Both 5' and 3' flanks regulate Zebrafish brain-derived neurotrophic factor gene expression

Background  

Precise control of developmental and cell-specific expression of the brain-derived neurotrophic factor (BDNF) gene is essential for normal neuronal development and the diverse functions of BDNF in the adult organism. We previously showed that the zebrafish BDNF gene has multiple promoters. The complexity of the promoter structure and the mechanisms that mediate developmental and cell-specific expression are still incompletely understood.     

Treatment of trigeminal ganglion neurons in vitro with NGF,GDNF or BDNF: effects on neuronal survival,neurochemical properties and TRPV1-mediated neuropeptide secretion

Background  

Nerve growth factor (NGF), glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF) all play important roles in the development of the peripheral sensory nervous system. Additionally, these growth factors are proposed to modulate the properties of the sensory system in the adult under pathological conditions brought about by nerve injury or inflammation. We have examined the effects of NGF, GDNF and BDNF on adult rat trigeminal ganglion (TG) neurons in culture to gain a better understanding of how these growth factors alter the cytochemical and functional phenotype of these neurons, with special attention to properties associated with nociception.     

Difference in trafficking of brain-derived neurotrophic factor between axons and dendrites of cortical neurons,revealed by live-cell imaging

Background  

Brain-derived neurotrophic factor (BDNF), which is sorted into a regulated secretory pathway of neurons, is supposed to act retrogradely through dendrites on presynaptic neurons or anterogradely through axons on postsynaptic neurons. Depending on which is the case, the pattern and direction of trafficking of BDNF in dendrites and axons are expected to be different. To address this issue, we analyzed movements of green fluorescent protein (GFP)-tagged BDNF in axons and dendrites of living cortical neurons by time-lapse imaging. In part of the experiments, the expression of BDNF tagged with cyan fluorescent protein (CFP) was compared with that of nerve growth factor (NGF) tagged with yellow fluorescent protein (YFP), to see whether fluorescent protein-tagged BDNF is expressed in a manner specific to this neurotrophin.     

Structural effects and potential changes in growth factor signalling in penis-projecting autonomic neurons after axotomy

Background  

The responses of adult parasympathetic ganglion neurons to injury and the neurotrophic mechanisms underlying their axonal regeneration are poorly understood. This is especially relevant to penis-projecting parasympathetic neurons, which are vulnerable to injury during pelvic surgery such as prostatectomy. We investigated the changes in pelvic ganglia of adult male rats in the first week after unilateral cavernous (penile) nerve axotomy (cut or crush lesions). In some experiments FluoroGold was injected into the penis seven days prior to injury to allow later identification of penis-projecting neurons. Neurturin and glial cell line-derived neurotrophic factor (GDNF) are neurotrophic factors for penile parasympathetic neurons, so we also examined expression of relevant receptors, GFRα1 and GFRα2, in injured pelvic ganglion neurons.     

A novel BDNF gene promoter directs expression to skeletal muscle

Background  

Cell-specific expression of the gene that encodes brain-derived neurotrophic factor (BDNF) is required for the normal development of peripheral sensory neurons and efficient synaptic transmission in the mature central and peripheral nervous system. The control of BDNF gene expression involves multiple tissue and cell-specific promoters that are differentially regulated. The molecular mechanisms that are responsible for tissue and cell-specific expression of these promoters are still incompletely understood.     

Regionalization of Plasma Membrane-Bound Flavoproteins of Cerebellar Granule Neurons in Culture by Fluorescence Energy Transfer Imaging

Flavoproteins are components of plasma membrane redox chains, which have been suggested to play major roles in neuronal activity and survival. We found that the red/orange autofluorescence of mature primary cultures of cerebellar granule neurons (8–9 days in vitro) was largely quenched by millimolar concentrations of dithionite added to the extracellular medium, and pointed out that nearly 50% of this autofluorescence was due to plasma membrane-bound flavoproteins. We report in this work that the lipophilic neuronal plasma membrane markers N-(3-triethylammoniumpropyl)-4-(4-(4-(diethylamino)phenyl)butadienyl)-pyridinium dibromide (RH-414) and N-(3-triethylammoniumpropyl)-4-(6-(4-(diethylamino)phenyl)hexatrienyl)pyridinium dibromide (FM4-64) can form fluorescence energy transfer donor–acceptor pairs with flavoproteins with calculated R ₀ values between 3.7 and 4.2 nm. The quantification of the efficiency of fluorescence energy transfer with different concentrations of acceptor dyes has been worked out with re-suspended neurons. Using quantitative images of the neurons in culture, acquired with a CCD camera attached to an epifluorescence microscope, regionalization of the plasma membrane-bound flavoproteins of cerebellar granule neurons has been achieved from the quenching by dithionite of the fluorescence of the acceptor dye. The results unraveled that plasma membrane-bound flavoproteins are largely enriched in interneuronal contact sites forming clusters of 0.5–1 μm diameter size, which appears largely regionalized in the neuron's cell body.     

Tissue-specific and neural activity-regulated expression of human BDNF gene in BAC transgenic mice

Background  

Brain-derived neurotrophic factor (BDNF) is a small secreted protein that has important roles in the developing and adult nervous system. Altered expression or changes in the regulation of the BDNF gene have been implicated in a variety of human nervous system disorders. Although regulation of the rodent BDNF gene has been extensively investigated, in vivo studies regarding the human BDNF gene are largely limited to postmortem analysis. Bacterial artificial chromosome (BAC) transgenic mice harboring the human BDNF gene and its regulatory flanking sequences constitute a useful tool for studying human BDNF gene regulation and for identification of therapeutic compounds modulating BDNF expression.     

Exercise-induced motor improvement after complete spinal cord transection and its relation to expression of brain-derived neurotrophic factor and presynaptic markers

Background  

It has been postulated that exercise-induced activation of brain-derived neurotrophic factor (BDNF) may account for improvement of stepping ability in animals after complete spinal cord transection. As we have shown previously, treadmill locomotor exercise leads to up-regulation of BDNF protein and mRNA in the entire neuronal network of intact spinal cord. The questions arise: (i) how the treadmill locomotor training, supplemented with tail stimulation, affects the expression of molecular correlates of synaptic plasticity in spinal rats, and (ii) if a response is related to BDNF protein level and distribution.     

Transfection using hydroxyapatite nanoparticles in the inner ear via an intact round window membrane in chinchilla

Hydroxyapatite nanoparticles (nHAT) are known to have excellent biocompatibility, and have attracted increasing attention as new candidates of non-viral vectors for gene therapy. In our previous studies, nHAT carrying a therapeutic gene and a reporter gene were successfully transfected into the spiral ganglion neurons in the inner ear of guinea pigs in vivo as well as in the cultured cell lines, although the transfection efficiencies were never higher than 30%. In this study, the surface modification of nHAT with polyethylenimine (PEI) was made (PEI–nHAT, diameter = 73.09 ± 27.32 nm) and a recombinant plasmid carrying enhanced green fluorescent protein (EGFP) gene and neurotrophin-3 (NT-3) gene was constructed as pEGFPC2–NT3. The PEI modified nHAT and the recombinant plasmid was then connected to form the nHAT-based vector–gene complex (PEI–nHAT–pEGFPC2–NT3). This complex was then placed onto the intact round window membranes of the chinchillas for inner ear transfection. Auditory brainstem response (ABR) was tested to evaluate auditory function. Green fluorescence of EGFP was observed using confocal microscopy 48 h after administering vector–gene complexes. There was no significant threshold shift in tone burst-evoked ABR at any tested frequency. Abundant, condensed green fluorescence was found in dark cells on both sides of the crista and around the macula of the utricle. Scattered EGFP signals were also detected in vestibular hair cells, some Schwann cells in the cochlear spiral ganglion region, some outer pillar cells in the organ of Corti, and a few cells in the stria vascularis. The density of green fluorescence-marked cells was obviously higher in the vestibular dark cell area than in other areas of the inner ear, suggesting that vestibular dark cells may have the ability to actively engulf the nHAT-based vector–gene complexes. Considering the high transfection efficiency in the vestibular system, PEI–nHAT may be a potential vector for gene therapy of inner ear diseases, especially vestibular disorders, and deserves further study.     

Long-term neprilysin gene transfer is associated with reduced levels of intracellular Abeta and behavioral improvement in APP transgenic mice

Background  

Proteolytic degradation has emerged as a key pathway involved in controlling levels of the Alzheimer's disease (AD)-associated amyloid-β (Aβ) peptide in the brain. The endopeptidase, neprilysin, has been implicated as a major Aβ degrading enzyme in mice and humans. Previous short and intermediate term studies have shown the potential therapeutic application of neprilysin by delivering this enzyme into the brain of APP transgenic mice using gene transfer with viral vectors. However the effects of long-term neprilysin gene transfer on other aspects of Aβ associated pathology have not been explored yet in APP transgenic mice.     

The targeted gene (KDRP-CD/TK) therapy of breast cancer mediated by SonoVue and ultrasound irradiation in vitro

Suicide gene therapy has become an effective therapy for breast cancer, and ultrasound targeted microbubble destruction (UTMD) has become a popular topic in the gene therapy field. In this study, MCF-7 cells with the KDR promoter and LSl74T cells without the KDR promoter were transfected with the recombinant plasmid pEGFP-KDRP-CD/TK using UTMD. The recombinant plasmid pEGFP-KDRP-CD/TK was transfected into MCF-7 and LS174T cells successfully with no significant difference in transfection efficiency (p > 0.05). By RT-PCR, the CD/TK fusion gene was shown to be expressed in MCF-7 cells but not expressed in LS174T cells. In a cytotoxicity experiment, transgenic MCF-7 cells were sensitive to the prodrugs 5-FC and GCV. When both 5-FC and GCV were administered, the rate of cellular inhibition was significantly greater than that achieved when only one of the prodrugs was administered (p < 0.001). Moreover, the inhibition rates achieved administering 5-FC, GCV and both 5-FC and GCV were all significantly greater than the gene transfection rate of 21.92 ± 3.64% (p < 0.001). However, transgenic LS174T cells were not sensitive to any prodrug. These results demonstrated that UTMD is a safe, effective and targeted gene delivery system. Also, the KDR promoter can drive expression of the CD/TK double suicide gene target in MCF-7 cells, and the targeted killing effect of the KDRP-CD/TK gene on MCF-7 cells in vitro has good synergy with expression of the CD/TK fusion gene.     

Targeted adenoviral vectors

The practical implementation of gene therapy in the clinical setting mandates gene delivery vehicles, or vectors, capable of efficient gene delivery selectively to the target disease cells. The utility of adenoviral vectors for gene therapy is restricted by their dependence on the native adenoviral primary cellular receptor for cell entry. Therefore, a number of strategies have been developed to allow CAR-independent infection of specific cell types, including the use of bispecific conjugates and genetic modifications to the adenoviral capsid proteins, in particular the fibre protein. These targeted adenoviral vectors have demonstrated efficient gene transfer in vitro, correlating with a therapeutic benefit in preclinical animal models. Such vectors are predicted to possess enhanced efficacy in human clinical studies, although anatomical barriers to their use must be circumvented.     

Effects of dietary Na+ deprivation on epithelial Na+ channel (ENaC), BDNF, and TrkB mRNA expression in the rat tongue

Background  

In rodents, dietary Na⁺ deprivation reduces gustatory responses of primary taste fibers and central taste neurons to lingual Na⁺ stimulation. However, in the rat taste bud cells Na⁺ deprivation increases the number of amiloride sensitive epithelial Na⁺ channels (ENaC), which are considered as the "receptor" of the Na⁺ component of salt taste. To explore the mechanisms, the expression of the three ENaC subunits (α, β and γ) in taste buds were observed from rats fed with diets containing either 0.03% (Na⁺ deprivation) or 1% (control) NaCl for 15 days, by using in situ hybridization and real-time quantitative RT-PCR (qRT-PCR). Since BDNF/TrkB signaling is involved in the neural innervation of taste buds, the effects of Na⁺ deprivation on BDNF and its receptor TrkB expression in the rat taste buds were also examined.     

16-month periodicity of cosmic-ray intensity

Summary Data from four neutron monitors, Climax (1953–1976), Deep River (1964–1977), Mt. Norikura (1957–1983) and Tokyo (1970–1983) are analysed. Power spectra reveal peaks at frequency 1c/16 months for all the stations. Harmonic analysis for the 16-month period is performed after eliminating the long-term solar-cycle variation. There is general agreement in the phases of the mean vectors for various neutron monitors. The amplitude averaged over all the four stations is 0.41±0.01, while it is higher for the two stations having lower cut-off rigidity as compared to that of the other two. The vectors are seen to become more consistent in phase for periods of minimum solar activity than those for maximum-activity period. The summation dials reveal abrupt changes in the directions of the 16-month wave vector corresponding to the reversal of polarity of the solar magnetic field. Comparison of the 16-month wave of sunspot activity with that of cosmic-ray intensity, however, does not show any significant correlation.

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Riassunto Si analizzano dati da quattro monitor di neutroni, Climax (1953–1976), Deep River (1964–1977), Mt. Norikura (1957–1983) e Tokyo (1970–1983). Gli spettri di frequenza rivelano picchi alla frequenza di 1 ciclo ogni 16 mesi per tutte le stazioni. Si effettua un'analisi armonica per un periodo di 16 mesi dopo aver eliminato la variazione del ciclo solare a lungo termine. Esiste un accordo generale riguardo alle fasi dei vettori medi per vari monitor di neutroni. L'ampiezza media rispetto a tutte e quattro le stazioni è di 0.41±0.01, mentre risulta superiore per le due stazioni che presentano una rigidità di taglio inferiore alle altre due. Si osserva una crescente coerenza di fase dei vettori in periodi di attività solare minima, rispetto a quella in periodi di attività massima. I grafici della somma rivelano cambiamenti repentini nelle direzioni del vettore d'onda di 16 mesi, corrispondenti all'inversione di polarità del campo magnetico del Sole. Il confronto di un'onda di 16 mesi dell'attività delle macchie solari con quella dell'intensità dei raggi cosmici, tuttavia, non mostra nessuna correlazione significativa.

     

Improved laser-induced forward transfer of organic semiconductor thin films by reducing the environmental pressure and controlling the substrate–substrate gap width

Laser-induced forward transfer (LIFT) has been investigated for bilayer transfer material systems: silver/organic film (Alq₃ or PFO). The LIFT process uses an intermediate dynamic release layer of a triazene polymer. This study focuses on the effect of introducing a controlled donor–receiver substrate gap distance and the effect of doing the transfer at reduced air pressures, whilst varying the fluence up to ∼200 mJ/cm². The gap between ‘in-contact’ substrates has been measured to be a minimum of 2–3 μm. A linear variation in the gap width from ‘in contact’ to 40 μm has been achieved by adding a spacer at one side of the substrate–substrate sandwich. At atmospheric pressure, very little transfer is achieved for Alq₃, although PFO shows some signs of successful doughnut transfer (with a large hole in the middle) in a narrow fluence range, at gaps greater than 20 μm. For the transfer of Ag/PFO bilayers at atmospheric pressure, the addition of a PFO layer onto the receiver substrate improved the transfer enormously at smaller gaps and higher fluences. However, the best transfer results were obtained at reduced pressures where a 100% transfer success rate is obtained within a certain fluence window. The quality of the pixel morphology at less than 100 mbar is much higher than at atmospheric pressure, particularly when the gap width is less than 20 μm. These results show the promise of LIFT for industrial deposition processes where a gap between the substrates will improve the throughput.     

General construction for topological singular vectors

A general construction is found for “topological” singular vectors (BRST-invariant singular vectors of the twisted N=2 superconformal algebra which are constructed over chiral primary states). The new construction, employed in parallel with the well-known MFF construction for the singular vectors of the sℓ(2) Kac-Moody algebra, implements an isomorphism of topological and sℓ(2)-singular vectors. Pis’ma Zh. éksp. Teor. Fiz. 63, No. 2, 129–134 (25 January 1996)     

Early raise of BDNF in hippocampus suggests induction of posttranscriptional mechanisms by antidepressants

Background  

The neurotrophin BDNF has been implicated in the regulation of neuroplasticity, gene expression, and synaptic function in the adult brain, as well as in the pathophysiology of neuropsychiatric disorders and the mechanism of action of antidepressants. Antidepressant treatments have been shown to increase the expression of BDNF mRNA, although the changes measured were found to be different depending on various factors. A few studies only have measured levels of BDNF protein after antidepressant treatments, and poor correlation was found between mRNA and protein changes. We studied the time course of expression of BDNF mRNA and protein during drug treatments, in order to elucidate the temporal profile of regulation of this effector and whether mRNA and protein levels correlate. Rat groups were treated for 1, 2 or 3 weeks with fluoxetine or reboxetine; in additional groups drug treatment was followed by a washout week (3+1). Total BDNF mRNA was measured by Real Time PCR, pro- and mature BDNF proteins were measured by Western blot.     

Absorption spectra and the nature of complexes in the polyvinyl Alcohol-Nickel (II) chloride system

We have used electronic spectroscopy in the 160–1100 nm range to study the polyvinyl alcohol-nickel(II) chloride system. Based on the results obtained, we hypothesize formation of mixed-ligand complexes of the type [Ni(H₂O)_6-nCl_n]^2-n (n = 0, 1,..., 5) in the polyvinyl alcohol matrix. Transformation of the coordination sphere as the NiCl₂ concentration changes is apparent both in the region of the d-d transition bands (350–1100 nm) and in the region of the charge transfer bands (160–250 nm). We propose assigning the absorption bands separated by mathematical treatment to complexes of specific compositions. __________ Translated from Zhurnal Prikladnoi Spektroskopii, Vol. 73, No. 1, pp. 136–138, January–February, 2006.