Inflammation-induced increase in nicotinic acetylcholine receptor current in cutaneous nociceptive DRG neurons from the adult rat.

The goals of the present study were to determine (1) the properties of the nicotinic acetylcholine receptor (nAChR) currents in rat cutaneous dorsal root ganglion (DRG) neurons; (2) the impact of nAChR activation on the excitability of cutaneous DRG neurons; and (3) the impact of inflammation on the density and distribution of nAChR currents among cutaneous DRG neurons. Whole-cell patch-clamp techniques were used to study retrogradely labeled DRG neurons from naïve and complete Freund's adjuvant inflamed rats. Nicotine-evoked currents were detectable in ∼70% of the cutaneous DRG neurons, where only one of two current types, fast or slow currents based on rates of activation and inactivation, was present in each neuron. The biophysical and pharmacological properties of the fast current were consistent with nAChRs containing an α7 subunit while those of the slow current were consistent with nAChRs containing α3/ß4 subunits. The majority of small diameter neurons with fast current were IB4- while the majority of small diameter neurons with slow current were IB4+. Preincubation with nicotine (1 µM) produced a transient (1 min) depolarization and increase in the excitability of neurons with fast current and a decrease in the amplitude of capsaicin-evoked current in neurons with slow current. Inflammation increased the current density of both slow and fast currents in small diameter neurons and increased the percentage of neurons with the fast current. With the relatively selective distribution of nAChR currents in putative nociceptive cutaneous DRG neurons, our results suggest that the role of these receptors in inflammatory hyperalgesia is likely to be complex and dependent on the concentration and timing of acetylcholine release in the periphery.

TRAF family member-associated NF-kappa B activator (TANK) expression increases in injured sensory neurons and is transcriptionally regulated by Sox11.

Peripheral nerve injury evokes rapid and complex changes in gene transcription and cellular signaling pathways. Understanding how these changes are functionally related is essential for developing new approaches that accelerate and improve nerve regeneration. Toward this goal we found that nerve injury induces a rapid and significant up-regulation of the transcription factor Sox11 in dorsal root ganglia (DRG) neurons. Gain and loss of function studies have shown this increase is essential for normal axon regeneration. To determine how Sox11 impacts neuronal gene expression, DRG neurons were treated with Sox11 siRNA to identify potential transcriptional targets. One gene significantly reduced by Sox11 knockdown was TRAF (tumor necrosis factor (TNF) receptor-associated factor)-associated NF-κB activator (TANK). Here we show that TANK is expressed in DRG neurons, that TANK expression is increased in response to peripheral nerve injury and that Sox11 overexpression in vitro increases TANK expression. Injury and in vitro overexpression were also found to preferentially increase TANK transcript variant 3 and a larger TANK protein isoform. To determine if Sox11 regulates TANK transcription bioinformatic analysis was used to identify potential Sox-binding motifs within 5kbp of the TANK 5' untranslated region (UTR) across several mammalian genomes. Two sites in the mouse TANK gene were examined. Luciferase expression assays coupled with site-directed mutagenesis showed each site contributes to enhanced TANK promoter activity. In addition, chromatin immunoprecipitation assays showed direct Sox11 binding in regions containing the two identified Sox motifs in the mouse TANK 5'-UTR. These studies are the first to show that TANK is expressed in DRG neurons, that TANK is increased by peripheral nerve injury and that the regulation of TANK expression is, at least in part, controlled by the injury-associated transcription factor Sox11.

Mitochondrial fission proteins in peripheral blood lymphocytes are potential biomarkers for Alzheimer's disease.

BACKGROUND AND PURPOSE: Expression of the mitochondrial fission proteins dynamin-related protein 1 (Drp1), S-nitrosylated Drp1 (SNO-Drp1), and Fis1 has been found to be altered in brain tissues and skin fibroblasts from patients with Alzheimer's disease (AD). The aim of this study was to determine whether these proteins are also changed in peripheral blood lymphocytes (PBL) of AD patients and whether these changes are specific and sensitive enough for AD diagnosis. METHODS: Western blot analysis and enzyme-linked immunosorbent assay (ELISA) were employed to quantify relative levels of Drp1, SNO-Drp1, and Fis1 in PBL obtained from 91 controls, 82 AD, 26 mild cognitive impairment (MCI), 12 Parkinson's disease (PD), and 36 vascular dementia (VaD) patients. Logistic regression and receiver operating characteristic (ROC) curve analysis were used to measure diagnostic accuracy of these proteins. RESULTS: Compared with controls, SNO-Drp1 and Fis1 levels were remarkably increased in PBL of AD and MCI patients, and Drp1 was significantly decreased in AD, MCI, and PD. None of these proteins were changed in VaD patients. Disease severity or duration had no major effects on levels of these proteins in AD PBL. ROC curve analysis showed that the specificity and sensitivity were 81% and 73% for Drp1, 84% and 82% for SNO-Drp1, and 89% and 80% for Fis1 in identifying AD patients from control subjects. CONCLUSIONS: Altered mitochondrial fission proteins Drp1, SNO-Drp1, and Fis1 in PBL were relatively sensitive and specific in identifying AD patients and could be serving as a biomarker in the procedure of diagnosis.

Modulation of visceral hypersensitivity by glial cell line-derived neurotrophic factor family receptor α-3 in colorectal afferents.

Irritable bowel syndrome is characterized by colorectal hypersensitivity and contributed to by sensitized mechanosensitive primary afferents and recruitment of mechanoinsensitive (silent) afferents. Neurotrophic factors are well known to orchestrate dynamic changes in the properties of sensory neurons. Although pain modulation by proteins in the glial cell line-derived neurotrophic factor (GDNF) family has been documented in various pathophysiological states, their role in colorectal hypersensitivity remains unexplored. Therefore, we investigated the involvement of the GDNF family receptor α-3 (GFRα3) signaling in visceral hypersensitivity by quantifying visceromotor responses (VMR) to colorectal distension before and after intracolonic treatment with 2,4,6-trinitrobenzene sulfonic acid (TNBS). Baseline responses to colorectal distension did not differ between C57BL/6 and GFRα3 knockout (KO) mice. Relative to intracolonic saline treatment, TNBS significantly enhanced the VMR to colorectal distension in C57BL/6 mice 2, 7, 10, and 14 days posttreatment, whereas TNBS-induced visceral hypersensitivity was significantly suppressed in GFRα3 KO mice. The proportion of GFRα3 immunopositive thoracolumbar and lumbosacral colorectal dorsal root ganglion neurons was significantly elevated 2 days after TNBS treatment. In single fiber recordings, responses to circumferential stretch of colorectal afferent endings in C57BL/6 mice were significantly increased (sensitized) after exposure to an inflammatory soup, whereas responses to stretch did not sensitize in GFRα3 KO mice. These findings suggest that enhanced GFRα3 signaling in visceral afferents may contribute to development of colorectal hypersensitivity.

SRY-box containing gene 11 (Sox11) transcription factor is required for neuron survival and neurite growth.

The transcription factor Sox11 is expressed at high levels in developing sensory neurons and injured adult neurons but little is known about its transcriptional targets and function. In this study we examined the role of Sox11 using Neuro2a neuroblastoma cells and cultured mouse dorsal root ganglia (DRG) neurons. Results show Sox11 has an essential role in regulation of neuron survival and neurite outgrowth in Neuro2a cells and primary sensory neurons. Neuro2a cells increase expression of Sox11 as they differentiate in culture. Following addition of 20 microM retinoic acid (RA), a stimulus for differentiation that enhances neurite growth and differentiation, Sox11 level rises. RNAi-mediated knockdown of Sox11 in RA-differentiated Neuro2a cells caused a decrease in neurite growth and an increase in the percent of apoptotic cells. RNA expression analysis showed that Sox11 knockdown modulated the level of mRNAs encoding several genes related to cell survival and death. Further validation in the Neuro2a model showed Sox11 knockdown increased expression of the pro-apoptotic gene BNIP3 (BclII interacting protein 1 NIP3) and decreased expression of the anti-apoptotic gene TANK (TNF receptor-associated factor family member-associated NFkappaB activator). Cultured primary DRG neurons also express Sox11 and treatment with Sox11 small interfering RNA (siRNA) caused a significant decrease in neurite growth and branching and a decrease in mRNA encoding actin-related protein complex 3 (Arpc3), an actin organizing protein that may be involved in axon growth. The percent of apoptotic neurons also increased in cultures of DRG neurons treated with Sox11 siRNA. Similar to Neuro2a cells, a decrease in TANK gene expression occurred, suggesting at least some overlap in Sox11 transcriptional targets in Neuro2a and DRG neurons. These data are consistent with a central role for Sox11 in regulating events that promote neurite growth and neuron survival.

Skin-derived nerve growth factor blocks programmed cell death in the trigeminal ganglia but does not enhance neuron proliferation.

Development of the cutaneous sensory nervous system is dependent on the production of neurotrophic factors, such as nerve growth factor (NGF), by the skin. Limited synthesis of NGF in developing skin is thought to underlie programmed cell death and cause a 50% neuronal loss. This loss does not occur in transgenic mice that overexpress NGF in the skin, which have double the number of neurons (J. Neurosci. 14 (1994) 1422). To determine whether increased NGF blocks neuronal death and/or increases neuronal precursor replication, we analyzed the trigeminal ganglia at embryonic days E12.5, E14.5 and E16.5 using transferase-mediated dUTP nick-end labeling (TUNEL) and bromodeoxyuridine labeling. Results show that excess target-derived NGF causes a major decrease in the percent of TUNEL-labeled neurons without affecting the percent of replicating neurons. Analysis of RNA and protein expression suggests this block in cell death is mediated via the anti-apoptotic protein bcl-2.

Sympathetic innervation of lymphoreticular organs is rate limiting for prion neuroinvasion.

Transmissible spongiform encephalopathies are commonly propagated by extracerebral inoculation of the infectious agent. Indirect evidence suggests that entry into the central nervous system occurs via the peripheral nervous system. Here we have investigated the role of the sympathetic nervous system in prion neuroinvasion. Following intraperitoneal prion inoculation, chemical or immunological sympathectomy delayed or prevented scrapie. Prion titers in spinal cords were drastically reduced at early time points after inoculation. Instead, keratin 14-NGF transgenic mice, whose lymphoid organs are hyperinnervated by sympathetic nerves, showed reduction in scrapie incubation time and, unexpectedly, much higher titers of prion infectivity in spleens. We conclude that sympathetic innervation of lymphoid organs is rate limiting for prion neuroinvasion and that splenic sympathetic nerves may act as extracerebral prion reservoirs.

Epithelial overexpression of BDNF or NT4 disrupts targeting of taste neurons that innervate the anterior tongue.

Brain-derived neurotrophic factor (BDNF) and neurotrophin-4 (NT4) are essential for the survival of geniculate ganglion neurons, which provide the sensory afferents for taste buds of the anterior tongue and palate. To determine how these target-derived growth factors regulate gustatory development, the taste system was examined in transgenic mice that overexpress BDNF (BDNF-OE) or NT4 (NT4-OE) in basal epithelial cells of the tongue. Overexpression of BDNF or NT4 caused a 93 and 140% increase, respectively, in the number of geniculate ganglion neurons. Surprisingly, both transgenic lines had severe reduction in fungiform papillae and taste bud number, primarily in the dorsal midregion and ventral tip of the tongue. No alterations were observed in taste buds of circumvallate or incisal papillae. Fungiform papillae were initially present on tongues of newborn BDNF-OE animals, but many were small, poorly innervated, and lost postnatally. To explain the loss of nerve innervation to fungiform papillae, the facial nerve of developing animals was labeled with the lipophilic tracer DiI. In contrast to control mice, in which taste neurons innervated only fungiform papillae, taste neurons in BDNF-OE and NT4-OE mice innervated few fungiform papillae. Instead, some fibers approached but did not penetrate the epithelium and aberrant innervation to filiform papillae was observed. In addition, some papillae that formed in transgenic mice had two taste buds (instead of one) and were frequently arranged in clusters of two or three papillae. These results indicate that target-derived BDNF and NT4 are not only survival factors for geniculate ganglion neurons, but also have important roles in regulating the development and spatial patterning of fungiform papilla and targeting of taste neurons to these sensory structures.

Effect of peripheral nerve lesion and lumbar sympathectomy on peptide regulation in dorsal root ganglia in the NGF-overexpressing mouse.

Galanin is a peptide normally expressed at low levels both in sensory and in sympathetic neurons. It is strongly upregulated after peripheral nerve lesions, and it has been proposed that nerve growth factor (NGF) plays a role in this regulation. In the present study the effect of both sciatic nerve transection and lumbar sympathectomy on galanin in lumbar dorsal root ganglia (DRGs) was examined in mice overexpressing NGF (NGFOE) in the skin under the keratin promoter. The superior cervical ganglia (SCG) were also studied. In the DRG pericellular baskets containing tyrosine hydroxylase- (TH) and galanin-like immunoreactivity (LI) were found, mostly in the same fibers. Galanin-positive baskets were also found in the trigeminal ganglia. However, only single neuropeptide Y (NPY)-positive baskets were observed within the DRGs. No marked difference in number of galanin-positive neurons was seen between wild-type and NGFOE mice. After sciatic nerve transection galanin was upregulated in DRG neurons to about the same extent in NGFOE mice as in wild-type mice. Galanin-, but not TH-LIs decreased in the pericellular baskets. After lumbar sympathectomy both galanin- and TH-immunoreactive baskets disappeared, suggesting a sympathetic origin. In the SCG the very low galanin mRNA levels were strongly increased after lesion of the carotid nerves, both in wild-type and in NGFOE mice. However, whereas NPY mRNA levels decreased in the SCG after axotomy in the wild-type mice, there was a distinct increase in the NGFOE mice. Our results show that high NGF levels in skin induce formation of pericellular baskets in DRGs expressing galanin- and TH-LI and that galanin in these baskets is strongly influenced by peripheral axotomy. However, overexpression of NGF did not markedly influence galanin expression in DRG neurons, neither normally nor after nerve lesions. Finally, expression of NPY in sympathetic ganglia is differently regulated in NGFOE compared to wild-type mice.

A role for p75 neurotrophin receptor in the control of apoptosis-driven hair follicle regression.

To examine the mechanisms that underlie the neurotrophin-induced, apoptosis-driven hair follicle involution (catagen), the expression and function of p75 neurotrophin receptor (p75NTR), which is implicated in apoptosis control, were studied during spontaneous catagen development in murine skin. By RT-PCR, high steady-state p75NTR mRNA skin levels were found during the anagen-catagen transition of the hair follicle. By immunohistochemistry, p75NTR alone was strongly expressed in TUNEL+/Bcl2- keratinocytes of the regressing outer root sheath, but both p75NTR and TrkB and/or TrkC were expressed by the nonregressing TUNEL-/Bcl2+ secondary hair germ keratinocytes. To determine whether p75NTR is functionally involved in catagen control, spontaneous catagen development was compared in vivo between p75NTR knockout (-/-) and wild-type mice. There was significant catagen retardation in p75NTR knockout mice as compared to wild-type controls (P<0.05). Instead, transgenic mice-overexpressing NGF (promoter: K14) showed substantial acceleration of catagen (P<0.001). Although NGF, brain-derived neurotrophic factor (BDNF), and neurotrophin 3 (NT-3) accelerated catagen in the organ-cultured skin of C57BL/6 mice, these neurotrophins failed to promote catagen development in the organ-cultured p75NTR null skin. These findings suggest that p75NTR signaling is involved in the control of kerotinocyte apoptosis during catagen and that pharmacological manipulation of p75NTR signaling may prove useful for the treatment of hair disorders that display premature entry into catagen.

Levels of nerve growth factor and neurotrophin-3 are affected differentially by the presence of p75 in sympathetic neurons in vivo.

The development and survival of sympathetic neurons is critically dependent on the related neurotrophic factors nerve growth factor (NGF) and neurotrophin-3 (NT3), the actions of which must be executed appropriately despite spatial and temporal overlaps in their activities. The tyrosine receptor kinases, trkA and trkC, are the cognate receptors for NGF and NT3, respectively. The p75 neurotrophin receptor has been implicated in neurotrophin binding and signaling for both NGF and NT3. In this study, the authors used mice that overexpressed NGF (NGF-OE) or NT3 (NT3-OE) in skin and mice that lacked p75 (p75(-/-)) to understand the dynamics of sympathetic neuron response to each neurotrophin and to address the role of p75. NGF and NT3 were measured in sympathetic ganglia and skin (a major target of sympathetic neurons) by using the enzyme-linked immunosorbent assay (ELISA) technique. A three- to four-fold increase in skin NT3 was seen in both NT3-OE and p75(-/-) mice. Moreover, both mouse lines exhibited a three-fold increase in ganglionic NT3. However, the increase in ganglionic NT3 was accompanied by a decrease in ganglionic NGF in p75(-/-) mice but not in NT3-OE mice. This indicated that p75 plays an important role in determining the level of NGF within sympathetic neurons. In NGF-OE mice, the overexpression of NGF was correlated with increased ganglionic NGF and increased ganglionic expression of p75 mRNA. In addition, in NGF-OE mice, ganglionic trkC expression was decreased, as was the amount of NT3 present within sympathetic ganglia. These results indicate that the level of p75 is integral in determining the level of sympathetic NGF and that NGF competes with NT3 by increasing the expression of p75 and decreasing the expression of trkC.

Cutaneous overexpression of neurotrophin-3 (NT3) selectively restores sensory innervation in NT3 gene knockout mice.

Neurotrophin-3 (NT3) is essential for development of sensory innervation to the skin. NT3 supports the postnatal survival of primary sensory neurons that mediate mechanoreception and their Merkel cell containing touch dome end organs (Airaksinen et al., 1996). In this study we determined whether NT3 overexpressed in the skin could restore innervation lost when endogenous NT3 levels were reduced. Hybrid mice that overexpress NT3 in basal keratinocytes but lack one endogenous NT3 allele (K14-NT3/NT3(+/-)) were compared to NT3 overexpresser (K14-NT3) mice, heterozygous knockout (NT3(+/-)) mice, and littermate control mice. In line with previous analyses, NT3(+/-) mice lost 63% of the Merkel cells associated with touch domes, 67% of touch dome units and the associated SAI innervation. All of these parameters were restored to overexpresser levels in K14-NT3/NT3(+/-) mice. Knockout NT3(+/-) mice also had a 31% reduction of L4/L5 dorsal root ganglion cells and a 24% reduction of myelinated axons in the saphenous cutaneous nerve. These losses were also restored in hybrid K14-NT3/NT3(+/-) mice, though only to control mouse values. These results indicate that overexpression of NT3 in skin of NT3(+/-) knockout mice rescued most cutaneous neurons lost in NT3(+/-) mice, but was unable to rescue NT3-dependent neurons that project to noncutaneous sensory targets.

Distinct roles for nerve growth factor and brain-derived neurotrophic factor in controlling the rate of hair follicle morphogenesis.

Increasing evidence suggests that neurotrophins play an important part in the control of the development of ectodermal derivatives, such as the hair follicle. Here, we show that, during hair follicle morphogenesis in C57BL/6 mice, nerve growth factor, brain-derived neurotrophic factor and their corresponding high-affinity tyrosine kinase receptors, TrkA and TrkB, show stringently controlled spatiotemporal expression patterns in the follicular epithelium and mesenchyme. Constitutive overexpression of nerve growth factor in mice is associated with a discrete, but significant acceleration of hair follicle morphogenesis, whereas this is not seen in brain-derived neurotrophic factor transgenic mice. In neonatal skin organ culture, nerve growth factor and brain-derived neurotrophic factor differentially influence hair follicle development: nerve growth factor accelerates late stages of hair follicle morphogenesis, whereas brain-derived neurotrophic factor does not show significant effects. This suggests that the morphogenetic properties of locally generated neurotrophins in the skin, similar to their classical neurotrophic functions, are quite distinct and depend on the response patterns of the corresponding neurotrophin target receptor-expressing cells in the developing hair follicle. These data further strengthen the concept that neurotrophin signaling is an important element in controlling the rate of hair follicle morphogenesis, yet also highlight the complexity of this signaling system.

Overexpression of nerve growth factor in skin selectively affects the survival and functional properties of nociceptors.

Mice that overexpress nerve growth factor (NGF-OE) in the skin have double the normal number of cutaneous sensory neurons, have increased innervation of the skin and spinal cord, and are hyperalgesic. Here, we have asked whether the increased cutaneous NGF level results in a selective survival of only certain functional types of neurons and whether it changes the properties of cutaneous neurons. Using electron microscopy, we show that the number of both myelinated and unmyelinated nociceptors increases substantially in NGF-OE mice by a factor of 3.3 and 1.5, respectively. Using extracellular recordings from single units, we demonstrate that large myelinated (Abeta) fibers are unchanged in prevalence and receptive properties. In contrast, among thin myelinated (Adelta) fibers, the percentage of nociceptors increased from a normal 65 to 97%, consistent with a selective survival of nociceptors during embryogenesis. These afferents showed a twofold increase in their mechanical responsiveness, but their heat responsiveness remained normal. Among unmyelinated (C) fibers, there was a profound increase in the percentage of heat responsive neurons from a normal 42 to 96%. This change cannot be accounted for by a selective survival of heat-sensitive neurons. Unmyelinated nociceptors increased fourfold in their thermal responsiveness but decreased in mechanical responsiveness. Therefore, target-derived NGF selectively rescues nociceptors during the period of programmed cell death with different efficacy for thin myelinated or unmyelinated fibers. NGF also affects the response to noxious heat or mechanical stimuli in each group differently, implying specific regulations of transduction processes rather than general changes of excitability.

Sodium channel expression in NGF-overexpressing transgenic mice.

Dorsal root ganglion (DRG) neurons depend on nerve growth factor (NGF) for survival during development, and for the maintenance of phenotypic expression of neuropeptides in the adult. NGF also plays a role in the regulation of expression of functional sodium channels in both PC12 cells and DRG neurons. Transgenic mice that overexpress NGF under the keratin promoter (hyper-NGF mice) show increased levels of NGF in the skin from embryonic day 11 through adulthood, hypertrophy of the peripheral nervous system and mechanical hyperalgesia. We show here that mRNA levels for specific sodium channel isotypes are greater in small (< 30 microm diameter) DRG neurons from hyper-NGF mice compared to wild-type mice. Hybridization signals for sodium channel subunits alphaII and beta2 displayed the most substantial enhancement in hyper-NGF mice, compared to wild-type mice DRG, and mRNA levels for alphaI, NaG, Na6, SNS/PN3, NaN, and beta1 were also greater in hyper-NGF DRG. In contrast, the levels of alphaII and PN1 mRNAs were similar in neurons from hyper-NGF and wild-type DRG. Whole-cell patch-clamp studies showed no significant differences in the peak sodium current densities in hyper-NGF vs. wild-type DRG neurons. These data demonstrate that DRG neurons in wild-type mice have a heterogeneous pattern of sodium channel expression, which is similar to that previously described in rat, and suggest that transcripts of some, but not all, sodium channel mRNAs can be modulated by long-term overexpression of NGF.

Overexpression of brain-derived neurotrophic factor enhances sensory innervation and selectively increases neuron number.

Target-derived neurotrophin growth factors have significant effects on the development and maintenance of the mammalian somatosensory system. Studies of transgenic mice that overexpress neurotrophins NGF and neurotrophin 3 (NT-3) at high levels in skin have shown increased sensory neuron number and enhanced innervation of specific sensory ending types. The effects of two other members of this family, BDNF and NT-4, on sensory neuron development are less clear. This study examined the role of brain-derived neurotrophic factor (BDNF) using transgenic mice that overexpress BDNF in epithelial target tissues of sensory neurons. BDNF transgenic mice had an increase in peripheral innervation density and showed selective effects on neuron survival. Neuron number in trigeminal ganglia, DRG, and SCG were unchanged, although a 38% increase in neurons comprising the placode-derived nodose-petrosal complex occurred. BDNF transgenic skin showed notable enhancement of innervation to hair follicles as detected by PGP9.5 immunolabeling. In nonhairy plantar skin, Meissner corpuscle sensory endings were larger, and the number of Merkel cells with associated innervation was increased. In trigeminal ganglia, neurons expressing trkB receptor were increased threefold, whereas trkA-positive neurons doubled. Analysis of trkB by Northern, reverse transcription-PCR, and Western assays indicated a modest increase in the expression of the T1 truncated receptor and preferential distribution to the periphery. These data indicate that skin-derived BDNF does not enhance survival of cutaneous sensory neurons, although it does promote neurite innervation of specific sites and sensory end organs of the skin.

Neurotrophins, nociceptors, and pain.

It is now well established that neurotrophins play a crucial role in the development of the nervous system. However, there is increasing evidence that the function of neurotrophins persists throughout adulthood. The broad scope of neurotrophin action is well documented in the case of nerve growth factor (NGF) and its effect on nociceptors and nociception. Here, we review the evidence for these multiple roles for NGF. Two manipulations influencing NGF levels are discussed in detail. The first involves the use of transgenic mice that overexpress or underexpress neurotrophins. A second strategy involves administration of NGF or its antibody in vivo to increase or decrease its level. During prenatal development, NGF is required for survival of nociceptors. In the early postnatal period, NGF is required for expression of the appropriate nociceptor phenotype. In adults, NGF acts as an important intermediate in inflammatory pain, contributing to both peripheral and central sensitization. The sensitization of peripheral nociceptors can be very rapid and can involve non-neural cells such as mast cells, neutrophils, fibroblasts, and macrophages. Recent evidence indicates that other neurotrophins also play key supporting roles in the development of nociceptors (e.g., NT-3) and in inflammatory pain (e.g., BDNF, NT-4/5). Furthermore, molecules from other superfamilies (e.g., GDNF) also are required to assure survival of certain classes of nociceptors. The diverse effects of neurotrophins on nociceptive processing emphasize their broad importance in the development and function of the nervous system.

A new role for neurotrophin-3: involvement in the regulation of hair follicle regression (catagen).

Nervous system and hair follicle epithelium share a common ectodermal origin, and some neurotrophins (NTs) can modulate keratinocyte proliferation and apoptosis. Therefore, it is reasonable to ask whether NTs are also involved in hair growth control. Here, we show that the expression of NT-3 and its high-affinity receptor, tyrosine kinase C, in the skin of C57BL/6 mice is strikingly hair cycle-dependent, with maximal transcript and protein expression seen during spontaneous hair follicle regression (catagen). During catagen, NT-3 and tyrosine kinase C are co-expressed by terminal deoxynucleotidyl transferase-mediated in situ nick end labeling-positive keratinocytes in the club hair and secondary germ. NT-3-overexpressing transgenic mice show precocious catagen development during the postnatal initiation of hair follicle cycling, whereas heterozygous NT-3 knockout (+/-) mice display a significant catagen retardation. Finally, NT-3 stimulates catagen development in organ culture of normal C57BL/6 mouse skin. These observations suggest that the hair follicle is both a source and target of NT-3 and that NT-3/tyrosine kinase C signaling is functionally important in the control of hair follicle regression. Therefore, tyrosine kinase C agonists and antagonists deserve systematic exploration for the management of hair growth disorders that are related to premature (alopecia/effluvium) or retarded catagen (hirsutism/hypertrichosis).

Neurotrophin-3 involvement in the regulation of hair follicle morphogenesis.

Hair follicle epithelium and nervous system share a common ectodermal origin, and some neurotrophins can modulate keratinocyte proliferation and apoptosis. It is therefore reasonable to ask whether growth factors that control neural development are also involved in the regulation of hair follicle morphogenesis. Focusing on neurotrophin-3 (NT-3) and its high-affinity-receptor [tyrosine kinase C (TrkC)], we show that hair placode keratinocytes express TrkC mRNA and immunoreactivity early during murine hair follicle morphogenesis. In later stages of hair follicle development, TrkC mRNA, TrkC-, and NT-3-immunoreactivity are seen in keratinocytes of the proximal hair bulb as well as in dermal papilla fibroblasts. Compared with the corresponding wild-type animals, early stages of hair follicle morphogenesis are significantly accelerated in newborn NT-3 overexpressing mice, whereas these are retarded in newborn heterozygous NT-3 knockout (+/-) mice. These observations suggest that NT-3 is an important growth modulator during morphogenesis and remodeling of neuroectodermal-mesenchymal interaction systems like the hair follicle.