Glial cell line‐derived neurotrophic factor in genetically defined fear‐induced aggression

Glial cell line‐derived neurotrophic factor (GDNF) plays an important role in maintenance of neuronal system throughout life. However, there is a lack of data on the involvement of GDNF in the regulation of different kinds of behavior. In this study, GDNF, its precursor (proGDNF) and GDNF mRNA levels were investigated in the brain of rats selectively bred for 85 generations for either high level or for the lack of affective aggressiveness toward human. It was found that GDNF mRNA level was decreased in the frontal cortex, increased in the raphe nuclei area of the midbrain of aggressive rats compared to tame animals and was not detected in the amygdala and hypothalamus. The level of proGDNF was reduced in the raphe nuclei area of the midbrain of highly aggressive rats and was not detected in the striatum, nucleus accumbens of investigated animals. Two forms of mature GDNF – monomer and dimer – were revealed. GDNF monomer level was increased in the raphe nuclei area, substantia nigra and amygdala of aggressive rats and it was not found in the frontal cortex and nucleus accumbens of investigated rats. Dimer GDNF level was found in all investigated brain structures. It was reduced in the hippocampus and increased in amygdala of highly aggressive rats. Thus, considerable structure‐specific differences in GDNF expression between highly aggressive and nonaggressive rats were shown. The data suggested the implication of both mature GDNF monomer and dimer as well as proGDNF in the mechanism underlying genetically defined aggressiveness.     

Effect of microgravity on glial cell line‐derived neurotrophic factor and cerebral dopamine neurotrophic factor gene expression in the mouse brain

Mice were exposed to 1 month of space flight on the Russian biosatellite BION‐M1 to determine its effect on the expression of genes involved in the maintenance of the mouse brain dopamine system. The current article focuses on the genes encoding glial cell line‐derived neurotrophic factor (GDNF) and cerebral dopamine neurotrophic factor (CDNF). Space flight reduced expression of the GDNF gene in the striatum and hypothalamus but increased it in the frontal cortex and raphe nuclei area. At the same time, actual space flight reduced expression of the gene encoding CDNF in the substantia nigra but increased it in the raphe nuclei area. To separate the effects of space flight from environmental stress contribution, we analyzed expression of the investigated genes in mice housed for 1 month on Earth in the same shuttle cabins that were used for space flight and in mice of the vivarium control group. Shuttle cabin housing failed to alter the expression of the GDNF and CDNF genes in the brain structures investigated. Thus, actual long‐term space flight produced dysregulation in genetic control of GDNF and CDNF genes. These changes may be related to downregulation of the dopamine system after space flight, which we have shown earlier. © 2015 Wiley Periodicals, Inc. Significance: Our results provide the first evidence of microgravity effects on expression of the GDNF and CDNF neurotrophic factor genes. A considerable decrease in mRNA level of GDNF and CDNF in the nigrostriatal dopamine system was found. Because both GDNF and CDNF play a significant role in maintenance and survival of brain dopaminergic neurons, we can assume that this dysregulation in genetic control of GDNF and CDNF genes in substantia nigra could be among the reasons for the deleterious effects of space flight on the dopamine system.     

Antiapoptotic and antiautophagic effects of glial cell line‐derived neurotrophic factor and hepatocyte growth factor after transient middle cerebral artery occlusion in rats

Glial cell line‐derived neurotrophic factor (GDNF) and hepatocyte growth factor (HGF) are strong neurotrophic factors, which function as antiapoptotic factors. However, the neuroprotective effect of GDNF and HGF in ameliorating ischemic brain injury via an antiautophagic effect has not been examined. Therefore, we investigated GDNF and HGF for changes of infarct size and antiapoptotic and antiautophagic effects after transient middle cerebral artery occlusion (tMCAO) in rats. For the estimation of ischemic brain injury, the infarct size was calculated at 24 hr after tMCAO by HE staining. Terminal deoxynucleotidyl transferase‐mediated dUTP‐biotin in situ nick end labeling (TUNEL) was performed for evaluating the antiapoptotic effect. Western blot analysis of microtubule‐associated protein 1 light chain 3 (LC3) and immunofluorescence analysis of LC3 and phosphorylated mTOR/Ser²⁴⁴⁸ (p‐mTOR) were performed for evaluating the antiautophagic effect. GDNF and HGF significantly reduced infarct size after cerebral ischemia. The amounts of LC3‐I plus LC3‐II (relative to β‐tubulin) were significantly increased after tMCAO, and GDNF and HGF significantly decreased them. GDNF and HGF significantly increased p‐mTOR‐positive cells. GDNF and HGF significantly decreased the numbers of TUNEL‐, LC3‐, and LC3/TUNEL double‐positive cells. LC3/TUNEL double‐positive cells accounted for about 34.3% of LC3 plus TUNEL‐positive cells. This study suggests that the protective effects of GDNF and HGF were greatly associated with not only the antiapoptotic but also the antiautophagic effects; maybe two types of cell death can occur in the same cell at the same time, and GDNF and HGF are capable of ameliorating these two pathways. © 2010 Wiley‐Liss, Inc.     

Differential regulation of glial cell line–derived neurotrophic factor (GDNF) expression in human neuroblastoma and glioblastoma cell lines

Human SK‐N‐AS neuroblastoma and U‐87MG glioblastoma cell lines were found to secrete relatively high levels of glial cell line–derived neurotrophic factor (GDNF). In response to growth factors, cytokines, and pharmacophores, the two cell lines differentially regulated GDNF release. A 24‐hr exposure to tumor necrosis factor‐α (TNFα; 10 ng/ml) or interleukin‐1β (IL‐1β; 10 ng/ml) induced GDNF release in U‐87MG cells, but repressed GDNF release from SK‐N‐AS cells. Fibroblast growth factors (FGF)‐1, ‐2, and ‐9 (50 ng/ml), the prostaglandins PGA₂, PGE₂, and PGI₂ (10 μM), phorbol 12,13‐didecanoate (PDD; 10 nM), okadaic acid (10 nM), dexamethasone (1 μM), and vitamin D₃ (1 μm) also differentially effected GDNF release from U‐87MG and SK‐N‐AS cells. A result shared by both cell lines, was a two‐ to threefold increase in GDNF release by db‐cAMP (1 mM), or forskolin (10 μM). In general, analysis of steady‐state GDNF mRNA levels correlated with changes in extracellular GDNF levels in U‐87MG cells but remained static in SK‐N‐AS cells. The data suggest that human GDNF synthesis/release can be regulated by numerous facto, signaling through multiple and diverse secondary messenger systems. Furthermore, we provide evidence of differential regulation of human GDNF synthesis/release in cells of glial (U‐87MG) and neuronal (SK‐N‐AS) origin. J. Neurosci. Res. 55:187–197, 1999. © 1999 Wiley‐Liss, Inc.     

Glial cell line‐derived neurotrophic factor is expressed by inflammatory cells in the sciatic nerves of Lewis rats with experimental autoimmune neuritis

Neurotrophic factors, including glial cell line‐derived neurotrophic factor (GDNF), have been known to play a role in neuroprotection in the injured peripheral nervous system (PNS). To evaluate the involvement of GDNF in experimental autoimmune neuritis (EAN) pathogenesis, the expression of GDNF in rat sciatic nerves with EAN was studied. Western blot analysis showed that the level of GDNF protein significantly increased 1.8‐fold at the paralytic stage of EAN at day 12 post‐immunization (PI) (p < 0.01), and its level further increased approximately 2.5‐fold at day 21 PI (p < 0.001) in the sciatic nerves of EAN‐affected rats compared with those of control rats, and then declined thereafter at day 28 PI. Immunohistochemical analysis showed that axons and Schwann cells constitutively contained GDNF in normal controls. In sciatic nerves with EAN at day 12 PI, GDNF was immunostained in infiltrating inflammatory cells including macrophages and T cells. Collectively, we postulate that GDNF plays a regulatory role in EAN paralysis. A paradoxical role of inflammatory cells to ameliorate PNS inflammation remains to be further studied in EAN, an animal model of human demyelinating disease.     

Glial cell line‐derived neurotrophic factor–secreting genetically modified human bone marrow‐derived mesenchymal stem cells promote recovery in a rat model of Parkinson's disease

Parkinson's disease (PD) is a neurodegenerative disease characterized by progressive degeneration of nigrostriatal dopaminergic (DA) neurons. The therapeutic potential of glial cell line‐derived neurotrophic factor (GDNF), the most potent neurotrophic factor for DA neurons, has been demonstrated in many experimental models of PD. However, chronic delivery of GDNF to DA neurons in the brain remains an unmet challenge. Here, we report the effects of GDNF‐releasing Notch‐induced human bone marrow‐derived mesenchymal stem cells (MSC) grafted into striatum of the 6‐hydroxydopamine (6‐OHDA) progressively lesioned rat model of PD. Human MSC, obtained from bone marrow aspirates of young, healthy adult volunteers, were transiently transfected with the intracellular domain of the Notch1 gene (NICD) to generate SB623 cells. SB623 cells expressing GDNF and/or humanized Renilla green fluorescent protein (hrGFP) following lentiviral transduction or nontransduced cells were stereotaxically placed into rat striatum 1 week after a unilateral partial 6‐OHDA striatal lesion. At 4 weeks, rats that had received GDNF‐transduced SB623 cells had significantly decreased amphetamine‐induced rotation compared with control rats, although this effect was not observed in rats that received GFP‐transduced or nontransduced SB623 cells. At 5 weeks, rejuvenated tyrosine hydroxylase‐immunoreactive (TH‐IR) fibers that appeared to be host DA axons were observed in and around grafts. This effect was more prominent in rats that received GDNF‐secreting cells and was not observed in controls. These observations suggest that human bone‐marrow derived MSC, genetically modified to secrete GDNF, hold potential as an allogeneic or autologous stem cell therapy for PD. © 2010 Wiley‐Liss, Inc.     

Furin mediates brain‐derived neurotrophic factor upregulation in cultured rat astrocytes exposed to oxygen–glucose deprivation

This study investigated the changes in brain‐derived neurotrophic factor (BDNF) expression and the role of furin in BDNF maturation in reactive astrocytes from rats exposed to oxygen–glucose deprivation (OGD). Furin, a proprotein convertase, is upregulated and cleaves certain substrates during hypoxia in cancer cells. In addition, during hypoxia in the central nervous system, astrocytes become reactive and release BDNF to protect neurons. Maturation of BDNF in astrocytes requires furin‐mediated endoproteolytic processing of the precursor protein pro‐BDNF to BDNF. To expand our knowledge about the role of furin in BDNF maturation in astrocytes, these cells were exposed to OGD, and expression of furin and BDNF was detected by Western blot analysis. Changes in BDNF expression were observed when furin activity was inhibited by furin prosegment. We found that protein expression of BDNF and furin was upregulated, and this upregulation correlated with OGD stimulation. Furin inhibition reduced BDNF maturation and secretion. These results indicate that furin mediates the upregulation of BDNF in reactive astrocytes exposed to OGD and that furin may impact the biological effect of reactive astrocytes. © 2014 Wiley Periodicals, Inc.     

Higher levels of brain derived neurotrophic factor but similar nerve growth factor in human milk in women with preeclampsia

Children born to mothers with preeclampsia have consistently been suggested to be at risk for cognitive and behavioral disorders in later life. Breastfeeding is said to be associated with better neurodevelopment outcomes. Our earlier studies indicated higher levels of docosahexaenoic acid (DHA) in human milk in women with preeclampsia. DHA is known to regulate the expression of neurotrophins and together they play a vital role in neurodevelopment and cognitive performance. The present study examines the levels of maternal plasma and milk neurotrophins [(nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF)] in women with preeclampsia and compares them with normotensive women who served as controls. Singleton pregnant women diagnosed with preeclampsia (n = 72) and controls (n = 102) were recruited for this study from Bharati Hospital, Pune. Plasma and milk samples were analyzed for NGF and BDNF levels using the Emax Immuno Assay System using promega kits. Maternal plasma NGF and BDNF levels were lower (p < 0.01 for both) in women with preeclampsia as compared to the control women. Milk NGF levels were similar while milk BDNF levels were higher (p < 0.05) in the preeclampsia group as compared to controls. Plasma NGF levels were positively correlated with milk NGF levels in the control group. Our results indicate the differential regulation of milk NGF and BDNF levels in women with preeclampsia. The present study suggests a role for both NGF and BDNF in human milk for postnatal brain development. Further studies need to examine the associations of DHA and BDNF in human milk with cognition at later ages.     

Motoneurons crave glial cell line-derived neurotrophic factor

This is a commentary on the developmental and therapeutic relevance of recent studies in the glial fibrillary acid protein (GFAP)-glial cell line-derived neurotrophic factor (GDNF) transgenic mouse reported by Zhao et al. (2004). This interesting study demonstrated that increased expression of GDNF in astrocytes increases the number of neighboring motoneurons of certain motoneuron subpopulations by diminishing programmed cell death during development. In addition, astrocyte-derived GDNF was shown to protect facial motoneurons from injury-induced cell death. Since this is the first direct demonstration that secretion of GDNF from astrocytes in the CNS can affect motoneuron development in utero and motoneuron survival after axotomy, novel approaches for motor neuron disease are suggested. The known target neurons that respond to GDNF are reviewed, as are studies using GDNF gene delivery in animal models of amyotrophic lateral sclerosis (ALS). It is postulated that GDNF is a factor to which many motoneurons respond along their whole extent from soma to axon to terminal.     

Brain‐derived neurotrophic factor interacts with astrocytes and neurons to control respiration

Respiratory rhythm is generated and modulated in the brainstem. Neuronal involvement in respiratory control and rhythmogenesis is now clearly established. However, glial cells have also been shown to modulate the activity of brainstem respiratory groups. Although the potential involvement of other glial cell type(s) cannot be excluded, astrocytes are clearly involved in this modulation. In parallel, brain‐derived neurotrophic factor (BDNF) also modulates respiratory rhythm. The currently available data on the respective roles of astrocytes and BDNF in respiratory control and rhythmogenesis lead us to hypothesize that there is BDNF‐mediated control of the communication between neurons and astrocytes in the maintenance of a proper neuronal network capable of generating a stable respiratory rhythm. According to this hypothesis, progression of Rett syndrome, an autism spectrum disease with disordered breathing, can be stabilized in mouse models by re‐expressing the normal gene pattern in astrocytes or microglia, as well as by stimulating the BDNF signaling pathway. These results illustrate how the signaling mechanisms by which glia exerts its effects in brainstem respiratory groups is of great interest for pathologies associated with neurological respiratory disorders.     

Secretion of nerve growth factor,brain-derived neurotrophic factor,and glial cell-line derived neurotrophic factor in co-culture of four cell types in cerebrospinal fluid-containing medium

The present study co-cultured human embryonic olfactory ensheathing cells, human Schwann cells, human amniotic epithelial cells and human vascular endothelial cells in complete culture medium- containing cerebrospinal fluid. Enzyme linked immunosorbent assay was used to detect nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor secretion in the supernatant of co-cultured cells. Results showed that the number of all cell types reached a peak at 7-10 days, and the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor peaked at 9 days. Levels of secreted nerve growth factor were four-fold higher than brain-derived neurotrophic factor, which was three-fold higher than glial cell line-derived neurotrophic factor. Increasing concentrations of cerebrospinal fluid (10%, 20% and 30%) in the growth medium caused a decrease of neurotrophic factor secretion Results indicated co-culture of human embryonic olfactory ensheathing cells, human Schwann cells human amniotic epithelial cells and human vascular endothelial cells improved the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor. The reduction of cerebrospinal fluid extravasation at the transplant site after spinal cord injury is beneficial for the survival and secretion of neurotrophic factors from transplanted cells.     

Electrically induced brain‐derived neurotrophic factor release from schwann cells

Regulating the production of brain‐derived neurotrophic factor (BDNF) in Schwann cells (SCs) is critical for their application in traumatic nerve injury, neurodegenerative disorders, and demyelination disease in both central and peripheral nervous systems. The present study investigated the possibility of using electrical stimulation (ES) to activate SCs to release BDNF. We found that short‐term ES was capable of promoting BDNF production from SCs, and the maximal BDNF release was achieved by ES at 6 V (3 Hz, 30 min). We further examined the involvement of intracellular calcium ions ([Ca₂₊]_i) in the ES‐induced BDNF production in SCs by pharmacological studies. We found that the ES‐induced BDNF release required calcium influx through T‐type voltage‐gated calcium channel (VGCC) and calcium mobilization from internal calcium stores, including inositol triphosphate‐sensitive stores and caffeine/ryanodine‐sensitive stores. In addition, calcium‐calmodulin dependent protein kinase IV (CaMK IV), mitogen‐activated protein kinase (MAPK), and cAMP response element‐binding protein (CREB) were found to play important roles in the ES‐induced BDNF release from SCs. In conclusion, ES is capable of activating SCs to secrete BDNF, which requires the involvement of calcium influx through T‐type VGCC and calcium mobilization from internal calcium stores. In addition, activation of CaMK IV, MAPK, and CREB were also involved in the ES‐induced BDNF release. The findings indicate that ES can improve the neurotrophic ability in SCs and raise the possibility of developing electrically stimulated SCs as a source of cell therapy for nerve injury in both peripheral and central nervous systems. © 2014 Wiley Periodicals, Inc.     

Glial cell line‐derived neurotrophic factor is a key neurotrophin in the postnatal enteric nervous system

Background The enteric nervous system (ENS) continues its structural and functional growth after birth, with formation of ganglia and the innervation of growing smooth muscle. However, little is known about factors in the postnatal intestine that influence these processes. Methods We examined the presence and potential role of glial cell line‐derived nerve growth factor (GDNF) in the rat postnatal ENS using neonatal tissue, primary co‐cultures of the myenteric plexus, smooth muscle, and glial cells as well as cell lines of smooth muscle or glial cells. Key Results Western blot analysis showed that GDNF and its co‐receptors rearranged during transfection (RET) and GDNF family receptor alpha‐1 were expressed in the muscle layer of the neonatal and adult rat intestine. Immunohistochemistry localized the receptors for GDNF to myenteric neurons, while GDNF was localized to smooth muscle cells. In a co‐culture model, GDNF but not nerve growth factor, brain derived neurotrophic factor or neurotrophin‐3 significantly increased neuronal survival and more than doubled the numbers of neurites in vitro. RT‐PCR, qPCR, Western blotting, ELISA, and immunocytochemistry as well as bioassays of neuronal survival and of RET phosphorylation all identified intestinal smooth muscle as the source of GDNF in vitro. GDNF also induced morphological changes in the structure and organization of neurons and axons, causing marked aggregation of neuronal cell bodies and collinear development of axons. As well, GDNF (50–150 ng mL^?1) significantly increased [³H]‐choline uptake and stimulated [³H]‐acetylcholine release. Conclusions & Inferences We conclude that GDNF derived from intestinal smooth muscle cells is a key factor influencing the structural and functional development of postnatal myenteric neurons.