Nerve growth factor stimulates development of substance P in the embryonic spinal cord

Development of the putative neurotransmitter, substance P (SP), in the embryonic rat dorsal root ganglion (DRG) and spinal cord was defined in vivo. SP was not detectable by radioimmunoassay before day 17 of gestation (E17). On E17, cervical sensory ganglia contained 4 pg SP/ganglion, rising to 49 pg/ganglion at birth. The dorsal cervical spinal cord contained 0.75 ng SP/mg protein on E17, rising to 6 ng SP/mg protein on postnatal day 3. The ventral spinal cord contained approximately 20% of the SP content in the dorsal cord at each gestational age. Intrauterine forelimb amputation partially prevented the normal development increase of SP in sensory ganglia destined to innervate that limb, suggesting that target structures regulate the development of peptidergic neruons. Conversely, treatment with nerve growth factor (NGF) stimulated development of SP in the DRG. Moreover, NGF treatment increased SP in the dorsal spinal cord, suggesting that NGF can modulate development within the CNS, as well as peripheral structures. It is likely that the CNS effect reflects NGF peptidergic neruons. Conversely, treatment with nerve growth factor (NGF) stimulated development of SP in the DRG. Moreover, NGF treatment increased SP in the dorsal spinal cord, suggesting that NGF can modulate development within the CNS, as well as peripheral structures. It is likely that the CNS effect reflects NGF peptidergic neruons. Conversely, treatment with nerve growth factor (NGF) stimulated development of SP in the DRG. Moreover, NGF treatment increased SP in the dorsal spinal cord, suggesting that NGF can modulate development within the CNS, as well as peripheral structures. It is likely that the CNS effect reflects NGF action on peripheral ganglia, but a direct effect on the spinal cord has not been excluded. However, treatment with antiserum to NGF failed to significantly inhibit development of ganglion SP. The system of SP-containing neurons in the DRG may provide a convenient model for defining events regulating peptidergic maturation.     

Projection of nodose ganglion cells to the upper cervical spinal cord in the rat

Afferent fibers mediating pain from myocardial ischemia classically are believed to travel in sympathetic nerves to enter the thoracic spinal cord. After sympathectomies, angina pectoris still may radiate to the neck and inferior jaw. Sensory fibers from those regions are thought to enter the central nervous system through upper spinal cord segments. We postulated that axons from nodose ganglion cells might project to cervical cord segments. The purpose of this study was to determine the density and pathway of vagal afferent innervation to the upper cervical spinal cord. Following an injection of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) into the upper cervical spinal cord, approximately 5.8% of cells in the nodose ganglion contained reaction product. Cervical vagotomy did not diminish the density of WGA-HRP labeled cells in the nodose ganglion. However, a spinal cord hemisection cranial to the injection site eliminated labeling of nodose cells. These data indicate that a portion of vagal afferent neurons project from the nodose ganglion to the upper cervical spinal cord. In addition, vagal afferent fibers reach the spinal cord via a central route rather than through dorsal root ganglia.     

Vagal and gastric connections to the central nervous system determined by the transport of horseradish peroxidase

Horseradish peroxidase (HRP, Sigma Type VI) crystals were encased in a parafilm envelope and applied to the transected central ends of the left and right cervical vagus nerves and the anterior and posterior esophageal vagus nerves of adult male hooded rats. Injections of 30% HRP were made into the muscle wall of the fundus and antrum regions of the stomach. After 48 hr survival time, animals were perfused intracardially with a phosphate buffer plus sucrose wash followed by glutaraldehyde and paraformaldehyde fixative. The brain stem, spinal cord and corresponding dorsal root ganglia, superior cervical sympathetic ganglion, and the nodose ganglion were removed and cut into 50 micron sections. All tissue was processed with tetramethylbenzidine (TMB) for the blue reaction according to Mesulum and counterstained with neutral red. Sequential sections were examined under a microscope. Labeled neurons and nerve terminals were identified using bright and dark field condensers and polarized light. In tissue from animals that had HRP applied to the cervical vagus nerves, retrogradely labeled neurons were identified ipsilaterally in the medulla located in the dorsal motor nucleus of the vagus (DMN) and the nucleus ambiguus (NA). Labeled cells extended from the DMN into the spinal cord in ventral-medial and laminae X regions C1 and C2 of cervical segments. Many neurons were labeled in the nodose ganglion. Anterogradely labeled terminals were observed throughout and adjacent to the solitary nucleus (NTS) dorsal to the DMN and intermixed among labeled neurons located in the DMN. In tissue from animals that had HRP applied to the esophageal vagus nerves, similar labeling was observed. However, fewer neurons were identified in the NA, the nodose ganglion, and only in laminae X of the cervical spinal cord segments C1 and C2. Also, very little terminal labeling was observed in and adjacent to the NTS. Labeled neurons in tissue from animals that had HRP injected into the stomach wall were observed bilaterally in the DMN, nodose ganglion, and only in laminae X at the C1 and C2 levels of the spinal cord. Labeled neurons also were observed in the dorsal root ganglia of the thoracic cord. These data indicate that cervical cord and NA neurons are important in the supradiaphragmatic motor innervation by the vagus. Also, many afferents to the NTS originate above the diaphragm. In addition, some afferents from the stomach enter the central nervous system via the thoracic spinal cord.     

Effects of nerve growth factor on sensory neurons in the chick embryo: a stereological study

Chicken embryos on days 6-13 of incubation received injections of purified beta NGF (80 micrograms/day) for 3 or 4 days and were then killed. Sensory ganglia were fixed and taken for embedding and sectioning. A stereological method based on unfolding of cell-diameter frequencies was used to determine the number of neurons of different size in the spinal, trigeminal and nodose ganglia. The total volume of the ganglia was also determined. NGF induced increases in diameter of the neural crest-derived dorsomedial (DM) neurons in spinal and trigeminal ganglia. Injected NGF did not influence ventrolateral (VL) neurons of neural crest origin in the spinal ganglia nor the ventrolateral neurons of placodal origin in the trigeminal ganglion. The volumes of spinal and trigeminal ganglia increased by 50 and 100%, respectively. The volume of the nodose ganglion and the total number and size of the placodal nodose neurons were unaffected by NGF. The results demonstrate a clear difference in the response to NGF in vivo between smaller and larger sensory neurons.     

Secretoneurin-like immunoreactivity in rat sympathetic, enteric and sensory ganglia

Distribution of secretoneurin-like immunoreactivity (SN-LI) was studied in the rat sympathetic ganglia/adrenal gland, enteric and sensory ganglia by immunohistochemical methods. SN-LI nerve fibers formed basket-like terminals surrounding many of the postganglionic neurons of the superior cervical, stellate, paravertebral chain ganglia, coeliac/superior mesenteric and inferior mesenteric ganglia. Postganglionic neurons of the superior cervical and other sympathetic ganglia exhibited low-to-moderate levels of SN-LI. In all these sympathetic ganglia, clusters of small diameter (<10 μm) cells, which may correspond to the small intensely fluorescent (SIF) cells, were found to be intensely labeled. Surgical sectioning or ligation of the cervical sympathetic trunk for 7–10 days resulted in a nearly total loss of SN-LI fibers in the superior cervical ganglia, whereas immunoreactivity in the postganglionic neurons and small diameter cells remained essentially unchanged. In the thoracolumbar and sacral segments of the spinal cord, SN-LI nerve fibers were detected in the superficial layers of the dorsal horn as well as in the intermediolateral cell column (IL_p). Occasionally, SN-LI somata were noted in the IL_p. SN-LI nerve fibers formed a delicate plexus underneath the capsule of the adrenal gland, some of which traversed the adrenal cortex and reached the adrenal medulla. While heavily invested with SN-LI nerve terminals, chromaffin cells seemed to express a low level of SN-LI. In the enteric plexus, varicose SN-LI nerve fibers and terminals formed a pericellular network around many myenteric and submucous ganglion cells; the ganglionic neurons were lightly to moderately labeled. A population of ganglion cells in the dorsal root, nodose and trigeminal ganglia exhibited moderate-to-strong SN-LI. The detection of SN-LI in nerve fibers and somata of various sympathetic ganglia, enteric plexus and adrenal medulla and in somata of the sensory ganglia implies an extensive involvement of this peptide in sympathetic, enteric and sensory signal processing.     

Pituitary adenylate cyclase activating polypeptide-immunoreactive sensory neurons innervate rat adrenal medulla

Rat adrenal chromaffin cells were invested by a dense network of nerve fibers immunoreactive to pituitary adenylate cyclase activating polypeptide-38 (PACAP-IR). Immunohistochemical studies demonstrated the presence of PACAP-IR in nodose and dorsal root ganglion cells, but not in neurons of the intermediolateral cell column and other autonomic nuclei of the thoracic and upper lumbar spinal cord. Somata of the T7 to T12 paravertebral ganglia were PACAP-negative. A few lightly labeled neurons were occasionally noted in the dorsal motor nucleus of the vagus. Injection of the retrograde tracer Fluorogold into the left adrenal medulla 3 days prior to sacrifice resulted in the labeling of a population of neurons in the ipsilateral spinal cord intermediolateral cell column (T1 to L1), ipsilateral and contralateral nodose ganglia and ipsilateral dorsal root ganglia from T7 to T10 inclusive. A small number of lightly labeled somata was occasionally noted in the dorsal motor nucleus of the vagus. Combined retrograde tracing and PACAP immunohistochemistry showed that a population of Fluorogold-containing nodose and dorsal root ganglion cells were also PACAP-positive. Pre-treatment of the rats with capsaicin caused a marked reduction of the PACAP-IR in the adrenal gland as well as in the superficial layers of the dorsal horn and caudal spinal trigeminal nucleus. These findings, in conjunction with the apparent absence of PACAP-IR in spinal sympathetic preganglionic neurons, sympathetic postganglionic neurons, and dorsal motor nucleus of the vagus, raise the possibility that PACAP-IR fibers observed in the adrenal medulla are primarily sensory in origin. As a corollary, catecholamine secretion from chromaffin cells may be modulated by the peptidergic sensory afferents in addition to the cholinergic sympathetic preganglionic nerve fibers.     

NGF mRNA is expressed in the dorsal root ganglia after spinal cord injury in the rat

Nerve growth factor (NGF) mRNA is expressed in a variety of cell types in the injured spinal cord and its protein implicated in both positive and negative neurological outcomes of cord injury. Here we demonstrate that NGF mRNA is also upregulated in dorsal root ganglion (DRG) neurons after spinal cord injury and that the percentage of sensory neurons expressing NGF mRNA correlates with proximity to the lesion epicenter. Our data suggest that, in DRG, NGF gene expression may be upregulated by damage to the central processes of sensory neurons.     

Visualizing sensory transmission between dorsal root ganglion and dorsal horn neurons in co-culture with calcium imaging

Sensory information is conveyed to the central nervous system by primary afferent neurons within dorsal root ganglia (DRG), which synapse onto neurons of the dorsal horn of the spinal cord. This synaptic connection is central to the processing of both sensory and pain stimuli. Here, we describe a model system to monitor synaptic transmission between DRG neurons and dorsal horn neurons that is compatible with high-throughput screening. This co-culture preparation comprises DRG and dorsal horn neurons and utilizes Ca(2+) imaging with the indicator dye Fura-2 to visualize synaptic transmission. Addition of capsaicin to co-cultures stimulated DRG neurons and led to activation of dorsal horn neurons as well as increased intracellular Ca(2+) concentrations. This effect was dose-dependent and absent when DRG neurons were omitted from the culture. NMDA receptors are a critical component of synapses between DRG and dorsal horn neurons as MK-801, a use-dependent non-competitive antagonist, prevented activation of dorsal horn neurons following capsaicin treatment. This model system allows for rapid and efficient analysis of noxious stimulus-evoked Ca(2+) signal transmission and provides a new approach both for investigating synaptic transmission in the spinal cord and for screening potential analgesic compounds.     

Apoptosis of spinal interneurons induced by sciatic nerve axotomy in the neonatal rat is counteracted by nerve growth factor and ciliary neurotrophic factor

We have previously shown that not only motoneurons and dorsal root ganglion cells but also small neurons, presumably interneurons in the spinal cord, may undergo apoptotic cell death as a result of neonatal peripheral nerve transection in the rat. With the aid of electron microscopy, we have here demonstrated that apoptosis in the spinal cord is confined to neurons and does not involve glial cells at the survival time studied (24 hours). To define the relative importance of the loss of a potential target (motoneuron) and a potential afferent input (dorsal root ganglion cell) for the induction of apoptosis in interneurons in this situation, we have compared the distributions and time courses for TUNEL labeling, which detects apoptotic cell nuclei, in the L5 segment of the spinal cord and the L5 dorsal root ganglion after sciatic nerve transection in the neonatal (P2) rat. In additional experiments, we studied the effects on TUNEL labeling of interneurons after treatment of the cut sciatic nerve with either ciliary neurotrophic factor (CNTF) to rescue motoneurons or nerve growth factor (NGF) to rescue dorsal root ganglion cells. The time courses of the TUNEL labeling in motoneurons and interneurons induced by the lesion show great similarities (peak at 8-48 hours postoperatively), whereas the labeling in dorsal root ganglion cells occurs later (24-72 hours). Both CNTF and NGF decrease the number of TUNEL-labeled interneurons, but there is a regional difference, in that CNTF preferentially saves interneurons in deep dorsal and ventral parts of the spinal cord, whereas the rescuing effects of NGF are seen mainly in the superficial dorsal horn. The results are interpreted as signs of a trophic dependence on both the target and the afferent input for the survival of interneurons neonatally.     

Activation of transient receptor potential vanilloid 2‐expressing primary afferents stimulates synaptic transmission in the deep dorsal horn of the rat spinal cord and elicits mechanical hyperalgesia

Probenecid, an agonist of transient receptor vanilloid (TRPV) type 2, was used to evaluate the effects of TRPV2 activation on excitatory and inhibitory synaptic transmission in the dorsal horn (DH) of the rat spinal cord and on nociceptive reflexes induced by thermal heat and mechanical stimuli. The effects of probenecid were compared with those of capsaicin, a TRPV1 agonist. Calcium imaging experiments on rat dorsal root ganglion (DRG) and DH cultures indicated that functional TRPV2 and TRPV1 were expressed by essentially non‐overlapping subpopulations of DRG neurons, but were absent from DH neurons and DH and DRG glial cells. Pretreatment of DRG cultures with small interfering RNAs against TRPV2 suppressed the responses to probenecid. Patch‐clamp recordings from spinal cord slices showed that probenecid and capsaicin increased the frequencies of spontaneous excitatory postsynaptic currents (sEPSCs) and spontaneous inhibitory postsynaptic currents in a subset of laminae III–V neurons. In contrast to capsaicin, probenecid failed to stimulate synaptic transmission in lamina II. Intrathecal or intraplantar injections of probenecid induced mechanical hyperalgesia/allodynia without affecting nociceptive heat responses. Capsaicin induced both mechanical hyperalgesia/allodynia and heat hyperalgesia. Activation of TRPV1 or TRPV2 in distinct sets of primary afferents increased the sEPSC frequencies in a largely common population of DH neurons in laminae III–V, and might underlie the development of mechanical hypersensitivity following probenecid or capsaicin treatment. However, only TRPV1‐expressing afferents facilitated excitatory and/or inhibitory transmission in a subpopulation of lamina II neurons, and this phenomenon might be correlated with the induction of thermal heat hyperalgesia.     

The difference of osteocalcin-immunoreactive neurons in the rat dorsal root and trigeminal ganglia: co-expression with nociceptive transducers and central projection

The co-expression of osteocalcin (OC) with the capsaicin receptor (VR1) and vanilloid receptor 1-like receptor (VRL-1) was examined in the dorsal root (DRG) and trigeminal ganglia (TG). Virtually all OC-immunoreactive (ir) DRG neurons were devoid of VR1- and VRL-1-immunoreactivity (ir). In the TG, 14.1% of OC-ir neurons were also immunoreactive for VR1. Only 1.7% of OC-ir TG neurons co-expressed VRL-1-ir. The distribution of OC-ir was also examined in the spinal cord and trigeminal sensory nuclei. In the spinal cord, the superficial laminae of the dorsal horn were devoid of OC-ir. The neuropil was weakly stained in other regions of the spinal horns. The medullary dorsal horn (MDH) contained numerous OC-ir varicose fibers in laminae I and II. These fibers were occasionally observed originating from the spinal trigeminal tract. The neuropil was weakly stained in deeper laminae of the MDH, and the rostral parts of the trigeminal sensory nuclei. The present study suggests that OC-ir TG nociceptors send their unmyelinated axons to the superficial laminae of the MDH.     

Human and Rat Primary C-Fibre Afferents Store and Release Secretoneurin, a Novel Neuropeptide

Secretoneurin is a recently discovered neuropeptide derived from secretogranin II (SgII). Since this peptide could be detected in the dorsal horn of the spinal cord we studied whether it is localized in and released from primary afferent neurons. Secretoneurin was investigated with immunocytochemistry and radioimmunoassay in spinal cord, dorsal root ganglia and peripheral organs. SgII mRNA was determined in dorsal root ganglia. Normal rats and rats pre-treated neonatally with capsaicin to destroy selectively polymodal nociceptive (C-) fibres were used. Slices of dorsal spinal cord were perfused in vitro for release experiments. Immunocytochemistry showed a distinct distribution of secretoneurin-immunoreactivity (IR) in the spinal cord and lower brainstem. A particularly high density of fibres was found in lamina I and outer lamina II of the caudal trigeminal nucleus and of the spinal cord. This distribution was qualitatively identical in rat and human post-mortem tissue. Numerous small diameter and some large dorsal root ganglia neurons were found to contain SgII mRNA. Capsaicin treatment led to a marked depletion of secretoneurin-IR in the substantia gelatinosa, but not in other immunopositive areas of the spinal cord and to a substantial loss of small (<25 μm) SgII-mRNA-containing dorsal root ganglia neurons. Radioimmunoassay revealed a significant decrease of secretoneurin-IR in the dorsal spinal cord, the trachea, heart and urinary bladder of capsaicin-treated rats. Perfusion of spinal cord slices with capsaicin as well as with 60 mM potassium led to a release of secretoneurin-IR. In conclusion, secretoneurin is a neuropeptide which is stored in and released from capsaicin-sensitive, primary afferent (C-fibre) neurons. It may, therefore, be a novel peptidergic modulator of pain transmission or of C-fibre mediated non-nociceptive information.     

Effects of nerve growth factor on TTX- and capsaicin-sensitivity in adult rat sensory neurons

We have investigated the effects of nerve growth factor (NGF, 2.5 ng/ml for 1–2 weeks) on enriched adult rat dorsal root ganglion (DRG) neurons maintained in cell culture in defined media. Whole-cell recordings in cells cultured in the absence and presence of NGF revealed no significant difference in resting membrane potential and input resistance. However, the threshold for spike generation was significantly lower in untreated cells than in treated cells; −25 ± 1.1mV vs−19 ± 2.2mV, respectively. The sensitivity of the Na⁺ spike to tetrodotoxin (TTX, 1 μM) was different in cells cultured in the absence or presence of NGF. For example, spikes were abolished by TTX in 100% of untreated cells, while in NGF-treated cells the spike was abolished in only 41% of the neurons. Chemosensitivity of DRG neurons was also different in the absence and presence of NGF. For example, the percent of neurons in which a current activated by 8-methyl-N-vanillyl-6-nonenamide (capsaicin, 500 nM) was detected, increased from 18% in untreated cells to 55% in NGF-treated cells. NGF did not influence the number of cells surviving. The results indicate that NGF can regulate TTX and capsaicin sensitivity in these adult rat sensory neurons. Our experimental protocol indicates that this effect is not mediated by a factor in the serum or released from non-neuronal cells.     

神经生长因子对培养的大鼠背根神经节神经元P物质释放的调节作用

目的观察神经生长因子（nerve growth factor, NGF）对原代培养的背根神经节（dorsal root ganglion, DRG）神经元中P物质（substance P, SP）的基础释放量和辣椒素诱发释放量的调节效应。方法将15 天胚龄的Wistar大鼠DRG神经元培养于含有不同浓度NGF的DMEM/F12培养液中,不含NGF的培养液培养的神经元作为对照。72小时后,用RT-PCR检测神经元中SP mRNA和辣椒素受体（vanilloid receptor 1, VR1）mRNA的表达,用放射免疫分析（radioimmunoassay,RIA）法检测SP的基础释放量和辣椒素（100 nmol/L）刺激10 min后的诱发释放量。结果SPmRNA和VR1 mRNA在NGF孵育的标本中表达增加,并与孵育液中NGF的浓度呈剂量依赖关系。SP的基础释放量和辣椒素诱发释放量在NGF孵育的标本中均增加,而且诱发释放量与NGF的浓度呈剂量依赖关系。结论NGF使DRG神经元SP的基础释放量和诱发释放量增加,表明NGF能增加初级传入神经元感受伤害刺激的敏感性,该效应可能与SP和VR1的mRNA表达增加有关。     

神经生长因子对培养的大鼠背根神经节神经元P物质释放的调节作用(英文)

目的观察神经生长因子(nerve growth factor, NGF)对原代培养的背根神经节(dorsal root ganglion, DRG)神经元中P物质(substance P, SP)的基础释放量和辣椒素诱发释放量的调节效应。方法将15 天胚龄的Wistar大鼠DRG神经元培养于含有不同浓度NGF的DMEM/F12培养液中,不含NGF的培养液培养的神经元作为对照。72小时后,用RT-PCR检测神经元中SP mRNA和辣椒素受体(vanilloid receptor 1, VR1)mRNA的表达,用放射免疫分析(radioimmunoassay,RIA)法检测SP的基础释放量和辣椒素(100 nmol/L)刺激10 min后的诱发释放量。结果SPmRNA和VR1 mRNA在NGF孵育的标本中表达增加,并与孵育液中NGF的浓度呈剂量依赖关系。SP的基础释放量和辣椒素诱发释放量在NGF孵育的标本中均增加,而且诱发释放量与NGF的浓度呈剂量依赖关系。结论NGF使DRG神经元SP的基础释放量和诱发释放量增加,表明NGF能增加初级传入神经元感受伤害刺激的敏感性,该效应可能与SP和VR1的mRNA表达增加有关。     

The differential contribution of capsaicin-sensitive afferents to behavioral and cardiovascular measures of brief and persistent nociception and to Fos expression in the formalin test

Intraplantar injection of dilute formalin evokes brief (Phase 1) and persistent (Phase 2) increases in primary afferent activity, pain behavior, and cardiovascular responses, and induces spinal cord Fos-like immunoreactivity (Fos-LI). Although previous studies demonstrated that the destruction of small diameter primary afferents with neonatal capsaicin treatment decrease formalin-evoked nociception, these studies only evaluated behavioral responses, and did not distinguish between Phase 1 and 2. To address these questions, we simultaneously evaluated formalin-evoked pain behavior (flinching of the afflicted paw), cardiovascular responses (heart rate and mean arterial pressure), and lumbar spinal cord Fos expression in control rats and in rats treated with capsaicin (100 mg/kg) one day postpartum. We found that neonatal capsaicin-treated rats, compared to controls, exhibited similar cardiovascular responses and slightly less flinching behavior during Phase 1. During Phase 2, however, capsaicin-treated rats exhibited 59% less flinching and 45% smaller heart rate responses. Also, in capsaicin-treated rats, we counted 59% fewer Fos-labeled neurons in the spinal cord. These results indicate that capsaicin-sensitive afferents contribute to formalin-evoked behavioral and cardiovascular responses and to spinal cord neuronal responses. The differential effect of neonatal capsaicin on nociception during Phase 1 and Phase 2 suggests that sensitization mechanisms during Phase 1 do not contribute to the magnitude of nociceptive responses during Phase 2.     

The coexistence of TrkA with putative transmitter agents and calcium-binding proteins in the vagal and glossopharyngeal sensory neurons of the adult rat

The presence of the neurotrophin receptor, TrkA, in neurochemically identified vagal and glossopharyngeal sensory neurons of the adult rat was examined. TrkA was colocalized with calcitonin gene-related peptide (CGRP), parvalbumin, or calbindin D-28k in neurons of the nodose, petrosal and/or jugular ganglia. In contrast, no TrkA-immunoreactive (ir) neurons in these ganglia colocalized tyrosine hydroxylase-ir. About one-half of the TrkA-ir neurons in the jugular and petrosal ganglia contained CGRP-ir, whereas only a few of the numerous TrkA-ir neurons in the nodose ganglion contained CGRP-ir. Although 43% of the TrkA-ir neurons in the nodose ganglion contained calbindin D-28k-ir, few or no TrkA-ir neurons in the petrosal or jugular ganglia were also labeled for either calcium-binding protein. These data show distinct colocalizations of TrkA with specific neurochemicals in vagal and glossopharyngeal sensory neurons, and suggest that nerve growth factor (NGF), the neurotrophin ligand for TrkA, plays a role in functions of specific neurochemically defined subpopulations of mature vagal and glossopharyngeal sensory neurons.     

The Janus role of c-Jun: cell death versus survival and regeneration of neonatal sympathetic and sensory neurons

We investigated the functional outcome of c-Jun activation in sympathetic and sensory neurons of neonatal rat superior cervical ganglion (SCG) and dorsal root ganglion (DRG), respectively. Distinctly different roles of c-Jun activation have been suggested for these two types of neurons. In dissociated sympathetic neurons, c-Jun has been demonstrated to promote apoptosis, whereas in sensory neurons it stimulates axonal outgrowth. In organ-cultured ganglia, we found that c-Jun was activated within 24 h of explantation in both types of neurons, and that the JNK inhibitor SP600125 could mitigate this response. In both types of neurons, c-Jun activation was also reduced by NGF treatment. Inhibition of c-Jun activation did not affect the viability of sympathetic neurons, whereas the number of apoptotic sensory neurons increased. Furthermore, inhibition of c-Jun reduced axonal outgrowth from both SCG and DRG. Thus, in organ culture, c-Jun activation may be required for axonal outgrowth and, at least in sensory neurons, it promotes survival. The role of ATF3, a neuronal marker of injury and a c-Jun dimerization partner, was also examined. We found an ATF3 induction in both SCG and DRG neurons, a response, which was reduced by JNK inhibition. The reduction of ATF3 upon JNK inhibition was much larger in DRG than in SCG, a result which might account for the higher number of apoptotic neurons in JNK inhibitor exposed DRG. Taken together, and contrary to our expectations, neonatal sympathetic and sensory neurons seem to respond to axonal injury similarly with respect to c-Jun activation, and in no case was this activation pro-apoptotic.     

Embryonic sensory development: Local expression of neurotrophin-3 and target expression of nerve growth factor

Development and maintenance of peripheral sensory and sympathetic neurons are regulated by target-derived neurotrophins, including nerve growth factor (NGF). To determine whether trophins are potentially critical prior to and during target innervation, for neuronal survival or axon guidance, in situ hybridization was performed in the rat embryo. We examined the expression of genes encoding NGF, neurotrophin-3 (NT-3), and their putative high-affinity receptors, trk A and trk C, respectively. Trks A and C were detected in dorsal root sensory ganglia (DRG) on embryonic day 12.5 (E12.5), implying early responsiveness to NGF and NT-3. NGF mRNA was expressed in the central spinal cord target and by the peripheral somite, at this early time, which thereby may function as a transient “guidepost” target for sensory fibers. Somitic expression was transient and was undetectable by E17.5. NT-3 was expressed in the DRG itself from E13.5 to 17.5, suggesting local transient actions on sensory neurons. NT-3 was also expressed in the ventral spinal cord at low levels on E13.5. We examined the trigeminal ganglion to determine whether cranial sensory neurons are similarly regulated. Trk A was detected in the trigeminal ganglion, while NGF was expressed in the central myelencephalon target, paralleling observations in the DRG and spinal cord. However, NT-3 and trk C were undetectable, in contrast to DRG, suggesting that the environment or different neural crest lineages govern expression of different trophins and trks. Apparently, multiple trophins regulate sensory neuron development through local as well as transient target mechanisms prior to innervation of definitive targets.