Brain-derived Neurotrophic Factor Treatment Research Articles

Rapid enrichment of presynaptic protein in boutons undergoing classical conditioning is mediated by brain-derived neurotrophic factor

Presynaptic structural modifications are thought to accompany activity-dependent synaptic plasticity and learning. This may involve the conversion of nonfunctional synapses into active ones or the generation of entirely new synapses. Here, using an in vitro neural analog of classical conditioning, we investigated presynaptic structural changes restricted to auditory nerve synapses that convey the conditioned stimulus (CS) by tract tracing using fluorescent tracers combined with immunostaining for the synaptic vesicle-associated protein synaptophysin. The results show that the size of presynaptic auditory boutons increased and the area and fluorescence intensity of punctate staining for synaptophysin were enhanced after conditioning. This occurred only for auditory nerve boutons apposed to the dendrites but not the somata of abducens motor neurons. Conditioning increased the percentage of boutons that were immunopositive for synaptophysin and enhanced the number of synaptophysin puncta they contained. Pretreatment with antibodies against brain-derived neurotrophic factor (BDNF) inhibited these conditioning-induced structural changes. There was also a net increase in the number of boutons apposed to abducens motor neurons after conditioning or BDNF treatment. These data indicate that the rapid enrichment of presynaptic boutons with proteins required for neurotransmitter recycling and release occurs during classical conditioning and that these processes are mediated by BDNF.

BDNF Promotes EGF-Induced Proliferation and Migration of Human Fetal Neural Stem/Progenitor Cells via the PI3K/Akt Pathway

Neurogenesis is a complex process, which contributes to the ability of the adult brain to function normally and adapt to diseases. Epidermal growth factor (EGF) is known to play an important role in neurogenesis; however, the underlying mechanism is still unclear. Here, we hypothesized that brain-derived neurotrophic factor (BDNF) can enhance the effect of EGF on neurogenesis. Using in vitro cell culture of aborted human fetal brain tissues, we investigated proliferation and migration of neural stem/progenitor cells (NSPCs) after treatment with EGF and different concentrations of BDNF. EGF stimulated proliferation and migration of NSPCs, and this effect was significantly enhanced by co-incubation with BDNF. In the NSPCs treated with 50 ng/mL BDNF, BrdU incorporation was significantly increased (from 7.91% to 17.07%), as compared with that in the control. Moreover, the number of migrating cells was at least 2-fold higher than that in the control. Furthermore, phosphorylation of Akt-1 was increased by BDNF treatment, as well. By contrast, the enhancing effect of BDNF on EGF-induced proliferation and migration of NSPCs were abolished by an inhibitor of PI3K, LY294002. These findings suggest that BDNF promotes EGF-induced proliferation and migration of NSPC through the PI3K/Akt pathway, providing significant insights into not only the mechanism underlying EGF-induced neurogenesis but also potential neuronal replacement strategies to treat brain damage.

Protection by [6]-shogaol against lipopolysaccharide-induced toxicity in murine astrocytes is related to production of brain-derived neurotrophic factor

[6]-Shogaol has beneficial effects in spinal neuronal regeneration, but associated molecules and mechanisms are not identified. Neurotrophic factors, including brain-derived neurotrophic factor (BDNF), are associated with proliferation and differentiation of neuronal cells and exert a neuroprotective effect in neurodegenerative models. We investigated whether treatment with [6]-shogaol increases BDNF expression in lipopolysaccharide (LPS)-treated astrocytes, and examined the effect of [6]-shogaol on neuronal protection. [6]-Shogaol significantly attenuated the cell death induced by LPS. Western blotting showed that [6]-shogaol treatment reduced Bax expression and increased B-cell lymphoma (Bcl)-2 and BclxL expression in LPS-treated cells, consistent with the effects of BDNF treatment. Furthermore, K252a, a blocker of neurotrophic factors, attenuated the cellular protective effects of [6]-shogaol and BDNF. This study provides the first evidence that [6]-shogaol increases the expression of BDNF in LPS-treated astrocytes. Furthermore, these experimental results indicate that production of BDNF in astrocytes might be related to altered cell viability following [6]-shogaol treatment. Thus, the neuroprotective effects of [6]-shogaol is mediated by up-regulation of BDNF.

BDNF regulates GLAST and glutamine synthetase in mouse retinal Müller cells

This study investigated whether brain-derived neurotrophic factor (BDNF) regulates the L-glutamate/L-aspartate transporter (GLAST) and glutamine synthetase (GS) in mouse retinal Müller cells (RMCs) under normal and hypoxic conditions. Mouse RMCs were treated with recombinant human BDNF (50, 75, 100, 125, or 150 ng/ml) for 24 h or underwent hypoxia induced by CoCl(2) (125 µM; 6, 12, 24, 48, or 72 h). An additional group underwent combined treatment with BDNF (100 ng/ml; 24, 48, 72, or 96 h) and CoCl(2) (125 µM/ml; 72 h). GLAST and GS mRNA and protein expression, L-[3,4-3H]-glutamic acid uptake, and apoptosis were assessed. BDNF dose-dependently up-regulated GLAST and GS mRNA and protein and increased glutamate uptake. Similarly, in early-stage CoCl(2)-induced hypoxia, GLAST and GS were up-regulated and glutamate uptake increased, but these decreased over time. BDNF also up-regulated GLAST and GS and increased glutamate uptake when RMCs under CoCl(2) induced hypoxic condition. However, BDNF treatment 24 h before CoCl(2) had no effect on GLAST or GS expression. CoCl(2) alone or combined with BDNF did not induce apoptosis. Hypoxia rapidly increased GLAST and GS expressions. This effect was transient, perhaps due to compensatory mechanisms that reduce GLAST and GS by 72 h. BDNF can up-regulate GLAST and GS and increase glutamate uptake during hypoxia, and these functions may underlie its neuroprotective effects.

Brain-Derived Neurotrophic Factor Modulates Auditory Function in the Hearing Cochlea

Neurotrophins prevent spiral ganglion neuron (SGN) degeneration in animal models of ototoxin-induced deafness and may be used in the future to improve the hearing of cochlear implant patients. It is increasingly common for patients with residual hearing to undergo cochlear implantation. However, the effect of neurotrophin treatment on acoustic hearing is not known. In this study, brain-derived neurotrophic factor (BDNF) was applied to the round window membrane of adult guinea pigs for 4 weeks using a cannula attached to a mini-osmotic pump. SGN survival was first assessed in ototoxically deafened guinea pigs to establish that the delivery method was effective. Increased survival of SGNs was observed in the basal and middle cochlear turns of deafened guinea pigs treated with BDNF, confirming that delivery to the cochlea was successful. The effects of BDNF treatment in animals with normal hearing were then assessed using distortion product otoacoustic emissions (DPOAEs), pure tone, and click-evoked auditory brainstem responses (ABRs). DPOAE assessment indicated a mild deficit of 5 dB SPL in treated and control groups at 1 and 4 weeks after cannula placement. In contrast, ABR evaluation showed that BDNF lowered thresholds at specific frequencies (8 and 16 kHz) after 1 and 4 weeks posttreatment when compared to the control cohort receiving Ringer's solution. Longer treatment for 4 weeks not only widened the range of frequencies ameliorated from 2 to 32 kHz but also lowered the threshold by at least 28 dB SPL at frequencies ≥16 kHz. BDNF treatment for 4 weeks also increased the amplitude of the ABR response when compared to either the control cohort or prior to treatment. We show that BDNF applied to the round window reduces auditory thresholds and could potentially be used clinically to protect residual hearing following cochlear implantation.

BDNF-Induced Increase of PSD-95 in Dendritic Spines Requires Dynamic Microtubule Invasions

Microtubules (MTs) are capable of entering dendritic spines in mature hippocampal neurons through dynamic polymerization. Although these MT invasions are directly associated with neuronal activity, their function remains unknown. Here we demonstrate in mouse hippocampal neurons that MT entries into spines regulate the increase in postsynaptic density-95 (PSD-95) protein after brain-derived neurotrophic factor (BDNF) treatment. Using multiwavelength total internal reflectance fluorescence microscopy, we show that BDNF prolonged the average MT dwell time in spines and that this effect was dependent on TrkB receptor activation. Further examination revealed that peaks of MT polymerization into spines corresponded to rapid PSD-95 increases in the spine head. Over time, spines targeted by MTs after BDNF application, but not before, showed a robust increase in PSD-95. Conversely, spines completely devoid of MT invasions showed no significant change in the level of PSD-95. Pharmacological inhibition of MT dynamics abolished the BDNF-induced increase in PSD-95. Together, these results support the hypothesis that the well known increase in PSD-95 within spines after BDNF treatment is dependent on MT invasions of dendritic spines. Thus, our study provides a direct link between dynamic MTs and the postsynaptic structure, and provides a functional role for MT invasion of dendritic spines.

Association analysis between the Val66Met polymorphism in the brain-derived neurotrophic factor (BDNF) gene and treatment response to venlafaxine XR in generalized anxiety disorder

While antidepressant drugs are used to treat generalized anxiety disorder (GAD), patients vary greatly in their treatment response. Evidence shows genetic factors may play a role in treatment response in GAD. We examined whether the BDNF gene, which has been shown to play a role in antidepressant treatment response in major depressive disorder (MDD), also has an effect in GAD. In our study, 155 patients diagnosed with GAD received venlafaxine XR treatment as part of an 18-month relapse prevention study. Genotypes were obtained for the BDNF functional variant rs6265 (Val66Met) for the entire sample (n=155); however, only the European American (EA) population was considered (n=111) for pharmacogenetic analysis. We did not find a significant association between rs6265 and antidepressant treatment response in our GAD population. Future studies in larger populations will need to be conducted to further elucidate the pharmacogenetic role of this variant in anxiety disorders.

Early reactions of brain-derived neurotrophic factor in plasma (pBDNF) and outcome to acute antidepressant treatment in patients with Major Depression

In Major Depressive Disorder, a growing data base suggests that the onset of antidepressants' action can be detected by improvement of depressive symptoms in the first 10-14 days of treatment. Previous studies showed that the mean concentration of the brain-derived neurotrophic factor (BDNF) in blood increases during antidepressant treatment and positively correlates with amelioration of MDD symptoms. We previously showed an association between very early changes of the serum BDNF concentration and treatment outcome (Tadić et al., 2011. Prog Neuropsychopharmacol Biol Psychiatry 35, 415-420). However, no study has yet investigated whether BDNF concentration in plasma increases in the early course of treatment and enables the prediction of final treatment outcome. The goal of this study was to investigate in MDD patients, whether the change of pBDNF in the early course of treatment is a specific and sensitive marker for final treatment outcome. For this purpose, we performed a naturalistic pilot study with 39 inpatients with MDD according to DSM-IV. Depression severity and pBDNF were measured in weekly intervals from baseline (EP) to endpoint (EP, max. week six) with the 21-item Hamilton Depression Rating Scale (HAMD-21) and enzyme-linked immunosorbent assay (ELISA), respectively. According to ROC-analysis, the best cut-off value for the prediction of response at EP is an increase of 338 pg/ml or 126%, respectively, of pBDNF between BL and day 7. The single markers pBDNF change and HAMD-21 improvement from BL-d7 predicted later treatment outcome with moderate to high sensitivity and specificity (pBDNF: 42% and 96%, resp.; HAMD improvement: 83% and 65%, resp.). The combined marker early pBDNF change plus HAMD-21 improvement at day 7 increased the specificity for response to 100%. Our data provide first preliminary evidence that an early change of pBDNF in conjunction with early improvement might be a peripheral marker predictive for treatment outcome in patients with MDD. This has to be confirmed in further investigations. This article is part of a Special Issue entitled 'Anxiety and Depression'.

Perifosine-induced inhibition of akt attenuates brain-derived neurotrophic factor/TrkB-induced chemoresistance in neuroblastoma in vivo

Neuroblastoma (NB) tumors expressing high levels of brain-derived neurotrophic factor (BDNF) and its receptor TrkB or activated Akt are associated with decreased event-free or overall survival in patients with NB. In the current study, the effect of perifosine, an Akt inhibitor, on the chemosensitivity of TrkB-expressing NB cells or tumors was evaluated. A tetracycline-regulated TrkB-expressing isogenic NB cell model system was tested. In this system, NB cells were treated with etoposide and/or perifosine both in vitro and in vivo. Inhibition of the target by perifosine was evaluated by Western blot analysis or kinase activity assay. Cell survival and tumor growth were investigated. In vitro BDNF treatment induced Akt phosphorylation and rescued cells from etoposide-induced cell death in cells with high TrkB expression, but not in cells with low TrkB expression. Pretreatment of high TrkB-expressing TB3 cells with perifosine blocked BDNF/TrkB-induced Akt phosphorylation and inhibited BDNF's protection of TB3 cells from etoposide treatment. In vivo, tumors with high TrkB expression were found to have elevated levels of phosphorylated Akt and were less sensitive to etoposide treatment compared with tumors with low TrkB expression. Mice treated with a combination of perifosine and etoposide were found to have a statistically significant decrease in tumor growth compared with mice treated with either etoposide or perifosine alone. Activation of Akt through the BDNF/TrkB signaling pathway induced chemoresistance in NB in vivo. Perifosine-induced inhibition of Akt increased the sensitivity of NB to chemotherapy. The results of the current study support the future clinical evaluation of an Akt inhibitor combined with cytotoxic drugs for the improvement of treatment efficacy.

Brain-derived neurotrophic factor promotes cochlear spiral ganglion cell survival and function in deafened, developing cats.

Postnatal development and survival of spiral ganglion (SG) neurons depend on both neural activity and neurotrophic support. Our previous studies showed that electrical stimulation from a cochlear implant only partially prevents SG degeneration after early deafness. Thus, neurotrophic agents that might be combined with an implant to improve neural survival are of interest. Recent studies reporting that brain-derived neurotrophic factor (BDNF) promotes SG survival after deafness have been conducted in rodents and limited to relatively short durations. Our study examined longer duration BDNF treatment in deafened cats that may better model the slow progression of SG degeneration in human cochleae, and this is the first study of BDNF in the developing auditory system. Kittens were deafened neonatally, implanted at 4-5 weeks with intracochlear electrodes containing a drug-delivery cannula, and BDNF or artificial perilymph was infused for 10 weeks from a miniosmotic pump. In BDNF-treated cochleae, SG cells grew to normal size and were significantly larger than cells on the contralateral side. However, their morphology was not completely normal, and many neurons lacked or had thinned perikaryl myelin. Unbiased stereology was employed to estimate SG cell density, independent of cell size. BDNF was effective in promoting significantly improved survival of SG neurons in these developing animals. BDNF treatment also resulted in higher density and larger size of myelinated radial nerve fibers, sprouting of fibers into the scala tympani, and improvement of electrically evoked auditory brainstem response thresholds. BDNF may have potential therapeutic value in the developing auditory system, but many serious obstacles currently preclude clinical application.

Cytoprotective effects of growth factors: BDNF more potent than GDNF in an organotypic culture model of Parkinson's disease

Parkinson's disease (PD) is a neurodegenerative disorder characterized by a preferential loss of dopaminergic (DAergic) neurons of the substantia nigra pars compacta (SNpc). Both glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF) play key roles in maintaining the DAergic phenotype and exert a cytoprotective effect on these neurons in vivo and in vitro. However, controversy still exists regarding the relative potency of the two factors and the extent to which they act synergistically. In this study, we used a refined version of organotypic cultures as a model for PD. The neurotoxin 6-hydroxydopamine (6-OHDA) was applied unilaterally in slices of rat mesencephalon, allowing for internal controls and enabling a precise comparison between the two sides of the midbrain. We evaluated the cytoprotective and regenerative effects of BDNF, GDNF and the combination of these in terms of surviving tyrosine hydroxylase positive (TH+) cells and TH mRNA expression. Pre-, co-, or post-treatment with neurotrophic factors clearly protects DAergic neurons from cell death. Cell survival is particularly pronounced in cultures pre-treated with BDNF and is not further increased when BDNF is applied in combination with GDNF in equimolar dose. On the lesion side, surviving TH+ cells exposed to neurotrophic factors showed extensive sprouting, and BDNF treatment resulted in a two-fold increase in TH mRNA. Such effects were not seen in the absence of toxin exposure. Thus, we observed that BDNF induced an upregulation of the DAergic phenotype, which suggest a cytoprotective and regenerative effect.

Visualization of intracellular trafficking of Math1 protein in different cell types with a newly‐constructed nonviral gene delivery plasmid

In recent years, Math1 gene therapy was indicated to be the future therapy for deafness in combination with other growth factors. However, Math1 delivery using adenovirus-mediated gene delivery or electroporation was impractical. The contribution of Math1 in the combined procedure was not clearly elucidated using the existing plasmids. Nonviral gene delivery vectors are expected to be extremely safe and convenient. The present study aimed to construct the pCDNA6.2/C-EmGFP-Math1 plasmid and evaluate its transfection efficiency and intracellular trafficking of Math1 protein corresponding to transcription regulation function. After constructing the pCDNA6.2/C-EmGFP-Math1 expression plasmid, the plasmid was transfected into different cell lines and primary cochlear cells using Lipofectamine 2000. Transfection efficiencies of the plasmid were evaluated. Transfection efficiencies using liposome nanoparticles containing Math1 plasmid were also assessed. Intracellular trafficking of Math1 was monitored using confocal microscopy. Different cell types can be transfected with high transfection efficiencies by the pcDNA6.2/C-EmGFP-Math1 plasmid using Lipofectamine 2000. Liposome nanoparticles containing the Math1 plasmid expressed the gene with variable efficiencies, depending on the particle size, surface charge and PEGylation status. Unique intracellular trafficking of Math1 was demonstrated in different cell types. The newly-constructed plasmid pcDNA6.2/C-EmGFP-Math1 was suitable for nonviral gene delivery of Math1. Unique intracellular trafficking of Math1 with dynamics from the cytoplasm to the nucleus was demonstrated. The modification of mesenchymal stem cells by Math1 gene delivery and by brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor treatments can potentially be applied to cell replacement for the treatment of cochlear spiral ganglion cell loss in deafness.

Phosphorylation of E3 Ligase Smurf1 Switches Its Substrate Preference in Support of Axon Development

Ubiquitin E3 ligases serve for ubiquitination of specific substrates, and its ligase efficacy is regulated by interacting proteins or substrate modifications. Whether and how the ligases themselves are modified by cellular signaling is unclear. Here we report that protein kinase A (PKA)-dependent phosphorylation of Smad Ubiquitin Regulatory Factor 1 (Smurf1) can switch its substrate preference between two proteins of opposing actions on axon development. Extracellular factors that promote axon formation elevated Smurf1 phosphorylation at a PKA site Thr³⁰⁶, and preventing this phosphorylation reduced axon formation in cultured hippocampal neurons and impaired polarization of cortical neurons in vivo. Thr³⁰⁶-phosphorylation changed the relative affinities of Smurf1 for its substrates, leading to reduced degradation of polarity protein Par6 and increased degradation of growth-inhibiting RhoA. Thus, PKA-dependent phosphorylation of the E3 ligase could switch its substrate preference, contributing to selective protein degradation required for localized cellular function.

Rapid brain-derived neurotrophic factor-dependent sequestration of amygdala and hippocampal GABAA receptors via different tyrosine receptor kinase B-mediated phosphorylation pathways

During the consolidation of fear memory, it has been shown that GABA(A) receptors (GABA(A)R) are rapidly downregulated in amygdala. This rapid decrease in GABA(A)R functioning may permit transient hyperexcitablity, contributing to cellular mechanisms of memory consolidation. Memory consolidation also requires brain-derived neurotrophic factor (BDNF) activation of tyrosine receptor kinase B (TrkB) receptors in the amygdala and hippocampus. We hypothesized that rapid internalization of GABA(A)Rα1 is mediated via TrkB activation of PKA and PKC-dependent processes. Primary neuronal cell cultures, from postnatal day 14-21 mouse amygdala and hippocampus, were analyzed with immunofluorescence using cell-surface, whole-cell permeabilization, and antibody internalization techniques, as well as with (3)H-muscimol binding assays. In both hippocampal and amygdala cultures, we found a >60% reduction in surface GABA(A)Rα1 within 5 min of BDNF treatment. Notably, the rapid decrease in surface GABA(A)Rα1 was confirmed biochemically using surface biotinylation assays followed by western blotting. This rapid effect was accompanied by TrkB phosphorylation and increased internal GABA(A)Rα1 immunofluorescence, and was blocked by k252a, a broad-spectrum tyrosine kinase antagonist. To further demonstrate TrkB specificity, we used previously characterized TrkB(F616A) mice, in which the highly selective TrkB-mutant specific antagonist, 1NMPP1, prevented the BDNF-dependent GABA(A)Rα1 internalization. In hippocampus, we found both PKA and PKC inhibition, using Rp-8-Br-cAMP and Calphostin C, respectively, blocked GABA(A)Rα1 internalization, whereas inhibition of MAPK (U0126) and PI3K (LY294002) did not prevent rapid internalization. By contrast in amygdala cultures, Rp-8-Br-cAMP had no effect. Together, these data suggest that rapid GABA(A)R internalization during memory consolidation is BDNF-TrkB dependent. Further, it appears that hippocampal GABA(A)R internalization is PKA and PKC dependent, while it may be primarily PKC dependent in amygdala, implying differential roles for TrkB-dependent kinase activation in BDNF-dependent memory formation.

Automated Sholl analysis of digitized neuronal morphology at multiple scales: Whole cell Sholl analysis versus Sholl analysis of arbor subregions

The morphology of dendrites and the axon determines how a neuron processes and transmits information. Neurite morphology is frequently analyzed by Sholl analysis or by counting the total number of neurites and branch tips. However, the time and resources required to perform such analysis by hand is prohibitive for the processing of large data sets and introduces problems with data auditing and reproducibility. Furthermore, analyses performed by hand or using course-grained morphometric data extraction tools can obscure subtle differences in data sets because they do not store the data in a form that facilitates the application of multiple analytical tools. To address these shortcomings, we have developed a program (titled "Bonfire") to facilitate digitization of neurite morphology and subsequent Sholl analysis. Our program builds upon other available open-source morphological analysis tools by performing Sholl analysis on subregions of the neuritic arbor, enabling the detection of local level changes in dendrite and axon branching behavior. To validate this new tool, we applied Bonfire analysis to images of hippocampal neurons treated with 25 ng/ml brain-derived neurotrophic factor (BDNF) and untreated control neurons. Consistent with prior findings, conventional Sholl analysis revealed that global exposure to BDNF increases the number of neuritic intersections proximal to the soma. Bonfire analysis additionally uncovers that BDNF treatment affects both root processes and terminal processes with no effect on intermediate neurites. Taken together, our data suggest that global exposure of hippocampal neurons to BDNF results in a reorganization of neuritic segments within their arbors, but not necessarily a change in their number or length. These findings were only made possible by the neurite-specific Sholl data returned by Bonfire analysis.

Spiral ganglion cell survival after round window membrane application of brain-derived neurotrophic factor using gelfoam as carrier

Several studies have shown that treatment with various neurotrophins protects spiral ganglion cells (SGCs) from degeneration in hair-cell deprived cochleas. In most of these studies the neurotrophins are delivered by means of intracochlear delivery methods. Recently, other application methods that might be more suited in cochlear implant patients have been developed. We have examined if round window membrane application of gelfoam infiltrated with a neurotrophin resulted in SGC survival in deafened guinea pigs. Two weeks after deafening, gelfoam cubes infiltrated with 6μg of brain-derived neurotrophic factor (BDNF) were deposited onto the round window membrane of the right cochleas. Electric pulses were delivered through an electrode positioned within the round window niche to electrically evoke auditory brainstem responses (eABRs). Two or four weeks after deposition of the gelfoam all cochleas were histologically examined. We found that local BDNF treatment enhances the survival of SGCs in the basal cochlear turn after two and four weeks. The treatment had no effect on SGC size or shape. In animals treated with BDNF, eABR amplitudes were smaller than in normal-hearing control animals and similar to those in deafened controls. We conclude that BDNF delivered by means of local gelfoam application provides a protective effect, which is limited compared to intracochlear delivery methods.

The selective Trk inhibitor AZ623 inhibits brain‐derived neurotrophic factor–mediated neuroblastoma cell proliferation and signaling and is synergistic with topotecan

TrkB expression is associated with poor prognosis for patients with neuroblastoma. AZ623 is a novel potent and selective inhibitor of the Trk family of tyrosine kinases. The authors hypothesized that AZ623 would inhibit TrkB-mediated signaling in neuroblastoma tumor cells and would be synergistic when combined with chemotherapy. Neuroblastoma cell lines were screened for TrkB receptor mRNA expression and for their proliferation rates in response to brain-derived neurotrophic factor (BDNF). The effects of AZ623 on Trk receptor phosphorylation, signaling, and cell growth were evaluated in BDNF-treated neuroblastoma cells. Mice with human neuroblastoma xenograft tumors were treated with AZ623 alone and in combination with topotecan, and tumor growth rates were determined during and after treatment. Neuroblastoma cell lines expressed various levels of the TrkB receptor and demonstrated increased proliferation in response to BDNF. BDNF treatment stimulated TrkB phosphorylation and downstream signaling that could be inhibited by AZ623. Neuroblastoma cells demonstrated in vitro sensitivity to AZ623, with concentration that inhibits 50% (IC50) values between 0.8 to 7 μM. AZ623 treatment was found to inhibit BDNF-mediated neuroblastoma cell proliferation. Mice with human neuroblastoma xenograft tumors demonstrated tumor growth inhibition when treated with AZ623 and with AZ623 combined with topotecan. Limited tumor regrowth was noted in mice with tumors treated with AZ623 combined with topotecan after treatment discontinuation. AZ623 is a novel selective Trk inhibitor that inhibits BDNF-mediated signaling and neuroblastoma cell proliferation. AZ623 treatment inhibits the growth of human neuroblastoma xenograft tumors, and treatment with AZ623 combined with topotecan results in the prolonged inhibition of tumor regrowth. On the basis of these results, further preclinical development is warranted.

MiR-375 Inhibits Differentiation of Neurites by Lowering HuD Levels

Neuronal development and plasticity are maintained by tightly regulated gene expression programs. Here, we report that the developmentally regulated microRNA miR-375 affects dendrite formation and maintenance. miR-375 overexpression in mouse hippocampus potently reduced dendrite density. We identified the predominantly neuronal RNA-binding protein HuD as a key effector of miR-375 influence on dendrite maintenance. Heterologous reporter analysis verified that miR-375 repressed HuD expression through a specific, evolutionarily conserved site on the HuD 3' untranslated region. miR-375 overexpression lowered both HuD mRNA stability and translation and recapitulated the effects of HuD silencing, which reduced the levels of target proteins with key functions in neuronal signaling and cytoskeleton organization (N-cadherin, PSD-95, RhoA, NCAM1, and integrin alpha1). Moreover, the increase in neurite outgrowth after brain-derived neurotrophic factor (BDNF) treatment was diminished by miR-375 overexpression; this effect was rescued by reexpression of miR-375-refractory HuD. Our findings indicate that miR-375 modulates neuronal HuD expression and function, in turn affecting dendrite abundance.

Trophic factors GDNF and BDNF improve function of retinal sheet transplants

The aim of this study was to compare glial-derived neurotrophic factor (GDNF) treatment with brain-derived neurotrophic factor (BDNF) treatment of retinal transplants on restoration of visual responses in the superior colliculus (SC) of the S334ter line 3 rat model of rapid retinal degeneration (RD). RD rats (age 4-6 weeks) received subretinal transplants of intact sheets of fetal retina expressing the marker human placental alkaline phosphatase (hPAP). Experimental groups included: (1) untreated retinal sheet transplants, (2) GDNF-treated transplants, (3) BDNF-treated transplants, (4) none surgical, age-matched RD rats, (5) sham surgery RD controls, (6) progenitor cortex transplant RD controls, and (7) normal pigmented rat controls. At 2-8 months after transplantation, multi-unit visual responses were recorded from the SC using a 40 ms full-field stimulus (-5.9 to +1 log cd/m(2)) after overnight dark-adaptation. Responses were analyzed for light thresholds, spike counts, response latencies, and location within the SC. Transplants were grouped into laminated or rosetted (more disorganized) transplants based on histological analysis. Visual stimulation of control RD rats evoked no responses. In RD rats with retinal transplants, a small area of the SC corresponding to the position of the transplant in the host retina, responded to light stimulation between -4.5 and -0.08 log cd/m(2), whereas the light threshold of normal rats was at or below -5 log cd/m(2) all over the SC. Overall, responses in the SC in rats with laminated transplants had lower response thresholds and were distributed over a wider area than rats with rosetted transplants. BDNF treatment improved responses (spike counts, light thresholds and responsive areas) of rats with laminated transplants whereas GDNF treatment improved responses from rats with both laminated and rosetted (more disorganized) transplants. In conclusion, treatment of retinal transplants with GDNF and BDNF improved the restoration of visual responses in RD rats; and GDNF appears to exert greater overall restoration than BDNF.

A high-fat diet impairs neurogenesis: Involvement of lipid peroxidation and brain-derived neurotrophic factor

Obesity is a growing global health problem that contributes to diabetes, hypertension, cardiovascular diseases, dementia, and cancer. The increased consumption of saturated fats in a high-fat diet (HFD) contributes to obesity, neurodegenerative diseases, long-term memory loss, and cognitive impairment. We tested whether HFD influences adult hippocampal neurogenesis. Male C57BL/6 mice were divided into two groups and maintained on either a normal diet (ND) or HFD. Seven weeks of HFD significantly decreased the numbers of newly generated cells in the dentate gyrus of the hippocampus without neuronal loss. HFD also increased the level of malondialdehyde (MDA) and decreased the level of brain-derived neurotrophic factor (BDNF) in the hippocampus. The toxic effects of MDA were evaluated on neural progenitor cells (NPCs). MDA reduced the growth of NPCs, but BDNF treatment restored NPCs proliferation. The present data indicate that a HFD impairs hippocampal neurogenesis and NPCs proliferation through increased lipid peroxidation and decreased BDNF.