Gain- and loss-of-function studies in chick embryos reveal that the status of Runx3 expression is a major determinant of the dorso-ventral position of termination of proprioceptive and cutaneous sensory axons. In addition, the level of expression and/or activity of Runx3 in individual proprioceptive sensory neurons appears to specify whether their axons terminate in intermediate or ventral regions. Our findings suggest that the selectivity of Runx3 expression, and its level of activity, control sensory afferent targeting in the developing spinal cord.

A Role for Runx Transcription Factor Signaling in Dorsal Root Ganglion Sensory Neuron Diversification

Subpopulations of sensory neurons in the dorsal root ganglion (DRG) can be characterized on the basis of sensory modalities that convey distinct peripheral stimuli, but the molecular mechanisms that underlie sensory neuronal diversification remain unclear. Here, we have used genetic manipulations in the mouse embryo to examine how Runx transcription factor signaling controls the acquisition of distinct DRG neuronal subtype identities. Runx3 acts to diversify an Ngn1-independent neuronal cohort by promoting the differentiation of proprioceptive sensory neurons through erosion of TrkB expression in prospective TrkC+ sensory neurons. In contrast, Runx1 controls neuronal diversification within Ngn1-dependent TrkA+ neurons by repression of neuropeptide CGRP expression and controlling the fine pattern of laminar termination in the dorsal spinal cord. Together, our findings suggest that Runx transcription factor signaling plays a key role in sensory neuron diversification.

IL-31: A new link between T cells and pruritus in atopic skin inflammation

IL-31 is a novel T-cell-derived cytokine that induces severe pruritus and dermatitis in transgenic mice, and signals through a heterodimeric receptor composed of IL-31 receptor A and oncostatin M receptor. To investigate the role of human IL-31 in pruritic and nonpruritic inflammatory skin diseases. The expression of IL-31 was analyzed by quantitative real-time PCR in skin samples of healthy individuals and patients with chronic inflammatory skin diseases. Moreover, IL-31 expression was analyzed in nonlesional skin of atopic dermatitis patients after allergen or superantigen exposure, as well as in stimulated leukocytes. The tissue distribution of the IL-31 receptor heterodimer was investigated by DNA microarray analysis. IL-31 was significantly overexpressed in pruritic atopic compared with nonpruritic psoriatic skin inflammation. Highest IL-31 levels were detected in prurigo nodularis, one of the most pruritic forms of chronic skin inflammation. In vivo, staphylococcal superantigen rapidly induced IL-31 expression in atopic individuals. In vitro, staphylococcal enterotoxin B but not viruses or T(H)1 and T(H)2 cytokines induced IL-31 in leukocytes. In patients with atopic dermatitis, activated leukocytes expressed significantly higher IL-31 levels compared with control subjects. IL-31 receptor A showed most abundant expression in dorsal root ganglia representing the site where the cell bodies of cutaneous sensory neurons reside. Our findings provide a new link among staphylococcal colonization, subsequent T-cell recruitment/activation, and pruritus induction in patients with atopic dermatitis. Taken together, these findings show that IL-31 may represent a novel target for antipruritic drug development.

The Editors' Choice

The Editors' Choice

Adaptation of trigeminal ganglion cells to periodic whisker deflections

Trigeminal ganglion neurons in adult rats adapt to periodic whisker deflections in the range of 1–40 Hz, manifested as a reduction in spike counts to progressively later stimuli in a train of pulsatile or sinusoidal deflections. For high velocity, pulsatile deflections, adaptation is time- and frequency-dependent; as in the case of thalamic and cortical neurons, adaptation is greater at higher stimulus frequencies. With slower velocity, sinusoidal movements, trigeminal ganglion cells differ from central neurons, however, by exhibiting strong adaptation even at low frequencies. For both types of stimuli, effects in trigeminal ganglion neurons were more pronounced in rats maintained during the recording session under neuromuscular blockade than in non-paralysed animals. Results are consistent with previous findings in other systems that frequency-dependent adaptation of cutaneous primary afferent neurons is affected by mechanical properties of the skin. Such effects are likely to vary depending on the nature of the whisker stimuli and physiological states that affect skin viscoelasticity.

Trigeminal P2X 3 receptor expression differs from dorsal root ganglion and is modulated by deep tissue inflammation

The distribution and modulation of the P2X 3 receptor was studied in trigeminal ganglion neurons to provide insight into the role of ATP in craniofacial sensory mechanisms. Binding to the d-galactose specific lectin IB4 was found in 73% of P2X 3-positive neurons while only 16% of IB4 neurons expressed P2X 3. Neurons expressing P2X 3 alone were significantly larger than IB4-or IB4/P2X 3-positive neurons. Investigation of target-specificity revealed that 22% of trigeminal ganglion muscle afferent neurons were positive for P2X 3 versus 16% of cutaneous afferent neurons. Muscle P2X 3 afferents were significantly smaller than the overall muscle afferent population while P2X 3 cutaneous afferent neurons were not. Presumptive heteromeric (P2X 2/3) muscle afferent neurons were also identified and comprised 77% of the P2X 3 muscle afferent population. Muscle afferent neurons co-expressed P2X 3 with either calcitonin gene-related peptide (15%) or substance P (4%). The number of P2X 3-positive muscle afferent neurons significantly increased one and four days following complete Freund's adjuvant-induced masseter muscle inflammation, but significantly decreased after 12 days. These results indicate that within trigeminal ganglia: (1) the P2X 3 receptor is expressed in both small and medium-sized neurons; (2) the P2X 3 receptor is not exclusively expressed in IB4 neurons; (3) P2X 3 is co-expressed with neuropeptides; (4) differences in the proportion of cutaneous versus muscle P2X 3 afferents are not apparent. Trigeminal P2X 3 neurons therefore differ markedly from dorsal root ganglion P2X 3 afferents. This study also shows that deep tissue inflammation modulates expression of the P2X 3 receptor and thus may warrant exploration as a target for therapeutic intervention.

Schwann Cell Engraftment Into Injured Peripheral Nerve Prevents Changes in Action Potential Properties

Peripheral nerve injury results in changes in action potential waveform, ion channel organization, and firing properties of primary afferent neurons. It has been suggested that these changes are the result of reduction in basal trophic support from skin targets. Subcutaneous injections of Fluro-Gold (FG) in the hind limb of the rat were used to identify cutaneous primary afferent neurons. Five days after FG injection, sciatic nerves were ligated and encapsulated in a silicon tube allowing neuroma formation. Green fluorescent protein (GFP)-expressing Schwann cells (SCs) were injected proximal to the cut end of the nerve. Thirteen to 22 days after injury and SC injection, the L4 and L5 dorsal root ganglia (DRG) were prepared for acute culture. Whole cell patch-clamp recordings in current clamp mode were obtained and action potential properties of medium-sized (34-45 microm) FG+ DRG neurons were characterized. In the neuroma group without cell transplantation, action potential duration and spike inflections were reduced as were the amplitude and duration of spike afterhyperpolarizations. These changes were not observed after transection by nerve crush where axons were allowed to regenerate to distal peripheral targets. In the transplantation group, GFP(+)-SCs were extensively distributed throughout the neuroma, and oriented longitudinally along axons proximal to the neuroma. Changes in action potential properties were attenuated in the GFP(+)-SC group. Thus the engrafted SC procedure ameliorated the changes in action potential waveform of cutaneous primary afferents associated with target disconnection and neuroma formation.

Properties of mouse cutaneous rapidly adapting afferents: relationship to skin viscoelasticity.

When skin is stretched, stimuli experienced by a cutaneous mechanoreceptor neuron are transmitted to the nerve ending through the skin. In these experiments, we tested the hypothesis that the viscoelastic response of the skin influences the dynamic response of cutaneous rapidly adapting (RA) neurons. Cutaneous RA afferent neurons were recorded in 3 species of mice (Tsk, Pallid, and C57BL6) whose skin has different viscoelastic properties. Isolated samples of skin and nerve were stimulated mechanically with a dynamic stretch stimulus, which followed a pseudo Gaussian waveform with a bandwidth of 0-60 Hz. The mechanical response of the skin was measured as were responses of single RA cutaneous mechanoreceptor neurons. For each neuron, the strength of association between spike responses and the dynamic and static components of stimuli were determined with multiple logistic regression analysis. The viscoelastic material properties of each skin sample were determined indirectly, by creating a nonlinear (Wiener-Volterra) model of the stress-strain relationship, and using the model to predict the complex compliance (i.e., the viscoelastic material properties). The dynamic sensitivity of RA mechanoreceptor neurons in mouse hairy skin was weakly related to the viscoelastic properties of the skin. Loss modulus and phase angle were lower (indicating a decreased viscous component of response) in Tsk and Pallid than in C57BL6 mice. However, RA mechanoreceptor neurons in Tsk and Pallid skin did not differ from those in C57 skin with regard to their sensitivity to the rate of change of stress or to the rate of change of incremental strain energy. They did have a decreased sensitivity to the rate of change of tensile strain. Thus the skin samples with lower dynamic mechanical response contained neurons with a somewhat lower sensitivity to dynamic stimuli.

The P2Y agonist UTP activates cutaneous afferent fibers

The majority of adenosine triphosphate (ATP)-induced nociceptive transduction and pain has been attributed to ionotropic P2X3 receptors. Metabotropic P2Y receptors, some of which bind pyrimidines as well as purines, have received little attention. Here we have examined the ability of P2Y receptor signaling to evoke action potential firing in functionally identified afferent fibers using the skin nerve preparation from adult mouse. The P2Y2/P2Y4 ligand UTP activated sustained action potential firing in 54% of C fibers in a concentration-dependent manner. The effect was specific for P2Y2/P2Y4 receptors, as the P2Y6 ligand UDP never activated C fibers. In comparison to C fibers, few thinly myelinated A-mechanoreceptors (AM) (12%) were activated by UTP. The majority (70–80%) of the UTP-sensitive C and Aδ fibers responded to the algogen capsaicin with a barrage of action potentials, whereas the UTP-insensitive fibers were largely unresponsive to capsaicin. Furthermore, 86% of the UTP-sensitive C fibers and 100% of the UTP-sensitive AM fibers also responded to the P2X agonist α,β-methylene ATP, indicating that P2Y and P2X receptors are widely co-expressed. Surprisingly, a significant proportion (20–40%) of low threshold slowly and rapidly adapting Aβ fibers were also activated by UTP and α,β-methylene ATP. These data indicate that P2Y receptors on the terminals of capsaicin-sensitive cutaneous sensory neurons effectively evoke nociceptive transmission, and support the hypothesis that UTP may be an endogenous nociceptive messenger. Furthermore, P2Y signaling may contribute to mechanotransduction in low threshold Aβ fibers under normal or pathological conditions.

Spinal nerve injury increases the percentage of cold-responsive DRG neurons

We tested the hypothesis that cold allodynia, observed following nerve injury reflects change(s) in the cold responsiveness of sensory neurons. To test this hypothesis we assessed the impact of the spinal nerve ligation (SNL) model of nerve injury on the responses of cutaneous sensory neurons to cooling in vitro. Nerve injury induced a significant increase in the incidence of cold responsive cutaneous neurons in uninjured but not injured ganglia. Because an increase in the percentage of cold responsive neurons in uninjured ganglia should increase the total neuronal response to cooling of peripheral tissue, these findings suggest that cold allodynia reflects, at least in part, a change in sensory neurons.

Selective Reactions of Cutaneous and Muscle Afferent Neurons to Peripheral Nerve Transection in Rats

To determine whether peripheral nerve injury has similar effects on all functional types of afferent neuron, we retrogradely labeled populations of neurons projecting to skin and to muscle with FluoroGold and lesioned various peripheral nerves in the rat. Labeled neurons were counted after different periods and related to immunohistochemically identified ectopic terminals and satellite cells in lumbar dorsal root ganglia. After 10 weeks, 30% of cutaneous afferent somata labeled from transected sural nerves had disappeared but, if all other branches of the sciatic nerve had also been cut, 60% of cutaneous neurons were lost. Small-diameter sural neurons preferentially disappeared. In contrast, the number of muscle afferent somata was not affected by transection of various nerves. p75 was downregulated in axotomized cutaneous neurons but in not axotomized muscle neurons. Conversely, p75 was upregulated in satellite cells around cutaneous but not muscle neurons. Consistent with this, perineuronal rings containing tyrosine hydroxylase, calcitonin gene-related peptide, galanin, or synaptophysin were formed preferentially around cutaneous neurons. Selective lesions of predominantly cutaneous nerves triggered the formation of rings, but none were detected after selective lesions of muscle nerves. We conclude that cutaneous neurons are both more vulnerable and more associated with ectopic nerve terminals than muscle neurons in dorsal root ganglia after transection and ligation of peripheral nerves.

Bladder and cutaneous sensory neurons of the rat express different functional p2x receptors

The expression and functional responses of P2X receptors in bladder and cutaneous sensory neurons of adult rats and mice have been studied using immunohistochemistry and patch clamp techniques. Cell bodies of bladder pelvic afferents were identified in L6 and S1 dorsal root ganglia (DRG), following Fast Blue injection into the muscle wall of the urinary bladder. Similarly, cutaneous sensory neurons were identified in L3 and L4 DRG, following Fast Blue injection into the saphenous nerve innervating the skin. Bladder sensory neurons contained only weak to moderate P2X 3-immunoreactivity (IR), in contrast to strong P2X 3-IR observed in a sub-population of cutaneous afferents. Whole-cell patch-clamp recordings revealed that approximately 90% of bladder afferent neurons responded to αβ-methylene ATP (αβmeATP) and ATP (30 μM) with persistent currents, which were inhibited by 2′,3′- O-trinitrophenyl-ATP (TNP-ATP) (0.3 μM) to 6.4±1.9% and 8.0±2.6% of control, respectively ( n=8). The remaining bladder sensory neurons demonstrated biphasic, transient or no response to P2X agonists. In contrast, only 24% of cutaneous afferent neurons gave persistent currents to αβmeATP (30 μM), with 66% of cells giving transient or biphasic currents and the remaining 10% being non-responsive. Our results suggest that, in contrast to DRG neurons in general, bladder sensory neurons projecting via pelvic nerves express predominantly P2X 2/3 heteromeric receptors, which are likely to mediate the important roles of ATP as a signaling molecule of urinary bladder filling and nociception.

Histological and Electrical Properties of Rat Dorsal Root Ganglion Neurons Innervating the Lower Urinary Tract

We investigated whether primary afferent neurons innervating different regions of the lower urinary tract have different histochemical and electrophysiological properties. Neurons in rat L6-S1 DRG were identified by axonal transport of a fluorescent dye. Neurofilament-negative C-fiber cells comprise approximately 70% of bladder and proximal urethral afferent neurons that send axons through the pelvic nerves, but comprise a smaller proportion (51%) of distal urethral neurons that send axons through the pudendal nerves. Isolectin-B4 (IB4) binding was detected in a higher percentage (49%) of C-fiber neurons innervating the distal urethra than in those innervating the bladder or proximal urethra (18-22%). Neurofilament-positive A-fiber neurons innervating the distal urethra had a larger average somal size than neurons innervating the bladder or proximal urethra. In patch-clamp recordings, the majority (70%) of bladder and proximal urethral neurons were sensitive to capsaicin and exhibited TTX-resistant, high-threshold action potentials, whereas a smaller proportion (53%) of distal urethral neurons exhibited TTX-resistant spikes. T-type Ca2+ currents were observed in 47% of distal urethral neurons with TTX-sensitive spikes, but not in TTX-sensitive bladder or proximal urethral neurons. In summary, afferent neurons innervating bladder or proximal urethra differ from those innervating distal urethra. The latter, which more closely resemble cutaneous afferent neurons, consist of a smaller number of C-fiber neurons containing a higher percentage of IB4-positive cells and a more diverse population of A-fiber neurons, some of which exhibit T-type Ca2+ channels. These differences may be related to different functions of respective target organs in the lower urinary tract.