Glial Cell Line-derived Neurotrophic Factor Protein Levels Research Articles

Evaluation of the Continuous Positive Airway Pressure Effect on Neurotrophins’ Gene Expression and Protein Levels

Neurotrophins (NT) might be associated with the pathophysiology of obstructive sleep apnea (OSA) due to concurrent intermittent hypoxia and sleep fragmentation. Such a relationship could have implications for the health and overall well-being of patients; however, the literature on this subject is sparse. This study investigated the alterations in the serum protein concentration and the mRNA expression of the brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), neurotrophin-3 (NTF3), and neurotrophin-4 (NTF4) proteins following a single night of continuous positive airway pressure (CPAP) therapy. This study group consisted of 30 patients with OSA. Venous blood was collected twice after a diagnostic polysomnography (PSG) and PSG with CPAP treatment. Gene expression was assessed with a quantitative real-time polymerase chain reaction. An enzyme-linked immunosorbent assay was used to determine the protein concentrations. After CPAP treatment, BDNF, proBDNF, GDNF, and NTF4 protein levels decreased (p = 0.002, p = 0.003, p = 0.047, and p = 0.009, respectively), while NTF3 increased (p = 0.001). Sleep latency was correlated with ΔPSG + CPAP/PSG gene expression for BDNF (R = 0.387, p = 0.038), NTF3 (R = 0.440, p = 0.019), and NTF4 (R = 0.424, p = 0.025). OSA severity parameters were not associated with protein levels or gene expressions. CPAP therapy could have an impact on the posttranscriptional stages of NT synthesis. The expression of different NTs appears to be connected with sleep architecture but not with OSA severity.

Effects of gestational inflammation on age-related cognitive decline and hippocampal Gdnf-GFRα1 levels in F1 and F2 generations of CD-1 Mice

BackgroundIt has been reported that age-associated cognitive decline (AACD) accelerated by maternal lipopolysaccharide (LPS) insult during late pregnancy can be transmitted to the second generation in a sex-specificity manner. In turn, recent studies indicated that glial cell line‐derived neurotrophic factor (GDNF) and its cognate receptor (GFRα1) are critical for normal cognitive function. Based on this evidence, we aimed to explore whether Gdnf-GFRα1 expression contributes to cognitive decline in the F1 and F2 generations of mouse dams exposed to lipopolysaccharide (LPS) during late gestation, and to evaluate also the potential interference effect of pro-inflammatory cytokines.MethodsDuring gestational days 15–17, pregnant CD-1 mice (8–10 weeks old) received a daily intraperitoneal injection of LPS (50 μg/kg) or saline (control). In utero LPS-exposed F1 generation mice were selectively mated to produce F2 generation mice. In F1 and F2 mice aged 3 and 15 months, the Morris water maze (MWM) was used to evaluated the spatial learning and memory ability, the western blotting and RT-PCR were used for analyses of hippocampal Gdnf and GFRα1 expression, and ELISA was used to analyse IL-1β, IL-6 and TNF-α levels in serum.ResultsMiddle-aged F1 offspring from LPS-treated mothers exhibited longer swimming latency and distance during the learning phase, lower percentage swimming time and distance in targe quadrant during memory phase, and lower hippocampal levels of Gdnf and GFRα1 gene products compared to age-matched controls. Similarly, the middle-aged F2 offspring from the Parents-LPS group had longer swimming latency and distance in the learning phase, and lower percentage swimming time and distance in memory phase than the F2-CON group. Moreover, the 3-month-old Parents-LPS and 15-month-old Parents- and Father-LPS groups had lower GDNF and GFRα1 protein and mRNAs levels compared to the age-matched F2-CON group. Furthermore, hippocampal levels of Gdnf and GFRα1 were correlated with impaired cognitive performance in the Morris water maze after controlling for circulating pro-inflammatory cytokine levels.ConclusionsOur findings indicate that accelerated AACD by maternal LPS exposure can be transmitted across at least two generations through declined Gdnf and GFRα1 expression, mainly via paternal linage.

The Polyunsaturated Fatty Acid EPA, but Not DHA, Enhances Neurotrophic Factor Expression through Epigenetic Mechanisms and Protects against Parkinsonian Neuronal Cell Death.

ω-3 Polyunsaturated fatty acids (PUFAs) have been found to exert many actions, including neuroprotective effects. In this regard, the exact molecular mechanisms are not well understood. Parkinson's disease (PD) is the second most common age-related neurodegenerative disease. Emerging evidence supports the hypothesis that PD is the result of complex interactions between genetic abnormalities, environmental toxins, mitochondrial dysfunction, and other cellular processes, such as DNA methylation. In this context, BDNF (brain-derived neurotrophic factor) and GDNF (glial cell line-derived neurotrophic factor) have a pivotal role because they are both involved in neuron differentiation, survival, and synaptogenesis. In this study, we aimed to elucidate the potential role of two PUFAs, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), and their effects on BDNF and GDNF expression in the SH-SY5Y cell line. Cell viability was determined using the MTT assay, and flow cytometry analysis was used to verify the level of apoptosis. Transmission electron microscopy was performed to observe the cell ultrastructure and mitochondria morphology. BDNF and GDNF protein levels and mRNA were assayed by Western blotting and RT-PCR, respectively. Finally, methylated and hydroxymethylated DNA immunoprecipitation were performed in the BDNF and GDNF promoter regions. EPA, but not DHA, is able (i) to reduce the neurotoxic effect of neurotoxin 6-hydroxydopamine (6-OHDA) in vitro, (ii) to re-establish mitochondrial function, and (iii) to increase BNDF and GDNF expression via epigenetic mechanisms.

TNF-α inhibits GDNF levels in Sertoli cells, through a NF-κB-dependent, HES1-dependent mechanism.

Glial cell line-derived neurotrophic factor (GDNF) is a soluble molecule crucial for the regulation of the spermatogonial stem cells (SSC) of the testis. The effects of GDNF on target cells have been extensively described, but mechanisms underlying GDNF regulation are currently under investigation. In the nervous system, GDNF expression is regulated by pro-inflammatory cytokines including lipopolysaccharide (LPS), interleukin 1 beta (IL-1β), and tumor necrosis factor alpha (TNF-α) but the effect of these cytokines on GDNF expression in the testis is unclear. The aim of the present study was to investigate the impact of TNF-α on GDNF expression levels using primary murine Sertoli cells as experimental model. The expression of TNF-α-regulated genes including Gdnf in different culture conditions was determined by real-time PCR. GDNF protein levels were determined by ELISA. The activation of the NF-κb pathway and HES1 levels were assessed by Western Blot analysis and immunofluorescence. HES1 expression was downregulated by RNAi. In primary Sertoli cells, TNF-α downregulates GDNF levels through a nuclear factor-κB (NF-κB)-dependent mechanism. Mechanistically, TNF-α induces the transcriptional repressor HES1 by a NF-Κb-dependent mechanism, which in turn downregulates GDNF. Under physiological conditions, TNF-α is secreted by germ cells suggesting that this cytokine plays a role in the paracrine control of SSC niche by modulating GDNF levels. HES1, a well-known target of the Notch pathway, is implicated in the regulation of GDNF expression. In Sertoli cells, TNF-α and Notch signaling may converge at molecular level, to regulate the expression of HES1 and HES1- target genes, including GDNF. Because of the importance of GDNF for spermatogonial stem cell self-renewal and proliferation, this data may give important insights on how cytokine signals in the testis modulate the expression of niche-derived factors.

The Effects of Aging and Exercise on Glial Cell Line‐Derived Neurotrophic Factor Content in Rat Hearts

Heart function is regulated by the sensory nervous system and the two branches of the autonomic nervous system, the sympathetic and parasympathetic nervous systems. Neural development, plasticity, and survival of these branches of the nervous system are controlled by neurotrophic factors such as the glial cell line‐derived neurotrophic factor (GDNF). Studies in our laboratory have shown that aging can shift the innervation pattern in mesenteric vessels from a balanced sensory/sympathetic pattern to a predominately sympathetic pattern and this is associated with an increase in blood pressure. However, the same studies suggest that exercise is associated with an increase in GDNF protein content in mesenteric vessels, a restored balance of innervation, and a lowering of blood pressure. Little is known about how aging and exercise impacts neurotrophic factor expression in cardiac muscle. Therefore, the purpose of this study was to measure GDNF protein content in the heart of rats at different ages and to investigate the effects of exercise on GDNF protein levels. Control groups consisted of 4‐week‐old, 14‐week‐old, 6‐month‐old, 12‐month‐old, and 18‐month‐old sedentary rats (5 or 6 animals per group). In the exercise study, 4‐week‐old, 6‐month‐old, and 12‐month‐old rats were placed in cages with access to voluntary running wheels and were allowed to exercise for 10 weeks (4‐week‐old), or 6 months (6‐ and 12‐month‐old). Sedentary and exercised animals were euthanized and cardiac tissues were removed. Each heart was divided into three parts: right atria (RA), left atria (LA), and ventricles (VT). Tissues were processed and neurotrophic factor protein content was measured using enzyme‐linked immunosorbent assay. GDNF protein content in right atria (0.95 ± 0.14pg of GDNF/mg of tissue), left atria (0.91 ± 0.29pg of GDNF/mg of tissue), and ventricles (0.54 ± 0.35pg of GDNF/mg of tissue) of 4‐week‐old animals was significantly higher than that in heart chambers of older animals (6‐month‐old sedentary RA: 0.17 ± 0.02pg, LA: 0.14 ± 0.07pg, VT: 0.03 ± 0.02pg of GDNF/mg of tissue; 12–month‐old sedentary RA: 0.02 ± 0.01pg, LA: 0.018 ± 0.012pg of GDNF/mg of tissue, VT: Non‐detectable; 18‐month‐old sedentary RA: 0.000172 ± 0.000170pg, LA: 0.00128 ± 0.00236pg of GDNF/mg of tissue, VT: Non‐detectable). The results show that exercise significantly increased GDNF protein content in all heart chambers, in all age‐matched groups (14‐week‐old sedentary: RA: 1.20 ± 0.17pg, LA: 1.30 ± 0.14pg, VT: 1.27 ± 0.10pg of GDNF/mg of tissue and 14‐week‐old exercised: RA: 2.97 ± 0.27pg, LA: 2.06 ± 0.23pg, VT: 2.57 ± 0.32pg of GDNF/mg of tissue; 12‐month‐old exercised RA: 0.25 ± 0.07pg, LA: 0.11 ± 0.03pg, VT: 0.02 ± 0.01pg of GDNF/mg of tissue; 18‐month‐old exercised RA: 0.18 ± 0.21pg, LA: 0.04 ± 0.03pg, VT: 0.006 ± 0.006pg of GDNF/mg of tissue). These results suggest that GDNF protein content decreases with aging in all heart chambers and that exercise increases GDNF protein content in all heart chambers. The observed lowering of GDNF content with age may contribute to altered neural function observed in aged individuals, while the increased GDNF expression observed following exercise could help explain beneficial effects of exercise on cardiac innervation.Support or Funding InformationNIH 1 R15 AG022908‐01A2/NSF 0552517This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Suppression of MicroRNA-383 Enhances Therapeutic Potential of Human Bone-Marrow-Derived Mesenchymal Stem Cells in Treating Spinal Cord Injury via GDNF

Background/Aims: Transplantation of bone-marrow-derived mesenchymal stem cells (MSCs) has been used to treat spinal cord injury (SCI) to enhance tissue repair and neural cell regeneration. Glial cell line derived neurotrophic factor (GDNF) is an identified neural growth and survival factor. Here, we examined whether modification of GDNF levels in MSCs may further increase the potential of MSCs in promoting neural cell regeneration and subsequently the therapeutic outcome. Methods: We examined the mRNA and protein levels of GDNF in human MSCs by RT-qPCR and Western blot, respectively. Bioinformatics analyses were done to predict microRNAs (miRNAs) that target GDNF in MSCs. The functional binding of miRNAs to GDNF mRNA was examined by a dual luciferase reporter assay. MSCs were transduced with adeno-associated virus (AAV) carrying null or antisense for miR-383 (as-miR-383), which were transplanted into nude rats that underwent SCI. The intact tissue, cavity volume, and recovery of locomotor activity were assessed. Results: MSCs expressed very low GDNF protein, but surprisingly high levels of GDNF mRNA. Bioinformatics analyses showed that miR-383 inhibited protein translation of GDNF, through binding to the 3’-UTR of the GDNF mRNA. MSCs transduced with AAV-as-miR-383 further increased the intact tissue percentage, decreased cavity volume, and enhanced the recovery of locomotor activity in nude rats that underwent SCI, compared to MSCs. Conclusions: Suppression of miR-383 may increase the therapeutic potential of human bone-marrow-derived MSCs in treating SCI via augmentation of GDNF protein levels.

Glial Cell Line Derived Neurotrophic Factor Protein Secretion by Skeletal Muscles is Altered by Activity of L‐Type Calcium Channels

Glial cell line‐derived neurotrophic factor (GDNF) is an important signaling molecule throughout the peripheral nervous system. GDNF protein is produced and secreted by skeletal muscle cells and helps maintain motor neuron innervation at the neuromuscular junction. Treatment with exogenous GDNF prevents denervation which occurs with aging and neurodegenerative disease. The therapeutic potential of GDNF cannot be fully explored without understanding the mechanisms by which GDNF protein production is regulated. Although the specific cellular events necessary for regulation of GDNF secretion remains to be elucidated, previous studies from our laboratory have shown that increased skeletal muscle activity significantly alters GDNF protein levels in vivo and in vitro. The primary objective of this study was to determine the role of calcium in regulating GDNF protein secretion by skeletal muscle cells. Skeletal muscle cells (C2C12) were grown in culture and allowed to differentiate into myotubes which display a contractile phenotype. The following treatments were given, Nifedipine to block the voltage‐gated L‐type calcium channels, and Bay K8644 to enhance calcium influx through L‐type calcium channels. Both Western Blot and ELISA were employed for protein identification and GDNF protein quantification respectively. Preliminary results indicate that treatment of myotubes with Bay K8644 (100μM) caused an increase in GDNF secretion to 170% of that in untreated controls. Treatment of cells with Nifedipine blocked the stimulatory effect of Bay K8644, and treatment with Nifedipine alone (10μM and 100μM) inhibited GDNF secretion to 77% of that in control cells. The results suggest that secretion of GDNF protein by C2C12 myotubes is regulated by intracellular calcium levels. Acquiring a greater understanding of the role that calcium plays in regulating GDNF production may help to identify potential sites for therapeutic intervention to increase or decrease GDNF production.Support or Funding InformationNSF Grant DBI‐1062883 and NIH Grant 1R15AG022908‐01A2

Acute and chronic effects of lithium on BDNF and GDNF mRNA and protein levels in rat primary neuronal, astroglial and neuroastroglia cultures.

The neuroprotective effect of lithium has been attributed to its therapeutic action. However, the role of glial cells particularly astrocytes, and the possible interactions between neurons and astrocytes in neuroprotective effects of lithium have been disregarded. Thus, the aim of this study was to evaluate the direct effects of lithium on brain derived neurotrophic factor (BDNF) and glial cell line derived neurotrophic factor (GDNF) in rat primary neuronal, astrocytes, and mixed neuro-astroglial cultures to assess the possible effects of lithium on astrocytes and neuro-astroglia interactions. Rat primary astrocyte, neuronal and mixed neuro-astrocyte cultures were prepared from cortices of 18-day embryos. Cell cultures were exposed to lithium (1 mM) or vehicle for 1 day (acute) or 7 days (chronic). BDNF and GDNF mRNA and protein levels were determined by RT-PCR and ELISA, respectively. Chronic but not acute lithium treatment increased intracellular BDNF and GDNF protein levels in rat primary neuronal and astrocyte cultures, respectively (P<0.05). However, chronic lithium treatment had no significant effect on intracellular BDNF protein level in astrocyte and mixed neuron-astrocyte cultures or GDNF protein levels in mixed neuron-astrocyte culture. Furthermore, acute and chronic lithium treatment had no significant effect on mRNA and extracellular BDNF and GDNF protein levels in three studied cultures. Present study showed that chronic lithium treatment affected neurotrophins both in neurons and astrocytes in a cell-type specific manner with no effect on neuron-astrocyte interactions. The findings of this study also highlighted the importance of astrocytes as drug targets involved in the neuroprotective action of lithium.

Temporal changes in neurotrophic factors and neurite outgrowth in the major pelvic ganglion following cavernous nerve injury.

Despite nerve-sparing radical prostatectomy, nerve damage and erectile dysfunction (ED) prevail, and preventing neurodegeneration is of great importance. Neurotrophic factors and neurite outgrowth were characterized in major pelvic ganglia (MPG) following bilateral cavernous nerve injury (BCNI). Young male Sprague-Dawley rats underwent sham or BCNI surgery, and the intracavernosal pressure to mean arterial pressure ratio was measured 2, 7, 14, 21, 30, and 60 days following injury (n = 8/group). MPG gene expression (qPCR) and Western blot were performed for glial cell line-derived neurotrophic factor (GDNF), nerve growth factor (NGF), neurturin, neurotrophin (NT)-3, NT4, brain-derived neurotrophic factor (BDNF), vascular endothelial growth factor, and activating transcription factor 3 (ATF3). Additional rats were injured, and MPGs were removed 24 hr, 48 hr, 3 days, and 7 days following BCNI (n = 3/group). MPGs were cultured in Matrigel, and neurite outgrowth was measured. Erections were impaired early and improved by 60 days in BCNI rats. GDNF, NGF, BDNF, and ATF3 gene expression was significantly increased and NT3 was decreased in MPGs following BCNI (48 hr to 21 days, P < 0.05). GDNF and NGF protein levels were elevated in 48-hr BCNI rats. MPG neurite outgrowth from 24-hr and 48-hr BCNI was higher than sham (658 ± 19 μm, 607 ± 24 μm, 393 ± 23 μm, respectively, P < 0.05). Further studies examining the roles of neurotrophic factors in modulating signaling pathways may provide therapeutic avenues for neurogenically mediated ED.

Selegiline rescues gait deficits and the loss of dopaminergic neurons in a subacute MPTP mouse model of Parkinson’s disease

The monoamine oxidase type-B (MAO-B) inhibitor, selegiline, is often recommended as a first-line treatment for Parkinson's disease (PD) and has been shwon to possess neuroprotective effects. The aim of the present study was to determine whether selegiline increases the levels of the neurotrophic factors (NTFs), glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF), and whether it rescues motor dysfunction and the loss of dopaminergic neurons in mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced lesions. We found that the oral administration of selegiline (1.0 mg/kg/day for 14 days) successfully suppressed the MPTP-induced reduction of nigral dopaminergic neurons and striatal fibers (192.68 and 162.76% of MPTP-exposed animals, respectively; both P<0.001). Moreover, improvements in gait dysfunction were observed after 7 and 14 days of a low dose of selegiline that is reported not to inhibit MAO‑B. Furthermore, there was a significant increase in GDNF and BDNF mRNA (2.10 and 2.75-fold) and protein levels (143.53 and 157.05%) in the selegiline-treated mice compared with the saline-treated MPTP-exposed mice. In addition, the Bax/Bcl-2 gene and protein expression ratios were significantly increased in the MPTP-exposed mice, and this effect was reversed by selegiline. Correlation analysis revealed that gait measurement and GDNF/BDNF levels positively correlated with the number of dopaminergic neurons. These findings demonstrate that selegiline has neurorescue effects that are possibly associated with the induction of NTFs and anti-apoptotic genes.

Short-term exercise increases GDNF protein levels in the spinal cord of young and old rats

Neurotrophic factors may play a role in exercise-induced neuroprotective effects, however it is not known if exercise mediates changes in glial cell line-derived neurotrophic factor (GDNF) protein levels in the spinal cord. The aim of the current study was to determine if 2weeks of exercise alters GDNF protein content in the lumbar spinal cord of young and old rats. GDNF protein was quantified via an enzyme-linked immunosorbent assay and Western blot. Immunohistochemical analysis localized GDNF in choline acetyltransferase (ChAT)-positive motor neurons and cell body areas were measured. Involuntary running in the young animals appeared to elicit the greatest increase in GDNF protein content (sixfold increase), followed by swimming (threefold increase) and voluntary running (twofold increase); however there was no significant difference between the modalities of exercise. Low-intensity running of the old animals significantly increased GDNF protein content in the spinal cord. Both young and old exercised animals showed a doubling in ChAT-positive motor neuron cell body areas. These results suggest that GDNF protein content in the spinal cord is modulated by exercise.

GDNF is a novel ethanol‐responsive gene in the VTA: implications for the development and persistence of excessive drinking

Glial cell line-derived neurotrophic factor (GDNF) is a potent inhibitor of ethanol consumption and relapse, and GDNF heterozygous knockout mice display increased reward sensitivity to ethanol and consume more ethanol after a period of abstinence than their wild-type littermates. Here, we tested whether ethanol alters GDNF expression in the ventral tegmental area (VTA; GDNF's site of action) and/or the nucleus accumbens (NAc; the main source of GDNF), and if so, determine the role of the endogenous growth factor in the regulation of ethanol consumption. Systemic administration of ethanol increased GDNF expression and protein levels in the VTA, but not the NAc. Additionally, GDNF levels were elevated after an ethanol-drinking session in rats that consumed ethanol in the intermittent-access two-bottle choice procedure for 1 week, but not 7 weeks. Deprivation following 7 weeks of excessive ethanol intake reduced GDNF levels, while a short ethanol binge drinking period following deprivation upregulated GDNF expression. Importantly, knockdown of GDNF within the VTA using adenovirus expressing short hairpin RNA facilitated the escalation of ethanol drinking by ethanol-naïve rats, but not by rats with a history of excessive ethanol consumption. These results suggest that during initial ethanol-drinking experiences, GDNF in the VTA is increased and protects against the development of excessive ethanol intake. However, the growth factor's protective response to ethanol breaks down after protracted excessive ethanol intake and withdrawal, resulting in persistent, excessive ethanol consumption.

Glial cell-line derived neurotrophic factor (GDNF) replacement attenuates motor impairments and nigrostriatal dopamine deficits in 12-month-old mice with a partial deletion of GDNF

Glial cell-line derived neurotrophic factor (GDNF) has been established as a growth factor for the survival and maintenance of dopamine (DA) neurons. In phase I clinical trials, GDNF treatment in Parkinson's disease patients led to improved motor function and GDNF has been found to be down regulated in Parkinson's disease patients. Studies using GDNF heterozygous (Gdnf+/−) mice have demonstrated that a partial reduction of GDNF leads to an age-related accelerated decline in nigrostriatal DA system- and motor-function and increased neuro-inflammation and oxidative stress in the substantia nigra (SN). Therefore, the purpose of the current studies was to determine if GDNF replacement restores motor function and functional markers within the nigrostriatal DA system in middle-aged Gdnf+/− mice. At 11months of age, male Gdnf+/− and wildtype (WT) mice underwent bilateral intra-striatal injections of GDNF (10μg) or vehicle. Locomotor activity was assessed weekly 1–4weeks after treatment. Four weeks after treatment, their brains were processed for analysis of GDNF levels and various DAergic and oxidative stress markers. An intrastriatal injection of GDNF increased motor activity in Gdnf+/− mice to levels comparable to WT mice (1week after injection) and this effect was maintained through the 4-week time point. This increase in locomotion was accompanied by a 40% increase in striatal GDNF protein levels and SN GDNF expression in Gdnf+/− mice. Additionally, GDNF treatment significantly increased the number of tyrosine hydroxylase (TH)-positive neurons in the SN of middle-aged Gdnf+/− mice, but not WT mice, which was coupled with reduced oxidative stress in the SN. These studies further support that long-term changes related to the dysfunction of the nigrostriatal pathway are influenced by GDNF expression and add that this dysfunction appears to be responsive to GDNF treatment. Additionally, these studies suggest that long-term GDNF depletion alters the biological and behavioral responses to GDNF treatment.

Injury-Induced Accumulation of Glial Cell Line-Derived Neurotrophic Factor in the Rostral Part of the Injured Rat Spinal Cord

The spinal cord of a 7-week-old female Wistar rat was hemi-transected at thoracic position 10 with a razor blade, and changes in glial cell line-derived neurotrophic factor (GDNF) protein and mRNA expression levels in the spinal cord were examined. GDNF protein and mRNA expression levels were evaluated by enzyme immunoassay and reverse transcription polymerase chain reaction, respectively. Although GDNF is distributed in the healthy spinal cord from 150 to 400 pg/g tissue in a regionally dependent manner, hemi-transection (left side) of the spinal cord caused a rapid increase in GDNF content in the ipsilateral rostral but not in the caudal part of the spinal cord. On the other hand, injury-induced GDNF mRNA was distributed limitedly in both rostral and caudal stumps. These observations suggest the possibility that increased GDNF in the rostral part is responsible for the accumulation of GDNF that may be constitutively transported from the rostral to caudal side within the spinal cord. Although such local increase of endogenous GDNF protein may not be sufficient for nerve regeneration and locomotor improvement, it may play a physiological role in supporting spinal neurons including motoneurons.

GDNF Contributes to Oestrogen‐Mediated Protection of Midbrain Dopaminergic Neurones

Parkinson's disease (PD) is characterised by the preferential loss of dopaminergic neurones from the substantia nigra (SN) that leads to the hallmark motor disturbances. Animal and human studies suggest a beneficial effect of oestrogen to the nigrostriatal system, and the regulation of neurotrophic factor expression by oestrogens has been suggested as a possible mechanism contributing to that neuroprotective effect. The present study was designed to investigate whether the neuroprotection exerted by 17β-oestradiol on nigrostriatal dopaminergic neurones is mediated through the regulation of glial cell line-derived neurotrophic factor (GDNF) expression. Using an in vivo rat model of PD, we were able to confirm the relevance of 17β-oestradiol in defending dopaminergic neurones against 6-hydroxydopamine (6-OHDA) toxicity. 17β-oestradiol, released by micro-osmotic pumps, implanted 10 days before intrastriatal 6-OHDA injection, prevented the loss of dopaminergic neurones induced by 6-OHDA. 17β-oestradiol treatment also promoted an increase in GDNF protein levels both in the SN and striatum. To explore the relevance of GDNF increases to 17β-oestradiol neuroprotection, we analysed, in SN neurone-glia cultures, the effect of GDNF antibody neutralisation and RNA interference-mediated GDNF knockdown. The results showed that both GDNF neutralisation and GDNF silencing abolished the dopaminergic protection provided by 17β-oestradiol against 6-OHDA toxicity. Taken together, these results strongly identify GDNF as an important player in 17β-oestradiol-mediated dopaminergic neuroprotection.

The Effects of Progesterone on Glial Cell Line-derived Neurotrophic Factor Secretion from C6 Glioma Cells.

Progesterone is a steroid hormone whose biology has been greatly studied within the confines of reproductive function. In recent years, the neuroprotective effects of progesterone have attracted growing interest. Glial cell line-derived neurotrophic factor (GDNF), is a neurotrophic factor which plays a crucial role in the development and maintenance of distinct sets of central and peripheral neurons. In the present study, we investigated the potential implication of GDNF in the neuroprotective action of progesterone. Cultured rat C6 glioma cells were treated with progesterone (100 nm, 1 µM, and 10 µM) or its vehicle. After 24, 36, 48 and 72 hr, GDNF protein levels were measured in the cell-conditioned media and cell lysates using a GDNF ELISA kit. Cell numbers were determined by a cell-counting assay kit. Forty-eight hr treatment with progesterone (10 µM) resulted in a significant elevation of GDNF secretion from C6 glioma cells that remained elevated up to 72 hr. The intracellular content of GDNF and cell numbers were not affected by progesterone treatment. Stimulation of GDNF release from glial cells appears as a potential mechanism through which progesterone exerts its neuroprotective effects.

Long‐term expression of glial cell line‐derived neurotrophic factor slows, but does not stop retinal degeneration in a model of retinitis pigmentosa

Retinitis pigmentosa is a group of diseases in which one of hundreds of mutations causes death of rod photoreceptor cells and then cones gradually die from oxidative damage. As different mutations cause rod cell death by different mechanisms, mutation-specific treatments are needed. Another approach is to use a neurotrophic factor to promote photoreceptor survival regardless of the mechanism of cell death, and previous studies have demonstrated encouraging short-term results with gene transfer of glial cell line-derived neurotrophic factor (GDNF). We generated rd10 mice with doxycycline-inducible expression of GDNF in photoreceptors (Tet/IRBP/GDNF-rd10 mice) or retinal pigmented epithelial cells (Tet/VMD2/GDNF-rd10 mice). In doxycycline-treated Tet/IRBP/GDNF-rd10 mice, there was a 9.3 × 10(4) -fold increase in Gdnf mRNA at P35 and although it decreased over time, it was still increased by 9.4 × 10(3) -fold at P70. Gdnf mRNA was increased 4.5 × 10(2) -fold in doxycycline-treated Tet/VMD2/GDMF-rd10 mice at P35 and was not significantly decreased at P70. GDNF protein levels were increased about 2.3-fold at P35 and 30% at P70 in Tet/IRBP/GDNF-rd10 mice, and in Tet/VMD2/GDNF-rd10 mice they were increased 30% at P35 and not significantly increased at P70. Despite the difference in expression, Tet/IRBP/GDNF-rd10 and Tet/VMD2/GDNF-rd10 mice had comparable significant increases in outer nuclear layer thickness and mean photopic and scotopic ERG b-wave amplitudes compared with rd10 mice at P35 which decreased, but was still significant at P70. Compared with rd10 mice, Tet/IRBP/GDNF-rd10 and Tet/VMD2/GDNF-rd10 mice had comparable significant improvements in cone density at P50 that decreased, but were still significant at P70. These data indicate that despite a large difference in expression of GDNF, Tet/IRBP/GDNF-rd10 and Tet/VMD2/GDNF-rd10 provide comparable slowing of photoreceptor degeneration, but cannot stop the degeneration.

Exercise-dependent regulation of glial cell line-derived neurotrophic factor (GDNF) expression in skeletal muscle and its importance for the neuromuscular system

The focus of this review is to highlight the importance of glial cell line-derived neurotrophic factor (GDNF) for the motor nervous system. GDNF is the most potent survival factor for motor neurons, where it enhances maintenance and survival of both developing and mature motor neurons in vivo and in vitro. GDNF aids in neuromuscular junction formation, maintenance, and plasticity, where skeletal muscle-derived GDNF may be responsible for this phenomenon. Increased levels of physical activity can increase GDNF protein levels in skeletal muscle, where alterations in acetylcholine and acetylcholine receptor activation may be involved in regulation of these changes observed. With inactivity and disuse, GDNF expression shows different patterns of regulation in the central and peripheral nervous systems. Due to its potent effects for motor neurons, GDNF is being extensively studied in neuromuscular diseases.

Positive autoregulation of GDNF levels in the ventral tegmental area mediates long-lasting inhibition of excessive alcohol consumption

Glial cell line-derived neurotrophic factor (GDNF) is an essential growth factor for the survival and maintenance of the midbrain dopaminergic (DA-ergic) neurons. Activation of the GDNF pathway in the ventral tegmental area (VTA), where the GDNF receptors are expressed, produces a long-lasting suppression of excessive alcohol consumption in rats. Previous studies conducted in the DA-ergic-like cells, SHSY5Y, revealed that GDNF positively regulates its own expression, leading to a long-lasting activation of the GDNF signaling pathway. Here we determined whether GDNF activates a positive autoregulatory feedback loop in vivo within the VTA, and if so, whether this mechanism underlies the long-lasting suppressive effects of the growth factor on excessive alcohol consumption. We found that a single infusion of recombinant GDNF (rGDNF; 10 μg) into the VTA induces a long-lasting local increase in GDNF mRNA and protein levels, which depends upon de novo transcription and translation of the polypeptide. Importantly, we report that the GDNF-mediated positive autoregulatory feedback loop accounts for the long-lasting inhibitory actions of GDNF in the VTA on excessive alcohol consumption. Specifically, the long-lasting suppressive effects of a single rGDNF infusion into the VTA on excessive alcohol consumption were prevented when protein synthesis was inhibited, as well as when the upregulation of GDNF expression was prevented using short hairpin RNA to focally knock down GDNF mRNA in the VTA. Our results could have implications for the development of long-lasting treatments for disorders in which GDNF has a beneficial role, including drug addiction, chronic stress and Parkinson's disease.

A Detoxified Extract ofRhus vernicifluaStokes Upregulated the Expression of BDNF and GDNF in the Rat Brain and the Human Dopaminergic Cell Line SH-SY5Y

Rhus verniciflua Stokes (RVS) has traditionally been used as a food supplement and a traditional herbal medicine for centuries in Korea. This study attempted to evaluate the effects of RVS on the expression of Brain derived neurotrophic factor (BDNF) and Glial cell line-derived neurotrophic factor (GDNF) in SH-SY5Y cells and the rat brain. The results indicated that RVS is a potent inducer of Neurotrophic factor (NTF) production both in vitro and in vivo. Treatment with 10 µg/ml and 10 mg/kg RVS for 4 h of SH-SY5Y cells and rats yielded significant increases in BDNF and GDNF protein levels. We also detected BDNF and GDNF immunoreactive neurons in the rat brain. Both BDNF and GDNF-immunohistochemical staining was markedly enhanced in the animals treated with RVS. These results suggest that RVS serves as an ideal adjuvant in regard to regulating NTF expression, and can contribute to neuroprotection in neurodegenerative diseases.