Effect of nerve growth factor and amizil on the viability and metabolism of cultured spinal ganglia

Spinal ganlia of a 9-day chick embryo were cultivated by the method of "floating rafts" in common medium (control) and in the medium containing amizyl (100 microgram/ml) or a neuregrowth factor (50 microgram/ml). With the action of amizyl there proved to be an increase in the number of surviving neurons; the majority of these neurons contained monoaminoxidase; there was a rise of NAD-diaphorase activity, and, to a lesser extent, of lactic dehydrogenase and isocitric dehydrogenase activities. The neurogrowth factor caused an increase in the number of nerve cells with acetylcholinesterase; there was an elevation of NAD-diaphorase and some rise of malic dehydrogenase activities; the activity of lactic dehydrogenase became maximal; as to succinic dehydrogenase--its activity was somewhat suppressed.     

Loss of nerve growth factor receptors in sympathetic ganglia from aged mice

[125I]-Nerve growth factor (NGF) binding was studied in superior cervical ganglia from mice 4-7 months and 24-27 months of age. Scatchard analysis demonstrated losses of both high and low affinity components of NGF receptor. These results indicate loss of NGF receptors may lead to the diminished responsiveness to NGF in aged sympathetic ganglion neurons.     

Macrophage dependence of peripheral sensory nerve regeneration: possible involvement of nerve growth factor.

The levels of NGF and NGF receptor mRNA, the degree of macrophage recruitment, and the ability of sensory and motor axons to regenerate were measured in C57BL/Ola mice, in which Wallerian degeneration following a nerve lesion is very slow. Results were compared with those from C57BL/6J and BALB/c mice, in which degeneration is normal. We found that in C57BL/Ola mice, apart from the actual lesion site, recruitment of macrophages was much lower, levels of mRNA for both NGF and its receptor were raised only slightly above normal, and sensory axon regeneration was much impaired. Motor axons regenerated quite well. These results provide in vivo evidence that macrophage recruitment is an important component of NGF synthesis and of sensory (but not motor) axon maintenance and regrowth.     

Physical properties of the detergent-extracted nerve growth factor receptor of sympathetic ganglia.

The nerve growth factor (NGF) receptor from microsomes of adult rabbit superior cervical ganglia has been solubilized with Triton X-100 and sodium deoxycholate. The physical properties of the detergent-extracted NGF receptor were assessed by Sepharose 6B chromatography and sucrose density gradient ultracentrifugation studies in H2O and D2O. The predominant form of the NGF receptor has a Stokes radius of 71 A, a partial specific volume of 0.74 ml/g, a sedimentation coefficient of 4.3 S, and a frictional ratio of 1.8. From these parameters, it can be calculated that the NGF receptor in Triton X-100 is a minimally hydrophobic, highly asymmetric, intrinsic membrane protein with a molecular weight of approximately 135,000. A form of the receptor with a sedimentation coefficient of 10.4 S was occasionally seen which appears to represent an aggregated form of the 4.3 S moiety.     

Effect of a bovine hypothalamic factor on the release and biosynthesis of growth hormone.

Partial purification of growth hormone (GH)-releasing factor (GRF) by acid extraction followed by gel filtration on Sephadex G-25 has been attained from bovine hypothalami. When rat pituitaries were incubated in 2 ml Krebs Ringer-bicarbonate-glucose (KRBG) medium, a stimulatory effect of the GRF fraction on immunoreactive GH (IR-GH) release was observed, while that of the factor neither on GH synthesis nor release of the synthesized GH was demonstrated. Stimulation of the GH release was exerted maximally within 30 min of incubation. Cycloheximide and actinomycin D, at a concentration which inhibited protein and RNA synthesis to less than 5 and 20% of the control, respectively, were without effect on the stimulatory action of the factor on GH release. On the other hand, stimulation of GH synthesis was observed under incubation in 0.3 ml medium with the factor and enhancing effect of the factor on the IR-GH release was undetectable. These results suggest that stimulation of the release and synthesis of GH mediated by the hypothalamic GRF fraction is under influence of the pool size of incubation media.     

Transforming growth factor beta has neurotrophic actions on sensory neurons in vitro and is synergistic with nerve growth factor.

Transforming growth factor beta (TGF beta) influences the growth and differentiation of a wide variety of nonneuronal cells (nnc) during embryogenesis and in response to wounding. In the present study TGF beta 1 and TGF beta 2 were examined for their neurotrophic actions on neonatal rat dorsal root ganglion (DRG) neurons with ganglionic nnc in dissociated cultures. TGF beta 1 and TGF beta 2 each increased both neuronal survival and levels of the peptide neurotransmitter substance P (SP) expressed per neuron as well as per culture. TGF beta 1 was maximally effective at a concentration of 40 pM, whereas TGF beta 2 was about 10-fold less potent. Survival effects promoted by simultaneous treatment with both factors were not additive. TGF beta 1 also changed the morphology and distribution of DRG nnc which resulted in clustering of DRG neurons on top of the nnc. Cotreatment of the cultures with two different anti-nerve growth factor (NGF) antibodies eliminated the neurotrophic effects of TGF beta 1. However, treatment with TGF beta 1 did not alter NGF mRNA expression in the cultures nor did it change the amount of NGF in the medium. Further, TGF beta 1 greatly enhanced survival effects and SP stimulation promoted by exogenous NGF at concentrations up to 100 ng/ml. The neurotrophic effects of TGF beta 1 were significantly attenuated by decreasing the proportion of the ganglionic nnc, suggesting a role for these cells in mediating TGF beta 1 action on the neurons. It is hypothesized that the neurotrophic activity of TGF beta depended upon the presence of molecules immunologically related to NGF and that the effects of TGF beta were synergistic with NGF. These observations suggest that TGF beta may play a role in the differentiation and regeneration of DRG neurons in vivo.     

Presence and disappearance of nerve growth factor receptors on sensory neurons in culture

The age-dependent presence of nerve growth factor (NGF) receptors on neurites of sensory neurons of dorsal root ganglia removed from chicks of different embryonic ages and subsequently kept in culture with NGF and/or brain extract has been investigated by autoradiography. Most of the neurons removed at embryonic Day 10 (E10), (82–95%) can be labeled with iodinated NGF, irrespective of whether they are selected for survival by means of NGF, brain extract, or both. However, when neurons are isolated from E16 chicks and maintained in culture with brain extract, only about 28% of the neurons have NGF receptors at reduced density. This percentage is higher than that expected if the number of neurons surviving with NGF would be exactly correlated with the number of neurons displaying NGF receptors: at E16 only about 5% of the neurons survive with NGF alone. In order to determine if the decrease in the number of neurons displaying NGF receptors also occurs in vitro, E10 neurons were cultured for various periods of time either with NGF or brain extract. Most of the neurons grown with NGF do not lose their NGF receptors. In contrast, the majority of the neurons grown in the presence of brain extract lose their receptors: after 6 days in culture, only about 25% of the neurons can be labeled with NGF. It is concluded that in vitro, a maturation with regard to the NGF receptor occurs in the presence of brain extract similar to that observed in vivo.     

Prediction of the secondary structure of mouse nerve growth factor and its comparison with insulin

Five secondary structure prediction methods based on amino acid sequence have been used to predict the secondary structure of mouse nerve growth factor (NGF). The regions predicted helical $\text{do}\text{not}$ correlate well with the proposal, based on the alignment of primary sequences, that the NGF peptide chain is structurally and evolutionarily related to proinsulin.     

Ionic behaviors and nerve growth factor dependence in developing chick ganglia. II. Studies with neurons of dorsal root ganglia

Using intact dorsal root ganglia (DRG) from embryonic (E) chick and measuring 22Na+ accumulation, the authors have recently shown that (i) ionic control by the ganglia has a complete requirement for exogenous NGF between E6 and E10, and (ii) control of ion pump mechanisms independent of exogenous NGF is progressively acquired by these ganglia from E10 to E16. Similar experiments have now been carried out using enriched suspensions of ganglionic neurons to test whether the acquisition of endogenous control by older ganglia was (1) due to the close association between neurons and nonneurons, and (2) correlated with a decreasing need by these neurons for exogenous NGF for survival in culture. In this enriched neuronal population, Na+ accumulation in the absence of NGF increases from E7 to E10, paralleling the increase in Na+ accessible space under ouabain, but then decreases conspicuously between E10 and E16, despite little change in the ouabain-sensitive Na+ space. NGF prevents Na+ accumulation during the early period, and becomes increasingly irrelevant for this behavior in later (after E10) development. K+ movements (traced with 86Rb+) behaved similarly. Active K+ influx (Na+, K+-pump mediated) also increases severalfold between E7 and E10. This K+ influx is sensitive to NGF at E7 and E10 but not at E14, paralleling the observed Na+ and K+ behaviors. These data suggest that the control of Na+, K+-pump performances acquired by these neurons between E10 and E16 represents the development of a neuronal self-sufficiency. This increase in ionic control is not due to an increase in pump molecules or pumping efficiency. No increases in the binding of [3H]ouabain or active K+ influx occur between E10 and E16, when ionic control is developing. The ionic dependence on NGF by the DRG neurons changes with their developmental age along the same temporal pattern displayed by their survival response to NGF in culture.     

Gestational changes in calcitonin gene-related peptide, nerve growth factor, and its receptors in rat dorsal root ganglia.

In dorsal root ganglia (DRG) cell cultures, levels of calcitonin gene-related peptide (CGRP) are increased in the presence of ovarian hormones and nerve growth factor (NGF). In addition, injection of ovariectomized rats with ovarian hormones led to an increase in levels of two NGF receptors, TrkA and p75(NTR), in DRG. Thus, we hypothesized that increased levels of ovarian hormones during pregnancy may elevate the synthesis of CGRP and NGF receptors in the DRG. DRG harvested from rats on specific days of pregnancy, on Day 2 postpartum, and after ovariectomy were subjected to radioimmunoassay, Western blot analysis, and NGF immunoassay to determine levels of CGRP, TrkA and p75(NTR), and NGF, respectively. CGRP levels in rat DRG were significantly higher during pregnancy than at Day 2 postpartum or in ovariectomized rats. Levels of both TrkA and p75(NTR) in DRG increased during pregnancy and remained elevated at Day 2 postpartum, but CGRP levels declined. Levels of NGF reached a statistically significant peak at Day 18 of gestation, and were not significantly reduced at Day 2 postpartum. Increased levels of ovarian steroid hormones during pregnancy may be involved in the synthesis of CGRP, however, the postpartum decreases in CGRP synthesis appear to be unrelated to NGF and its receptors.     

Effect of nerve growth factor on lesioned PC12 cells.

The protecting effect of nerve growth factor (NGF) from hydrogen peroxide was studied on PC12 cells conditioned at 1 mM hydrogen peroxide with NGF and without NGF in comparison with cells treated with neither hydrogen peroxide nor NGF. NGF treatment of PC12 cells increased significantly the activity of catalase representing induction of free radical detoxifying mechanisms. The protection effect of NGF was reflected also on enhanced activities of choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) in the cells.     

steve bAccumulation of nerve growth factor and its receptors in the uterus and dorsal root ganglia in a mouse model of adenomyosis

Background  

Adenomyosis is a common gynecological disease, which is accompanied by a series of immunological and neuroendocrinological changes. Nerve growth factor (NGF) plays a critical role in producing pain, neural plasticity, immunocyte aggregation and release of inflammatory factors. This study aimed to investigate the expression of NGF and its two receptors in uteri and dorsal root ganglia (DRG) in an adenomyosis mouse model, as well as their relationship with the severity of adenomyosis.