Distribution Of Brain-derived Neurotrophic Factor Research Articles

Distribution of Brain-Derived Neurotrophic Factor in the Brain of the Small-Spotted Catshark Scyliorhinus canicula, and Evolution of Neurotrophins in Basal Vertebrates.

Neurotrophins (NTFs) are structurally related neurotrophic factors essential for differentiation, survival, neurite outgrowth, and the plasticity of neurons. Abnormalities associated with neurotrophin-signaling (NTF-signaling) were associated with neuropathies, neurodegenerative disorders, and age-associated cognitive decline. Among the neurotrophins, brain-derived neurotrophic factor (BDNF) has the highest expression and is expressed in mammals by specific cells throughout the brain, with particularly high expression in the hippocampus and cerebral cortex. Whole genome sequencing efforts showed that NTF signaling evolved before the evolution of Vertebrates; thus, the shared ancestor of Protostomes, Cyclostomes, and Deuterostomes must have possessed a single ortholog of neurotrophins. After the first round of whole genome duplication that occurred in the last common ancestor of Vertebrates, the presence of two neurotrophins in Agnatha was hypothesized, while the monophyletic group of cartilaginous fishes, or Chondrichthyans, was situated immediately after the second whole genome duplication round that occurred in the last common ancestor of Gnathostomes. Chondrichthyans represent the outgroup of all other living jawed vertebrates (Gnathostomes) and the sister group of Osteichthyans (comprehensive of Actinopterygians and Sarcopterygians). We were able to first identify the second neurotrophin in Agnatha. Secondly, we expanded our analysis to include the Chondrichthyans, with their strategic phylogenetic position as the most basal extant Gnathostome taxon. Results from the phylogenetic analysis confirmed the presence of four neurotrophins in the Chondrichthyans, namely the orthologs of the four mammalian neurotrophins BDNF, NGF, NT-3, and NT-4. We then proceeded to study the expression of BDNF in the adult brain of the Chondrichthyan Scyliorhinus canicula. Our results showed that BDNF is highly expressed in the S. canicula brain and that its expression is highest in the Telencephalon, while the Mesencephalic and Diencephalic areas showed expression of BDNF in isolated and well-defined cell groups. NGF was expressed at much lower levels that could be detected by PCR but not by in situ hybridization. Our results warrant further investigations in Chondrichthyans to characterize the putative ancestral function of neurotrophins in Vertebrates.

Music Therapy Alleviates Motor Dysfunction in Rats With Focal Cerebral Ischemia-Reperfusion Injury by Regulating BDNF Expression.

Background/Aim: Music-based therapy plays a role in central nervous system diseases. We aimed to explore the effect of different doses and durations of music therapy on motor function recovery after stroke and the underlying molecular mechanisms.Methods: Adult male Sprague–Dawley rats were subjected to middle cerebral artery occlusion (MCAO) for 1 h, which was followed by reperfusion. In experiment 1, the rats that survived 1 week after MCAO surgery were randomly allocated into four groups (n = 10 per group): MCAO group, 1 h music group (Mozart K.448 music therapy 1 h per day for 2 weeks), 12 h music group (Mozart K.448 music therapy 12 h/day for 2 weeks), and accelerated music group (reversely accelerated music therapy 12 h for 2 weeks, AM group). In experiment 2, the survived rats were randomly divied into three groups: MCAO group, 12 h music group (music therapy 12 h/day for 3 weeks), and 12 h music-R group (music therapy 12 h/day for 2 weeks and rest for 1 week). Three neuroscores were evaluated daily, starting on the first day after surgery until the end of the experiment. The rats were killed 3 weeks after MCAO surgery in experiment 1 or 4 weeks after surgery in experiment 2. Nissl staining of infart core, peri-infarct zone, and motor cortex was performed to assess neuronal survival and regeneration. Western blot and immunofluorescence were used to detect the expression and distribution of brain-derived neurotrophic factor (BDNF) and glial fibrillary acidic protein (GFAP) in ipsilateral hemispheres.Results: In the experiment of different music therapy doses, the motor function in the 12-h music group but not in the 1-h music group and AM group was significantly improved compared with that of the MCAO group. The BDNF protein level of the ipsilateral hemisphere motor cortex in the 12-h music group and the 1-h music group was higher than that of the MCAO group. The neurons and Nissl bodies were more in the 12-h music group than in the MCAO group. Immunofluorescence assay showed that a 12 h music therapy induces BDNF and GFAP accumulation at the damage boundary. In the experiment of different music therapy durations, 3 weeks music therapy (12 h music group) induced more longer cell synapses and more clearer cell-to-cell connections than 2 weeks music intervention (12 h music-R group). Moreover, the GFAP morphology in the 12-h music group was more similar to mature activated astrocytes than that in the 12-h music-R group.Conclusions: Music therapy may improve poststroke motor function and promote neuronal repair in the long term. The mechanism may be through stimulating BDNF and GFAP secretion in the injured motor cortex.

Repairing effect of Schwann cells combined with mesenchymal stem cells on optic nerve injury in rats.

To investigate the effect and the underlying mechanisms of combined transplantation of Schwann cells (Scs) and Mesenchymal stem cells (MSCs) on optic nerve injury in rats. A total of 160 normal healthy adult rats were randomly divided into 4 groups: optic nerve injury group, optic nerve injury + Sc transplantation group, optic nerve injury + MSC transplantation group and optic nerve injury + Sc + MSC transplantation group. The optic nerve in the left eye of each rat was damaged via clamping to establish a model of optic nerve injury, and the right eye was used as self-control. Scs + MSCs, Scs alone, MSCs alone and normal saline were injected into the vitreous space, respectively. After the treatment, the optic nerve tissues were collected and subjected to hematoxylin-eosin (HE) staining. Next, the morphologic and pathological changes of rats in each group were observed. Retrograde labeling was utilized to count the number of retinal ganglion cells (RGCs) in the optic nerve tissues. The apoptosis of RGCs was detected using flow cytometry. Western blot was carried out to measure the protein expression level of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax). The expression and distribution of brain-derived neurotrophic factor (BDNF) and growth associated protein-43 (GAP-43) in the optic nerve of rats in each group were detected by immunohistochemistry. Transplantation of Scs and MSCs could maintain the morphological structures of the retina and optic nerve of rats, increase the amount of RGCs in optic nerve tissues, reduce the apoptosis of RGCs, promote the expression of the Bcl-2 protein and decrease the expression of Bax protein. In addition, our joint transplantation strategy also showed an important role in repairing optic nerve injury by clearly promoting the secretion and expression of BDNF and GAP-43, which indicated a better curative effect than that of separate application of Scs or MSCs. Therapy with combined use of Scs and MSCs has a significant therapeutic effect in repairing optic nerve injury.

Distribution and expression of brain-derived neurotrophic factor, nerve growth factor, and neurotrophic factor-3 in refractory epilepsy-associated focal cortical dysplasia.

To investigate the expression and distribution of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and neurotrophic factor-3 (NT-3) in refractory epilepsy-associated type I focal cortical dysplasia (FCD I) and FCD IIA patients, and to explore their effects on pathogenesis of FCD I and FCD IIA. 19 subjects who received surgery at the Department of Neurosurgery, First Affiliated Hospital, Fujian Medical University, China between June 2010 and May 2012, were enrolled in this study. They were pathologically diagnosed as FCD IIA (n=7) and FCD I (n=12) after surgery and were considered as two experimental groups. Temporal lobe samples of 10 subjects who had suffered craniocerebral injury but did not have nervous system disease were collected as a control group. Immunohistochemical methods and Western blot assays were used to detect the expression and distribution of BDNF, NGF, and NT-3 in temporal lobes, and differences in their expression and distribution were compared between experimental and control groups. BDNF expression was slightly higher in the FCD IIA group compared to that in the FCD I group and was significantly greater when compared with the control group. Compared with the control group, NGF and NT-3 expression was higher in the FCD groups. However, no significant difference was observed between the FCD IIA and FCD I group. Abnormal distribution and expression of BDNF, NGF, and NT-3 may play an important role in the mechanism of FCD I and FCD IIA-induced epilepsy. .

3D Presentation of A Neurotrophic Factor for The Regulation of Neural Progenitor Cells

Adequate cell-scaffold interactions and neurotrophin support are essential factors for neural regeneration. To provide insight into the biofunctionalization of complex 3D scaffolds with nanoscale precision, as well as the effect of spatial distribution of brain-derived neurotrophic factor (BDNF) and its prolonged stimulation in combination with enhanced cell affinity of nanofibrous scaffolds on the survival/proliferation and neurite outgrowth. We developed a versatile approach using layer-by-layer self-assembly to incorporate cell adhesion and spatial representation of neurotrophic factors into complex nanofibrous scaffolds. Heparin/poly-L-lysine (PLL) polyelectrolyte multilayers (PEMs) were deposited on electrospun poly-ε-caprolatone nanofibers. Well-controlled amounts of BDNF were immobilized on the PEM-modified nanofibers. In addition, longer neurite outgrowth was observed in neural progenitor cells cultured on PLL-terminating PEM scaffolds. The immobilized BDNF on PLL-terminated PEM scaffolds resulted in significantly longer neurites and higher cell numbers (p < 0.01) compared with BDNF-free and BDNF-adsorbed PLL-terminating scaffolds. Interestingly, there was no upregulation of TrkB-FL, TrkB-T1 or GAP-43 mRNAs with immobilized BDNF in day 5 cultures. This work reinforces the importance of the combinatorial effects of biomaterial scaffold nanostructure and spatial presentation of neurotrophins in directing neural progenitor cell fates.

Changes in the serotonergic system and in brain-derived neurotrophic factor distribution in the main olfactory bulb of pcd mice before and after mitral cell loss

The serotonergic centrifugal system innervating the main olfactory bulb (MOB) plays a key role in the modulation of olfactory processing. We have previously demonstrated that this system suffers adaptive changes under conditions of a lack of olfactory input. The present work examines the response of this centrifugal system after mitral cell loss in the Purkinje cell degeneration ( pcd) mutant mice. The distribution and density of serotonergic centrifugal axons were studied in the MOB of control and pcd mice, both before and after the loss of mitral cells, using serotonin (5-HT) and 5-HT transporter immunohistochemistry. Studies of the amount of 5-HT and its metabolite, 5-hydroxyindole acetic acid (5-HIAA), were performed by means of high-performance liquid chromatography (HPLC), and the relative amounts of brain-derived neurotrophin factor, BDNF, and its major receptor, tropomyosin-related kinase B (TrkB), were measured by Western blot. Our study revealed that the serotonergic system develops adaptive changes after, but not before, mitral cell loss. The lack of the main bulbar projection cells causes a decrease in the serotonergic input received by the MOB, whereas the number of serotonergic cells in the raphe nuclei remains constant. In addition, one of the molecules directly involved in serotonergic sprouting, the neurotrophin BDNF and its main receptor TrkB, underwent alterations in the MOBs of the pcd animals even before the loss of mitral cells. These data indicate that serotonergic function in the MOB is closely related to olfactory activity and that mitral cell loss induces serotonergic plastic responses.

Masseteric nerve injury increases expression of brain-derived neurotrophic factor in microglia within the rat mesencephalic trigeminal tract nucleus.

The distribution of brain-derived neurotrophic factor was examined in the rat mesencephalic trigeminal tract nucleus after transection and crush of the masseteric nerve. In the intact mesencephalic trigeminal tract nucleus, brain-derived neurotrophic factor was detected in small cells with fine processes. These cells and processes were occasionally located adjacent to tyrosine kinase B receptor-immunoreactive sensory neurons. The transection and crush of the masseteric nerve increased expression of brain-derived neurotrophic factor in the nucleus. The number and size of brain-derived neurotrophic factor-immunoreactive cells and processes were dramatically elevated by the nerve injury. As a result, the density of brain-derived neurotrophic factor-immunoreactive profiles in the mesencephalic trigeminal tract nucleus at 7 days after the injury was significantly higher compared with the intact nucleus. Double immunofluorescence method also revealed that brain-derived neurotrophic factor-immunoreactive cells were mostly immunoreactive for OX-42 but not glial fibrillary acidic protein. In addition, the retrograde tracing method demonstrated that brain-derived neurotrophic factor-immunoreactive cells and processes surrounded retrogradely labeled neurons which showed tyrosine kinase B receptor-immunoreactivity. These findings indicate that the nerve injury increases expression of brain-derived neurotrophic factor in microglia within the mesencephalic trigeminal tract nucleus. The glial neurotrophic factor may be associated with axonal regeneration of the injured primary proprioceptor in the trigeminal nervous system.

BDNF and TrkB in the Preterm and Near-term Ovine Fetal Brain and the Effect of Intermittent Umbilical Cord Occlusion

We have determined the developmental change in immunoreactivity for brain-derived neurotrophic factor and its high-affinity tyrosine kinase receptor, TrkB, in the ovine fetal brain with advancing gestation and in response to intermittent umbilical cord occlusion, which might then contribute to adverse neurodevelopment. Fetal sheep (control and experimental groups at 0.75 and 0.90 of gestation) were studied over 4 days with umbilical cord occlusions performed in the experimental group animals by complete inflation of an occluder cuff for 90 seconds every 30 minutes for 3 to 5 hours each day. Animals were then euthanized and the fetal brains perfusion fixed and prepared for subsequent histology with the distribution of brain-derived neurotrophic factor and tyrosine kinase receptor immunoreactivity determined by immunohistochemistry. In the control group animals brain-derived neurotrophic factor immunoreactivity decreased in the gray matter, thalamus, and hippocampus but increased in the white matter, while tyrosine kinase receptor immunoreactivity decreased in all regions (most P < .01), with advancing gestation consistent with the developmental change from neurogenesis/gliagenesis to myelination over this time period. Intermittent umbilical cord occlusion as studied with severe but limited hypoxemia resulted in a variable decrease in brain-derived neurotrophic factor and tyrosine kinase receptor immunoreactivity for all brain regions in the preterm animals (most P < .01) when protein turnover is higher, but a selective increase in brain-derived neurotrophic factor immunoreactivity in the hippocampus of the near-term animals consistent with a heightened vulnerability for necrotic/apoptotic injury at this time. As such, brain-derived neurotrophic factor-tyrosine kinase receptor in the ovine fetal brain may be altered with intermittent hypoxic insults over the latter part of pregnancy with potential for longer term neurologic consequences.

High-pressure freezing followed by cryosubstitution as a tool for preserving high-quality ultrastructure and immunoreactivity in the Xenopus laevis pituitary gland

Subcellular localisation of proteins and peptides yields fundamental information about cell functioning. Immunoelectron microscopy is a powerful tool to achieve this goal, but combining good tissue preservation with strong immunoreactivity is a great challenge in electron microscopy. We have applied a novel approach, using high-pressure freezing (HPF) followed by cryosubstitution, to prepare the pituitary gland of the amphibian Xenopus laevis for immunogold-electron microscopy. In this way, we investigated the subcellular distribution of brain-derived neurotrophic factor and the amphibian neurohormone mesotocin in the pituitary neural lobe, and the peptide hormone α-melanophore-stimulating hormone and its protein precursor proopiomelanocortin in melanotrope cells of the pituitary intermediate lobe. In contrast to conventional chemical fixation (followed by cryosubstitution), HPF not only revealed strong immunoreactivity of the secretory products, but also provided excellent ultrastructural preservation of cell organelles, including secretory granule subtypes. We conclude that HPF followed by cryosubstitution provides a preparation technique of choice when both optimal tissue ultrastructure and strong immunoreactivity are required.

BDNF is upregulated by postnatal development and visual experience: quantitative and immunohistochemical analyses of BDNF in the rat retina.

This study sought to elucidate changes in the levels and distribution of brain-derived neurotrophic factor (BDNF) in the retina throughout aging and depending on visual experience. Protein and mRNA levels of BDNF were quantified by enzyme-linked immunosorbent assay (ELISA) and semiquantitative reverse transcription-polymerase chain reaction (RT-PCR), respectively. Levels were assayed in the retinas of rats on postnatal day (P)2, P7, and P14 (approximate time of eye opening) and at 1 month (M), 3M, 8M, and 18M of age. Changes in BDNF expression and localization in the retina were assessed by immunohistochemistry. The effect of monocular deprivation during infancy on retinal BDNF expression was also examined, by ELISA and immunohistochemistry. Both protein and mRNA levels of BDNF in the rat retina increased after P14. Immunohistochemical analyses revealed that the increase in BDNF protein levels occurred in retinal ganglion cells (RGCs) between P14 and 1M. BDNF immunoreactivity in Müller cell processes was observed in the inner nuclear layer at 1M, but not at P14. The levels of BDNF protein in the retinas of visually deprived eyes were lower than those of control eyes, as quantified by ELISA. Immunohistochemistry showed that BDNF immunoreactivity in RGCs was diminished by visual deprivation, whereas Müller cells were unaffected. These observations indicate that BDNF expression in RGCs is upregulated in an activity-dependent manner, whereas that in Müller cells is regulated only by development.

Changes in brain-derived neurotrophic factor and trkB receptor in the adult Rana pipiens retina and optic tectum after optic nerve injury.

In this study we used immunocytochemistry to investigate the distribution of brain-derived neurotrophic factor (BDNF) and its receptor tyrosine kinase (trkB) in retina and optic tectum of the frog Rana pipiens during regeneration after axotomy. We also measured changes in BDNF mRNA in retina and tectum. Retrograde labeling was used to identify retinal ganglion cells (RGCs) prior to quantification of the BDNF immunoreactivity. In control animals, BDNF was found in the majority of RGCs and displaced amacrine cells and in some cells in the inner nuclear layer (INL). After axotomy, BDNF immunoreactivity was reduced in RGCs but increased in the INL. BDNF mRNA levels in the retina remained high before and after axotomy. Three months after axotomy, after reconnection to the target, the staining intensity of many of the surviving RGCs had partially recovered. In the control tectum, BDNF staining was present in ependymoglial cells and in neurons throughout layers 4, 6, 8, and 9. After axotomy, BDNF staining in tectal neurons became more intense, even though mRNA synthesis was transiently down-regulated. In control retinas, trkB receptor immunostaining was present in most RGCs; no significant changes were observed after axotomy. In control tectum, trkB was detected only in ependymoglial cells. After axotomy, many neuronal cell bodies were transiently labeled. Our data are consistent with the hypothesis that a considerable fraction of the BDNF normally present in RGCs is acquired from their targets in the tectum. However, there are also intraretinal sources of BDNF that could contribute to the survival of RGCs.

Distribution of brain-derived neurotrophic factor and TrkB receptor proteins in the fetal and postnatal hippocampus and cerebellum of the guinea pig.

This study investigates the distribution of brain-derived neurotrophic factor protein (BDNF) and its receptor, TrkB, during the development of hippocampus and cerebellum in a long-gestation species, the guinea pig. In the granule cell populations of both structures, BDNF immunoreactivity (-IR) was exclusive to postmigratory, mature neurons. In dentate granule cells, TrkB-IR was coexpressed with BDNF-IR, suggesting that the ligand-receptor interaction could occur by means of an autocrine/paracrine mechanism. In cerebellar granule cells, TrkB-IR was detected in both pre- and postmigratory cells, indicating that immature neurons are also BDNF-responsive. With advancing gestational age an increase in the intensity of BDNF-IR in granule cells was accompanied by concomitant increases in the staining and areal growth of the associated mossy fiber layer in the hippocampus, and the molecular layer in the cerebellum. The developmental increase in BDNF- and TrkB-IR in the neuropil of both structures coincided with periods of significant growth in all strata, indicating a role for BDNF and TrkB in process outgrowth. In the hippocampus, CA2, CA3, and hilar, neurons demonstrated both BDNF- and TrkB-IR during development and maturation, whereas CA1 neurons showed TrkB-IR throughout this period but only transient BDNF-IR in early gestation. In the fetal cerebellum, Purkinje cell bodies coexpressed BDNF-IR and TrkB-IR. In the postnatal period, BDNF-IR was down-regulated but TrkB-IR persisted, indicating that mature Purkinje cells might retain their responsiveness to BDNF. Thus, we have demonstrated in both the hippocampus and cerebellum that the spatiotemporal distribution of BDNF-IR and TrkB-IR coincides with the maturation of granule cells prenatally and with significant periods of neuropil growth, both prenatally and in the immediate postnatal period.

An immunohistochemical study of the distribution of brain-derived neurotrophic factor in the adult human brain, with particular reference to Alzheimer's disease

Brain-derived neurotrophic factor is a member of the family of neuronal differentiation and survival-promoting molecules called neurotrophins. Neuronal populations known to show responsiveness to the action of brain-derived neurotrophic factor include the cholinergic forebrain, mesencephalic dopaminergic, cortical, hippocampal and striatal neurons. This fact has aroused considerable interest in the possible contribution of an abnormal brain-derived neurotrophic factor function to the aetiology and physiopathology of different neurodegenerative disorders, such as Alzheimer's disease. This report describes the cellular and regional distribution of brain-derived neurotrophic factor in post mortem control human brain and in limited regions of the brain in patients with Alzheimer's disease, as was revealed by immunohistochemistry. Brain-derived neurotrophic factor is widely expressed in the control human brain, both by neurons and glia. In neurons, brain-derived neurotrophic factor was localized in the cell body, dendrites and axons. Among the structures showing the most intense immunohistochemical labeling were the hippocampus, claustrum, amygdala, bed nucleus of the stria terminalis, septum and the nucleus of the solitary tract. In the striatum, immunoreactivity was more intense in striosomes than in the matrix. Many labeled neurons were found in the substantia nigra pars compacta. The large putatively cholinergic neurons in the basal forebrain showed no immunoreactivity. The general pattern of labeling was similar in individuals with Alzheimer's disease. Brain-derived neurotrophic factor-immunoreactive material was found in senile plaques, and some immunoreactive cortical pyramidal neurons showed neurofibrilary tangles, suggesting that brain-derived neurotrophic factor may be involved in the process of neuronal degeneration and/or compensatory mechanisms which occur in this illness.

Cortical spreading depression activates trophic factor expression in neurons and astrocytes and protects against subsequent focal brain ischemia

We recently reported that cortical spreading depression (CSD), used to precondition rat brain, reduced cortical infarction volume resulting from focal cerebral ischemia by middle cerebral artery occlusion (MCAO) 3 days later. The mechanisms underlying this protective effect by CSD remains to be explored. In this study, we confirm that CSD is neuroprotective when KCl is applied epidurally rather than intracortically. Neocortical infarct volume was 101.3±48.5 mm 3 and 45.3±44.1 mm 3 in the sham and CSD group, respectively ( p<0.05). Using image analysis, we identified the cortical region spared from infarction by the prior CSD. We then determined the distribution of brain-derived neurotrophic factor (BDNF) and basic fibroblast growth factor (bFGF) mRNA and the time course of their expression in groups of animals treated with CSD and their controls. We also examined the response of astrocytes to CSD using glial fibrillary acidic protein (GFAP) as a marker. In situ hybridization (done at 0, 3, 12, 24, 72 or 168 h after CSD) showed significant elevation of BDNF mRNA in the cortex immediately after CSD in a distribution surrounding the spared cortex, while bFGF mRNA rose 12 h after CSD and appeared more within the core of the ischemic region. Immunohistochemistry (done at 1, 3 or 7 days after CSD) demonstrated GFAP in the neocortex, with a peak at 3 days after CSD. Heat shock protein 72 (HSP72) expression was not affected by CSD. We concluded that upregulation of trophic factors and activation of glial cells may contribute to the neuroprotection induced by CSD.

Transient expression and transport of brain-derived neurotrophic factor in the male zebra finch's song system during vocal development.

The distribution of brain-derived neurotrophic factor (BDNF) in the song system of male zebra finches changes with posthatching age. At day 20, the hyperstriatum ventrale, pars caudale is the only song nucleus in which neurons showed BDNF immunoreactivity. At day 45, the staining in hyperstriatum ventrale, pars caudale was denser than at day 20 and the robust nucleus of the archistriatum, another song nucleus, showed BDNF labeling. By day 65, two additional song nuclei, area X and the lateral magnocellular nucleus of the anterior neostriatum, have become immunoreactive. In the adult, however, the amount of BDNF labeling in all of these brain nuclei is sharply reduced. These sequential events, the anatomical connections between these song nuclei, and the labeling of relevant axons and terminals suggest anterograde transport of BDNF. Furthermore, the timing of BDNF expression coincident with the development of singing behavior suggests that this neurotrophin may be directly involved with the differentiation of the song system.

Co-infusion with a TrkB-Fc Receptor Body Carrier Enhances BDNF Distribution in the Adult Rat Brain

Fusion proteins comprising the Fc domain of human IgG and extracellular domains of receptor tyrosine kinases can neutralize the activity of their cognate ligands when administered in molar excess. We have generated a fusion protein using the ectodomain of TrkB (TrkB-Fc). Although the ability of TrkB-Fc to neutralize the activity of brain-derived neurotrophic factor (BDNF)in vitrohas been demonstrated, there have been no conclusive demonstrations of its ability to neutralize the activity of BDNFin vivo.We co-infused TrkB-Fc with BDNF into the cortex and hippocampus of adult rats to determine whether TrkB-Fc would interfere with the ability of BDNF to upregulate neuropeptide Y (NPY). We report here that rather than neutralizing the activity of exogenous BDNF, co-infusion with the TrkB-Fc fusion protein greatly increased the volume of tissue in which neuropeptide Y immunostaining was upregulated. In addition, TrkB-Fc greatly enhanced BDNF's distribution through adult brain parenchyma. TrkB-Fc also markedly increased the otherwise limited diffusion of BDNF into brain parenchyma following intraventricular infusion. These results show that rather than neutralizing or sequestering BDNF, the TrkB-Fc, at close to molar equivalence to BDNF, can function as a carrier for BDNF and thus enhance the delivery or penetration of this polypeptide into the brain.

Distribution of brain-derived neurotrophic factor in cranial and spinal ganglia.

In a previous study we have shown that a subpopulation of primary sensory neurons contain brain-derived neurotrophic factor immunoreactivity. In the present study we investigated the distribution of brain-derived neurotrophic factor and its mRNA in cranial and spinal ganglia at different segmental levels, using immunohistochemical and quantitative reverse transcriptase-polymerase chain reaction techniques. Our results show that there is no significant difference in the percentage of brain-derived neurotrophic factor-immunoreactive neurons in spinal ganglia of different segmental levels. In contrast, more brain-derived neurotrophic factor-immunoreactive neurons were found in placode-derived than neural crest-derived ganglia. The percentage of brain-derived neurotrophic factor-immunoreactive neurons is consistent with the percentage of neurons lost after deletion of brain-derived neurotrophic factor or trkB genes. However, there is no correlation between brain-derived neurotrophic factor mRNA levels and the number of brain-derived neurotrophic factor immunoreactive neurons in these ganglia, suggesting that some neurons synthesize brain-derived neurotrophic factor while others accumulate the factor following its retrograde transport within nerve fibers. In particular, the proportion of brain-derived neurotrophic factor that is derived from extraganglionic sources in the placode-derived ganglia appears greater than that in the neural crest-derived ganglia.

Distribution of brain-derived neurotrophic factor in rats and its changes with development in the brain.

A newly established, sensitive, two-site enzyme-immunoassay system for brain-derived neurotrophic factor (BDNF) is described. Using this system, we investigated the tissue distribution of BDNF and developmental changes in tissue levels of BDNF in rats. The minimal limit of detection of the assay was 3 pg/0.2 ml of assay mixture. BDNF was successfully solubilized from tissues in the presence of guanidine hydrochloride but not in any of the other buffers examined. In the rat brain at 1 month of age, the highest level of BDNF was detected in the hippocampus (5.41 ng/g of wet weight), followed by the hypothalamus (4.23 ng/g) and the septum (1.68 ng/g). In other regions, levels of BDNF ranged between 0.9 and 1.7 ng/g. The level of BDNF in the posterior lobes of the cerebellum from rats at 30 days of age was slightly higher than that in the anterior lobes. The concentration of BDNF increased in all regions of the brain with postnatal development. In peripheral tissues, BDNF was found at very low concentrations (0.65 ng/g in the spleen, 0.21 ng/g in the thymus, and 0.06 ng/g in the liver). The subfractionation of the hippocampal homogenate indicated that approximately 50% of BDNF was contained in the crude nuclear fraction. Immunoblots of BDNF-immunoreactive proteins extracted from the hippocampus, hypothalamus, and cerebellum contained doublet bands of protein of approximately 14 kDa, a value close to the molecular mass of recombinant human BDNF. Immunocytochemical investigations showed that, in the hippocampus, BDNF was localized in the nucleus of the granule cells in the dentate gyrus and of the cells in the pyramidal cell layer. The frequency of cells that were stained in the dentate gyrus was greater than that of cells in the pyramidal cell layer.

Distribution of Brain-Derived Neurotrophic Factor (BDNF) Protein and mRNA in the Normal Adult Rat CNS: Evidence for Anterograde Axonal Transport

A sensitive immunohistochemical technique was used, along with highly specific affinity-purified antibodies to brain-derived neurotrophic factor (BDNF), to generate a detailed mapping of BDNF immunoreactivity (BDNF-ir) throughout the adult rat CNS. A parallel analysis of sites of BDNF synthesis was performed with in situ hybridization techniques using a cRNA probe to the exon encoding mature rat BDNF protein. These combined data revealed (1) groups of cell bodies containing diffuse BDNF-ir throughout the CNS that were strongly correlated with fields of cells containing BDNF mRNA; (2) varying degrees of BDNF-ir outside of cell bodies, in what appeared to be fibers and/or terminals; and (3) many regions containing extremely heavy BDNF-immunoreactive fiber/terminal labeling that lacked BDNF mRNA (e.g., medial habenula, central nucleus of the amygdala, bed nucleus of stria terminalis, lateral septum, and spinal cord). The latter observation suggested that in these regions BDNF was derived from anterograde axonal transport by afferent systems. In the two cases in which this hypothesis was tested by the elimination of select afferents, BDNF immunostaining was completely eliminated. These data, along with the observation that BDNF-ir was rarely found within dendrites or fibers en passage, suggest that BDNF protein produced in adult CNS neurons is polarized primarily along axonal processes and is preferentially stored in terminals within the innervation target.

Distribution of brain-derived neurotrophic factor in the rat pituitary gland.

Immunohistochemical techniques were used to determine the distribution of brain-derived neurotrophic factor (BDNF) in the adult rat pituitary gland. In the posterior lobe, BDNF staining was found only within fibers. Within the intermediate lobe, nearly all cells were labeled intensely for BDNF. In the anterior lobe, varying degrees of BDNF immunoreactivity were observed exclusively in cells shown by double-labeling techniques to contain thyroid stimulating hormone (TSH), although not all TSH-positive cells contained detectable BDNF labeling. These results are consistent with and extend information from previous studies demonstrating BDNF expression in the pituitary gland and further support the idea that this neurotrophin plays a role in endocrine function.