Biomedical subjects

Clifford J Woolf

Publications and source records attributed to Clifford J Woolf.

At least 19 recordsLinked to original sources

PubMed2006

The voltage-gated sodium channel Na(v)1.9 is an effector of peripheral inflammatory pain hypersensitivity.

We used a mouse with deletion of exons 4, 5, and 6 of the SCN11A (sodium channel, voltage-gated, type XI, alpha) gene that encodes the voltage-gated sodium channel Na(v)1.9 to assess its contribution to pain. Na(v)1.9 is present in nociceptor sensory neurons that express TRPV1, bradykinin B2, and purinergic P2X3 receptors. In Na(v)1.9-/- mice, the non-inactivating persistent tetrodotoxin-resistant sodium TTXr-Per current is absent, whereas TTXr-Slow is unchanged. TTXs currents are unaffected by the mutation of Na(v)1.9. Pain hypersensitivity elicited by intraplantar administration of prostaglandin E2, bradykinin, interleukin-1beta, capsaicin, and P2X3 and P2Y receptor agonists, but not NGF, is either reduced or absent in Na(v)1.9-/- mice, whereas basal thermal and mechanical pain sensitivity is unchanged. Thermal, but not mechanical, hypersensitivity produced by peripheral inflammation (intraplanatar complete Freund's adjuvant) is substantially diminished in the null allele mutant mice, whereas hypersensitivity in two neuropathic pain models is unchanged in the Na(v)1.9-/- mice. Na(v)1.9 is, we conclude, an effector of the hypersensitivity produced by multiple inflammatory mediators on nociceptor peripheral terminals and therefore plays a key role in mediating peripheral sensitization.

Animals↗

PubMed2006

GTP cyclohydrolase and tetrahydrobiopterin regulate pain sensitivity and persistence.

We report that GTP cyclohydrolase (GCH1), the rate-limiting enzyme for tetrahydrobiopterin (BH4) synthesis, is a key modulator of peripheral neuropathic and inflammatory pain. BH4 is an essential cofactor for catecholamine, serotonin and nitric oxide production. After axonal injury, concentrations of BH4 rose in primary sensory neurons, owing to upregulation of GCH1. After peripheral inflammation, BH4 also increased in dorsal root ganglia (DRGs), owing to enhanced GCH1 enzyme activity. Inhibiting this de novo BH4 synthesis in rats attenuated neuropathic and inflammatory pain and prevented nerve injury-evoked excess nitric oxide production in the DRG, whereas administering BH4 intrathecally exacerbated pain. In humans, a haplotype of the GCH1 gene (population frequency 15.4%) was significantly associated with less pain following diskectomy for persistent radicular low back pain. Healthy individuals homozygous for this haplotype exhibited reduced experimental pain sensitivity, and forskolin-stimulated immortalized leukocytes from haplotype carriers upregulated GCH1 less than did controls. BH4 is therefore an intrinsic regulator of pain sensitivity and chronicity, and the GTP cyclohydrolase haplotype is a marker for these traits.

Adult↗

PubMed2006

Prostaglandin E2 receptor EP4 contributes to inflammatory pain hypersensitivity.

Prostaglandin E(2) (PGE(2)) is both an inflammatory mediator released at the site of tissue inflammation and a neuromodulator that alters neuronal excitability and synaptic processing. The effects of PGE(2) are mediated by four G-protein-coupled EP receptors (EP1-EP4). Here we show that the EP4 receptor subtype is expressed by a subset of primary sensory dorsal root ganglion (DRG) neurons, and that its levels, but not that of the other EP1-3 subtypes, increase in the DRG after complete Freund' adjuvant-induced peripheral inflammation. Administration of both an EP4 antagonist [AH23848, (4Z)-7-[(rel-1S,2S,5R)-5-((1,1'-biphenyl-4-yl)methoxy)-2-(4-morpholinyl)-3-oxocyclopentyl]-4-heptenoic acid] and EP4 knockdown with intrathecally delivered short hairpin RNA attenuates inflammation-induced thermal and mechanical behavioral hypersensitivity, without changing basal pain sensitivity. AH23848 also reduces the PGE(2)-mediated sensitization of capsaicin-evoked currents in DRG neurons in vitro. These data suggest that EP4 is a potential target for the pharmacological treatment of inflammatory pain.

Animals↗

PubMed2006

The transcription factor ATF-3 promotes neurite outgrowth.

Dorsal root ganglion (DRG) neurons regenerate after a peripheral nerve injury but not after injury to their axons in the spinal cord. A key question is which transcription factors drive the changes in gene expression that increase the intrinsic growth state of peripherally injured sensory neurons? A prime candidate is activating transcription factor-3 (ATF-3), a transcription factor that we find is induced in all DRG neurons after peripheral, but not central axonal injury. Moreover, we show in adult DRG neurons that a preconditioning peripheral, but not central axonal injury, increases their growth, correlating closely with the pattern of ATF-3 induction. Using viral vectors, we delivered ATF-3 to cultured adult DRG neurons and find that ATF-3 enhances neurite outgrowth. Furthermore, ATF-3 promotes long sparsely branched neurites. ATF-3 overexpression did not increase c-Jun expression. ATF-3 may contribute, therefore, to neurite outgrowth by orchestrating the gene expression responses in injured neurons.

Activating Transcription Factor 3↗

PubMed2006

Persistent postsurgical pain: risk factors and prevention.

Acute postoperative pain is followed by persistent pain in 10-50% of individuals after common operations, such as groin hernia repair, breast and thoracic surgery, leg amputation, and coronary artery bypass surgery. Since chronic pain can be severe in about 2-10% of these patients, persistent postsurgical pain represents a major, largely unrecognised clinical problem. Iatrogenic neuropathic pain is probably the most important cause of long-term postsurgical pain. Consequently, surgical techniques that avoid nerve damage should be applied whenever possible. Also, the effect of aggressive, early therapy for postoperative pain should be investigated, since the intensity of acute postoperative pain correlates with the risk of developing a persistent pain state. Finally, the role of genetic factors should be studied, since only a proportion of patients with intraoperative nerve damage develop chronic pain. Based on information about the molecular mechanisms that affect changes to the peripheral and central nervous system in neuropathic pain, several opportunities exist for multimodal pharmacological intervention. Here, we outline strategies for identification of patients at risk and for prevention and possible treatment of this important entity of chronic pain.

Analgesics↗

PubMed2006

TRPA1 contributes to cold, mechanical, and chemical nociception but is not essential for hair-cell transduction.

TRPA1, a member of the transient receptor potential (TRP) family of ion channels, is expressed by dorsal root ganglion neurons and by cells of the inner ear, where it has proposed roles in sensing sound, painful cold, and irritating chemicals. To test the in vivo roles of TRPA1, we generated a mouse in which the essential exons required for proper function of the Trpa1 gene were deleted. Knockout mice display behavioral deficits in response to mustard oil, to cold ( approximately 0 degrees C), and to punctate mechanical stimuli. These mice have a normal startle reflex to loud noise, a normal sense of balance, a normal auditory brainstem response, and normal transduction currents in vestibular hair cells. TRPA1 is apparently not essential for hair-cell transduction but contributes to the transduction of mechanical, cold, and chemical stimuli in nociceptor sensory neurons.

Animals↗

PubMed2006

Bone morphogenetic protein signaling by hemojuvelin regulates hepcidin expression.

Hepcidin is a key regulator of systemic iron homeostasis. Hepcidin deficiency induces iron overload, whereas hepcidin excess induces anemia. Mutations in the gene encoding hemojuvelin (HFE2, also known as HJV) cause severe iron overload and correlate with low hepcidin levels, suggesting that hemojuvelin positively regulates hepcidin expression. Hemojuvelin is a member of the repulsive guidance molecule (RGM) family, which also includes the bone morphogenetic protein (BMP) coreceptors RGMA and DRAGON (RGMB). Here, we report that hemojuvelin is a BMP coreceptor and that hemojuvelin mutants associated with hemochromatosis have impaired BMP signaling ability. Furthermore, BMP upregulates hepatocyte hepcidin expression, a process enhanced by hemojuvelin and blunted in Hfe2-/- hepatocytes. Our data suggest a mechanism by which HFE2 mutations cause hemochromatosis: hemojuvelin dysfunction decreases BMP signaling, thereby lowering hepcidin expression.

Amino Acid Sequence↗

PubMed2006

Runx1 determines nociceptive sensory neuron phenotype and is required for thermal and neuropathic pain.

In mammals, the perception of pain is initiated by the transduction of noxious stimuli through specialized ion channels and receptors expressed by nociceptive sensory neurons. The molecular mechanisms responsible for the specification of distinct sensory modality are, however, largely unknown. We show here that Runx1, a Runt domain transcription factor, is expressed in most nociceptors during embryonic development but in adult mice, becomes restricted to nociceptors marked by expression of the neurotrophin receptor Ret. In these neurons, Runx1 regulates the expression of many ion channels and receptors, including TRP class thermal receptors, Na+-gated, ATP-gated, and H+-gated channels, the opioid receptor MOR, and Mrgpr class G protein coupled receptors. Runx1 also controls the lamina-specific innervation pattern of nociceptive afferents in the spinal cord. Moreover, mice lacking Runx1 exhibit specific defects in thermal and neuropathic pain. Thus, Runx1 coordinates the phenotype of a large cohort of nociceptors, a finding with implications for pain therapy.

Analysis of Variance↗

PubMed2006

Bradykinin and peripheral sensitization.

Pain hypersensitivity after tissue injury and inflammation is contributed to by a reduction in the threshold and an increase in the responsiveness of the peripheral terminals of high-threshold nociceptor neurons, the phenomenon of peripheral sensitization. Bradykinin, acting via G-protein-coupled receptors expressed by the sensory neurons, links to multiple intracellular signaling pathways that in turn interact with voltage-gated and ligand-gated ion channels, changing their properties in such a way as to enhance the response to peripheral stimuli.

Animals↗

PubMed2005

Detection of cold pain, cold allodynia and cold hyperalgesia in freely behaving rats.

BACKGROUND: Pain is elicited by cold, and a major feature of many neuropathic pain states is that normally innocuous cool stimuli begin to produce pain (cold allodynia). To expand our understanding of cold induced pain states we have studied cold pain behaviors over a range of temperatures in several animal models of chronic pain. RESULTS: We demonstrate that a Peltier-cooled cold plate with +/- 1 degrees C sensitivity enables quantitative measurement of a detection withdrawal response to cold stimuli in unrestrained rats. In naïve rats the threshold for eliciting cold pain behavior is 5 degrees C. The withdrawal threshold for cold allodynia is 15 degrees C in both the spared nerve injury and spinal nerve ligation models of neuropathic pain. Cold hyperalgesia is present in the spared nerve injury model animals, manifesting as a reduced latency of withdrawal response threshold at temperatures that elicit cold pain in naïve rats. We also show that following the peripheral inflammation produced by intraplantar injection of complete Freund's adjuvant, a hypersensitivity to cold occurs. CONCLUSION: The peltier-cooled provides an effective means of assaying cold sensitivity in unrestrained rats. Behavioral testing of cold allodynia, hyperalgesia and pain will greatly facilitate the study of the neurobiological mechanisms involved in cold/cool sensations and enable measurement of the efficacy of pharmacological treatments to reduce these symptoms.

Afferent Pathways↗

PubMed2005

RNAi blocks DYT1 mutant torsinA inclusions in neurons.

Early onset generalized dystonia is a dominantly inherited movement disorder caused by neuronal dysfunction without an apparent loss of neurons. The same single mutation (GAG deletion) causes most cases and results in loss of a glutamic acid (E) in the carboxy terminal region of torsinA (Delta302/303). To model the neuronal involvement, adult rat primary sensory dorsal root ganglion neurons in culture were infected with lentivirus vectors expressing human wild-type or mutant torsinA. Expression of the mutant protein resulted in formation of torsinA-positive perinuclear inclusions. When the cells were co-infected with lentivirus vectors expressing the mutant torsinA message and a shRNA selectively targeting this message, inclusion formation was blocked. Vector-delivered siRNAs have the potential to decrease the adverse effects of this mutant protein in neurons without affecting wild-type protein.

Animals↗

PubMed2005

Upregulation of the voltage-gated sodium channel beta2 subunit in neuropathic pain models: characterization of expression in injured and non-injured primary sensory neurons.

The development of abnormal primary sensory neuron excitability and neuropathic pain symptoms after peripheral nerve injury is associated with altered expression of voltage-gated sodium channels (VGSCs) and a modification of sodium currents. To investigate whether the beta2 subunit of VGSCs participates in the generation of neuropathic pain, we used the spared nerve injury (SNI) model in rats to examine beta2 subunit expression in selectively injured (tibial and common peroneal nerves) and uninjured (sural nerve) afferents. Three days after SNI, immunohistochemistry and Western blot analysis reveal an increase in the beta2 subunit in both the cell body and peripheral axons of injured neurons. The increase persists for >4 weeks, although beta2 subunit mRNA measured by real-time reverse transcription-PCR and in situ hybridization remains unchanged. Although injured neurons show the most marked upregulation,beta2 subunit expression is also increased in neighboring non-injured neurons and a similar pattern of changes appears in the spinal nerve ligation model of neuropathic pain. That increased beta2 subunit expression in sensory neurons after nerve injury is functionally significant, as demonstrated by our finding that the development of mechanical allodynia-like behavior in the SNI model is attenuated in beta2 subunit null mutant mice. Through its role in regulating the density of mature VGSC complexes in the plasma membrane and modulating channel gating, the beta2 subunit may play a key role in the development of ectopic activity in injured and non-injured sensory afferents and, thereby, neuropathic pain.

Animals↗

PubMed2005

Mutant SPTLC1 dominantly inhibits serine palmitoyltransferase activity in vivo and confers an age-dependent neuropathy.

Mutations in enzymes involved in sphingolipid metabolism and trafficking cause a variety of neurological disorders, but details of the molecular pathophysiology remain obscure. SPTLC1 encodes one subunit of serine palmitoyltransferase (SPT), the rate-limiting enzyme in sphingolipid synthesis. Mutations in SPTLC1 cause hereditary sensory and autonomic neuropathy (type I) (HSAN1), an adult onset, autosomal dominant neuropathy. HSAN1 patients have reduced SPT activity. Expression of mutant SPTLC1 in yeast and mammalian cell cultures dominantly inhibits SPT activity. We created transgenic mouse lines that ubiquitously overexpress either wild-type (SPTLC1(WT)) or mutant SPTLC1 (SPTLC1(C133W)). We report here that SPTLC1(C133W) mice develop age-dependent weight loss and mild sensory and motor impairments. Aged SPTLC1(C133W) mice lose large myelinated axons in the ventral root of the spinal cord and demonstrate myelin thinning. There is also a loss of large myelinated axons in the dorsal roots, although the unmyelinated fibers are preserved. In the dorsal root ganglia, IB4 staining is diminished, whereas expression of the injury-induced transcription factor ATF3 is increased. These mice represent a novel mouse model of peripheral neuropathy and confirm the link between mutant SPT and neuronal dysfunction.

Aging↗

PubMed2005

Bradykinin produces pain hypersensitivity by potentiating spinal cord glutamatergic synaptic transmission.

Bradykinin, an inflammatory mediator, sensitizes nociceptor peripheral terminals reducing pain threshold. We now show that the B2 kinin receptor is expressed in rat dorsal horn neurons and that bradykinin, a B2-specific agonist, augments AMPA- and NMDA-induced, and primary afferent-evoked EPSCs, and increases the frequency and amplitude of miniature EPSCs in superficial dorsal horn neurons in vitro. Administration of bradykinin to the spinal cord in vivo produces, moreover, an NMDA-dependent hyperalgesia. We also demonstrate that nociceptive inputs result in the production of bradykinin in the spinal cord and that an intrathecal B2-selective antagonist suppresses behavioral manifestations of central sensitization, an activity-dependent increase in glutamatergic synaptic efficacy. Primary afferent-evoked central sensitization is, in addition, reduced in B2 receptor knock-out mice. We conclude that bradykinin is released in the spinal cord in response to nociceptor inputs and acts as a synaptic neuromodulator, potentiating glutamatergic synaptic transmission to produce pain hypersensitivity.

Animals↗

PubMed2005

Constitutive cyclo-oxygenase-2 does not contribute to the development of human visceral pain hypersensitivity.

BACKGROUND AND AIMS: Central sensitisation (CS), contributes to the development and maintenance of gastrointestinal pain hypersensitivity. Constitutive cyclo-oxygenase-2 (COX-2) contributes to central sensitisation in somatic pain hypersensitivity but its role in mediating visceral pain hypersensitivity is unknown. We therefore conducted a study to determine if COX-2 inhibition with Valdecoxib attenuates the development or early maintenance of CS in a validated human oesophageal pain hypersensitivity model. METHODS: Healthy volunteers were studied in two randomised, double blind, crossover studies in which pain thresholds (PT) to electrical stimulation were assessed in the proximal oesophagus, chest wall and foot, prior to and following a distal oesophageal acid infusion. Protocol 1: Valdecoxib, (40 mg) or matching placebo was given orally for 4 days prior to oesophageal acid infusion. Protocol 2: IV Parecoxib (40 mg) or saline was given 120 min after oesophageal acid infusion. RESULTS: Valdecoxib did not prevent the induction of secondary allodynia in the proximal oesophagus nor did it attenuate it following its establishment. Chest wall PT fell following oesophageal acid but foot PT remained unchanged; highlighting the development viscero-somatic convergence due to CS. Valdecoxib had no analgesic or anti-hyperalgesic effect on chest wall or foot PT. CONCLUSIONS: Neither the induction nor initial maintenance of acid induced oesophageal pain hypersensitivity is prevented by Valdecoxib, suggesting that constitutive spinal COX-2 does not contribute to the development or early maintenance of acute visceral central sensitisation.

Adult↗

PubMed2005

Blocking caspase activity prevents transsynaptic neuronal apoptosis and the loss of inhibition in lamina II of the dorsal horn after peripheral nerve injury.

We show that transsynaptic apoptosis is induced in the superficial dorsal horn (laminas I-III) of the spinal cord by three distinct partial peripheral nerve lesions: spared nerve injury, chronic constriction, and spinal nerve ligation. Ongoing activity in primary afferents of the injured nerve and glutamatergic transmission cause a caspase-dependent degeneration of dorsal horn neurons that is slow in onset and persists for several weeks. Four weeks after spared nerve injury, the cumulative loss of dorsal horn neurons, determined by stereological analysis, is >20%. GABAergic inhibitory interneurons are among the neurons lost, and a marked decrease in inhibitory postsynaptic currents of lamina II neurons coincides with the induction of apoptosis. Blocking apoptosis with the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone (zVAD) prevents the loss of GABAergic interneurons and the reduction of inhibitory currents. Partial peripheral nerve injury results in pain-like behavioral changes characterized by hypersensitivity to tactile or cold stimuli. Treatment with zVAD, which has no intrinsic analgesic properties, attenuates this neuropathic pain-like syndrome. Preventing nerve injury-induced apoptosis of dorsal horn neurons by blocking caspase activity maintains inhibitory transmission in lamina II and reduces pain hypersensitivity.

Afferent Pathways↗

PubMed2005

Repulsive guidance molecule (RGMa), a DRAGON homologue, is a bone morphogenetic protein co-receptor.

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor beta (TGF-beta) superfamily of ligands, which regulate many mammalian physiologic and pathophysiologic processes. BMPs exert their effects through type I and type II serine/threonine kinase receptors and the Smad intracellular signaling pathway. Recently, the glycosylphosphatidylinositol (GPI)-anchored protein DRAGON was identified as a co-receptor for BMP signaling. Here, we investigate whether a homologue of DRAGON, repulsive guidance molecule (RGMa), is similarly involved in the BMP signaling pathway. We show that RGMa enhances BMP, but not TGF-beta, signals in a ligand-dependent manner in cell culture. The soluble extracellular domain of RGMa fused to human Fc (RGMa.Fc) forms a complex with BMP type I receptors and binds directly and selectively to radiolabeled BMP-2 and BMP-4. RGMa mediates BMP signaling through the classical BMP signaling pathway involving Smad1, 5, and 8, and it up-regulates endogenous inhibitor of differentiation (Id1) protein, an important downstream target of BMP signals. Finally, we demonstrate that BMP signaling occurs in neurons that express RGMa in vivo. These data are consistent with a role for RGMa as a BMP co-receptor.

Amino Acid Sequence↗

PubMed2005

Localization and action of Dragon (repulsive guidance molecule b), a novel bone morphogenetic protein coreceptor, throughout the reproductive axis.

Bone morphogenetic proteins (BMPs) play important roles in reproduction including primordial germ cell formation, follicular development, spermatogenesis, and FSH secretion. Dragon, a recently identified glycosylphosphatidylinositol-anchored member of the repulsive guidance molecule family, is also a BMP coreceptor. In the present study, we determined the tissue and cellular localization of Dragon in reproductive organs using immunohistochemistry and in situ hybridization. Among reproductive organs, Dragon was expressed in testis, epididymis, ovary, uterus, and pituitary. In the testis of early postnatal mice, Dragon was found in gonocytes and spermatogonia, whereas in immature testes, Dragon was only weakly expressed in spermatogonia. Interestingly, pregnant mare serum gonadotropin treatment of immature mice robustly induced Dragon production in spermatocytes. In adult testis, Dragon was found in spermatocytes and round spermatids. In the ovary, Dragon was detected exclusively within oocytes and primarily those within secondary follicles. In the pituitary, Dragon-expressing cells overlapped FSH-expressing cells. Dragon was also expressed in a number of cell lines originating from reproductive tissues including Ishikawa, Hela, LbetaT2, MCF-7, and JEG3 cells. Immunocytochemistry and gradient sucrose ultracentrifugation studies showed Dragon was localized in lipid rafts within the plasma membrane. In reproductive cell lines, Dragon expression enhanced signaling of exogenous BMP2 or BMP4. The present studies demonstrate that Dragon expression is dynamically regulated throughout the reproductive tract and that Dragon protein modulates BMP signaling in cells from reproductive tissues. The overlap between Dragon expression and the functional BMP signaling system suggests that Dragon may play a role in mammalian reproduction.

Animals↗