Influence Of Nerve Growth Factor Research Articles

Influence of nerve growth factor on neurofilament gene expression in mature primary sensory neurons

To analyze the possible influence of nerve growth factor (NGF) on neurofilament synthesis in primary sensory neurons, adjacent cryostat sections of lumbar dorsal root ganglia (DRG) from adult rats were processed for either NGF-receptor radioautography or in situ hybridization with a neurofilament cDNA probe. Labeling by both procedures was quantified with computer assistance for approximately 300 neurons in each of selected ganglia. For uninjured neurons, no correction was detected between NGF binding and neurofilament mRNA, even after infusion of NGF into the lumbar subarachnoid space for 1 week. One or 3 weeks after sciatic nerve transection, neurofilament labeling densities in large DRG neurons were sharply reduced and the normal bimodal pattern in frequency histograms had become unimodal. Intrathecal infusion of NGF counteracted this injury-induced reduction of neurofilament mRNA but only in neurons with high-affinity NGF receptors. To explain the effects of NGF on axotomized neurons and the normal diversity of neurofilament gene expression among neurons with NGF receptors, we postulate that NGF permits NGF-sensitive DRG neurons to respond differentially to a second factor stimulating neurofilament synthesis.

Characterization of sodium currents in mammalian sensory neurons cultured in serum-free defined medium with and without nerve growth factor

The influence of nerve growth factor (NGF) on Na currents of rat dorsal root ganglia (DRG) was studied in neurons obtained from newborns and cultured for 2-30 hr in serum-free defined medium (SFM). Cell survival for the period studied was 78-87% both with and without NGF. Na currents were detected in all cells cultured for 6-9 hr. They were also detected after 2 hr in culture in 21.5% of the cells cultured without NGF (-NGF cells), and in 91.5% of the cells cultured with NGF (+NGF cells). Current density of the -NGF cells was 2.3 and 2 pA/microns 2 after growth for 2 and 6-9 hr, respectively, compared to 3.0 and 3.9 pA/microns 2 for the +NGF cells. The +NGF cells were separated into fast (F), Intermediate (I) and slow (S) cells, based on the Na current they expressed, while -NGF cells were all of the I type. F, I and S currents differed in their voltage-dependent inactivation (Vh50 = -79, -28 and -20 mV), kinetics of inactivation (tau h = 0.55, 1.3 and 7.75 msec), and TTX sensitivity (Ki = 60, 550 and 1100 nM). All currents were depressed by [Ca]0 with a KdCa of 22, 17 and 8 mM for F, I and S currents, respectively. Current density of F and S currents was 5.5 and 5 pA/micron 2 for the I current. The concentration-dependent curve of I current vs. TTX indicated that I current has two sites: one with F-like and another with S-like Ki for TTX. Hybridization of F and S currents yielded I-like currents. Thus, the major effect of NGF on Na currents in SFM is the acceleration of Na current acquisition and diversity, reflected in an increase of either the S or F type in a cell.

Distribution of nerve growth factor receptor-like immunoreactivity in the adult rat central nervous system. Effect of colchicine and correlation with the cholinergic system—II. Brainstem, cerebellum and spinal cord

Multiple nuclei and fiber tracts in the adult rat brainstem and spinal cord were found to contain nerve growth factor receptor-related protein, as recognized by the monoclonal antibody 192-IgG. Both cholinergic and non-cholinergic sensory and motor regions demonstrated immunoreactive cell bodies and fibers. Nerve growth factor receptor-immunoreactive cells were seen in the mesencephalic nucleus of trigeminal nerve, superior colliculus, parabrachial, prepositus hypoglossal, raphe, dorsal and ventral cochlear, interstitial nucleus of the vestibular nerve, ambiguus and reticular nuclei, cerebellum and ventral spinal cord. Immunoreactive cells resembling neuroglia were distributed subpially along the superior colliculus. Intracerebroventricular injection of colchicine resulted in significantly increased nerve growth factor receptor immunoreactivity in all previously positive neurons and especially in certain neurons of the cochlear and ambiguus nuclei. It also resulted in the visualization of receptor immunoreactivity in certain neurons which were normally non-immunoreactive including cerebellar Purkinje cells, neurons of the central gray, locus coeruleus, facial, dorsal motor vagal and hypoglossal nuclei. In normal animals, nerve growth factor receptor-immunoreactive fibers and varicosities occurred in the trigeminal nerve nuclei, pontine, vestibular, parabrachial, facial, hypoglossal, dorsal motor vagal, solitary, gracile and cuneate nuclei and spinal cord. Although most fiber-like immunoreactive structures were probably axons and nerve terminals, neuroglial or extracellular localizations could not be excluded in some areas. For example, the medial nucleus of the inferior olive and most cerebellar nuclei contained diffuse non-fibrillar receptor immunoreactivity. The presence of nerve growth factor receptor-like immunoreactivity in cell bodies and fibers of several sensory and motor areas of the adult rat brainstem, cerebellum and spinal cord suggests multifocal actions of nerve growth factor or a nerve growth factor-like substance. Although the degree of overlap between nerve growth factor receptor- and choline acetyltransferase-containing regions in the brainstem is not as great as in the forebrain, our findings suggest a potential influence of nerve growth factor or nerve growth factor-like substances on cholinergic systems outside the forebrain. Furthermore, the disparities which occur imply that non-cholinergic nerve growth factor receptor-containing neurons of the brainstem, cerebellum and spinal cord may be affected by such trophic substances.

The influence of nerve growth factor on the activities of adenylate cyclase and high-affinity GTPase in pheochromocytoma PC12 cells

The influence of nerve growth factor (NGF) on the activities of adenylate cyclase and high-affinity GTPase in pheochromocytoma PC12 cells was studied. Incubation of cells with nerve growth factor led to a rapid activation of adenylate cyclase accompanied by an inhibition of high-affinity GTPase. By the 10th min of incubation the activity of adenylate cyclase had been reduced 2-fold when compared to the control. The activity of GTPase, however, increased. No significant changes in the cAMP level were detected. The data obtained indicate that NGF interaction with PC12 cells induces changes in the adenylate cyclase system and this process involves G-proteins that regulate the adenylate cyclase activity.

Facial nerve regeneration in the silicone chamber: The influence of nerve growth factor

The role of nerve growth factor (NGF) was examined in the neural repair of adult rabbit facial nerves using an in vivo preparation. A 35-μl nerve growth chamber was created by suturing the proximal and distal ends of a transected facial nerve (superior buccal branch) into a silicone tube. A gap of 8 mm in the chamber remained after removal of a 5-mm piece of nerve and insertion of the proximal and distal stumps into the tube. Animals were operated bilaterally; one side of the chamber was filled with NGF and the contralateral side was filled with Ringer's solution. Regeneration of the nerves was examined 1 to 5 weeks following the surgery. The caliber of the nerve bundle, the distribution pattern of regenerating motoneurons, axon number per fascicle, size distribution, and the total number of cells were compared to the preoperative morphology pattern found for that animal. Each buccal branch served as its own control. The NGF-filled chambers demonstrated an overall larger caliber of nerve regeneration at 5 weeks and a higher density distribution of axon growth at 3 and 5 weeks. In the early regeneration case (3 weeks), the axon growth profile exhibited more fascicles and less axons than the preoperative controls. In the more advanced stage (5 weeks), the fascicle number was reduced and the axon number was increased. After 5 weeks of regeneration the number of fascicles was still more than that found in the preoperative state. Axon size at 5 weeks was 80% that of the preoperative controls and the thickness of the myelin sheath was less than the preoperative level. The histogram of the size distribution revealed the same distribution as in the preoperative control section.

Comparison of nerve growth factor's effects on development of septum, striatum, and nucleus basalis cholinergic neurons in vitro.

In the central nervous system, nerve growth factor (NGF) affects basal forebrain cholinergic neurons during early development and in the adult mammalian brain. These neurons are located in medial septum, diagonal band of Broca, and nucleus basalis of Meynert. While the effects of NGF on the development of septal cholinergic neurons are well documented, only little is known about the influence of NGF on development of cholinergic neurons in the nucleus basalis. In addition to the basal forebrain cholinergic neurons, there are cholinergic interneurons in the corpus striatum, which form an anatomically and functionally distinct population of cholinergic neurons. These striatal interneurons have been reported to respond to NGF during early development; however, it is not known whether the effects of NGF on their development are similar to those on septal cholinergic neurons. We prepared cultures of dissociated cells from fetal rat septum, striatum, and nucleus basalis and investigated the development of cholinergic neurons localized in these three different areas in the presence or absence of NGF. We now report that, first, cholinergic neurons of striatum and nucleus basalis develop a more extensive fiber network and contain more acetylcholinesterase (AChE) per neuron than do cholinergic neurons of septum. The amount of choline acetyltransferase (ChAT) per cholinergic neuron is approximately the same in all three culture types when grown in the absence of NGF. Second, NGF treatment increases and anti-NGF treatment decreases the number of AChE-positive neurons in cultures of low plating density, suggesting that NGF is able to promote survival of cholinergic neurons of all three areas studied. Third, NGF increases the total length of fibers and the number of branching points of cholinergic neurons in septal cultures but not in cultures of striatum and nucleus basalis. Fourth, NGF treatment increases AChE activity in septal but not in nucleus basalis or striatal cultures, suggesting that AChE activity reflects the extent of the fiber network of cholinergic neurons of all areas. Fifth, NGF treatment produces severalfold elevations in ChAT activity in septal cultures and more modest increases in cultures of nucleus basalis and striatum, suggesting that NGF is able to stimulate ChAT activity also in the absence of a stimulatory effect on survival and fiber growth. Our results demonstrate that, during early development, NGF is able to affect survival and differentiation of all three populations of forebrain cholinergic neurons.(ABSTRACT TRUNCATED AT 400 WORDS)

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.

The influence of nerve growth factor on the in vitro proliferative response of rat spleen lymphocytes.

Splenic lymphocytes and accessory cells express receptors for nerve growth factor (NGF), a well-characterized neurotropic peptide that influences the development and survival of neuronal elements in the central and peripheral nervous systems. In the present study, we report that when rat splenic mononuclear cells (MC) are incubated in the presence of NGF, a dose-dependent increase in DNA synthesis occurs during 96-120 hours of culture as measured by 3H thymidine (3H-Thd) uptake. The minimal molar concentration of NGF at which the increased proliferative response of the cells (3.7 nM) is seen corresponded to the equilibrium disassociation binding constant of the MC (Kd = 2.5 nM), suggesting that the response was a consequence of receptor-ligand interaction. In addition, NGF was able to potentiate the lymphoproliferative response to several T-cell and B-cell mitogens. Significantly increased 3H-Thd uptake by NGF-stimulated cells was noted for concanavalin A (Con A), phytohemagglutinin (PHA), and lipopolysaccharide (LPS), particularly at suboptimal dosages of mitogen. Thus it appears that the NGF receptors on rat splenic MC are physiologically relevant and that NGF can modulate proliferation of T- and B- cells.

Peptide expression is altered when afferent nerves reinnervate inappropriate tissue

Neuropeptides are found in specific subpopulations of primary afferent neurones. Peptide expression can be altered following axotomy or under the influence of nerve growth factor. Here we have examined the consequence of altering the peripheral target of afferent neurones. Many unmyelinated afferents from skin contain substance P-like immunoreactivity (SPLI) whilst those from muscle do not. We have found that fibres will innervate inappropriate tissue types. We have therefore cut and cross-anastomosed a skin and muscle hindlimb nerve in the rat and 10 12 weeks later analysed the regenerated nerves immunocytochemically for SPLI. Muscle afferents inappropriately reinnervating skin were found to contain many SPLI fibres in contrast to control nerves resutured to their own distal stumps. Conversely, skin afferents made to innervate muscle showed reduced levels of peptide staining. These results demonstrate the plasticity of peptide expression and suggest that factors in peripheral tissue or perhaps distal nerve sheaths exert a trophic influence on nervous system function.

Influence of nerve growth factor and thyroid hormone on the development of subcortical cholinergic cells in vitro

Influence of nerve growth factor and thyroid hormone on the development of subcortical cholinergic cells <i>in vitro</i>

Short-latency local actions of nerve growth factor at the growth cone.

Cultures of neurite-bearing pheochromocytoma (PC12) cells and of sympathetic neurons have been examined by time-lapse video microscopy. In the presence of nerve growth factor (NGF), the neurites of such cultures elongated and their growth cones changed geometry, via microspike and lamellipodial motion, on a time scale of minutes. Withdrawal of NGF caused process extension to cease and a progressive reduction in growth-cone area as a result of retraction of lamellipodia and microspikes. By approximately equal to 4 hr after NGF withdrawal, most neurite tips were smooth sided, devoid of conical expansions at their termini, and virtually immobile. Addition of NGF to cultures from which it had been withdrawn induced motion of microspikes and projections from the upper surface of growth cones within 2 min, while lamellipodial spreading and neurite reextension were induced after approximately equal to 20 min. For PC12 cells, these responses to replacement of NGF could not be mimicked by addition of dibutyryl cAMP (less than or equal to 2 mM) or the Ca2+ ionophore A23187 (less than or equal to 5 microM) to NGF-deprived cultures nor inhibited by the presence of EGTA (less than or equal to 2 mM) or calcium antagonists in the culture medium. Since neurite fragments formed by transection of processes of PC12 cells deprived of NGF responded to its replacement in a manner similar to intact neurites, it is concluded that the effects are focal to the neurite and growth cone and independent of the cell body. This influence of NGF on growth-cone shape and motility represents short-term local activation of this structure and has significance for control of neurite extension.

Replication of thymidine kinase deficient herpes simplex virus type 1 in neuronal cell culture: Infection of the PC 12 cell

Replication of a thymidine kinase deficient (TK-) mutant of herpes simplex virus type 1 (HSV-1) was compared to replication of its parental TK+ strain in the PC 12 cell. This is a cell which ceases cell division and undergoes neuron-like morphological and physiological differentiation in the presence of nerve growth factor (NGF). No difference between mutant and parental strain replication was detected either when these cells were infected in the proliferative state or while maintained under the influence of NGF. Neither viral TK nor enhanced cellular TK activity was detected during TK- HSV-1 replication, which proceeded in the presence of selective antiviral drugs that inhibited TK+ HSV-1 viral replication. Moreover, thymidylate synthetase was inhibited early in TK- infection, and reutilization of thymine nucleotides derived from degraded cellular DNA was not detected. Under the conditions of these in vitro studies, increased production of dTTP as a result of enhanced TK activity did not appear to be rate-limiting, despite the non-dividing "differentiated" state of the PC 12 cell.

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.

Glial-released proteins: III. Influence on neuronal morphological differentiation

In vitro model systems in neurobiology are available to detect and help characterize various intercellular developmental signals. The presence of such active substances in conditioned medium (CM) from rat C6 glioma cells (2BD clone) was examined using the pheochromocytoma-derived clonal cell lines PC12 and PCG2. Conditioned medium from C6 monolayers induces neurite outgrowth in PC12 cells and extensive alteration in PCG2 cell morphology by 24 h. This ability of CM was found (1) to remain after dialysis and lyophilization, (2) to be modulated by steroid treatment of C6 monolayers (PC12 cells only and (3) to be distinct from the influence of NGF. Combinations of CM, nerve growth factor (NGF) and dibutyryl cyclic AMP when given together in pairs to PC12 or PCG2 cells revealed facilitation of neurite formation and cell morphology, respectively. These results suggest possible synergistic interaction between the three factors in the neuroglial regulation of neuronal morphological differentiation.

Influence of nerve growth factor on the embryonic mouse trigeminal ganglion in culture.

Developing trigeminal ganglia have been excised from mouse embryos of 9, 10, 11 and 12 days gestation and grown in tissue culture. A quantitative method was used to assess the effect of nerve growth factor (NGF) and an antiserum to NGF (anti-NGF) on fiber outgrowth from the explanted ganglia. Fiber outgrowth from 9-day ganglia (E9 ganglia) appeared to be unaffected by the presence of NGF. However, the ganglia became increasingly responsive to NGF from embryonic day 10 through 12. The increased responsiveness to NGF in vitro coincided with the establishment of fiber contacts with peripheral target tissues in vivo. Anti-NGF produced no significant reduction of fiber outgrowth from E9 ganglia, but from later ganglia significant reduction were observed.

The influence of nerve growth factor, potassium depolarization and dibutyryl (cyclic) adenosine 3′,5′-monophosphate on explant cultures of chick embryo sympathetic ganglia

Optimal nerve growth factor (NGF) and potassium concentrations for the culture of explants of single chick embryo sympathetic ganglia in Leighton tubes are described. NGF increases ganglionic monoamine oxidase (MAO; E.C.1.4.3.4) activity in a dose dependent fashion with maximum effects at a concentration of about 8 U/ml NGF. Peak activity compares closely with values seen in chick ganglia in vivo. Potassium-induced depolarization (45 mM K-+) also increases MAO activity; but the extent of the increase depends upon (NGF concentration, for there is little or no increase at high NGF concentrations. Dibutyryl adenosine 3',5'-cyclic monophosphoric acid (db cyclic AMP) can mimic the potassium-induced increase in MAO.

Factors influencing the development of the sympathetic nervous system: trans-synaptic regulation and the influence of nerve growth factor.

Factors influencing the development of the sympathetic nervous system: trans-synaptic regulation and the influence of nerve growth factor.