Extension of sympathetic neurites in vitro towards explants of embryonic and neonatal mouse heart and stomach: ontogeny of neuronotrophic factors

In order to establish when target organs first produce neuronotrophic factors, extension of neurites in vitro from sympathetic ganglia (superior cervical and coeliac) of 1-day neonatal mice towards explants of 10-, 11-, 14- and 17-day embryonic and 1-day neonatal atrium and stomach was examined in co-cultures. Longer neurites extended from ganglia towards, than away from, atrial targets at all stages examined, and was most marked towards 17-day embryonic and neonatal explants. Treatment of atrial co-cultures with antiserum to nerve growth factor (NGF) almost totally blocked preferential neurite outgrowth. Directional growth of neurites towards stomach explants in co-cultures was not as pronounced as that towards atrium; extension of neurites was most marked when stomach was provided by 11-, 14- and 17-day embryos. Such outgrowth was only partially blocked by antiserum to NGF, significant preferential extension of neurites towards stomach persisting in the presence of the antiserum. These results indicate that atrium and stomach produce neuronotrophic factors from the earliest ages studied; the evidence indicates that in the case of atrium, NGF predominates but that stomach produces NGF as well as another factor immunologically distinct from NGF. It is of interest that both types of target explanted before they receive sympathetic innervation show evidence of producing NGF in culture.     

NGF and anti-NGF: evidence against effects on fiber growth in locus coeruleus from cultures of perinatal CNS tissues.

The present study examines whether the developing noradrenergic neurons of locus coeruleus depend on endogenous nerve growth factor (NGF) for nerve fiber production and if exogenous NGF stimulates fiber growth in this nucleus, using a collagen gel tissue culture technique. Lucus coeruleus from perinatal rat brain was used in three culture experiments: (1) lucus coeruleus, parietal cerebral cortex, and the superior cervical ganglion, prepared from newborn rats and cultured in different sectors of the same dishes; (2) locus coeruleus and parietal cerebral cortex from 17-day-old rat fetuses cultured in the same manner, and (3) locus coeruleus from 17-day-old rat fetuses co-cultured with spinal, sympathetic and ciliary ganglia from 8-day chick embryos. Experiments 1 and 2 were run with and without addition of NGF and anti-NGF, experiment 3 with and without anti-NGF. Total fiber production in all cultured tissues was evaluated daily by dark field and phase contrast microscopy during 4 days. Adrenergic nerve fiber production was then studied in the same locus coeruleus and superior cervical ganglia from the rats by Falck-Hillarp fluorescence histochemistry. Locus coeruleus and cortex cerebri from fetal rats produced dense fiber halos in culture. Locus coeruleus from newborn rats produced considerably less fibers, newborn cortex only few fibers. Superior cervical ganglia from the same newborn animals produced no or almost no fibers. Addition of NGF was not able to stimulate fiber growth in locus coeruleus nor in cortex cerebri as observed both in the living cultures and by fluorescence microscopy. Likewise, addition of anti-NGF did not affect fiber production in the CNS areas. The negative results with NGF on newborn locus coeruleus and cortex cerebri were in sharp contrast to the strong, highly significant fiber growth response demonstrated by the superior cervical ganglion from the same animals cultured in the same dishes. The third experiment tested whether locus coeruleus in tissue culture contained or produced nerve growth factor or any one of the three chick embryo ganglia. No response whatsoever in these three ganglia was observed. It is concluded that the developing locus coeruleus area does not contain or produce NGF, does not depend on NGF for fiber production, and is not stimulated by exogenous NGF.     

Nerve growth activities in rat peripheral nerve

Nerve growth activities in rat sciatic nerves were assayed by recording the neuritic outgrowth from chick embryonic ganglia cultured in collagen gels beside nerve fragments for two days. Living nerve explants released activity that resembled nerve growth factor (NGF) in its effect on sympathetic ganglia and that was almost totally blocked by an antiserum to 2.5 S mouse NGF. Frozen and thawed specimens from normal nerves elicited responses from sympathetic ganglia that were only partially suppressed by anti-NGF and also induced neuritic outgrowth from ciliary ganglia. Thus, from observations on normal nerves, at least two agents promoting axonal extension in vitro were deduced to exist; one substance similar to NGF plus another, non-NGF factor. The level of NGF-like activity was low in killed segments of normal nerves but higher in autologous nerve grafts and degenerating nerves two days after grafting or cutting. However, one or two weeks after nerve transection, distal nerve segments contained little nerve growth activity of either kind. Furthermore, when endoneurial fragments from chronically denervated stumps were cultured, they appeared to have lost some of their capacity to produce NGF-like activity in vitro although the production of activity had, if anything, increased in the perineurial region. In summary, rat peripheral nervous tissue releases two or more soluble substances that stimulate neuritic outgrowth. The level of one or both activities in the endoneurium can be altered by manipulation of nerves in vivo.     

Tropic interactions between sympathetic nerves and vascular smooth muscle

Explants of sympathetic ganglia from 3-9 day old rats were grown on collagen-coated coverslips in modified Rose chambers for 5 days, either alone or 2 mm away from explants of 6 week old rat caudal artery and aorta. Nerve fibre growth was stimulated on the side of the ganglion explant near the caudal artery explants but was not stimulated near the aorta. To determine the source of the nerve growth stimulation, explants of whole intact wall of the caudal artery, separated adventitia and medial layers, enzyme-dispersed cells, homogenized cells and medium pre-conditioned by caudal artery explants, were combined with ganglia. Explants of whole caudal artery and dispersed cells were also precultured prior to combination with ganglia. These combinations allowed analysis of the role of smooth muscle cells, existing nerve fibres, necrotic cells and connective tissue. Results suggested that degenerating nerve fibres within the blood vessels caused the increase in the number and the 'attraction' of the nerve fibres growing from the sympathetic ganglia. In contrast, both caudal artery and aorta from 3-9 day old rats caused stimulation of nerve fibre growth. Since these vessels were not yet innervated, the effect cannot be due to degenerating nerve terminals and a different mechanism must be involved.     

Characteristics of neurite outgrowth from rat spinal ganglia: effects of serum and segmental level

The outgrowth of neurites from spinal ganglia in tissue culture may be affected by many variables including medium constituents and the age of ganglia. If E15 rats were used, the rate of outgrowth from ganglion explants in response to nerve growth factor was independent of the cranial-caudal position from which the ganglia were derived. This contrasts with the cranial-to-caudal gradient present during embryonic development. The effect of serum on this outgrowth was also studied and ten of 17 lots of human placental serum, two of three lots of pooled human placental serum, and one of two lots of fetal calf serum significantly inhibited the outgrowth of neurites. This has relevance to the design of tissue culture studies and may be important in the interpretation of in vitro studies of disease mechanisms.     

Influence of nerve growth factor on developing dorso-medial and ventro-lateral neurons of chick and mouse trigeminal ganglia

Trigeminal ganglia have been removed from five, six, seven and eight day chick embryos and explants of the dorso-medial (DM) and ventro-lateral (VL) parts of the maxillomandibular lobe were grown in tissue culture. Quantitative methods were used to assess the influence of nerve growth factor (NGF) on fiber outgrowth from these explants. At all ages outgrowth from DM explants was significantly greater than from VL explants, the difference being most pronounced between the extreme DM and VL poles of the maxillomandibular lobe. These observations are interpreted as indicating the existence of two distinct populations of neurons in terms of their response to NGF rather than the consequence of the asynchronous differentiation and maturation of the VL and DM neurons. A similar study of 10, 11 and 12 day embryonic mouse trigeminal ganglia revealed no significant difference in neurite outgrowth between DM and VL regions grown in the presence or absence of NGF.     

Neuronal growth factors produced by adult peripheral nerve after injury

Dorsal root ganglion neurons from embryonic rats, co-cultured with endoneurial explants from transected, adult rat sciatic nerve, extended neurites in the absence of exogenous nerve growth factor (NGF). The effect was seen with endoneurial explants from normal adult sciatic nerves or from nerves which had been permanently transected up to 51 days prior to explantation. The rate of outgrowth decreased at 5 and 7 days and reached a minimum at 14 days after transection. A second phase of increased neurite-promoting activity appeared in 28-, 35-, 41- and 51-day posttransection tissue. The early phase, but not the late phase, was partially inhibited by antisera to NGF.     

Retinal ganglion cell response to axotomy and nerve growth factor antiserum treatment in the regenerating visual system of the goldfish (Carassius auratus): An in vivo and in vitro analysis

In vitro nerve growth factor (NGF) antiserum (anti-NGF) treatment was found to significantly depress retinal ganglion cell neurite outgrowth in goldfish explant culture. Goldfish retinas, conditioned by a 14-day prior optic nerve crush, demonstrated a significant dose response inhibition of neurite outgrowth if incubated with various concentrations of the antiserum (i.e. concentrations from full strength to 1:100) before explanation for tissue culture. NGF added to the incubation medium containing antiserum partially eliminated the inhibition of neurite outgrowth during the first 4 days of explant culture. Antiserum treatment at the higher concentrations (i.e. full strength and 1:1.5 dilution) caused a cessation of nerve growth from explants between culture days 3 and 4. However, controls at this time still exhibited vigorous neurite outgrowth.In vivo treatment with anti-NGF administered intraocularly at 7 days after optic nerve crush (i.e. 7 DPA) was found to significantly reduce the size and complexity of retinal ganglion cell nucleoli when analyzed morphometrically at 14 DPA. No other cell parameters measured (i.e. cell size, nuclear size, cell/nuclear ratios and mitochondrial, Golgi and RER densities) were found to be affected by the single antiserum treatment.     

Growth of adult rat superior cervical ganglion explants in serum-free media

Neurite outgrowth from explants of superior cervical ganglion from adult rats can be achieved in a serum-free medium Extensive neurite outgrowth occurred from ganglion explants maintained in Eagle's minimum essential medium supplemented with either 10% (v/v) fetal calf serum or 1% (w/v) bovine serum albumin and nerve growth factor. After one week in culture, the ATP content of explants maintained in the serum-free medium was slightly higher than that noted in explants cultured in the presence of fetal calf serum and amounts of phosphocreatine were significantly lower. Despite these differences in high energy phosphate content, the abundance and morphology of neuritic outgrowth were essentially the same from explants cultured in the two types of media.Comparable activities of a number of NADP⁺-dependent dehydrogenases were noted in explants maintained in the two types of media. Increases in the activities of the oxidative enzymes of the pentose pathway, which occur in axotomized ganglia in vivo, were observed in the cultured ganglion explants. NADP⁺-dependent isocitrate dehydrogenase activity remained constant in ganglion explants in vitro, and measurements of this activity were employed in a new method to quantitate neurite outgrowth. The activity of isocitrate dehydrogenase in lyophilized neuritic processes that had grown out onto a Millipore filter substrate correlated well with visual estimates of neuritic outgrowth. Substitution of delipidated for normal bovine serum albumin in the growth medium resulted in a significant decrease in neuritic outgrowth from ganglion explants from both adult and weanling rats. Addition of fatty acids to media containing delipidated bovine serum albumin enhanced neuritic outgrowth in explants of weanling rats. Thus, lipophilic substances bound to bovine serum albumin including fatty acids appear necessary for optimal growth of neurites from explants of the rat superior cervical ganglion.     

不同载体及年龄对大鼠视神经组织体外培养的影响

目的 探讨不同载体及年龄对大鼠视神经组织体外培养的影响. 方法 取出生4d和3月龄SD大鼠的视神经,分别使用鼠尾胶原玻片、多聚赖氨酸玻片和Bioeoat小室进行组织培养,观察组织生长情况.培养48 h后观察组织块贴壁率,培养5 d时使用图像分析仪测量细胞最大迁移距离.使用乳酸脱氢酶(LDH)试剂盒对Biocoat小室组不同年龄大鼠视神经组织培养液LDH活性进行动态比较.HE染色形态学观察,透射电镜观察组织块超微结构变化. 结果 Bioeoat小室组的组织贴壁率明显高于鼠尾胶原组、多聚赖氨酸组;在Bioeoat小室中培养的成年和新生大鼠视神经组织的细胞最大迁移距离均明显大于鼠尾胶原组和多聚赖氨酸组;在相同培养载体上.新生大鼠细胞迁移距离均明显高于成年大鼠,差异均有统计学意义(P<0.05).视神经组织培养液LDH活性在接种3 d后开始下降,成年大鼠LDH水平在培养9 d后再次上升.而新生大鼠在培养12 d时仍保持较低水平.视神经在培养早期即有细胞从组织块边缘游出,逐渐发出突起.新生大鼠较早出现细胞迁移.随着培养时间延长,组织出现结构紊乱和坏死.新生大鼠组织存活时间较成年大鼠明显延长. 结论 通过选择适宜的培养条件,视神经可以在体外较长时间存活.     

Fine structure of the superficial layers of the viper optic tectum. A golgi and electron-microscopic study

Peripheral nerve fiber outgrowth from developing spinal cord is proposed to be under the influence of the limb bud target which, at the time of nerve fiber invasion, is in an essentially premuscular, mesenchymal condition. Thus, the true target for elongating spinal nerve fibers in early development is mesenchyme rather than differentiated skeletal muscle. Spinal cord explants derived from stage V larval Rana pipiens were cultured in a defined medium in the presence or absence of mesenchymal limb tissue or limbconditioned medium (LCM). Analysis of quantified neuritic outgrowth under these conditions demonstrated a dependency on the target tissue for enhanced nerve fiber density and oriented growth. The characteristics of neuritic growth in the presence of limb mesenchyme or LCM changed from the relatively sparse and straight outgrowth of control cords to dense, wavy arborizations. Areas of the cord explants nearest the limb tissue exhibited the greatest increases in nerve fiber density and morphologic complexity. Additionally, an inverse relationship existed between growth enhancement and the cord-to-target distance. Regulation of directed nerve growth in vitro is suggested to result from a diffusible, target-originated growth factor that binds to the attachment substratum as a concentration gradient guidance pathway with implications for mechanisms of in vivo nerve growth.     

Comparative studies on the effect of the nerve growth factor on sympathetic ganglia and adrenal medulla in newborn rats

The effect of nerve factor (NGF) and nerve growth factor antiserum (NGF-AS) on sympathetic ganglia and adrenal chromaffin cells of newborn rats was investigated in a comparative study.     

Effect of derivatives of substance P dipeptide on nerve fiber growth in tissue culture

Explants and cells from the peripheral (PNS) and central nervous system (CNS) were cultivated in MAXIMOW chambers. The following four derivatives of Substances P (SP) dipeptide were tested for their effects in concentrations between 10(-4) and 10(-8) mol/l: Lys-Pro X 2HBr, Lys(Z)-Pro X HCl, Lys[Z(NO2)]-Pro X HCl and cyclo(Lys-Pro) X HCl. The results suggest that SP and SP derivatives may modulate nerve fibre outgrowth in vitro. In ganglion trigeminale explants Lys(Z)-Pro X HCl stimulated the growth of nerve fibres. The explant covered areas increased significantly. In telencephalon explants and cell cultures the tested derivatives did not essentially promote neurite outgrowth. In retinal cell cultures incubated with cyclo(Lys-Pro) X HCl and Lys[Z(NO2)]-Pro X HCl the neurite outgrowth index was stimulated in a different way. The neurite length was affected and stimulated in the presence of Lys(Z)-Pro X HCl and cyclo(Lys-Pro) X HCl. Both for survival and differentiation PNS neurons depend on specific biofactors. In sensory ganglia explants SP and especially SP derivatives were shown to improve neuronal survival provided that serum or embryonic extract was simultaneously present in the medium. Even if the dipeptides need other biofactors for their full function the may be discussed as neuronotrophic factors because they ensure survival and general growth capability of the responsive neuroblasts.     

Triiodothyronine Exerts a Trophic Action on Rat Sensory Neuron Survival and Neurite Outgrowth through Different Pathways

Apart from several growth factors which play a crucial role in the survival and development of the central and peripheral nervous systems, thyroid hormones can affect different processes involved in the differentiation and maturation of neurons. The present study was initiated to determine whether triiodothyronine (T₃) affects the survival and neurite outgrowth of primary sensory neurons in vitro. Dorsal root ganglia (DRG) from 19-day-old embryos or newborn rats were plated in explant or dissociated cell cultures. The effect of T₃ on neuron survival was tested, either in mixed DRG cell cultures, where neurons grow with non-neuronal cells, or in neuron-enriched cultures where non-neuronal cells were eliminated at the outset. T₃, in physiological concentrations, promoted the growth of neurons in mixed DRG cell cultures as well as in neuron-enriched cultures without added nerve growth factor (NGF). Since neuron survival in neuron-enriched cultures cannot be promoted by endogenous neurotrophic factors synthesized by non-neuronal cells, the increased number of surviving neurons was due to a direct trophic action of T₃. Another trophic effect was revealed in this study: T₃ sustained the neurite outgrowth of sensory neurons in DRG explants. The stimulatory effect of T₃ on nerve fibre outgrowth was considerably reduced when non-neuronal cell proliferation was inhibited by the antimitotic agent cytosine arabinoside, and was completely suppressed when the great majority of non-neuronal cells were eliminated in neuron-enriched cultures. These results indicate that the stimulatory effect of T₃ on neurite outgrowth is mediated through non-neuronal cells. It is conceivable that T₃ up-regulates Schwann cell expression of a neurotrophic factor, which in turn stimulates axon growth of sensory neurons. Together, these results demonstrate that T₃ promotes both survival and neurite outgrowth of primary sensory neurons in DRG cell cultures. The trophic actions of T₃ on neuron survival and neurite outgrowth operate under two different pathways.     

Influence of nerve growth factor on chick trigeminal motor nucleus explants

Explants of the metencephalic basal plate from stage 11 (40-hour) chick embryos containing the trigeminal (V) motor nucleus were cultured in standard control medium, in medium supplemented with nerve growth factor (NGF), in medium supplemented with NGF and specific antibodies to NGF (anti-NGF), and in medium supplemented with anti-NGF alone. The explants grown in the presence of NGF displayed an enhanced density and complexity of neuritic outgrowth, with this growth significantly surpassing that seen in the control group (p less than .001). The explants grown in NGF plus anti-NGF and those grown in anti-NGF alone did not differ from controls. The results indicate that this early cholinergic population is specifically responsive to NGF. This finding is consistent with recent studies in which NGF receptor binding has been found in this and other early brainstem and spinal cord motor neuron populations. The possible relevance of these observations to the normal sequence involved in the development of the V motor nucleus is discussed, particularly as they may relate to the relationship between the V ganglion and the developing V motor population.     

Effect of the preparation baliz-2 on the growth of cultures of sympathetic ganglia from various strains of rats

The effect of the baliz-2 drug (0.01%, 0.001%, 0.0001%) on the growth and differentiation of nervous tissue was studied on organotypic cultures of the sympathetic ganglia of newborn Wistar, Wag, August rats. The growth zone of explants in living cultures and histochemical stained preparations were analyzed for catecholamine-containing neurites. It has been established that control value of growth intensity for August ganglia is 2.1 2.0 times lower than for Wistar and Wag. The baliz-2 drug at concentration 0.001% and 0.0001% is able to stimulate fibre outgrowth from the explant. Mean values of the reaction are different: they are maximal for August rats (2.3-2.6 times higher than the control), while for Wistar and Wag rats they are 1.8-2.0 times higher than the control. These results provide evidence that the baliz-2 drug is a neurotrophic agent for sympathetic ganglia in culture. The results are discussed in terms of the role of the sympathetic-adrenal system in the nervous tissue regeneration.     

Postnatal development of conduction velocity and fibre size in the rat tibial nerve

The maximum conduction velocity (CV) and fibre diameters (D) were determined in the tibial nerve of developing rats. In 1-day-old rats CV of the fastest motor and sensory fibres (assessed separately) was 1.4 m/sec on the average and increased to 35 m/sec by postnatal day 30. The maximum conduction rate in adult rats ranged from 60 to 84 m/sec. Diameters of at least 100 nerve fibres in each age group were measured in electronmicrographs. The calibre of myelinating fibres in 1-day-old rats was 0.5–1.5 μm. By day 90 after birth the range of myelinated fibre size extended to 1.5–12.5 μm. The factor relating conduction rate and total fibre diameter of the largest fibres (i.e. the value of ) was found to vary with age, increasing from 1.1 to 6.2 between postnatal days 1 and 90. These results indicate that functional and morphological properties of peripheral nerve fibres in the rat undergo considerable changes during postnatal ontogeny until they reach adult values.     

Lymphoid tissues induce NGF-dependent and NGF-independent neurite outgrowth from rat superior cervical ganglia explants in culture

Induction of neurite outgrowth from superior cervical ganglia (SCG) by rat lymphoid tissues was studied using a tissue culture model. Neonatal rat SCG were cultured with 6–12-week-old rat thymus, spleen, or mesenteric lymph node (MLN) explants in a Martrigel layer, in defined culture medium without exogenous nerve growth factor (NGF). SCG were also co-cultured with neonatal rat heart (as positive control) or spinal cord (SC; as negative control). To determine whether inflammation affects the ability of lymphoid tissues to induce neurite outgrowth, we also examined MLN at various times after infecting rats with Nippostrongylus brasiliensis (Nb-MLN). In one series of experiments, a single lymphoid tissue explant was surrounded by four SCG at a distance of 1 mm. The extent of neurite outgrowth was determinded by counting the number of neurites 0.5 mm away from each ganglion at several time points. Adult thymus and, to a lesser extent, spleen had strong stimulatory effects on neurite outgrowth from SCG after 12 hr or more in culture. For thymus tissue, this was similar to the positive control heart explants. MLN from normal rats had minimal effect on neurite outgrowth; however, Nb-MLN showed a time-dependent enhancement of the neurite outgrowth, maximal at 3 weeks after infection. The relative efficacy of neurite outgrowth induction (heart ≥ thymus ≥ Nb-MLN ≥ spleen ≥ MLN ≥ SC) was confirmed in a second series of experiments where one SCG was surrounded by three different tissue explants. We then examined the role of 2.5S NGF, a well-known trophic factor for sympathetic nerves, in the lymphoid tissue-induced neurite outgrowth. Anti-NGF treatment of co-cultures of SCG and heart almost completely blocked the neurite outgrowth. Anti-NGF also significantly inhibited thymus- and spleen-induced neurite outgrowth, but not as effectively as heart-induced neuritogenesis (93,80, and 77% inhibition at 24 hr; 86,70, and 68% inhibition at 48 hr for heart, thymus, and spleen, respectively). On the other hand, anti-NGF inhibited only 8% of neurite outgrowth induced by 3-week post-infection Nb-MLN at 24 hr, and 41% at 48 hr. These data show that several adult rat lymphoid tissues exert neurotrophic/tropic effects. The predominant growth factor in thymus and spleen is NGF, while Nb-MLN produces factor(s) which is (are) immunologically distinguishable from NGF. These neurotrophic/tropic factors are produced during the reactive lymphoid hyperplasia that forms part of the inflammatory response against the nematode, N. brasiliensis. This suggests the possibility that cytokines produced by lymphocytes or other inflammatory cells may stimulate sympathetic neurite outgrowth in vivo. © 1994 Wiley-Liss, Inc.     

Unilateral sciatic nerve injury stimulates contralateral nerve regeneration.

Axonal outgrowth in tissue cultures was measured to determine whether unilateral peripheral nerve injuries affect contralateral nerve regeneration. The right sciatic nerves of young male Wistar rats were cut at mid-thigh level. Sham operation as a control was limited to the exposure of the nerve without cutting. At day 6 post-surgery, bilateral L5 dorsal root ganglia (DRG) with attached nerve stumps were resected and cultured. Axonal outgrowth from the nerve stump was measured in situ. The contralateral preparations showed longer outgrowths than controls. Therefore the conditioning effect was not merely restricted to the ipsilateral neurons but also affected undamaged sensory neurons of the contralaretal DRG.     

Subpopulations of sympathetic neurones differ in their sensitivity to nerve growth factor antiserum

Sympathetic postganglionic nerve fibres supplying mesenteric arteries and intrinsic ileal neurones differ in their characteristics of regeneration. Since the latter population of neurones occurs predominantly in prevertebral ganglia, which have been reported to be spared to some extent after treatment with antiserum to nerve growth factor (anti-NGF), we have investigated whether the two populations were differentially sensitive to anti-NGF. Newborn rats were treated daily for the first postnatal week with either anti-NGF or 154 mM NaCl solution. At 4 and 8 weeks of age, the presence of a functional sympathetic innervation to the mesenteric arteries and the gut was determined and correlated with the fluorescence histochemical demonstration of noradrenergic fibres. At both ages, stimulation of extrinsic sympathetic fibres caused an inhibition of gut motility, while the mesenteric arteries completely lacked a sympathetic innervation. Retrograde labelling of nerve cell bodies in control and antiserum treated rats confirmed that the sympathetic neurones supplying the ileal neurones were located in the prevertebral, superior mesenteric and coeliac ganglia and in the splanchnic ganglia lying along the greater splanchnic nerves. By interference from retrograde labelling in control animals, sympathetic neurones supplying the mesenteric arteries were present in all these ganglia, as well as in the thoracic and lumbar paravertebral sympathetic chains. The results suggest that two functionally distinct populations of sympathetic neurones, which overlap considerably in their distributions, are differentially sensitive to the immunological postnatal removal of NGF.