Infusion Of Nerve Growth Factor Research Articles

Intracerebroventricular Infusion of Nerve Growth Factor in Three Patients with Alzheimer’s Disease

Nerve growth factor (NGF) is important for the survival and maintenance of central cholinergic neurons, a signalling system impaired in Alzheimer’s disease. We have treated 3 patients with Alzheimer’s disease with a total of 6.6 mg NGF administered continuously into the lateral cerebral ventricle for 3 months in the first 2 patients and a total of 0.55 mg for 3 shorter periods in the third patient. The patients were extensively evaluated with clinical, neuropsychological, neurophysiological and neuroradiological techniques. Three months after the NGF treatment ended, a significant increase in nicotine binding was found in several brain areas in the first 2 patients and in the hippocampus in the third patient as studied by positron emission tomography. A clear cognitive amelioration could not be demonstrated, although a few neuropsychology tests showed slight improvements. The amount of slow-wave cortical activity as studied by electroencephalography was reduced in the first 2 patients. Two negative side effects occurred with NGF treatment: first, a dull, constant back pain was observed in all 3 patients, which in 1 patient was aggravated by axial loading resulting in sharp, shooting pain of short duration. When stopping the NGF infusion, the pain disappeared within a couple of days. Reducing the dose of NGF lessened the pain. Secondly, a marked weight reduction during the infusion with a clear weight gain after ending the infusion was seen in the first 2 patients. We conclude from this limited trial that, while long-term intracerebroventricular NGF administration may cause certain potentially beneficial effects, the intraventricular route of administration is also associated with negative side effects that appear to outweigh the positive effects of the present protocol. Alternative routes of administration, and/or lower doses of NGF, perhaps combined with low doses of other neurotrophic factors, may shift this balance in favor of positive effects.

Uninjured aged sympathetic neurons sprout in response to exogenous NGF in vivo

The extent to which the loss of plasticity by aged neurons is due to changes in the neuronal environment or to a loss of growth potential of the neurons has not been determined. In previous studies we observed that young adult cerebrovascular axons undergo a sprouting response following a 2-week intracerebroventricular infusion of nerve growth factor (15 μg; NGF). The present study used electron microscopy to examine the innervation of the intradural segment of the internal carotid artery of the aged rat and to determine whether aged sympathetic perivascular axons would respond to in vivo infusion of NGF. Young adult and aged Fischer 344 female rats received a 2-week intracranial infusion of NGF (15 μg) or vehicle (VEH) and were perfused for electron microscopy. Although there was no change in the total number of perivascular axons associated in aged VEH when compared with young adult VEH, a significant reduction was observed in aged VEH when total axons and sympathetic axons were expressed per μm 2 vascular wall, reflecting an age-related increase in blood vessel size. Following NGF infusion, aged sympathetic axons were significantly increased by 192% compared with aged VEH cases. These results suggest that there is a proportional reduction in sympathetic cerebrovascular neurons with aging but that they exhibit robust sprouting in response to an exogenous neurotrophin.

NGF delays rather than prevents the cholinergic terminal damage and delayed neuronal death in the hippocampus after ischemia

Cerebral ischemia induces damage of cholinergic terminals in the hippocampus, which preceded the delayed neuronal death (DND) of the CA1 pyramidal cells. We investigated the effects of nerve growth factor (NGF) on the cholinergic terminal damage after ischemia. Continuous NGF infusion (0.5 μg/7 days) into the lateral ventricle before and after 5 min ischemia prevented a decrease in choline acetyltransferase (ChAT)-immunoreactivity and disturbance of acetylcholine (ACh) release on the 4th day after ischemia, but not on day 7, i.e., NGF infusion caused delay in the progress of the cholinergic terminal damage. These findings show that the cholinergic terminal damage may result from deficiency of endogenous NGF in an ischemic brain. In addition, we investigated whether NGF would prevent the DND after ischemia. NGF infusion also caused delay in the progress of the DND until day 14. Our results suggested that the neuroprotective effect of NGF on the DND may be secondarily yielded by maintenance of communication between cholinergic terminal and the target CA1 cell, and that prevention of cholinergic terminal damage may be useful for the treatment of cerebrovascular disease.

Improvement of Cognitive Deficits and Decreased Cholinergic Neuronal Cell Loss and Apoptotic Cell Death Following Neurotrophin Infusion after Experimental Traumatic Brain Injury

✓ This study explores the effects of infusion of nerve growth factor (NGF) on behavioral outcome and cell death in the septal region using the clinically relevant model of fluid-percussion brain injury in the rat. Animals were subjected to fluid-percussion brain injury and 24 hours later a miniosmotic pump was implanted to infuse NGF (12 animals) or vehicle (12 animals) directly into the region of maximum injury for 2 weeks. Four weeks postinjury the animals were tested for cognitive function using a Morris Water Maze paradigm. Neurological motor function was evaluated over a 4-week postinjury period. The rats receiving NGF infusions had significantly higher memory scores than vehicle-treated animals. Examination of the cholinergic neurons in the medial septal region using choline acetyltransferase immunohistochemistry demonstrated significant cell loss after injury. Infusion of NGF significantly attenuated loss of these cholinergic neurons. A second group of animals was subjected to fluid-percussion brai...

Characterization of a prolonged regenerative attempt by diffusely injured axons following traumatic brain injury in adult cat: a light and electron microscopic immunocytochemical study.

Traumatic brain injury in animals and humans is well known to cause axonal damage diffusely scattered throughout the brain without evidence of other brain parenchymal change. This observation has prompted some to posit that such damaged axons are well positioned to mount a regenerative attempt. The present study uses an immunocytochemical marker specific for regenerating neurites to explore this issue. Further, in an attempt to expedite and enhance any potential regenerative effort, this study evaluates the efficacy of intrathecally applied nerve growth factor. Three sets of experiments were performed in adult cats. One group of animals was subjected to moderate fluid percussion brain injury and followed for 7 or 14 days post injury, with the continuous intraventricular infusion of nerve growth factor delivered by implanted osmotic pumps. These animals were compared to a second group of time-matched, sham-operated animals receiving artificial cerebrospinal fluid infusion. To assess axonal damage immunohistochemical staining for the low molecular weight neurofilament subunit (NF-L) was carried out using an NR4 monoclonal antibody. To localize axons exhibiting a regenerative response immunohistochemical staining for the growth associated protein GAP43 was employed. In sham controls, at the light microscopic level NF-L-immunoreactive axonal swellings were numerous at 7 days, but by 14 days post injury their frequency declined markedly. In contrast, GAP43-immunoreactive, disconnected reactive axonal swellings were rarely observed at 7 days but were numerous at 14 days. Ultrastructural analysis at 14 days post injury of carefully matched sections revealed reactive axons demonstrating sprouting consistent with a regenerative effort. Analysis of tissue from animals of 14 days of survival indicated that supplementation with nerve growth factor did not appear to enhance the capacity of damaged brain axons to mount a regenerative attempt. Rather, it appears that regenerative efforts seen reflect a spontaneous response. A third group of adult cats, subjected to the same injury but not subjected to osmotic pump implantation, was allowed to survive for 22-28 days. Animals in this group also demonstrated GAP43 immunoreactivity in reactive axonal swellings in the brain stem. This study demonstrates that diffusely injured axons can mount a sustained regenerative attempt that is associated with a reorganization of their cytoskeleton and accompanied by an up-regulation of growth-associated proteins.

Disruption of a Single Allele of the Nerve Growth Factor Gene Results in Atrophy of Basal Forebrain Cholinergic Neurons and Memory Deficits

Administration of nerve growth factor (NGF) to aged or lesioned animals has been shown to reverse the atrophy of basal forebrain cholinergic neurons and ameliorate behavioral deficits. To examine the importance of endogenous NGF in the survival of basal forebrain cholinergic cells and in spatial memory, mice bearing a disruption mutation in one allele of the NGF gene were studied. Heterozygous mutant mice (ngf+/-) have reduced levels of NGF mRNA and protein within the hippocampus and were found to display significant deficits in memory acquisition and retention in the Morris water maze. The behavioral deficits observed in NGF-deficient mice were accompanied by both shrinkage and loss of septal cells expressing cholinergic markers and by a decrease in cholinergic innervation of the hippocampus. Infusions of NGF into the lateral ventricle of adult ngf+/- mice abolished the deficits on the water maze task. Prolonged exposure to NGF may be required to induce cognitive effects, because reversal of the acquisition deficit was seen after long (5 weeks) but not short (3 d) infusion. Although NGF administration did not result in any improvement in the number of septal cells labeled for choline acetyltransferase, this treatment did effectively correct the deficits in both size of cholinergic neurons and density of cholinergic innervation of the hippocampus. These findings demonstrate the importance of endogenous NGF for survival and function of basal forebrain cholinergic neurons and reveal that partial depletion of this trophic factor is associated with measurable deficits in learning and memory.

Chronic intraventricular infusion with NGF improves LTP in old cognitively-impaired rats

Aged (21 months) cognitively-impaired male Sprague–Dawley rats received intraventricular infusion of nerve growth factor (NGF) or cytochrome C (Cit C) for 14 or 28 days using miniosmotic pumps and were evaluated either 1 week or 3 months after treatment. Groups of untreated young, aged-impaired and aged non-impaired rats were also evaluated. Under narcose recording and stimulating electrodes were stereotactically implanted in the dentate gyrus and the perforant path. The stimulation intensity was individually adjusted to obtain a half-maximal population spike (P) for test stimuli and a quarter-maximal for tetanization. The amplitude and latency of P and the slope (S) of the field EPSP were determined before and at 2, 5, 15, 30 and 60 min after tetanization at 400 Hz. Paired stimuli at 30 ms interval were also applied before and after tetanization. Aged, cognitively impaired rats showed an absent S potentiation and a delayed P potentiation, both in amplitude and latency, while non-impaired rats behaved like the young controls. Paired pulse inhibition showed no difference among groups before or after tetanization suggesting that the impaired potentiation is not due to an increased retroactive inhibition. NGF treatment ameliorates LTP deficits to levels equivalent to non-impaired rats, while Cit C controls showed no improvement. No differences appear among NGF treated groups, but evidence suggest that the animals evaluated 3 months after treatment developed a stronger potentiation.

Nerve Growth Factor Attenuates Cholinergic Deficits Following Traumatic Brain Injury in Rats

Traumatic brain injury (TBI) results in chronic derangements in central cholinergic neurotransmission that may contribute to posttraumatic memory deficits. Intraventricular cannula (IVC) nerve growth factor (NGF) infusion can reduce axotomy-induced spatial memory deficits and morphologic changes observed in medial septal cholinergic neurons immunostained for choline acetyltransferase (ChAT). We examined the efficacy of NGF to (1) ameliorate reduced posttraumatic spatial memory performance, (2) release of hippocampal acetylcholine (ACh), and (3) ChAT immunoreactivity in the rat medial septum. Rats (n=36) were trained prior to TBI on the functional tasks and retested on Days 1–5 (motor) and on Day 7 (memory retention). Immediately following injury, an IVC and osmotic pump were implanted, and NGF or vehicle was infused for 7 days. While there were no differences in motor performance, the NGF-treated group had significantly better spatial memory retention (P<0.05) than the vehicle-treated group. The IVC cannula was then removed on Day 7, and a microdialysis probe was placed into the dorsal hippocampus. After a 22-h equilibration period, samples were collected prior to and after administration of scopolamine (1 mg/kg), which evoked ACh release by blocking autoreceptors. The posttraumatic reduction in scopolamine-evoked ACh release was completely reversed with NGF. Injury produced a bilateral reduction in the number and cross-sectional area of ChAT immunopositive medial septal neurons that was reversed by NGF treatment. These data suggest that cognitive but not motor deficits following TBI are, in part, mediated by chronic deficits in cholinergic systems that can be modulated by neurotrophic factors such as NGF.

O-30-441 - Regeneration of the injured acoustic nerve promoted by intrathecal treatment with the antibody IN-1 against neurite growth inhibitors

We have investigated the effects of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) on the intraspinal regeneration of anterogradely labeled axotomized ascending primary sensory fibers in the adult rat. These fibers were allowed to grow across a predegenerated peripheral nerve graft and back into the thoracic spinal cord. In control animals that had been infused with vehicle for two weeks into the dorsal column, 3 mm rostral to the nerve graft, essentially no fibers had extended from the nerve graft back into the spinal cord. The number of sensory fibers in the rostral end of the nerve graft was not significantly different between control and neurotrophin-infused animals. With infusion of NGF, 37±2% of the fibers at the rostral end of the graft had grown up to 0.5 mm into the dorsal column white matter, 30±2% up to 1 mm, 19±3% up to 2 mm and 8±2% up to 3 mm, i.e., the infusion site. With infusion of NT-3, sensory fiber outgrowth was similar to that seen with NGF, but with BDNF fewer fibers reached farther distances into the cord. Infusion of a mixture of all three neurotrophins did not increase the number of regenerating sensory fibers above that seen after infusion of the individual neurotrophins. These findings suggest that injured ascending sensory axons are responsive to all three neurotrophins and confirm our previous findings that neurotrophic factors can promote regeneration in the adult central nervous system.

Hyperplastic Changes within the Leptomeninges of the Rat and Monkey in Response to Chronic Intracerebroventricular Infusion of Nerve Growth Factor

Recombinant human nerve growth factor (rhNGF) was delivered for up to 6 months by continuous intracerebroventricular (icv) infusion to CD (Sprague– Dawley derived) rats and cynomolgus monkeys. Rats (n=15/sex/group) received doses of 0 (vehicle), 6, 60, or 300 ng/day; monkeys (n=5/sex/group) received 0, 0.6, 6, or 60 μg/day of rhNGF. Animals tolerated icv infusion with no behavioral signs attributable to rhNGF. Body weight was transiently decreased in female rats at the highest dose. At the completion of dosing, histological examination in both species revealed an increase in the thickness of the leptomeninges along the ventral and lateral surfaces of the hindbrain and extending over the dorsal aspect of the spinal cord. The change was present to varying degrees at all doses of rhNGF and tended to be more severe at higher doses. At the light microscopic level, the leptomeninges contained layers of well-differentiated, spindle-shaped cells and a plexus of axonal fibers. Cells were immunoreactive for S-100 protein and were associated with an accumulation of Type IV collagen, suggesting Schwann cell origin. Electron microscopy revealed numerous fine caliber axons ensheathed by the presumptive Schwann cells, with myelination of individual axonal segments. These findings suggest that chronic icv delivery of rhNGF has stimulated axonal sprouting and secondary hyperplasia of Schwann or Schwann-like support cells within the pia-arachnoid.

The Effects of Nerve Growth Factor on Spatial Recent Memory in Aged Rats Persist after Discontinuation of Treatment

Nerve growth factor (NGF) infusion significantly reduces spatial recent memory deficits in aged rats, an effect that has great relevance to the treatment of memory impairments characteristic of patients with Alzheimer's disease. The present study was designed to examine whether this NGF-induced improvement in spatial recent memory persists after the discontinuation of NGF treatment, an issue of crucial importance for the potential clinical use of this compound. Spatial recent memory was tested in a Morris water maze delayed nonmatch-to-position task. In addition to memory, sensorimotor skills were also examined. Four- and 22-month-old rats were tested preoperatively, infused intraventricularly with recombinant human NGF or vehicle, and tested both during the 4 week infusion period and during the 4 weeks after discontinuation of the infusion. NGF significantly improved spatial recent memory in 22-month-old rats only, during the 4th week of infusion and for up to 4 weeks after discontinuation of the infusion. Although NGF did not affect overall sensorimotor skills during infusion in either age group, sensorimotor skills were significantly improved both 2 and 4 weeks after discontinuation of infusion in 22-month-old rats. These findings demonstrate that the beneficial effects of NGF on spatial recent memory can persist for up to 1 month after discontinuation of infusion and suggest that NGF can be used intermittently for the treatment of age-associated memory dysfunction and Alzheimer's disease.

Nerve growth factor induces galanin gene expression in the rat basal forebrain: Implications for the treatment of cholinergic dysfunction

Nerve growth factor (NGF) is a potential treatment for cholinergic dysfunction associated with Alzheimer's disease (AD). In rats, NGF activates gene expression of the acetylcholine synthetic enzyme choline acetyltransferase (ChAT) and prevents age- and lesion-induced degeneration of basal forebrain (BF) cholinergic neurons. Cholinergic neurons in the BF coexpress galanin (GAL), a neuropeptide that has been shown to impair performance on memory tasks possibly through the inhibition of cholinergic memory pathways. NGF up-regulates both ChAT and GAL gene expression in cultured pheochromocytoma cells; however, the effect of chronic in vivo NGF administration on GAL gene expression within the BF has not been studied. We used in situ hybridization and quantitative autoradiography to assess GAL and ChAT gene expression within the BF of adult male rats following chronic intracerebroventricular infusion of NGF or cytochrome c. We now report that, in addition to stimulating ChAT gene expression, NGF strongly up-regulated the GAL gene in the rat cholinergic BF. NGF had no effect on GAL gene expression in other noncholinergic forebrain regions. NGF induction of GAL gene expression in the BF was specific, because gene expression for another neuropeptide, neurotensin, present within noncholinergic BF neurons was unchanged. Our data provide the first evidence that in vivo NGF administration up-regulates GAL gene expression in the cholinergic BF. These results suggest that the concurrent induction of GAL in the BF could limit the ameliorating actions of NGF on cholinergic dysfunction.

Nerve growth factor stimulates growth of cortical pyramidal neurons in young adult rats

The cytoarchitectonics of pyramidal neurons in the cerebral cortex of non-lesioned rats can be re-modeled by i.c.v. infusions of nerve growth factor (NGF). 4 months after the application of NGF, the pyramidal neurons in layers III and V of the motor cortex and layer V of the anterior cingulate cortex were analyzed and compared with pyramidal neurons from vehicle-treated rats. NGF-treated brains showed: (1) significant increase in dendritic branching in the basilar fields of the layer V, but not layer III, neurons; and (2) a significant increase in spine density in the terminal, but not proximal, dendritic branches. These findings indicated that, besides its known effects on forebrain cholinergic neurons, NGF produces a very generalized synaptic re-modeling involving the cells responsible for the major output of the cerebral cortex in the intact adult brain. © 1979 Elsevier Science B.V. All rights reserved.

Improvement of cognitive deficits and decreased cholinergic neuronal cell loss and apoptotic cell death following neurotrophin infusion after experimental traumatic brain injury.

This study explores the effects of infusion of nerve growth factor (NGF) on behavioral outcome and cell death in the septal region using the clinically relevant model of fluid-percussion brain injury in the rat. Animals were subjected to fluid-percussion brain injury and 24 hours later a miniosmotic pump was implanted to infuse NGF (12 animals) or vehicle (12 animals) directly into the region of maximum injury for 2 weeks. Four weeks postinjury the animals were tested for cognitive function using a Morris Water Maze paradigm. Neurological motor function was evaluated over a 4-week postinjury period. The rats receiving NGF infusions had significantly higher memory scores than vehicle-treated animals. Examination of the cholinergic neurons in the medial septal region using choline acetyltransferase immunohistochemistry demonstrated significant cell loss after injury. Infusion of NGF significantly attenuated loss of these cholinergic neurons. A second group of animals was subjected to fluid-percussion brain injury alone (23 rats) or injury followed by NGF infusion (18 rats). These animals were killed between 24 hours and 2 weeks postinjury and the septal region was examined for the presence of apoptotic cells using the terminal deoxynucleotidyl transferase-mediated biotinylated-deoxyuridinetriphosphate nick-end labeling technique. Apoptotic cells were identified as early as 24 hours postinjury; their numbers peaked at 4 and 7 days, and then declined by 14 days. The NGF-treated animals had some apoptotic cells; however, even at 7 days there were significantly fewer of these cells. No significant motor differences were observed between the NGF- and vehicle-treated groups. These data indicate that NGF administration beginning 24 hours after fluid-percussion brain injury has a beneficial effect on cognition and results in sparing of cholinergic septal neurons. These improvements persist after cessation of NGF administration. The beneficial effects of NGF may be related to its ability to attenuate traumatically induced apoptotic cell death.

Axonal regeneration of sensory nerves is delayed by continuous intrathecal infusion of nerve growth factor

While it is well established that nerve growth factor is growth promoting for sensory neurons in culture, it is unclear whether it serves such a function in vivo. In fact, our previous studies led to the hypothesis that nerve growth factor could actually impair axonal regeneration by reducing the neuronal cell body response to injury. In the present study, the consequence of continuous intrathecal infusion of nerve growth factor on regeneration of sensory neurons was examined in rats given a bilateral sciatic nerve crush. Rats received nerve growth factor (125ng/h) as a continuous infusion into the subarachnoid space of the lumbar spinal cord via an osmotic minipump (Alzet); controls received cytochrome C. At seven or 10 days, the pump was removed and L4 or L5 dorsal root ganglion exposed and injected with 50μCi of [3H]leucine. Animals were killed 24h later, the sciatic nerves removed, cut into 3mm segments and the radioactivity in each segment determined by liquid scintillation spectrophotometry. Maximal regeneration distances (determined from the front of the resultant transport curves) were similarly reduced (by approximately 6mm) in nerve growth factor-infused compared to cytochrome C-infused rats. Thus, regeneration rates (determined between eight and 11 days) were unaltered by nerve growth factor infusion; regeneration rates from cytochrome C-infused and nerve growth factor-infused animals were 2.8mm/day and 3.1mm/day, respectively. However, nerve growth factor significantly (P<0.005) increased the delay to onset for regeneration by two days.Taken together, the present study demonstrates that nerve growth factor delays the onset of regeneration without affecting the rate of regeneration. The results implicate the involvement of at least two signals in the regulation of axonal regeneration in dorsal root ganglion neurons. It is suggested that the loss of nerve growth factor serves as an early, induction signal regulating the onset of regeneration and that a second, unidentified signal independently serves to maintain regeneration.

Dose-dependent response of mature cerebrovascular axons in vivo following intracranial infusion of nerve growth factor

The perivascular axons associated with the intradural segment of the internal carotid artery undergo a sprouting response following intracerebroventricular infusion of nerve growth factor (NGF). The objective of the present study was to determine whether a relationship exists between the number of sprouted axons and the amount of NGF infused into the ventricular system. Using regression analysis, we observed a significant log-log relationship between the dose of NGF and number of axons. No significant relationship was observed for the control (VEH) group. A significant increase in axonal number was observed following infusion of 3.0 μg NGF and higher when compared with VEH treatment of similar concentration. Results of this analysis suggest that a maximal response to NGF is approximated at doses of 15 μg or higher. These findings suggest a dose-dependent relationship between the response of mature sympathetic cerebrovascular axons in vivo and the dose of exogenous NGF.

The Role of NGF and Afferent Denervation in the Process of Sympathetic Fiber Ingrowth into the Hippocampal Formation

Nerve growth factor (NGF) has been postulated to play an important role in the process of sympathetic sprouting into the septally deafferented hippocampal formation. In the current investigation we have used transplants of NGF-dependent neonatal superior cervical ganglion (SCG) neurons to investigate the influence of NGF and septal denervation (either alone or in combination with one another) upon neuronal survival and intrahippocampal sprouting. In the current model, SCG transplants were placed onto the dorsal surface of the CA1 hippocampal subfield and a continuous infusion device was used to deliver either NGF or vehicle into the hippocampal parenchyma. Following 15 days of vehicle infusion, little or no sympathetic neuronal survival was observed and no hippocampal fiber outgrowth was detected. NGF infusions, however, promoted both neuronal survival and intrahippocampal fiber outgrowth directed mainly toward the location of the infusion cannula. Septal denervation, achieved by either a septal ablation or fimbria–fornix transection, had no discernible impact upon SCG neuronal survival or fiber outgrowth, with or without NGF infusions. Similar results were also obtained when SCG were transplanted directly within the cortex, with or without an intracortical infusion. It appears, therefore, that NGF may be a sufficient, as well as necessary, component for eliciting and guiding the invasion of a tissue by NGF-sensitive fibers.

Chronic infusion of nerve growth factor into rat striatum increases cholinergic markers and inhibits striatal neuronal discharge rate.

New strategies have recently been developed where infusion of neurotrophic factors into the brain can rescue different populations of neurons. Infusion of nerve growth factor (NGF) has been used in combination with transplants of chromaffin tissue to the striatum in the rat model of Parkinson's disease as well as to patients suffering from Alzheimer's disease. In this study we have evaluated the distribution of recombinant human NGF (rhNGF) in different brain areas and evaluated morphological and electrophysiological effects after continuous infusion for 2 weeks of rhNGF (500 micrograms/ml) into the striatum of normal rats. One week after termination of rhNGF infusion, NGF levels in the infused striata were 10-fold increased while in contralateral striata normal levels were found. Extracellular recordings from striatal neurons revealed a significantly decreased spontaneous firing rate (0.76 +/- 0.07 Hz) in rats infused with rhNGF compared to vehicle-infused control animals (1.36 +/- 0.16 Hz). Local application of rhNGF during recordings showed no direct inhibitory effect of NGF on neuronal discharge rate. Immunohistochemistry, using antibodies against acetyl cholinesterase (AChE) and glial fibrillary acidic protein (GFAP), revealed a 38.7 +/- 7.0% increase in optical density of AChE immunoreactivity close to the NGF source and an increase in GFAP-positive profiles that was restricted close to the implanted dialysis fibre. In situ hybridization showed an increase in mRNAs for choline acetyltransferase, trkA, p75 and muscarinic m2 receptor in the large neurons of rhNGF-infused striatum. Messenger RNAs for m1 and m4 receptors in striatal neurons were not changed. Thus, chronic infusion of rhNGF into the striatum caused a cholinergic hyperinnervation and reduced spontaneous activity of striatal neurons.

Nerve growth factor induced modification of presynaptic elements in adult visual cortex in vivo

Nerve growth factor (NGF) has been shown to play important roles in neuronal survival, growth and differentiation. Recently, we have found that intracortical infusion of NGF into adult cat visual cortex can recreate ocular dominance plasticity, suggesting that NGF is also involved in activity-dependent modification of synaptic connectivity in the adult brain. To further explore the mechanisms of NGF-induced plasticity in adult visual cortex, we studied two presynaptic markers: GAP-43 and synaptophysin. Immunocytochemical staining showed that NGF-treatment of adult visual cortex selectively increased the level of the phosphorylated form of GAP-43, while the total level of GAP-43 was not changed. These results demonstrate that NGF-treatment stimulates phosphorylation processes of GAP-43 in vivo. In addition, NGF-treatment of adult visual cortex increased the level of synaptophysin immunoreactivity. Since the phosphorylated form of GAP-43 is known to be enriched in the membrane skeleton of growth cones and of developing synapses, and the phosphorylation of GAP-43 has been linked with events that underlie synaptic plasticity, and since synaptophysin is a major component of presynaptic vesicles, our results suggest that NGF-treatment of adult visual cortex modulates presynaptic terminals, possibly by inducing axonal sprouting and formation of new synapses, and that these changes may play a role in the NGF-induced functional plasticity.

Expression of neuronal-NOS in developing basal forebrain cholinergic neurons: regulation by NGF.

Nerve growth factor (NGF) acts through the receptor tyrosine kinase trkA to serve as a trophic factor for cholinergic neurons in the medial septal nucleus and vertical limb of the diagonal band. We have previously shown that the neuronal isoform of nitric oxide synthase (NOS) is selectively expressed in a large fraction of trkA-expressing cholinergic neurons in these brain regions in the adult rat, and that NGF induces the expression of neuronal-NOS in these cells. Herein, we show that: 1) neuronal-NOS is also localized to these neurons in the developing septum; 2) the expression of neuronal-NOS is regulated in the developing medial septal nucleus and vertical limb of the diagonal band; 3) neuronal-NOS regulation parallels that for other markers of basal forebrain cholinergic neuron differentiation, such as cholineacetyltransferase; and 4) NGF infusion in the postnatal period induces robust increases in neuronal-NOS mRNA and in NOS activity in the basal forebrain. Taken together with earlier findings, our results suggest that neuronal-NOS has a role in the differentiation and mature function of septal cholinergic neurons. Through enhancing neuronal-NOS synthesis, endogenous NGF is likely to regulate NO functions in vivo.