Nerve Growth Factor Receptor mRNA Research Articles

Resveratrol Induces Autophagy and Apoptosis in Non-Small-Cell Lung Cancer Cells by Activating the NGFR-AMPK-mTOR Pathway.

Resveratrol (RSV) has been reported to induce autophagy and apoptosis in non-small-cell lung cancer A549 cells, and the nerve growth factor receptor (NGFR) regulates autophagy and apoptosis in many other cells. However, the effect of NGFR on autophagy and apoptosis induced by RSV in A549 cells remains unclear. Here, we found that RSV reduced the cell survival rate in time- and concentration-dependent manners, activating autophagy and apoptosis. Lethal autophagy was triggered by RSV higher than 55 μM. The relationship between autophagy and apoptosis depended on the type of autophagy. Specifically, mutual promotion was observed between apoptosis and lethal autophagy. Conversely, cytoprotective autophagy facilitated apoptosis but was unaffected by apoptosis. RSV enhanced NGFR by increasing mRNA expression and prolonging the lifespan of NGFR mRNA and proteins. RSV antagonized the enhanced autophagy and apoptosis caused by NGFR knockdown. As the downstream pathway of NGFR, AMPK-mTOR played a positive role in RSV-induced autophagy and apoptosis. Overall, RSV-induced autophagy and apoptosis in A549 cells are regulated by the NGFR-AMPK-mTOR signaling pathway.

Effects of n-hexane on learning and memory and the expressions of nerve growth factor mRNA and nerve growth factor receptor mRNA of brain tissue in mice exposed to N-hexane

Objective: To Effects of n-hexane on learning and memory and the expressions of nerve growth factor (NGF) mRNA and nerve growth factor receptor (NGFR) mRNA of brain tissue in mice exposed to N-hexane. Methods: 40 Kunming mice were randomly divided into low-dose group, meddle-dose group, high-dose dose group and control group, with 10 mice in each group. All the groups were orally exposed to n-hexane in different doses: low-dose group with 43.5 mg/kg, middle-dose group with 86.5 mg/kg and high-dose group with 173.0 mg/kg, 1 time per day for 20 d. After the poisoning, the Y-arm test and the expressions of NGF mRNA and NGFR mRNA and the concentrations of NGF and NGFR in the brain tissues of each group were measured. Results: In the first Y-arm test, there existed a significant difference in correct reaction rate generally in all groups (P<0.05), and correct reaction rate in the middle-dose group and the high-dose group were low significantly compared with that in the control group(P<0.05). In the second Y-arm test, there existed a significant differences in total electric shock time and correct reaction rate generally in all groups (P<0.01), and the total electric shock time prolonged significantly and the correct response rate decreased significantly in 3 dose groups compared with those of the control group(P<0.05). The expression levels of NGF mRNA in brain tissues of low, meddle and high dose-groups were 0.81±0.66, 0.67±0.37 and 0.69±0.26, and the expression levels of NGFR mRNA were 1.22±0.42, 1.98±0.84 and 2.01±2.01, respectively. Compared with the control group, the expressions of NGF mRNA in the 3 dose groups decreased significantly (P<0.05), and the expression of NGFR mRNA in middle-and high-dose groups increased significantly (P<0.05). The concentrations of NGF in brain tissues of low,meddle and high dose-groups were 39.97±7.24 ng/L, 39.26±7.88 ng/L,31.70±8.21 ng/L,and the concentrations of NGFR were 17.37±6.82 ng/L,21.37±7.16 ng/L, 22.46±7.70 ng/L, respectively. Compared with the control group, the concentrations of NGF in high-dose groups decreased significantly(P<0.05), and the concentrations of NGFR in middle-and high-dose groups increased significantly (P<0.05). Conclusion: N-hexane exposure can result in decrease of learning and memory in mice, which may be related to abnormal expression of NGF mRNA and NGFR mRNA in brain tissue.

Effects of Inhaled Lavender Essential Oil on Stress-Loaded Animals: Changes in Anxiety-Related Behavior and Expression Levels of Selected mRNAs and Proteins

Inhalation of various essential oils elicits behavioral changes as a consequence of a complex centrally coordinated response. To understand the molecular mechanisms of action of aromatic compounds on emotional responses, we evaluated the stress-induced changes in mouse brain and the efficacy of inhaled essential oil from Lavandula officinalis (LvEO) using two approaches: a behavioral test, and examining the expression levels of selected genes {fast nerve growth factor receptor (NGFR) mRNA, activity regulated cytoskeletal-associated protein (Arc) mRNA} and proteins {galactokinase 1 (GLK1) and brain-derived neurotrophic factor (BDNF)}. Animals were randomly divided into 4 groups depending on the treatment given: stress (-)/H20, stress (-)/LvEO, stress (+)/H2O, and stress (+)/LvEO group. For behavioral testing, using an elevated plus-maze test, significant anxiolytic-like effects were seen in both the stress (-)/LVEO and stress (+)/LvEO groups, indicating that LvEO exerts anxiolytic-like effects regardless of the administration of water immersion stress. On expression analysis, the levels of NGFR and Arc mRNA were significantly lower in animals subjected to stress. Inhalation of LvEO, however, reversed this change, thus suggesting that LvEO negates the impact of stress on gene expression levels. Meanwhile, significant decreases in expression levels were also observed in the stress (-)/LvEO group, which implies that LvEO, when given in a stress-free situation, may act as a stress stimulus. Taken together, our data suggest that inhalation of LvEO exerts bidirectional influences in the central nervous system (CNS) of animals, either attenuating the effects of stress or acting as a stressor, depending on the subject state.

The Akt-Nitric Oxide-cGMP Pathway Contributes to Nerve Growth Factor-Mediated Neurite Outgrowth in Apolipoprotein E Knockout Mice

Apolipoprotein E (apo)-deficient [apoE(-/-)] mice have peripheral sensory nerve defects and a reduced and delayed response to noxious thermal stimuli. However, to date, no report has focused on the influence of apoE deficiency on calcitonin gene-related peptide (CGRP)-containing nerve fiber extensions. We have shown that the density of CGRP-containing nerve fibers decreases in mesenteric arteries of apoE(-/-) mice compared with wild-type mice. Here, we investigated whether apoE deficiency is involved in nerve growth factor (NGF)-induced CGRP-containing nerve regeneration using apoE(-/-) mice. NGF-mediated CGRP-like immunoreactivity (LI)-neurite outgrowth in apoE(-/-) cultured dorsal root ganglia (DRG) cells was significantly lower than that in wild-type cultures. However, the level of NGF receptor mRNA in apoE(-/-) DRG cells was similar to that in wild-type mice. To clarify the mechanism of the impaired ability of NGF-mediated neurite outgrowth, we focused on the Akt-nitric oxide (NO)-cGMP pathway. Expression of phosphorylated Akt was significantly reduced in apoE(-/-) DRG. The NO donor, sodium nitroprusside or S-nitroso-N-acetylpenicillamine, did not affect NGF-mediated neurite outgrowth in apoE(-/-) cultured DRG cells. However, 8-bromoguanosine 3',5'-cyclic monophosphate sodium salt n-hydrate, a cGMP analog, induced NGF-mediated nerve facilitation similar to wild-type NGF-mediated neurite outgrowth levels. Furthermore, in apoE(-/-) DRG, soluble guanylate cyclase expression was significantly lower than that in wild-type DRG. These results suggest that in apoE(-/-) mice the Akt-NO-cGMP pathway is impaired, which may be caused by NGF-mediated CGRP-LI-neurite outgrowth defects.

ROLE OF NERVE GROWTH FACTOR IN ALLERGIC AND INFLAMMATORY LUNG DISEASES

INTRODUCTION: Nerve growth factor (NGF) is a neurotrophin that exerts an important role in the development and functions of the central and peripheral nervous system, as it was originally discovered for its properties of simulating growth and differentiation of neurons; however, it was recently documented that several immune cells, such as mast cells, lymphocytes, and eosinophils, produce, store, and release NGF. Neurotrophins, including NGF, are constitutively expressed by resident lung cells and produced in increasing quantities by immune cells that invade the airways under inflammatory conditions. Furthermore, NGF appears as a promoter of allergic airway inflammation by increasing eosinophil and lymphocyte recruitment into the lungs. Neurotrophin receptors are expressed on several immune cells, including mast cells, T cells, B cells, and macrophages. OBJECTIVE: The objective of this study was to clarify the role of NGF in allergic and inflammatory lung diseases. METHODS: Our study was conducted of 90 children who attended the outpatient pediatric clinic or were admitted to the inpatient pediatric department of Elminia University Hospital. They were classified into 3 groups as follows: group 1, 35 children with asthma during the acute attack; group 2, 35 children with severe inflammatory lung disease and bronchopneumonia; group 3, 20 seemingly healthy children who were age and gender matched to the children with disease. For all children, the following were done: careful history taking, thorough clinical examination, chest radiograph, complete blood count, erythrocyte sedimentation rate, and reverse-transcriptase polymerase chain reaction (RT-PCR) to detect NGF receptor mRNA expression on purified eosinophils that were obtained from peripheral blood. RESULTS: We found only 3 patients who had asthma and had positive NGF receptors on isolated eosinophils from the peripheral blood by RT-PCR; however, all studied patients with bronchopneumonia had negative results. Moreover, there was a statistically significant difference between patients with positive and negative results for NGF receptors on isolated eosinophils by RT-PCR regarding age, the frequency of recurrence of asthma attacks, and positive history of other atopic diseases such as allergic dermatitis and allergic rhinitis; however, there was no statistically significant difference between patients with positive and negative results regarding gender, type of feeding, or family history. CONCLUSIONS: There is a strong association between NGF receptors on isolated eosinophils and the severity of allergic lung diseases and bronchial asthma.

Effects of huperzine A on secretion of nerve growth factor in cultured rat cortical astrocytes and neurite outgrowth in rat PC12 cells

To study the effects of huperzine A (HupA) on neuritogenic activity and the expression of nerve growth factor (NGF). After being treated with 10 micromol/L HupA, neurite outgrowth of PC12 cells was observed and counted under phase-contrast microscopy. Mitogenic activity was assayed by [3H]thymidine incorporation. Cell cytotoxicity was evaluated by lactate dehydrogenase (LDH) release. AChE activity, mRNA and protein expression were measured by the Ellman method, RT-PCR, and Western blot, respectively. NGF mRNA and protein levels were determined by RT-PCR and ELISA assays. Treatment of PC12 cells with 10 micromol/L HupA for 48 h markedly increased the number of neurite-bearing cells, but caused no significant alteration in cell viability or other signs of cytotoxicity. In addition to inhibiting AChE activity, 10 micromol/L HupA also increased the mRNA and protein levels of this enzyme. In addition, following 2 h exposure of the astrocytes to 10 micromol/L HupA, there was a significant up-regulation of mRNA for NGF and P75 low-affinity NGF receptor. The protein level of NGF was also increased after 24 h treatment with HupA. Our findings demonstrate for the first time that HupA has a direct or indirect neurotrophic activity, which might be beneficial in treatment of neurodegenerative disorders such as Alzheimer disease.

Analysis of Distinct Tartrate-resistant Acid Phosphatase Promoter Regions in Transgenic Mice

The tartrate-resistant acid phosphatase (TRAP) is present in multiple tissues, including kidney, liver, lung, spleen, and bone. Recent study of (TRAP) gene expression has provided evidence for distinct promoters within the (TRAP) gene, suggesting that the gene has alternative, tissue-preferred mRNA transcripts. Examination of endogenous (TRAP) exon 1B and 1C mRNA transcripts revealed tissue-preferred transcript abundance with increased exon 1B transcripts detected in liver and kidney and increased exon 1C transcripts detected in bone and spleen. In this investigation, we have made transgenic mice that express a marker gene driven by two candidate promoters, designated BC and C, within the (TRAP) gene. The BC and C promoters are 2.2 and 1.6 kb, respectively, measured from the translation initiation site. Evaluation of BC transgenic lines demonstrated robust expression in multiple tissues. In contrast, significant transgene expression was not detected in C transgenic lines. Evaluation of transgene mRNAs in BC transgenic lines revealed that virtually all expression was in the form of B transcripts, suggesting that the tissue-preferred pattern of endogenous (TRAP) was not replicated in the BC transgenic line. Likewise, osteoclastogenic cultures from BC, but not C, transgenic bone marrow cells expressed the transgene following receptor activator of NFkappaB ligand/macrophage colony-stimulating factor stimulation. In conclusion, when compared with the 2.2-kb BC portion of the (TRAP) promoter region, the 1.6-kb C portion does not account for significant gene expression in vivo or in vitro; production of the bone- and spleen-preferred (TRAP) C transcript must depend on regulatory elements outside of the 2.2-kb promoter. As the majority of currently investigated transcription factors that influence transcriptional regulation of osteoclast gene expression bind within the 1.6-kb C portion of the (TRAP) promoter, it is likely that transcription binding sites outside of the 2.2-kb region will have profound effects on regulation of the gene in vivo and in vitro.

Expression of the low-affinity p75 nerve growth factor receptor in the developing rat pituitary gland.

We investigated, by means of in situ hybridization with a digoxigenin-labelled RNA probe, the expression of the low-affinity p75 nerve growth factor receptor (NGFR) in the developing pituitary primordium of the rat. In 13-day pc embryos, intense staining of p75 NGFR mRNA was present in the cytoplasm of all cells of Rathke's pouch. In day-17 pc embryos p75 NGFR expression was present primarily in the cells of the intermediate lobe. In the newborn rat pituitary only very weak staining was observed, predominantly in the intermediate lobe. In neural structures the staining at day 13 pc was comparable to that of day 17 pc. Since p75 expression is seen very early during pituitary development and declines during the time the expression of pituitary hormonal phenotypes are steadily increasing, we suggest that the p75 NGFR expression in Rathke's pouch may play a temporally defined role in the commitment rather than in the differentiation of the various pituitary cell types.

MRNA for NGF and p75 in the central nervous system of rats affected by experimental allergic encephalomyelitis.

In this study we measured the concentration of nerve growth factor (NGF) and the expression of NGF and the low affinity NGF-receptor (NGF-r) mRNA in the central nervous system (CNS) of rats affected by experimental allergic encephalomyelitis (EAE) during the acute phase of the disease. Significant levels of NGF protein were found in thalamus and cortex on day 13 post-immunization. Molecular analysis of the NGF gene expression and of its NGF-r revealed that they were enhanced in several regions of the CNS of EAE rats when compared with untreated animals. These results suggest a functional link between local NGF synthesis and this autoimmune inflammatory disease.

Attenuation and recovery of nerve growth factor receptor mRNA in dorsal root ganglion neurons following axotomy.

The actions of nerve growth factor (NGF) are mediated by two receptor proteins, trk and p75. Recent evidence indicates that NGF upregulates the expression of both trk and p75 in responsive neurons including rat dorsal root ganglion (DRG) neurons. Axotomy by disconnecting the neuron from its source of target-derived NGF is predicted to lead to the downregulation of trk and p75 expression. However, previous studies of the effects of axotomy on trk and p75 mRNA expression in rat DRG have yielded discrepant results. We report that following sciatic nerve crush, trk and p75 mRNA levels in L4-L6 DRG decrease to approximately 50% of control levels at 4-14 days, return to control levels by 30 days, and are increased by approximately 30% at 60 days. Similar changes are observed following nerve transection although mRNA levels are slower in returning to normal and do not exceed control levels at later timepoints. Thus, trk and p75 expression decline early following target disconnection and later recover irrespective of target reinnervation. These observations indicate that target derived NGF is required for the maintenance of NGF receptor expression in adult rat DRG neurons and that non-target derived factors can appropriate this function following peripheral nerve injury.

Mechanisms of estrogen action during neural development: Mediation by interactions with the neurotrophins and their receptors?

Estrogen enhances the growth and differentiation of neurites within the developing forebrain. A critical issue is whether these developmental actions of estrogen are mediated directly or indirectly by means of autocrine responses or local paracrine mechanisms, through interactions with growth factors, such as the neurotrophins, and their receptors. Support for the latter hypothesis comes from our recent observations of co-expression of estrogen receptor mRNA with the mRNAs for the neurotrophins and their receptors; differential and reciprocal up-regulation of estrogen and NGF receptor mRNA and protein expression by estrogen in adult female rat sensory neurons, PC12 cells; and cerebral cortical cultures; and putative estrogen response elements in the NGF, BDNF, trkA and p75 genes. Estrogen and the neurotrophins may influence each other's actions by regulating receptor and ligand availability or by reciprocal regulation at the level of signal transduction or gene transcription. The neurotrophins may serve as regulatory “switches” for the apparent developmentally-regulated, differential pattern of estrogen receptor regulation by its ligand, whereby their ability to increase estrogen receptor levels significantly may be sufficient to override the intrinsic suppressive action of estrogen on its receptor. Estrogen and the neurotrophins, acting in concert and reciprocally, may stimulate the synthesis of proteins required for neuronal differentiation, survival and maintenance of function.

Dexamethasone regulates the expression of neuronal properties of a rat insulinoma cell line.

Insulin producing cells of the pancreas (beta cells) and neuronal cells share a large number of similarities. For example, different molecules, thought to be specific of neuronal cells, are expressed by beta cells. The factors regulating the expression of these molecules in beta cells are poorly understood. In the present work, we have studied the effect of dexamethasone, a synthetic glucocorticoid, on the expression of three different neuronal traits expressed by INS-1 cells, a highly differentiated beta cell line. We demonstrate that dexamethasone treatment decreases the steady state levels of mRNAs coding for both the low- and the high-affinity NGF receptors and of mRNA coding for NF-H, an intermediate neurofilament specific of neurons. This effect was time-dependent, the decrease being detectable after 4-8 h treatment. The decrease in NGF receptors mRNAs steady state levels was paralleled by a decrease in the number of NGF binding sites as demonstrated after Scatchard analysis. We further focused on the mechanisms by which dexamethasone affects the expression of the low affinity NGF receptor. The effect is countered by the glucocorticoid antagonist RU486, indicating that it is mediated by the glucocorticoid receptor. Finally, the decrease in the low-affinity nerve growth factor receptor mRNA steady state level after dexamethasone treatment is not due to mRNA destabilization but can be rather explained through a change in gene transcription.

Loss of p75 nerve growth factor receptor mRNA containing neurons in rat forebrain after intraventricular IgG 192-saporin administration

Cholinergic neurons of the basal forebrain express the p75 (low affinity) nerve growth factor receptor (NGFr) on the cell surface. It has been previously shown that an immunotoxin that recognizes the p75 NGFr, IgG 192-saporin, eliminates these neurons as judged by a variety of techniques after intraventricular injection into rat brain. Here we show that this loss of neurons can also be identified by detecting the disappearance of the p75 NGFr mRNA utilizing a non-radioactive in situ hybridization method.

Localization of trkB and low-affinity nerve growth factor receptor mRNA in the developing rat retina

The localization of trkB and low-affinity nerve growth factor receptor (LNGFR) mRNAs in the developing rat retina was examined by in situ hybridization. TrkB mRNA was expressed in the ganglion cell layer (GCL), in the inner border of the neuroblastic layer (NBL), and the inner border of the inner nuclear layer (INL). LNGFR mRNA was expressed in the GCL, in almost full thickness of the NBL, and in the intermediate part of the INL. Although both trkB mRNA and LNGFR mRNA were expressed in the GCL, the expression pattern was different between these mRNAs; trkB mRNA was expressed in almost all cells in the GCL uniformly and the expression of LNGFR mRNA varied greatly from cell to cell. In addition, the expression of both mRNAs, especially LNGFR mRNA seemed to be down-regulated at P7, when programmed cell death of the RGCs was prominent. These observations indicate that LNGFR may modulate the function of trkB and that trkB and LNGFR play important roles in the development and maintenance of the RGCs.

Anti-NGF treatment blocks the upregulation of NGF receptor mRNA expression associated with collateral sprouting of rat dorsal root ganglion neurons

The collateral sprouting of intact cutaneous sensory neurons has been shown to be dependent upon the presence of nerve growth factor (NGF). We have examined NGF receptor (NGFR) mRNA expression in DRG neurons undergoing sprouting and in dorsal root ganglion (DRG) neurons whose sprouting had been prevented through exposure to anti-NGF antiserum during the course of the experiment. The results indicate that the low affinity p75 NGFR mRNA is increased by 4 days post-operatively in DRGs from control serum-treated animals, and that this increase is most pronounced in smaller neurons. In contrast, the expression of p75 mRNA was maintained at control levels in DRGs from anti-NGF-treated animals. Results for trkA expression indicate an increase in expression at days 4–6 post-op in both ;groups of animals, with the anti-NGF treatment having a delayed influence on mRNA levels. Examination of GAP-43 mRNA levels revealed an increased expression in sprouting DRG neurons, whereas this increase was not observed in DRGs from anti-NGF treated animals. Taken together, these results provide further evidence of NGF's role in the collateral sprouting of nociceptive neurons.

Effects of acetyl- l-carnitine treatment and stress exposure on the nerve growth factor receptor (p75 NGFR) mRNA level in the central nervous system of aged rats

1. 1. There is growing evidence that the nerve growth factor protein (NGF), a neurotrophic factor for peripheral and central nervous system (CNS) neurons, may play a role in the modulation of the hypothalamo-pituitary-adrenocortical axis (HPAA). While NGF binding is decreased in rodent CNS after stress exposure, this reduction is prevented by treatment with Acetyl-L-Carnitine (ALCAR), a chemical substance able to prevent some degenerative events associated with aging. 2. 2. The authors studied the effect of cold stress on the low-affinity NGF receptor (p75 NGFR) mRNA levels in the basal forebrain and cerebellum of aged rats chronically treated with ALCAR. 3. 3. The present results show that ALCAR abolished the age-associated reduction of p75 NGFR mRNA levels in the basal forebrain of old animals, but did not affect the response to stress stimuli. 4. 4. Also, treatment with ALCAR mantained p75 NGFR mRNA levels in the cerebellum of old animals at levels almost identical to those observed in young control animals. 5. 5. These results suggest a neuroprotective effect for ALCAR on central cholinergic neurons exerted at the level of transcription of p75 NGFR. The restoration of p75 NGFR levels could increase trophic support by NGF of these CNS cholinergic neurons which are implicated in degenerative events associated with aging.

Disruption of Fast Axonal Transport in Vivo Leads to Alterations in Schwann Cell Gene Expression

Following nerve injury, Schwann cells distal to the site of injury down-regulate genes associated with myelination. We hypothesized that at least some of these alterations were due to the loss of ongoing axon:Schwann cell homeostatic signals, as opposed to loss of physical contact and/or inflammatory responses. To directly test this hypothesis, we perturbed axonal physiology by selectively blocking fast axonal transport via locally cooling the sciatic nerve to 5-8°C (a cold block). Immunostaining with the monoclonal antibody ED1, which recognizes mononuclear phagocytic cells, demonstrated that macrophages did not invade the cold-blocked nerve, indicating the lack of an inflammatory response. Morphological studies demonstrated that the nerve distal to the cold block showed no signs of Wallerian degeneration, with maintenance of normal axon and myelin profiles, and confirmed the absence of invading macrophages. Thus, any effects of a cold-block treatment were not likely due to inflammatory responses or to loss of physical contact between axons and Schwann cells. To determine whether this treatment affected Schwann cell phenotype, we examined expression of the major myelin protein Po, and p75 NGF receptor, both of which are regulated as a function of axon:Schwann cell interactions. Levels of p75 NGF receptor mRNA were unaffected by the cold block, while p75 NGF receptor protein levels were increased in the region of the nerve immediately adjacent to the cold block, presumably reflecting protein accumulation as a consequence of the block to fast axonal transport. In contrast, levels of Po mRNA were decreased in the distal nerve in a fashion that indicated modulation of Schwann cell phenotype as a function of local axonal microenvironment. These data therefore suggest that Po and p75 NGF receptor are regulated as a function of two different aspects of Schwann cell:axon communication. Furthermore, these data demonstrate that the presence of axon:Schwann cell contact alone is insufficient to maintain Po gene expression and indicate that at least some myelin-specific Schwann cell responses are dependent upon ongoing biochemical signals generated by the axon and maintained by fast axonal transport.

Estrogen differentially regulates estrogen and nerve growth factor receptor mRNAs in adult sensory neurons

We have previously shown that neurons in the basal forebrain colocalize the neurotrophin receptor p75NGFR and estrogen receptors. The present study was designed to examine (1) if neural neurotrophin targets respond to estrogen as a general phenotypic feature and (2) if NGF receptor mRNAs are regulated by estrogen, using a prototypical target of NGF, the dorsal root ganglion (DRG) (sensory) neuron. We demonstrate, for the first time, the presence of estrogen receptor mRNA and protein (binding sites) in adult female rat DRG. Moreover, estrogen receptor mRNA expression, while present in DRG neurons from both the ovariectomized (OVX; estrogen deficient) and intact female rat, was downregulated, as in the adult CNS, during proestrus (high estrogen levels) and in OVX animals replaced with proestrus levels of estrogen, as compared to OVX controls. In contrast, although the mRNAs for the NGF receptors p75NGFR and trkA were also expressed in DRG neurons from OVX and intact animals, expression of both NGF receptor mRNAs was upregulated in sensory neurons during proestrus, as compared to the OVX condition. Estrogen replacement, on the other hand, resulted in a transient downregulation of p75NGFR mRNA and a time-dependent upregulation of trkA mRNA. Estrogen regulation of NGF receptor mRNA in adult peripheral neural targets of the neurotrophins supports the hypothesis that estrogen may regulate neuronal sensitivity to neurotrophins such as NGF and may be an important mediator of neurotrophin actions in normal neural function and following neural trauma.

Expression of NGF receptor and GAP‐43 mRNA in DRG neurons during collateral sprouting and regeneration of dorsal cutaneous nerves

The collateral sprouting of intact sensory axons and the regeneration of damaged ones differ in a number of respects. Regeneration is triggered by axotomy-induced damage, probably involves the loss of a peripheral signal, and appears to occur independently of NGF, while collateral sprouting is evoked and sustained by an increase in a target-derived signal, namely NGF. New findings strengthen the distinction between these two phenomena. Nerve growth factor receptor (NGFR) mRNA is increased in undamaged DRG neurons whose axons are sprouting into denervated skin. This response is related to an increased availability of target-derived NGF, a proposal supported by a number of findings including increased NGF mRNA in the denervated target. In contrast, we observed little or no change in the NGFR mRNA levels in regenerating neurons, consistent with the observations that NGF does not play a role in this process. However, increases in neuronal GAP-43 mRNA are found during both regeneration and collateral sprouting, a result in keeping with the proposal that GAP-43 is primarily associated with nerve growth, and the observation that GAP-43 expression is not especially influenced by NGF.

Nerve Growth Factor Derived from Terminals Selectively Increases the Ratio of p75 to trkA NGF Receptors on Mature Sympathetic Neurons

Exposure of neuronal terminals to target-derived NGF has been hypothesized to regulate cell body responses at a distance. To test this hypothesis and, more specifically, to determine whether NGF distally regulates the synthesis of its two characterized receptors, we developed the following paradigm. Sympathetic neurons of the superior cervical ganglion (SCG) that project either to the eye or to the ear were labeled, in adult animals, with the retrograde tracers Fast Blue and Fluoro-Gold, respectively. NGF was then injected daily into the anterior chamber of one eye, exposing the terminals of the ipsilateral eye neurons to increased NGF. To control for systemic and/or localized injury effects, cytochrome C or PBS were injected into the contralateral eye of the same animals. In situ hybridization and image analysis were then used to determine neuronal levels of p75 NGF receptor, trkA, and Tα1 α-tubulin mRNAs, with the latter providing a correlative measure of neuronal sprouting. Morphological measurements revealed that exogenous, terminally-derived NGF increased the mean cross-sectional area of eye neurons by 37%. Grain counts for p75 NGF receptor mRNA increased from 2- to 6-fold in the NGF-treated neurons, and grain densities, which accounted for neuronal hypertrophy, also increased significantly. In contrast, grain counts for trkA mRNA were not significantly increased by this treatment, while Tα1 α-tubulin mRNA levels increased only 1.5- to 2-fold. No increase in grain density was detected for either of these mRNAs. The NGF-induced increased in p75 NGF receptor mRNA levels was accompanied by terminal sprouting and by an increase in the density of p75 NGF receptors on terminal neurites, as indicated by IgG-192 immunostaining of the NGF-treated iris. These data therefore suggest that, in addition to promoting local sprouting, increased target-derived NGF increases the levels of p75 NGF receptor relative to trkA on terminal neurites, by differentially regulating receptor synthesis. Such a direct regulatory feedback loop may well play an integral role in precisely modulating neuronal responses as a function of the amount of available trophic support and/or target tissue.