High-affinity Receptor For Brain-derived Neurotrophic Factor Research Articles

Synergistic effects of brain-derived neurotrophic factor and ciliary neurotrophic factor on cultured basal forebrain cholinergic neurons from postnatal 2-week-old rats

Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, and ciliary neurotrophic factor (CNTF), a member of the neurocytokine family, are known to have synergistic effects on motoneurons, but such synergistic effect has not been studied in detail especially in the brain. In the present study, we examined the synergistic effects of BDNF and CNTF on the survival of basal forebrain cholinergic neurons cultured from postnatal 2-week-old (P2w) rats. Although BDNF is well-known to promote the survival of basal forebrain cholinergic neurons in P2w culture, CNTF had little effect on the survival of choline acetyltransferase (ChAT)-positive neurons and did not increase ChAT activity in the culture. However, CNTF enhanced BDNF-mediated promotion of cell survival of cholinergic neurons when added concomitantly. BDNF alone induced only a three-fold increase in ChAT activity in control cultures, but the concomitant addition of CNTF resulted in an eight-fold increase. CNTF did not enhance BDNF-mediated cell survival of total neurons from the basal forebrain, hippocampus or cerebellum, suggesting that the synergistic effects of CNTF on the BDNF-mediated increase of viability might be strong in basal forebrain cholinergic neurons. CNTF also enhanced the neurotrophin-4/5-mediated increase of ChAT activity, but not the nerve growth factor (NGF)-mediated one. Furthermore, the BDNF-mediated increase was also enhanced by leukemia inhibitory factor but not by interleukin-6. Similar synergistic pattern between neurotrophins and cytokines were also observed in the induction of ChAT activity in embryonic basal forebrain culture. These results suggest that TrkB, a functional high-affinity receptor of BDNF and NT-4/5, and LIFR β, a receptor component contained in CNTF and LIF receptor complex, might be involved in the observed synergistic effects.

Expression of trkB mRNA is altered in rat hippocampus after experimental brain trauma

Recent investigations have shown that expression of mRNAs for the neurotrophins brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) is differentially altered in the hippocampus following traumatic brain injury. In the present study, modulation of neurotrophin receptor expression was examined in the hippocampus in a rat model of traumatic brain injury using in situ hybridization. Messenger RNA for trkB, the high-affinity receptor for BDNF and neurotrophin-4 (NT-4), was increased between 3 and 6 h bilaterally in the dentate gyrus following a lateral fluid-percussion brain injury of moderate severity (2.0–2.1 atm). No time-dependent alterations were observed for trkB mRNA in hippocampal subfields CA1 and CA3. Levels of mRNA for trkC, the high-affinity receptor for NT-3, did not change in any region of the hippocampus. These data demonstrate that lateral fluid-percussion injury modulates expression of trkB mRNA in the hippocampus and support a role for BDNF/trkB signalling mechanisms in secondary events associated with traumatic brain injury.

Tyrosine kinase B messenger RNA expression in normal human brain and in the substantia nigra of parkinsonian patients: an in situ hybridization study

Tyrosine kinase B is a high-affinity receptor for brain-derived neurotrophic factor, a neurotrophin acting on numerous cells, including dopaminergic neurons. Yet, little is known of its expression in the human brain. We report an in situ hybridization analysis of tyrosine kinase B messenger RNA, encoding the catalytic form of the receptor, in the human brain post mortem. Its expression was found to be widespread but heterogenous among all the cerebral structures studied, the highest level being found in the cerebral cortex and the cerebellum. A strong but less intense staining was observed in the striatum, nucleus basalis of Meynert, hippocampus, tegmental pedonculopontinus nucleus and substantia nigra pars compacta. Combined immunohistochemistry for tyrosine hydroxylase and in situ hybridization for tyrosine kinase B messenger RNA showed that within the substantia nigra pars compacta a major proportion of dopaminergic neurons expressed tyrosine kinase B messenger RNA. We compared tyrosine kinase B messenger RNA expression in the mesencephalon of six control subjects and five patients with Parkinson's disease, a neurodegenerative disorder characterized by a severe loss of dopaminergic neurons. Despite the fact that the number of tyrosine kinase B messenger RNA-containing neurons was dramatically reduced in the substantia nigra pars compacta and ventral tegmental area of patients with Parkinson's disease, the level of tyrosine kinase B messenger RNA was unchanged in the remaining neurons in diseased brains. These results suggests that tyrosine kinase B is not involved in the process of neuronal death in Parkinson's disease. Furthermore, expression of brain-derived neurotrophic factor high-affinity receptor in patients could allow this neurotrophin to be used to prevent degeneration of surviving neurons at early stages of the disease.

Hippocampal mossy fiber sprouting is not impaired in brain-derived neurotrophic factor-deficient mice.

In human temporal lobe epilepsy, a loss of hilar neurons followed by the sprouting of recurrent mossy fiber collaterals and the reinnervation of free synaptic sites on granule cell dendrites are discussed as possible mechanisms underlying hippocampal hyperexcitability. Dentate granule cells have been shown to upregulate brain-derived neurotrophic factor (BDNF) as well as TrkB, the high-affinity receptor for BDNF, in response to limbic seizures. This raised the possibility that BDNF is an important factor in hippocampal mossy fiber sprouting. Here we have used slice cultures of hippocampus, in which mossy fibers sprout and form a supragranular plexus in response to granule cell deafferentation, and have compared cultures from early postnatal BDNF-deficient mice and wild-type mice. We demonstrate that there is sprouting of supragranular mossy fibers in cultured slices from both BDNF knock-out and wild-type mice. We conclude that BDNF is not an essential factor for mossy fiber sprouting. However, our data do not exclude a role for BDNF in mossy fiber sprouting in wild-type mice, as compensatory mechanisms might have become effective in the mutant.

A BRAIN-DERIVED NEUROTROPHIC FACTOR (BDNF) RELATED SYSTEM IS INVOLVED IN THE MAINTENANCE OF THE POLYINNERVATE TORPEDO ELECTRIC ORGAN

Target-derived molecules are essential for the maintenance of neuron survival. In the present work, we introduce the electric organ of Torpedo marmorata as a tool for the study of trophic interactions in a polyinnervate system. This electric organ maintains a large number of cholinergic terminals on the postsynaptic cell surface. We have observed that a soluble extract derived from the electric organ induces the maturation of Xenopus oocytes injected with presynaptic plasma membranes (PSPM), indicating that a trophic system may exist. Moreover, we have detected a p75NGFR related protein in PSPM by Western blot analysis. These results suggest the presence of a neurotrophin-related system maintaining the polyinnervate electric organ. Furthermore, molecular experiments showed that the brain-derived neurotrophic factor (BDNF) is the neurotrophin operating in our model. Using degenerate oligonucleotides which comprise a conserved fragment of all neurotrophins, we have only amplified by polymerase chain reaction a BDNF fragment. In a similar way, we have amplified and cloned a fragment of the TrkB/C high affinity BDNF receptor. The fact that degerate oligonucleotides only amplify BDNF allows us to conclude that the polyinnervation is maintained by this neurotrophin either alone or in combination with other trophic factors. © 1997 Elsevier Science Ltd

Low-level cadmium exposure of lactating rats causes alterations in brain serotonin levels in the offspring.

Effects on monoaminergic and cholinergic transmitter systems as well as neurotrophins were characterized in developing Sprague-Dawley rats directly exposed to 5 ppm cadmium in the drinking water or indirectly via exposed dams. Cadmium was given to dams during the lactation period, from parturition to postnatal day 17, and/or to the offspring until postnatal day 42. Cresyl violet staining and glial fibrillary acidic protein immunohistochemistry did not reveal any obvious neuropathology after cadmium exposure. Following high-power microwave fixation, concentrations of acetylcholine (ACh) and monoamines were determined in cerebral cortex, striatum, and hippocampus using HPLC with electro-chemical detection. ACh, dopamine, and noradrenaline levels were not significantly affected after the different cadmium exposures. Cortical levels of serotonin were significantly reduced in rats exposed to cadmium during lactation as well as in rats exposed to cadmium during both lactation and postweaning. A major decrease in 5-hydroxyindoleacetic acid was found in cortex and hippocampus in rats exposed to cadmium during lactation. The regional characteristics of cadmium toxicity as reflected in changes of neurotrophins were studied using in situ hybridization histochemistry with oligonucleotide probes and phosphoimaging evaluation. No significant changes in the mRNA expression of brain-derived neurotrophic factor (BDNF), neurotrophin-3, and the high-affinity tyrosine kinase receptor of BDNF, trkB, were detected. The present results demonstrate that exposure to levels of cadmium as low as 5 ppm in the drinking water leads to neurochemical disturbances of the serotonergic system in the offspring during the lactational period.

Functional trkB neurotrophin receptors are intrinsic components of the adult brain postsynaptic density

Neurotrophins have long been thought to act as target-derived factors that regulate the survival and differentiation of afferent neurons. Recently, brain-derived neurotrophic factor (BDNF) was shown to elicit rapid increases in synaptic activity of cultured hippocampal neurons by enhancing responsiveness to excitatory input. These findings suggest a postsynaptic localization of neurotrophin receptors. In this study, we examined the expression of trkB, a high-affinity receptor for BDNF, in the postsynaptic density (PSD), a proteinaceous specialization of the postsynaptic membrane. Western blot analyses with antibodies to trkB revealed localization to the PSD in adult rat cerebral cortex and hippocampus. Only the full-length, active form of trkB was detected in PSD samples. BDNF treatment of the adult cortical PSD resulted in a 5-fold increase in trkB autophosphorylation, supporting the contention that the PSD contains functional trkB. Truncated trkB, which does not contain the tyrosine kinase signaling domain, though present in membrane fractions, was undetectable in the PSD. The presence of trkB in the PSD is consistent with a role for neurotrophins in the regulation of synaptic activity via direct postsynaptic mechanisms.

Induced expression of neurotrophins in transgenic mice overexpressing ornithine decarboxylase and overproducing putrescine

Transgenic mice overexpressing ornithine decarboxylase (ODC) were recently generated (Halmekytö et al.: Biochem Biophys Res Commun 180:262-267, 1991). The dramatic ODC overexpression resulted in a very high accumulation of the polyamine putrescine in the brain. As elevated polyamine levels in the brain are believed to be associated with neuronal damage, we studied whether enhanced putrescine accumulation in the brain of these mice affects the expression of neurotrophins and their high affinity receptors. Northern blot analysis indicated that mRNA levels of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF) and neurotrophin-3 (NT-3) were significantly elevated about 1.5-fold in the hippocampus of ODC transgenic mice as compared with control animals. The levels of BDNF, NGF and NT-3 mRNA were also elevated in the kidneys of the transgenic mice. In eight other tissues examined there were no significant differences. The expression pattern of BDNF mRNA and of trkB and trkC (high affinity receptors for BDNF and NT-3 respectively) immunoreactivity in the hippocampal formation did not reveal significant differences. The induction of the expression of neurotrophins could belong to neuroprotective measures triggered by ODC overexpression.

Induced expression of neurotrophins in transgenic mice overexpressing ornithine decarboxylase and overproducing putrescine

Transgenic mice overexpressing ornithine decarboxylase (ODC) were recently generated (Halmekytö et al.: Biochem Biophys Res Commun 180:262–267, 1991). The dramatic ODC overexpression resulted in a very high accumulation of the polyamine putrescine in the brain. As elevated polyamine levels in the brain are believed to be associated with neuronal damage, we studied whether enhanced putrescine accumulation in the brain of these mice affects the expression of neurotrophins and their high affinity receptors. Northern blot analysis indicated that mRNA levels of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF) and neurotrophin-3 (NT-3) were significantly elevated about 1.5-fold in the hippocampus of ODC transgenic mice as compared with control animals. The levels of BDNF, NGF and NT-3 mRNA were also elevated in the kidneys of the transgenic mice. In eight other tissues examined there were no significant differences. The expression pattern of BDNF mRNA and of trkB and trkC (high affinity receptors for BDNF and NT-3 respectively) immunoreactivity in the hippocampal formation did not reveal significant differences. The induction of the expression of neurotrophins could belong to neuroprotective measures triggered by ODC overexpression. © 1996 Wiley-Liss, Inc.

S-34-3 - Neurotrophic factors: Biology & clinical potential

Development and maintenance of the mammalian nervous system require neurotrophic factors. The most well-characterized neurotrophic factor nerve growth factor (NGF) has recently been shown to be a member of a family of structurally related proteins that also includes brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4 (NT-4). An evolutionary study of these factors, collectively known as neurotrophins, in representative groups of vertebrates showed that the four factors are highly conserved from fishes to mammals. Chimeric molecules were constructed where different combinations of sequences from BDNF replaced the corresponding sequences in NGF. Assays for biological activity of such molecules showed that the biological specificities of the two proteins are obtained by specific combinations of a set of sequences that differ between the two molecules. In the rat embryo, expression of both the neurotrophins and the essential components of their high-affinity receptors (trk, trkB and trkC) are developmentally regulated. From their sites of synthesis, both target-derived and local modes of neurotrophic actions can be inferred and the development pattern of mRNA expression suggests the existence of serval neurotrophin dependent neuronal systems where these factors have not previously been implicated. In the adult rat brain, the highest levels of NGF, BDNF and NT-3 mRNAs are found within the hippocampus where they are expressed in a set of partially non-overlapping neurons. Expression of NGF and BDNF mRNAs in the brain appears to be regulated by neuronal activity and the levels of these mRNAs show a marked and transient increase following epileptic seizures, cerebral ischemia and hypoglycemic coma. The increases in NGF and BDNF mRNAs appear to be mediated via a release of glutamate activating AMPA receptors within the hippocampus as well as by a rise in the intracellular calcium levels. As a first step towards an understanding of the molecular mechanisms governing BDNF mRNA expression the rat BDNF gene was characterized. Multiple transcripts with different 5′-untranslated sequences are transcribed from this gene. These mRNAs are synthesized by a differential, tissue specific, usage of four different promoters within the BDNF gene. Two of these promoters appear to confirm a marked seizure-induced increase of BDNF mRNA in the brain. The product of the trkB gene encoding an essential component of the high-affinity BDNF receptor, is also expressed in the hippocampus where it is spatially and temporally regulated in a manner similar to BDNF. These data suggest that BDNF and its receptor could, in a paracrine/autocrine fashion, play a role in kindling-associated neural plasticity and in neuronal protection following ischemic and hypoglycemic insults. Alanine-scanning mutagenesis combined with receptor binding and biological assays were used to assess the functional importance of amino acid residues exposed on the surface of the NGF dimer. The results show that Lys-32, Lys-34 and Lys-95 form a positively charged interface involved in binding to the low-affinity NGF receptor (p75ngfr). Mutated molecules that do not bind to p75ngfr but still retain binding to p 140 trk and biological activity demonstrate for the first time a functional dissociation between the two NGF receptors.

S-34-4 - Opposing effects of stress andantidepressants of neurotrophic factors

Development and maintenance of the mammalian nervous system require neurotrophic factors. The most well-characterized neurotrophic factor nerve growth factor (NGF) has recently been shown to be a member of a family of structurally related proteins that also includes brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4 (NT-4). An evolutionary study of these factors, collectively known as neurotrophins, in representative groups of vertebrates showed that the four factors are highly conserved from fishes to mammals. Chimeric molecules were constructed where different combinations of sequences from BDNF replaced the corresponding sequences in NGF. Assays for biological activity of such molecules showed that the biological specificities of the two proteins are obtained by specific combinations of a set of sequences that differ between the two molecules. In the rat embryo, expression of both the neurotrophins and the essential components of their high-affinity receptors (trk, trkB and trkC) are developmentally regulated. From their sites of synthesis, both target-derived and local modes of neurotrophic actions can be inferred and the development pattern of mRNA expression suggests the existence of serval neurotrophin dependent neuronal systems where these factors have not previously been implicated. In the adult rat brain, the highest levels of NGF, BDNF and NT-3 mRNAs are found within the hippocampus where they are expressed in a set of partially non-overlapping neurons. Expression of NGF and BDNF mRNAs in the brain appears to be regulated by neuronal activity and the levels of these mRNAs show a marked and transient increase following epileptic seizures, cerebral ischemia and hypoglycemic coma. The increases in NGF and BDNF mRNAs appear to be mediated via a release of glutamate activating AMPA receptors within the hippocampus as well as by a rise in the intracellular calcium levels. As a first step towards an understanding of the molecular mechanisms governing BDNF mRNA expression the rat BDNF gene was characterized. Multiple transcripts with different 5′-untranslated sequences are transcribed from this gene. These mRNAs are synthesized by a differential, tissue specific, usage of four different promoters within the BDNF gene. Two of these promoters appear to confirm a marked seizure-induced increase of BDNF mRNA in the brain. The product of the trkB gene encoding an essential component of the high-affinity BDNF receptor, is also expressed in the hippocampus where it is spatially and temporally regulated in a manner similar to BDNF. These data suggest that BDNF and its receptor could, in a paracrine/autocrine fashion, play a role in kindling-associated neural plasticity and in neuronal protection following ischemic and hypoglycemic insults. Alanine-scanning mutagenesis combined with receptor binding and biological assays were used to assess the functional importance of amino acid residues exposed on the surface of the NGF dimer. The results show that Lys-32, Lys-34 and Lys-95 form a positively charged interface involved in binding to the low-affinity NGF receptor (p75ngfr). Mutated molecules that do not bind to p75ngfr but still retain binding to p 140 trk and biological activity demonstrate for the first time a functional dissociation between the two NGF receptors.

The effects of growth factors on the survival and differentiation of cultured dentate gyrus neurons

Dentate granule cells (DGCs) are the principal cell population of the hippocampal dentate gyrus, and granule cells provide the main excitation to the hippocampus proper via their mossy fibers axons. Although it is well established that granule cells express various growth factors and growth factor receptors, the functional effects of growth factors on the normal development and response to injury of granule cells are relatively unknown. To address this question, primary cultures enriched in DGCs were prepared by microdissecting hippocampal slices from neonatal rats and growing dissociated cells in defined media with added nerve growth factor, brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), neurotrophin-4/5 (NT4/5), ciliary neurotrophic factor, basic fibroblast growth factor (bFGF), or vehicle. The effects on cell survival and morphology were quantified by studying neuron-specific enolase-immunostained cells at various time points, plating densities, host ages, and growth factor concentrations. BDNF or bFGF significantly increased both neuronal survival and differentiation by 30-80% compared with control cultures. Maximal effects were observed at relatively longer time points (5-12 d), with younger cells (postnatal day 3-5) and at lowest plating densities. Addition of a trkB-IgG fusion protein that blocks the activity of BDNF or NT4/5 inhibited the effects of BDNF and attenuated the differentiation of cells cultured at high plating densities. Furthermore, treatment of cultures with the kinase inhibitor K252b specifically blocked the effects of BDNF, suggesting involvement of trkB (the high-affinity BDNF receptor) in BDNF-induced differentiation. These results show that growth properties of cultured neonatal DGCs are influenced by exogenously applied BDNF or bFGF in a time-, age-, and density-dependent manner. The effect of plating density suggests an endogenous expression of growth factors in these culture conditions, and this is mediated in part by endogenous BDNF acting via a tyrosine kinase receptor. Combined with previous work showing that various growth factors and their receptors are expressed by DGCs, these findings provide strong support for the hypothesis that BDNF and bFGF influence both the growth and development of DGCs in vivo.

1301 Different effects of neurotrophins on neurite regeneration from a DRG explant of an adult rat

Development and maintenance of the mammalian nervous system require neurotrophic factors. The most well-characterized neurotrophic factor nerve growth factor (NGF) has recently been shown to be a member of a family of structurally related proteins that also includes brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4 (NT-4). An evolutionary study of these factors, collectively known as neurotrophins, in representative groups of vertebrates showed that the four factors are highly conserved from fishes to mammals. Chimeric molecules were constructed where different combinations of sequences from BDNF replaced the corresponding sequences in NGF. Assays for biological activity of such molecules showed that the biological specificities of the two proteins are obtained by specific combinations of a set of sequences that differ between the two molecules. In the rat embryo, expression of both the neurotrophins and the essential components of their high-affinity receptors (trk, trkB and trkC) are developmentally regulated. From their sites of synthesis, both target-derived and local modes of neurotrophic actions can be inferred and the development pattern of mRNA expression suggests the existence of serval neurotrophin dependent neuronal systems where these factors have not previously been implicated. In the adult rat brain, the highest levels of NGF, BDNF and NT-3 mRNAs are found within the hippocampus where they are expressed in a set of partially non-overlapping neurons. Expression of NGF and BDNF mRNAs in the brain appears to be regulated by neuronal activity and the levels of these mRNAs show a marked and transient increase following epileptic seizures, cerebral ischemia and hypoglycemic coma. The increases in NGF and BDNF mRNAs appear to be mediated via a release of glutamate activating AMPA receptors within the hippocampus as well as by a rise in the intracellular calcium levels. As a first step towards an understanding of the molecular mechanisms governing BDNF mRNA expression the rat BDNF gene was characterized. Multiple transcripts with different 5′-untranslated sequences are transcribed from this gene. These mRNAs are synthesized by a differential, tissue specific, usage of four different promoters within the BDNF gene. Two of these promoters appear to confirm a marked seizure-induced increase of BDNF mRNA in the brain. The product of the trkB gene encoding an essential component of the high-affinity BDNF receptor, is also expressed in the hippocampus where it is spatially and temporally regulated in a manner similar to BDNF. These data suggest that BDNF and its receptor could, in a paracrine/autocrine fashion, play a role in kindling-associated neural plasticity and in neuronal protection following ischemic and hypoglycemic insults. Alanine-scanning mutagenesis combined with receptor binding and biological assays were used to assess the functional importance of amino acid residues exposed on the surface of the NGF dimer. The results show that Lys-32, Lys-34 and Lys-95 form a positively charged interface involved in binding to the low-affinity NGF receptor (p75ngfr). Mutated molecules that do not bind to p75ngfr but still retain binding to p 140 trk and biological activity demonstrate for the first time a functional dissociation between the two NGF receptors.

Differential distribution of exogenous BDNF, NGF, and NT‐3 in the brain corresponds to the relative abundance and distribution of high‐affinity and low‐affinity neurotrophin receptors

To evaluate effective means for delivering exogenous neurotrophins to neuron populations in the brain, we compared the distribution and transport of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and neurotrophin-3 (NT-3) following intracerebral delivery. Rats received an injection of radioiodinated or unlabeled neurotrophin into the lateral ventricle and were killed humanely after 1.5-24 hours. Other rats received continuous infusion of unlabeled neurotrophin into the lateral ventricle, the striatum, or the hippocampus for 3-14 days. The neurotrophins were detected by autoradiography or immunohistochemistry. There were striking differences between BDNF, NGF, and NT-3 in their penetration through brain tissue. These differences occurred regardless of the site or method of delivery, but were most pronounced following a bolus intracerebroventricular (ICV) injection. After ICV injection, NGF was widely distributed in tissues around the ventricles and at the surface of the brain, whereas the penetration of BDNF into brain tissue was distinctly less than that of NGF, and the penetration of NT-3 was intermediate. An ICV injection of NGF produced prominent but transient labeling of cells that contain the low-affinity NGF receptor, whereas an injection of BDNF prominently labeled the ventricular ependyma. During ICV infusion (12 micrograms/day), the distribution of BDNF was no greater than that observed after a 12-micrograms bolus injection. A sixfold increase in the amount of BDNF infused (72 micrograms/day) produced a more widespread distribution in the brain and doubled the depth of penetration into periventricular tissues near the cannula. Corresponding to their differences in penetration, NGF was retrogradely transported by basal forebrain cholinergic neurons after ICV or intrastriatal delivery, whereas NT-3 was transported by a few basal forebrain neurons after ICV delivery, and BDNF was rarely detected in neurons after ICV delivery. Delivery of BDNF directly to the striatum or the hippocampus labeled numerous neurons in nuclei afferent to these structures. In situ hybridization studies confirmed that the high-affinity BDNF receptor (TrkB) was much more widely expressed in neurons than was the high-affinity NGF receptor (TrkA). Moreover, mRNA for truncated forms of TrkB was expressed at high levels in the ependyma, the choroid epithelium, and the gray matter. It is likely that binding of BDNF to TrkB, which appears to be more abundant and ubiquitous than TrkA, restricts the diffusion of BDNF relative to that of NGF.

Expression of the proto-oncogene, trk, receptors in the developing rat retina.

The neurotrophins, including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and NT-4/5 are important in a variety of developmental processes in the peripheral and central nervous systems. These molecules bind to a low-affinity receptor and to distinct high-affinity receptors. The high-affinity receptor for NGF is the proto-oncogene product, p140trkA(trkA). Isoforms of p140trkA, p145trkB(trkB), and p140trkC(trkC), are the primary high-affinity receptors for BDNF and NT-3, respectively. We evaluated the developmental regulation of the high-affinity neurotrophin receptors in the rat retina using polyclonal antibodies directed to a highly conserved region of the C-terminus of the p140trkA isoforms (pantrk) and antibodies directed to unique amino-acid sequences of p140trkA, p145trkB, and p140trkC. Immunoreactivities for trkA and trkB, as well as pantrk, were detected in the developing retina and showed similar distributions. At similar antibody concentrations, trkC immunoreactivity was not detected. In the embryo, immunoreactivties were present in cells located throughout the neuroblastic retina, especially in the inner retinal layers, and in fibers in the nerve fiber layer and optic nerve. In the newborn retina, immunoreactivities for these two receptor isoforms were localized to numerous somata in the inner nuclear layer (INL), as well as to cells in the ganglion cell layer (GCL) and axons in the nerve fiber layer and optic nerve. A similar pattern of immunostaining persisted throughout the first postnatal week. By postnatal day-10, immunostaining was confined to large-diameter cells in the GCL, both heavily stained and lightly stained cells in the INL and a plexus of processes in the inner plexiform layer (IPL).(ABSTRACT TRUNCATED AT 250 WORDS)

Overlapping and distinct actions of the neurotrophins BDNF, NT-3, and NT-4/5 on cultured dopaminergic and GABAergic neurons of the ventral mesencephalon

The neurotrophins brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4/5 (NT-4/5) were compared for their effects in promoting the survival and/or regulation of expression of phenotypic markers of dopaminergic and GABAergic neurons in cultures derived from embryonic rat ventral mesencephalon. Dopaminergic neuron number and phenotypic expression were monitored by tyrosine hydroxylase (TH) immunocytochemistry, and measurement of high-affinity dopamine uptake activity and dopamine content, respectively. High-affinity GABA uptake, glutamic acid decarboxylase (GAD) activity, and endogenous GABA content were used to detect GABAergic neurons. Seven days of treatment with either BDNF or NT-3 resulted in dose-dependent increases in the number of TH-positive neurons, with maximal responses of 3-fold and 2.3-fold, respectively. Dopamine uptake activity and dopamine content were similarly increased. The effects of BDNF and NT-3 on dopamine uptake activity showed no additivity. NT-4/5 treatment elicited the greatest increase (7-fold) in the number of TH-positive neurons, as well as a 2.6-fold increase in dopamine content. In marked contrast to BDNF or NT-3, NT-4/5 had no effect on dopamine uptake capacity. BDNF, NT-3, or NT-4/5 also produced dose-dependent elevations of 2-3-fold in GABA uptake activity. These effects were not additive. GAD activity was increased by BDNF (1.8-fold) and NT-3 (threefold) treatment, but not by NT-4/5, whereas GABA content was increased to a similar extent by all three neurotrophins. NGF had no effect on any of the parameters measured in this study. Northern analyses indicated that the mRNAs encoding TrkB and TrkC, the functional high-affinity receptors for BDNF and NT-4/5, and NT-3, respectively, are expressed in the substantia nigra of adult rat brain, as well as in cultures of developing ventral mesencephalon. Taken together, our results indicate that BDNF and NT-3 have broadly similar effects in promoting the survival and differentiated phenotype of both dopaminergic and GABAergic neurons of the developing substantia nigra. Although BDNF and NT-4/5 are thought to act through the same high-affinity receptor, TrkB, it is evident that these two neurotrophins have distinct as well as overlapping actions toward mesencephalic dopaminergic or GABAergic neurons.

The neurotrophins BDNF, NT-3 and NT-4/5, but not NGF, up-regulate the cholinergic phenotype of developing motor neurons.

Although developing motor neurons express low-affinity nerve growth factor (NGF) receptors, there is no known biological effect of NGF on developing or adult motor neurons. In this study, we found that, unlike NGF, brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4/5 (NT-4/5) stimulated cholinergic phenotype by increasing choline acetyltransferase (CAT) activity in cultures enriched with embryonic rat motor neurons. Ciliary neurotrophic factor (CNTF) also stimulated CAT activity. The effects of BDNF and NT-4/5 on CAT activity appeared to be synergistic with that of CNTF. Cotreatment with BDNF and NT-3 resulted in an additive effect, suggesting that signal transduction was mediated through different high-affinity receptors tyrosine kinases B and C (Trk B and Trk C). However, cotreatment with BDNF and NT-4/5 did not result in an increase in CAT activity greater than that of either BDNF or NT-4/5 alone, suggesting that their effects were mediated via the same receptor Trk B. Supporting our findings that spinal cholinergic neurons are responsive to trophic actions of members of the neurotrophin family, motor neuron-enriched cultures were found to express mRNA for Trk B and Trk C, which have been identified as high-affinity receptors for BDNF and NT-4/5, and NT-3, respectively.

Juniors' hours: measuring the strength of feeling.

Brain-derived neurotrophic factor (BDNF) is transported anterogradely in neurons of the CNS and can be released by activity-dependent mechanisms to regulate synaptic plasticity. However, few neural networks have been identified in which the production, transport, and effects of BDNF on postsynaptic neurons can be analyzed in detail. In this study, we have identified such a network. BDNF has been colocalized by immunocytochemistry with tyrosine hydroxylase (TH) in nerve fibers and nerve terminals within the lateral septum of rats. BDNF-containing nerve fibers terminate on a population of calbindin-containing neurons in lateral septum that contain TrkB, the high-affinity receptor for BDNF. Overexpression of BDNF in noradrenergic neurons increased levels of calbindin in septum, as well as in whole-brain lysates. Septal levels of calbindin and BDNF partially decreased after unilateral lesions of the medial forebrain bundle (MFB), induced with 6-hydroxydopamine, a treatment that abolished TH staining. These data suggest that BDNF is anterogradely transported within the MFB in catecholaminergic neurons arising from brainstem nuclei. To determine whether BDNF affects the production of calbindin in lateral septal neurons directly, we tested the effects of BDNF on cultures of septal neurons from embryonic day 16–17 rats. BDNF promoted the expression of calbindin, as well as the arborization of calbindin-containing neurons, but BDNF had no effect on cell division or survival. Together, these results suggest that BDNF, anterogradely transported in catecholaminergic neurons, regulates calbindin expression within the lateral septum.