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Tissue damage is one of the major etiological factors in the emergence of chronic/persistent pain, although mechanisms remain enigmatic. Using incision of the back skin of adult rats as a model for tissue damage, we observed sensitization in a nociceptive reflex enduring to 28 days post-incision (DPI). To determine if the enduring behavioral changes corresponded with a long-term impact of tissue damage on sensory neurons, we examined the temporal expression profile of injury-regulated genes and the electrophysiological properties of traced dorsal root ganglion (DRG) sensory neurons. The mRNA for the injury/stress-hub gene Activating Transcription Factor 3 (ATF3) was upregulated and peaked within 4 DPI, after which levels declined but remained significantly elevated out to 28 DPI, a time when the initial incision appears healed and tissue-inflammation largely resolved. Accordingly, stereological image analysis indicated that some neurons expressed ATF3 only transiently (mostly medium-large neurons), while in others it was sustained (mostly small neurons), suggesting cell-type-specific responses. In retrogradely-traced ATF3-expressing neurons, Calcium/calmodulin-dependent protein kinase type IV (CAMK4) protein levels and isolectin-B4 (IB4)-binding were suppressed whereas Growth Associated Protein-43 (GAP-43) and Neuropeptide Y (NPY) protein levels were enhanced. Electrophysiological recordings from DiI-traced sensory neurons 28 DPI showed a significant sensitization limited to ATF3-expressing neurons. Thus, ATF3 expression is revealed as a strong predictor of single cells displaying enduring pain-related electrophysiological properties. The cellular injury/stress response induced in sensory neurons by tissue damage and indicated by ATF3 expression is positioned to contribute to pain which can occur after tissue damage. [ABSTRACT FROM AUTHOR]

Spinal sensory neurons innervating visceral and mucocutaneous tissues have unique microanatomic distribution, peripheral modality, and physiological, pharmacological, and biophysical characteristics compared with those neurons that innervate muscle and cutaneous tissues. In previous patch-clamp electrophysiological studies, we have demonstrated that small- and medium-diameter dorsal root ganglion (DRG) neurons can be subclassified on the basis of their patterns of voltage-activated currents (VAC). These VAC-based subclasses were highly consistent in their action potential characteristics, responses to algesic compounds, immunocytochemical expression patterns, and responses to thermal stimuli. For this study, we examined the VAC of neurons retrogradely traced from the distal colon and the glans penis/distal urethra in the adult male rat. The afferent population from the distal colon contained at least two previously characterized cell types observed in somatic tissues (types 5 and 8), as well as four novel cell types (types 15, 16, 17, and 18). In the glans penis/distal urethra, two previously described cell types (types 6 and 8) and three novel cell types (types 7, 14, and 15) were identified. Other characteristics, including action potential profiles, responses to algesic compounds (acetylcholine, capsaicin, ATP, and pH 5.0 solution), and neurochemistry (expression of substance P, CGRP, neurofilament, TRPV1, TRPV2, and isolectin B4 binding) were consistent for each VAC-defined subgroup. With identification of distinct DRG cell types that innervate the distal colon and glans penis/distal urethra, future in vitro studies related to the gastrointestinal and urogenital sensory function in normal as well as abnormal/pathological conditions may be benefitted. [ABSTRACT FROM AUTHOR]

Abstract: We have recently found that, following complete Freund’s adjuvant (CFA)-induced inflammation, cutaneous polymodal nociceptors (CPM) lacking the transient receptor potential vanilloid 1 (TRPV1) are sensitized to heat stimuli. In order to determine possible mechanisms playing a role in this change, we examined gene expression in the L2/L3 sensory ganglia following CFA injection into the hairy hind paw skin and found that G-protein-coupled purinoreceptor P2Y1 expression was increased. This receptor is of particular interest, as most CPMs innervating mouse hairy skin bind isolectin B4, which co-localizes with P2Y1. Additionally, our recent findings have shown that cutaneous CPMs in P2Y1−/− mice displayed significantly reduced thermal sensitivity. Together, these findings suggested a possible role for P2Y1 in inflammation-induced heat sensitization in these fibers. To test this hypothesis, we utilized our in vivo small interfering RNA technique to knock down the inflammation-induced increase in P2Y1 expression and then examined the functional effects using ex vivo recording. We found that the normal reduction of heat thresholds in CPM fibers induced by CFA was completely blocked by inhibition of P2Y1. Surprisingly, inhibition of P2Y1 during inflammation also significantly increased the number of CPM neurons expressing TRPV1 without a change in the total number of TRPV1-positive cells in the L2 and L3 dorsal root ganglia. These results show that the inflammation-induced enhanced expression of P2Y1 is required for normal heat sensitization of cutaneous CPM fibers. They also suggest that P2Y1 plays a role in the maintenance of phenotype in cutaneous afferent fibers containing TRPV1. [Copyright &y& Elsevier]

Damage to peripheral nerves is known to contribute to chronic pain states, including mechanical and thermal hyperalgesia and allodynia. It is unknown whether the establishment of these states is attributable to peripheral changes, central modifications, or both. In this study, we used several different approaches to assess the changes in myelinated (A) and unmyelinated (C) cutaneous nociceptors after transection and regeneration of the saphenous nerve. An ex vivo recording preparation was used to examine response characteristics and neurochemical phenotype of different types of functionally defined neurons. We found that myelinated nociceptors had significantly lower mechanical and thermal thresholds after regeneration, whereas C-polymodal nociceptors (CPMs) had lower heat thresholds. There was a significant increase in the percentage of mechanically insensitive C-fibers that responded to heat (CHs) after regeneration. Immunocytochemical analysis of identified afferents revealed that most CPMs were isolectin B4 (IB4) positive and transient receptor potential vanilloid 1 (TRPV1) negative, whereas CHs were always TRPV1 positive and IB4 negative in naive animals (Lawson et al., 2008). However, after regeneration, some identified CPMs and CHs stained positively for both markers, which was apparently attributable to an increase in the total number of IB4-positive neurons. Real-time PCR analysis of L2/L3 DRGs and hairy hindpaw skin at various times after saphenous nerve axotomy suggested multiple changes in neurotrophic factor signaling that correlated with either denervation or reinnervation of the cutaneous target. These changes may underlie the functional alterations observed after nerve regeneration and may explain how nerve damage leads to chronic pain conditions. [ABSTRACT FROM AUTHOR]