A neuroprotective agent that inactivates prodegenerative TrkA and preserves mitochondria.

Axon degeneration is an early event and pathological in neurodegenerative conditions and nerve injuries. To discover agents that suppress neuronal death and axonal degeneration, we performed drug screens on primary rodent neurons and identified the pan-kinase inhibitor foretinib, which potently rescued sympathetic, sensory, and motor wt and SOD1 mutant neurons from trophic factor withdrawal-induced degeneration. By using primary sympathetic neurons grown in mass cultures and Campenot chambers, we show that foretinib protected neurons by suppressing both known degenerative pathways and a new pathway involving unliganded TrkA and transcriptional regulation of the proapoptotic BH3 family members BimEL, Harakiri,and Puma, culminating in preservation of mitochondria in the degenerative setting. Foretinib delayed chemotherapy-induced and Wallerian axonal degeneration in culture by preventing axotomy-induced local energy deficit and preserving mitochondria, and peripheral Wallerian degeneration in vivo. These findings identify a new axon degeneration pathway and a potentially clinically useful therapeutic drug.

[Effect of Electroacupuncture Stimulation of Different Tissues at "Huantiao" (GB 30) Acupoint on Expression of Phosphorylated JNK and c-jun in Spinal Cord of Rats with Sciatic Nerve Injury].

OBJECTIVE: To observe the effects of electroacupuncture (EA) stimulation of different tissues (nerve stem, muscular layer) at "Huantiao" (GB 30) acupoint on expression of hosphorylated c-jun N-terminal kinase (p-JNK) and c-jun (p-c-jun) proteins in the lumbar spinal cord in rats with sciatic nerve injury, so as to explore its mechanism underlying improvement of peripheral neuropathic damage. METHODS: Forty-eight SD rats were randomly divided into normal, model (the left sciatic nerve severed), GB 30 deep needling (the acupuncture needle tip was inserted to the sciatic nerve trunk to elicit an instantaneous jerk of the hind limb) and GB 30 shallow needling (the needle tip was inserted to the muscle layer to evoke a local muscular contraction) groups (n = 12 rats in each group). EA stimuli were delivered at 2 Hz/100 Hz, 1 mA, 20 min in duration per treatment for 10 consecutive days. Histopathological changes were observed by Hematoxylin-eosin (HE) staining and immunohistochemical assay was carried out to examine the pathological change of spinal segments (L4-L5) and the expression of p-JNK and p-c-jun proteins, respectively. RESULTS: For rats with the sciatic nerve severed, the spinal neurons became swelling, degeneration or even apoptosis. Acupuncture intervention reduced the number of apoptosic neurons and improved the pathological change, which was relatively better in the.deep needling group than in the shallow needling group. Likewise, the elevated spinal p-JNK and p-c-jun expression levels of the model group were significantly reduced by EA intervention (deep needling vs shallow needling, P < 0.01. CONCLUSION: Acupuncture can improve the spinal pathological changes in rats with sciatic nerve injury, which is probably achieved by decreasing the p-JNK and p-c-jun expression and inhibiting the JNK signaling pathway, and thereby, reducing the apoptosis of the spinal neurons. Deep needling results in greater benefits than shallow needling.

Intrathecal injection of adenosine 2A receptor agonists reversed neuropathic allodynia through protein kinase (PK)A/PKC signaling.

A single intrathecal dose of adenosine 2A receptor (A2AR) agonist was previously reported to produce a multi-week reversal of allodynia in a chronic constriction injury (CCI) model of neuropathic pain. We aimed to determine if this long-term reversal was induced by A2AR agonism versus more generalized across adenosine receptor subtypes, and begin to explore the intracellular signaling cascades involved. In addition, we sought to identify whether the enduring effect could be extended to other models of neuropathic pain. We tested an A1R and A2BR agonist in CCI and found the same long duration effect with A2BR but not A1R agonism. An A2AR agonist (ATL313) produced a significant long-duration reversal of mechanical allodynia induced by long established CCI (administered 6 weeks after surgery), spinal nerve ligation and sciatic inflammatory neuropathy. To determine if ATL313 had a direct effect on glia, ATL313 was coadministered with lipopolysaccharide to neonatal microglia and astrocytes in vitro. ATL313 significantly attenuated TNFα production in both microglia and astrocytes but had no effect on LPS induced IL-10. Protein kinase C significantly reversed the ATL313 effects on TNFα in vitro in microglia and astrocytes, while a protein kinase A inhibitor only effected microglia. Both intrathecal PKA and PKC inhibitors significantly reversed the effect of the A2AR agonist on neuropathic allodynia. Therefore, A2AR agonists administered IT remain an exciting novel target for the treatment of neuropathic pain.

Peripherally injected linalool and bergamot essential oil attenuate mechanical allodynia via inhibiting spinal ERK phosphorylation.

Bergamot essential oil (BEO) is one of the most common essential oil containing linalool and linalyl acetate as major volatile components. This study investigated the effect of intraplantar (i.pl.) bergamot essential oil (BEO) or linalool on neuropathic hypersensitivity induced by partial sciatic nerve ligation (PSNL) in mice. The i.pl. injection of BEO or linalool into the ipsilateral hindpaw to PSNL reduced PSNL-induced mechanical allodynia in a dose-dependent manner. Peripheral (i.pl.) injection of BEO or linalool into the contralateral hindpaw did not yield anti-allodynic effects, suggesting a local anti-mechanical allodynic effect of BEO or linalool in PSNL mice. Anti-mechanical hypersensitivity of morphine was enhanced by the combined injection of BEO or linalool at an ineffective dose when injected alone. We also examined the possible involvement of spinal extracellular signal-regulated protein kinase (ERK) in BEO or linalool-induced anti-mechanical allodynia. In western blotting analysis, i.pl. injection of BEO or linalool resulted in a significant blockade of spinal ERK activation induced by PSNL. These results suggest that i.pl. injection of BEO or linalool may reduce PSNL-induced mechanical allodynia followed by decreasing spinal ERK activation.

The orally active glutamate carboxypeptidase II inhibitor E2072 exhibits sustained nerve exposure and attenuates peripheral neuropathy.

Peripheral neuropathy from nerve trauma is a significant problem in the human population and often constitutes a dose-limiting toxicity in patients receiving chemotherapy. (3-2-Mercaptoethyl)biphenyl-2,3-dicarboxylic acid (E2072) is a potent (K(i) = 10 nM), selective, and orally available inhibitor of glutamate carboxypeptidase II (GCPII). Here, we report that E2072 attenuates hyperalgesia and nerve conduction velocity deficits in preclinical rodent models of neuropathic pain and oxaliplatin-induced neuropathy. In the chronic constrictive injury model, orally administered E2072 reversed pre-existing thermal hyperalgesia in rats in a dose-dependent fashion with a minimally effective dose of 0.1 mg/kg/day. It is noteworthy that multiple days of dosing of E2072 were required before analgesia was realized even though GCPII inhibitory exposures were achieved on the first day of dosing. In addition, analgesia was found to persist for up to 7 days after cessation of dosing, consistent with E2072's pharmacokinetic profile and sustained exposure. Furthermore, in a chronic oxaliplatin-induced neuropathy model (6 mg/kg i.p. oxaliplatin twice weekly for 4 weeks), female BALB/c mice receiving daily oral E2072 at 1.0 and 0.1 mg/kg displayed no deficits in either caudal or digital velocity compared with significant deficits observed in mice treated with oxaliplatin alone (12 ± 3 and 9 ± 2%, respectively). Similar findings were seen with oxaliplatin-induced digital and caudal amplitude deficits. It is noteworthy that E2072 showed no interference with the antineoplastic efficacy of oxaliplatin in mice bearing leukemia (L1210), even at doses 100 times its neuroprotective/analgesic dose, indicating a selective effect on neuropathy. These data support the therapeutic utility of GCPII inhibitors in neuropathy and neuropathic pain.

Axonal outgrowth is associated with increased ERK 1/2 activation but decreased caspase 3 linked cell death in Schwann cells after immediate nerve repair in rats.

BACKGROUND: Extracellular-signal regulated kinase (ERK1/2) is activated by nerve damage and its activation precedes survival and proliferation of Schwann cells. In contrast, activation of caspase 3, a cysteine protease, is considered as a marker for apoptosis in Schwann cells. In the present study, axonal outgrowth, activation of ERK1/2 by phosphorylation (p-ERK 1/2 ) and immunoreactivity of cleaved caspase 3 were examined after immediate, delayed, or no repair of transected rat sciatic nerves. RESULTS: Axonal outgrowth, detected by neurofilament staining, was longer after immediate repair than after either the delayed or no repair conditions. Immediate repair also showed a higher expression of p-ERK 1/2 and a lower number of cleaved caspase 3 stained Schwann cells than after delayed nerve repair. If the transected nerve was not repaired a lower level of p-ERK 1/2 was found than in either the immediate or delayed repair conditions. Axonal outgrowth correlated to p-ERK 1/2, but not clearly with cleaved caspase 3. Contact with regenerating axons affected Schwann cells with respect to p-ERK 1/2 and cleaved caspase 3 after immediate nerve repair only. CONCLUSION: The decreased regenerative capacity that has historically been observed after delayed nerve repair may be related to impaired activation of Schwann cells and increased Schwann cell death. Outgrowing axons influence ERK 1/2 activation and apoptosis of Schwann cells.

The neuropathy-protective agent acetyl-L-carnitine activates protein kinase C-gamma and MAPKs in a rat model of neuropathic pain.

The gamma isoform of protein kinase C (PKCgamma) is an injury-activated intracellular modulator that boosts neuronal activity in algesic and neuroregenerative signalling pathways. Acetyl-L-carnitine (ALCAR), a physiological compound with role in bioenergetic functions, shows an antihyperalgesic effect and at the same time can exert neuroregenerative and neuroprotective effects. Aimed to explore the link between pain and neuroregeneration, the effect of ALCAR treatment (100 mg kg(-1) i.p. twice daily for 15 days) on PKCgamma and mitogen-activated protein kinases (MAPKs) expression has been evaluated in CCI (chronic constriction injury) rats. The sciatic nerve and the lumbar tract of the spinal cord were processed to evaluate the levels of the phosphorylated form of PKCgamma, ERK 1,2, SAP/JNK, p-38 and c-Jun; furthermore, the mRNA expression of the early genes c-Jun and c-Fos has been investigated. Fifteen days after injury, the analysis in the sciatic nerves highlighted a bilateral increase of the activated forms of PKCgamma, ERK 1,2 and SAP/JNK, whereas c-Jun showed an increase only ipsilaterally. ALCAR completely prevented mechanical hyperalgesia and provoked in the nerve a c-Jun increment only. In the lumbar tract of the spinal cord, higher levels of activated PKCgamma, ERK 1,2, p38, SAP/JNK and c-Jun proteins were detected in the ipsilateral side in respect of sham. ALCAR was able to stimulate this expression profile. At the transcriptional level c-Jun mRNA was increased in the ipsilateral side of spinal cord of CCI saline-treated rats, whereas c-Fos mRNA was unchanged. ALCAR had a stimulatory effect on both these early genes. These findings may represent a different approach in the study of the complex balance between pain and neuroregeneration and could constitute the basis for developing new disease modifying agents in the treatment of neuropathic pain.

Injured nerve-derived COX2/PGE2 contributes to the maintenance of neuropathic pain in aged rats.

Neuropathic pain (NeP) is a debilitating disease afflicting mostly the aged population. Inflammatory responses in injured nerves play a pivotal role in the pathogenesis of NeP. Injured nerve derived cyclooxygenase 2/prostaglandin E2 (COX2/PGE2) contributes to the genesis of NeP at the early stage in young rats. Here we show that COX2/PGE2 is involved in the maintenance of NeP at a chronic stage in aged rats. Eighteen months after partial sciatic nerve ligation (PSNL), NeP remained prominent in aged rats. COX2 expressing macrophages and PGE2 levels were increased in injured nerves. PGE2 receptors (EP1 and EP4) and pain-related ion channel transient receptor potential vanilloid-1 (TRPV1) were increased in the ipsilateral dorsal root ganglion (DRG) neurons of aged PSNL rats. Perineural injection of a selective COX2 inhibitor NS-398 relieved NeP, reversed PSNL increased expression of EP1, EP4 and TRPV1 and suppressed the levels of pain-related peptide substance P and calcitonin gene-related peptide in DRG neurons. These data suggest that injured nerve-derived PGE2 contributes to the maintenance of NeP at the chronic stage in aged rats. Chronically facilitating the synthesis of pain-related molecules in nociceptive DRG neurons is a novel mechanism underpinning the contribution of PGE2.

ERK1/2-mediated Schwann cell proliferation in the regenerating sciatic nerve by treadmill training.

Proliferation of Schwann cells in the injured peripheral nerve supports axonal regeneration, and physical training in experimental animals has been shown to promote nerve regeneration. Extracellular signal-regulated kinase 1/2 (ERK1/2) activity can mediate neuronal responses to lesion signals, but its role in non-neuronal cells in the injured area is largely unknown. Here we report that treadmill training (TMT) facilitates axonal regeneration via the upregulation of phospho-ERK1/2 protein levels in Schwann cells in the injured sciatic nerve. Low-intensity, but not high-intensity, TMT increased neurite outgrowth of dorsal root ganglion (DRG) sensory neurons and potentiated Schwann cell proliferation. TMT elevated levels of GAP-43 mRNA and protein, and phospho-ERK1/2 protein in the injured sciatic nerves. TMT also enhanced phospho-c-Jun protein levels in the injured nerve. In-vivo administration of the ERK1/2 inhibitor PD98059 eliminated phospho-c-Jun, suggesting ERK1/2 phosphorylation of the c-Jun protein. PD98059 treatment decreased levels of BrdU-labeled proliferating Schwann cells in the distal portion of the injured nerve, and delayed the axonal regrowth that was promoted by TMT. The present data suggest that increased ERK1/2 activity in Schwann cells may play an important role in TMT-mediated enhancement of axonal regeneration in the injured peripheral nerve.

Neurochemical characterization of neuronal populations expressing protein kinase C gamma isoform in the spinal cord and gracile nucleus of the rat.

Protein kinase C gamma (PKCgamma) is widely distributed throughout the CNS and is thought to play a role in long term hyper-excitability in nociceptive neurones. Here, we provide the first report of PKCgamma cells in the dorsal column nuclei of the adult rat. Retrograde labeling of PKCgamma cells from the thalamus with choleragenoid revealed that 25% of the PKCgamma positive gracile cells projected to the thalamus. Further, we have characterized the distribution of PKCgamma within gracile nucleus in terms of colocalization with various neurotransmitter receptors or enzymes and calcium binding proteins, and compared this with PKCgamma colocalization in cells of laminae I-III of the spinal cord. We show that approximately 90% of the PKCgamma cells in the gracile nucleus and 60% in the dorsal horn were immuno-positive for the AMPA receptor subunit glutamate 2/3 (GluR2/3). Little coexpression was seen with neurokinin 1 receptor, nitric oxide synthase (NOS) and the AMPA receptor subunit GluR1, markers of distinct neuronal subpopulations. In the spinal cord, a quarter of PKCgamma cells expressed calbindin, but very few cells did so in the gracile nucleus. Electrical stimulation at c-fiber strength of the normal or injured sciatic nerve was used to induce c-fos as a marker of postsynaptic activation in the spinal cord and gracile nucleus. Quantitative analysis of the number of PKCgamma positive gracile cells that expressed also c-fos increased from none to 24% after injury, indicating an alteration in the sensory activation pattern in these neurones after injury. C-fos was not induced in inner lamina II following c-fiber electrical stimulation of the intact or axotomized sciatic nerve, indicating no such plasticity at the spinal cord level. As dorsal column nuclei cells may contribute to allodynia after peripheral nerve injury, pharmacological modulation of PKCgamma activity may therefore be a possible way to ameliorate neuropathic pain after peripheral nerve injury.

Involvement of subtype 1 metabotropic glutamate receptors in apoptosis and caspase-7 over-expression in spinal cord of neuropathic rats.

The effect of the non-selective, 1-aminoindan-1,5-dicarboxylic acid (AIDA), and selective (3,4-dihydro-2H-pyrano[2,3-b]quinolin-7-yl)-(cis-4-methoxycyclohexyl) methanone (JNJ16259685), metabotropic glutamate subtype 1 (mGlu1) receptor antagonists, on rat sciatic nerve chronic constrictive injury (CCI)-induced hyperalgesia, allodynia, spinal dorsal horn apoptosis, and gliosis was examined at 3 and 7 days post-injury. RT-PCR analysis showed increased expression of bax, apoptotic protease-activating factor-1 (apaf-1), nestin, GFAP, and caspase-7 mRNA in the dorsal horn spinal cord by 3 days post-CCI. At 7 days post-CCI, only over-expression of bcl-2, nestin and GFAP mRNA was observed. Administration of AIDA reduced thermal hyperalgesia and mechanical allodynia at 3 and 7 days post-CCI; administration of JNJ16259685 reduced thermal hyperalgesia at 3 and 7 days post-CCI, but not mechanical allodynia. AIDA decreased the mRNA levels of bax, apaf-1, GFAP and caspase-7 genes. JNJ16259685 increased the mRNA levels of bcl-2 and GFAP gene, and decreased APAF-1 and caspases-7 genes. Inhibiting mGlu1 receptors also reduced TUNEL-positive profiles and immunohistochemical reactivity for caspase-7. We report here that despite inhibiting CCI-induced over-expression of pro-apoptotic genes in the spinal cord dorsal horn, the selective mGlu1 receptor antagonist JNJ16259685 exerted only a slight and transient allodynic effect. Moreover, JNJ16259685, but not the non-selective AIDA, increased astrogliosis which may account for its decreased analgesic efficacy. This study provides evidence that the contemporary and partial blockade of group I and likely ionotropic glutamate receptors may be a more suitable therapy than selective blockade of mGlu1 subtype receptors condition to decrease neuropathic pain symptoms.

Up-regulation of interleukin-6 induced by prostaglandin E from invading macrophages following nerve injury: an in vivo and in vitro study.

The mechanisms underlying neuropathic pain caused by nerve injury are not well understood. Inflammatory responses in injured nerves are likely to be key contributing factors in the generation and maintenance of neuropathic pain. The pro-inflammatory cytokine interleukin-6 (IL-6) is up-regulated in invading macrophages and has been implicated in the development of neuropathic pain. We previously demonstrated that invading macrophages up-regulate cyclooxygenase 2 (COX2) and prostaglandin E2 (PGE2) receptors EP1 and EP4, suggesting that PGE2 may affect macrophage function via autocrine or paracrine mechanisms. This study was undertaken to determine whether PGE2 is involved in the up-regulation of IL-6 in invading macrophages. Two weeks following partial sciatic nerve ligation, numerous IL-6 immunoreactive (IR) cell profiles were present in injured nerves. Colocalization of IL-6 with the invading macrophage marker ED1 or with COX2 was frequently observed. IL-6-IR, COX2-IR and ED1-IR cells were present only in cultures derived from injured nerve segments. PGE2 and IL-6 release from cultured cells derived from injured nerves was increased significantly compared with uninjured nerves. Non-selective and selective COX2 inhibitors suppressed PGE2 and IL-6 release. Treatment with PGE2 further enhanced IL-6 release in a concentration- and time-dependent manner. A selective EP4 receptor antagonist L-161982 was able to suppress IL-6 release, whereas an EP1 receptor antagonist, SC19220, was ineffective. Moreover, a protein kinase C inhibitor, calphostin C, dramatically suppressed IL-6 release, whereas a protein kinase A inhibitor H-89 and a Ca2+ chelator EGTA failed. Taken together, our data suggest that PGE2 is involved in mediating the up-regulation of IL-6 occurring in invading macrophages. This action is mediated through an EP4 receptor and the protein kinase C signaling pathway.

Differential involvement of spinal protein kinase C and protein kinase A in neuropathic and inflammatory pain in mice.

In the present study, we demonstrated the differential role of spinal protein kinases in neuropathic and inflammatory pain. Mice with sciatic nerve ligation exhibited a spinal protein kinase C (PKC)-dependent neuropathic pain-like state. In contrast, an intraplanter injection of inflammatory agent caused a protein kinase A (PKA)-related thermal hyperalgesia. These findings suggest that the substantial activation of spinal PKC and PKA may differentially contribute to the development of respective chronic pain-like state in mice.

Effect of 17 beta-estradiol on gene expression in lumbar spinal cord following sciatic nerve crush injury in ovariectomized mice.

Previously, we observed that estrogen treatment enhances regeneration of the sciatic nerve after crush injury [Brain Res. 943 (2002) 283]. In this research, we studied expression of estrogen receptors and effects of estrogen on gene expression in the lumbar spinal cord, following sciatic nerve crush injury. Using the Atlas Mouse 1.2 Array, changes in the expression of 267 of 1176 genes were registered 4 days after nerve injury. Those genes that exhibited a change in signal intensity ratios of 2-fold or greater were selected as up-regulated (42) or down-regulated (21). In estrogen treated mice, we have observed up-regulation of the genes known to control apoptosis, cell proliferation, and growth, which might account for the positive effects of estrogen on the regeneration of motor neurons. Immunohistochemical staining revealed estrogen receptor-alpha and estrogen receptor-beta localized in the nucleus and cytoplasm of lumbar motor neurons, and in the regenerating neurites of the sciatic nerve. Expression of estrogen receptor-alpha and estrogen receptor-beta mRNA in lumbar spinal cord was shown by traditional RT-PCR. Using real-time quantitative RT-PCR, we demonstrated increased expression of estrogen receptors-alpha and -beta mRNA on the injured side of the lumbar spinal cord. Western blot analysis showed the accumulation of ERs in regenerating sciatic nerve, and revealed a 40% increase of activated ERK1/2 in estrogen treated mice, compared to placebo. Our findings indicate that: (i). axotomized motor neurons increase expression of estrogen receptors-alpha and -beta mRNA, (ii). estrogen mediates the expression of genes which accelerate the growth and maturation of axons, and (iii). estrogen receptors are transported from the perikaryon into regenerating neurites, and estrogen promotes regeneration locally through the non-genomic ERK-activated signaling pathway.

Axonal transport of puromycin-sensitive aminopeptidase in rat sciatic nerves.

Axonal transport of puromycin-sensitive aminopeptidase (PSA), a putative neuropeptide degrading-enzyme which removes amino acid residues from the amino-terminal of neuropeptides, was examined in the proximal, middle, and distal segments of rat sciatic nerves using a double-ligation technique. The soluble fraction of each segment was partially purified by MonoQ column chromatography, and showed two peaks of aminopeptidase activity. One of the aminopeptidases was PSA. At 48 h after the ligations, a significant amount of the axonal transport of PSA activity was found in the proximal segment. Western blot analysis of the segments also showed that immunoreactive PSA in the proximal segment was 2.1-fold higher than that in the middle segment. Furthermore, the immunohistochemical analysis of the segments showed an increase of the immunoreactive PSA in the proximal segment in comparison with the enzyme in the distal segment, indicating that PSA is mainly transported by anterograde axonal flow. These results suggest that PSA plays a role in the metabolism of neuropeptides in nerve terminals or synaptic clefts.

Upregulation and interaction of TNFalpha and gelatinases A and B in painful peripheral nerve injury.

Chronic constriction injury (CCI) to peripheral nerve causes a painful neuropathy in association with a process of axonal degeneration and endoneural remodeling that involves macrophage recruitment and local increase in extracellular proteases and tumor necrosis factor alpha (TNF-alpha). Cell surface activation of TNF-alpha from its transmembrane precursor, as well as sequestration of TNF-alpha receptors II and I, is performed by the zinc-dependent endopeptidase family of matrix metalloproteinases (MMPs). Among TNF-alpha-converting MMPs, basal lamina degrading gelatinases are thought to play a role in sciatic nerve injury. In the present study, we determined the forms of TNF-alpha involved in the development of CCI neuropathy in rats, using Western blot analysis, and the temporal correlation of TNF-alpha and TNFRI protein profiles with gelatinases activity at the site of peripheral nerve injury. We observed two peaks in TNF-alpha protein during the first week of CCI that correspond to previously reported peaks in painful behavior. We propose that the first peak at 6 h post-CCI is due to the local expression of the cytotoxic transmembrane 26 kDa TNF-alpha protein released by resident Schwann cells, mast cells and macrophages. This peak in TNF-alpha protein expression corresponds to an increase in gelatinase B (MMP-9) activity, which is greatly upregulated as early as 3 h following CCI to rat sciatic nerve. The second peak occurs at 5 days post-CCI, and may represent TNF-alpha protein released by hematogenously recruited macrophages. This peak is marked by the increase in active soluble 17 kDa TNF-alpha and by gelatinase A (MMP-2) upregulation. These observations suggest that there is a pathogenic role for the TNF-alpha-converting function of MMP-2 in painful CCI neuropathy. We conclude that severe nerve injury induces MMPs, TNF-alpha and TNFRI, which interactively control the privileged endoneurial environment and the pathogenesis of the painful neuropathies associated with the macrophage-dependent processes of Wallerian degeneration.