Minoru Takebayashi

Decreased levels of whole blood glial cell line-derived neurotrophic factor (GDNF) in remitted patients with mood disorders

Recent post-mortem and imaging studies provide evidence for a glial reduction in different brain areas in mood disorders. This study was aimed to test whether glial cell line-derived neurotrophic factor (GDNF), a member of transforming growth factor (TGF)-beta superfamily, in blood levels was associated with mood disorders. We measured GDNF and TGF-beta levels in whole blood in remitted patients with mood disorders [n=56; major depressive disorders (MDD) 39, bipolar disorders (BD) 17] and control subjects (n=56). GDNF and TGF-beta were assayed with the sandwich ELISA method. Total GDNF levels were significantly lower in MDD and in BD than in control subjects (MDD, p=0.0003; BD, p=0.018), while no significant difference in total TGF-beta1 or total TGF-beta2 levels was found in these groups. Our study suggests that lower GDNF levels might be involved in the pathophysiology of mood disorders, although this preliminary study has several limitations.

Antidepressants Increase Glial Cell Line-Derived Neurotrophic Factor Production through Monoamine-Independent Activation of Protein Tyrosine Kinase and Extracellular Signal-Regulated Kinase in Glial Cells

Recent studies show that neuronal and glial plasticity are important for therapeutic action of antidepressants. We previously reported that antidepressants increase glial cell line-derived neurotrophic factor (GDNF) production in rat C6 glioma cells (C6 cells). Here, we found that amitriptyline, a tricyclic antidepressant, increased both GDNF mRNA expression and release, which were selectively and completely inhibited by mitogen-activated protein kinase kinase inhibitors. Indeed, treatment of amitriptyline rapidly increased extracellular signal-regulated kinase (ERK) activity, as well as p38 mitogen-activated protein kinase and c-Jun NH2-terminal kinase activities. Furthermore, different classes of antidepressants also rapidly increased ERK activity. The extent of acute ERK activation and GDNF release were significantly correlated to each other in individual antidepressants, suggesting an important role of acute ERK activation in GDNF production. Furthermore, antidepressants increased the acute ERK activation and GDNF mRNA expression in normal human astrocytes as well as C6 cells. Although 5-hydroxytryptamine (serotonin) (5-HT), but not noradrenaline or dopamine, increased ERK activation and GDNF release via 5-HT2A receptors, ketanserin, a 5-HT2A receptor antagonist, did not have any effect on the amitriptyline-induced ERK activation. Thus, GDNF production by amitriptyline was independent of monoamine. Both of the amitriptyline-induced ERK activation and GDNF mRNA expression were blocked by genistein, a general protein tyrosine kinase (PTK) inhibitor. Actually, we found that amitriptyline acutely increased phosphorylation levels of several phosphotyrosine-containing proteins. Taken together, these findings indicate that ERK activation through PTK regulates antidepressant-induced GDNF production and that the GDNF production in glial cells may be a novel action of the antidepressant, which is independent of monoamine.

Plasma Concentrations of Interleukin-1β, Interleukin-6, Soluble Interleukin-2 Receptor and Tumor Necrosis Factor α of Depressed Patients in Japan

There are a number of investigations which indicate the important relationship between depression and cytokines. In this study, we investigated plasma interleukin (IL)-1β, IL-6, soluble IL-2 receptor (sIL-2R) and tumor necrosis factor (TNF)-α of depressed patients whose clinical evaluation was performed by the Hamilton Rating Scale for Depression (HAM-D) and the Profile of Mood States (POMS). They were compared with those of the control subjects, and before and after treatment with antidepressants. Before the treatment, plasma IL-1β, IL-6, sIL-2R and TNF-α of the patients were not significantly different from those of the control subjects. sIL-2R was positively correlated with the POMS-tension-anxiety subscale and tended to have a positive correlation with HAM-D. After pharmacotherapy, TNF-α levels of the depressed patients increased, without any relationship between the change in the HAM-D or the POMS and the change in TNF-α. These results suggest that the plasma sIL-2R concentration is associated with mood state, and that the plasma TNF-α concentration is increased after pharmacotherapy in Japanese depressed patients.

Plasma dehydroepiandrosterone sulfate in unipolar major depression

Summary. We investigated plasma dehydroepiandrosterone sulfate (DHEAS) and cortisol levels in 12 patients with unipolar depression and 11 matched normal controls. The depressed patients showed significantly higher values of plasma DHEAS and cortisol than the controls. After 4 weeks of treatment with antidepressants (mainly clomiplamine), the high plasma DHEAS levels recovered.This finding showed the possible relationship between plasma DHEAS levels and depression, as well as cortisol levels.

Effect of β-estradiol on voltage-gated Ca2+ channels in rat hippocampal neurons: a comparison with dehydroepiandrosterone

We investigated the effects of beta-estradiol, dehydroepiandrosterone and dehydroepiandrosterone sulfate on intracellular calcium concentration ([Ca(2+)](i)) increases induced by gamma-aminobutyric acid (GABA), high K(+) and N-methyl-D-aspartate acid (NMDA) in cultured hippocampal neurons. Acute treatment with beta-estradiol, dehydroepiandrosterone and dehydroepiandrosterone sulfate inhibited the GABA-induced [Ca(2+)](i) increases to the similar extent. Tamoxifen, an estrogen receptor antagonist, did not block the inhibitory effects of beta-estradiol. On the other hand, GABA type A (GABA(A)) receptor antagonists, picrotoxin and bicuculline, blocked the GABA-induced [Ca(2+)](i) increases. Previously, we demonstrated that GABA- and high K(+)-induced [Ca(2+)](i) increases were commonly mediated by voltage-gated calcium channels (VGCCs). Therefore, we examined the effects of these steroids on the high K(+)-induced [Ca(2+)](i) increases. The inhibitory effect of beta-estradiol on the high K(+)-induced [Ca(2+)](i) increases was much greater than that of dehydroepiandrosterone and dehydroepiandrosterone sulfate. beta-Estradiol inhibited the NMDA-induced [Ca(2+)](i) increases with an IC(50) of 51.8 microM and NMDA responses were reduced to half in the presence of 10 micro M nifedipine, indicating that the NMDA-induced [Ca(2+)](i) increases also involved VGCCs. Further, we examined the inhibitory effect of beta-estradiol on the high K(+)-induced [Ca(2+)](i) increases in the presence of a N-type VGCCs antagonist, 1 microM omega-conotoxin, or a L-type VGCCs antagonist, 10 microM nifedipine. The IC(50) value of beta-estradiol alone (45.5 microM) was similar to that of omega-conotoxin (33.1 microM), while the value combined with nifedipine was reduced to 2.2 microM. beta-Estradiol also abolished the positive modulatory effect of L-type VGCCs agonist, 1,4-dihydro-2,6-dimethyl-5-nitro-4-[2-(trifluoromethyl)phenyl]pyridine-3-carboxylic acid methyl ester (Bay K 8644). Our results showed that the inhibitory mechanism of beta-estradiol is different from that of dehydroepiandrosterone and dehydroepiandrosterone sulfate and beta-estradiol may act primarily at L-type VGCCs.

Plasma levels of vascular endothelial growth factor and fibroblast growth factor 2 in patients with major depressive disorders.

We investigated the plasma levels of VEGF and FGF-2, important factors for regulation of neuroplasticity such as neurogenesis, in patients in remission from major depressive disorders (MDD). The plasma VEGF levels were significantly higher in the MDD patients than in the matched control subjects, while no significant difference in plasma FGF-2 levels was found. In particular, the MDD patients with family history of psychiatric disorders, but not patients without such a family history, showed significantly higher values of plasma VEGF than the controls. Although this is a preliminary study, altered VEGF levels might be involved in the pathophysiology of MDD.

Serotonin (5-HT) induces glial cell line-derived neurotrophic factor (GDNF) mRNA expression via the transactivation of fibroblast growth factor receptor 2 (FGFR2) in rat C6 glioma cells

We previously reported that serotonin (5‐HT) increased glial cell line‐derived neurotrophic factor (GDNF) release in a 5‐HT2 receptor (5‐HT2R) and mitogen‐activated protein kinase kinase/extracellular signal‐related kinase (MEK/ERK)‐dependent manner in rat C6 glioma cells (C6 cells), a model of astrocytes. We herein found that 5‐HT‐induced rapid ERK phosphorylation was blocked by 5‐HT2R antagonists in C6 cells. We therefore examined 5‐HT‐induced ERK phosphorylation to reveal the mechanism of 5‐HT‐induced GDNF mRNA expression. As 5‐HT‐induced ERK phosphorylation was blocked by inhibitors for Gαq/11 and fibroblast growth factor receptor (FGFR), but not for second messengers downstream of Gαq/11, 5‐HT2R‐mediated FGFR transactivation was suggested to be involved in the ERK phosphorylation. Although FGFR1 and 2 were functionally expressed in C6 cells, 5‐HT selectively phosphorylated FGFR2. Indeed, small interfering RNA for FGFR2, but not for FGFR1, blocked 5‐HT‐induced ERK phosphorylation. As Src family tyrosine kinase inhibitors and microtubule depolymerizing agents blocked 5‐HT‐induced FGFR2 phosphorylation, Src family tyrosine kinase and stabilized microtubules were suggested to act upstream of FGFR2. Finally, 5‐HT‐induced GDNF mRNA expression was also inhibited by the blockade of 5‐HT2R, FGFR, and Src family tyrosine kinase. In conclusion, our findings suggest that 5‐HT induces GDNF mRNA expression via 5‐HT2R‐mediated FGFR2 transactivation in C6 cells.

Antidepressants induce acute CREB phosphorylation and CRE-mediated gene expression in glial cells: a possible contribution to GDNF production.

Recently, the changes of neuronal and glial plasticity related gene expression following the increase of monoamine are suggested to be important for the therapeutic effect of antidepressants. We previously showed that antidepressants increased glial cell line-derived neurotrophic factor (GDNF) expression, which was dependent on acute activation of protein tyrosine kinase (PTK) and extracellular signal-regulated kinase (ERK) in rat C6 glioma cells (C6 cells) and normal human astrocytes (NHA). Transcription of many genes including GDNF is directed by the cAMP responsive element (CRE) and its cognate transcription factor CRE binding protein (CREB). In this study, we showed that amitriptyline, a tricyclic antidepressant, acutely increased phosphorylation of CREB, without altering the level of total CREB in C6 cells as well as in NHA. In contrast, acute amitriptyline treatment did not affect phosphorylation of CREB in SH-SY5Y cells, a human neuroblastoma cell line. Different classes of antidepressants as well as amitriptyline acutely increased phosphorylation of CREB, but haloperidol and diazepam did not. The amitriptyline-induced phosphorylation of CREB was completely blocked by U0126 [a mitogen-activated protein (MAP) kinase kinase 1 inhibitor] and genistein (a PTK inhibitor), but not by inhibitors of protein kinase A, p38 MAP kinase, or Ca(2+)/calmodulin-dependent kinase. Amitriptyline treatment also increased the expression of luciferase reporter gene regulated by CRE elements. The amitriptyline-induced luciferase activity was completely inhibited by U0126 in the same as phosphorylation of CREB. These results suggest that antidepressants acutely increase CREB activity in PTK and ERK-dependent manners, which might contribute to gene expression including GDNF in glial cells.

Serotonin increases glial cell line-derived neurotrophic factor release in rat C6 glioblastoma cells.

Antidepressants, which increase monoamine levels, induce glial cell line-derived neurotrophic factor (GDNF) release in C6 cells. Thus, we examined whether monoamines affect on GDNF release in C6 cells. We found that serotonin (5-HT) specifically increased GDNF mRNA expression and GDNF release in a dose- and time-dependent manner. The 5-HT-induced GDNF release was mediated through the MEK/mitogen-activated protein kinase (MAPK) pathway and, at least, 5-HT(2A) receptors. The action of 5-HT on GDNF release may provide important insights into the mechanism of antidepressants.

γ-Aminobutyric acid increases intracellular Ca2+ concentration in cultured cortical neurons: role of Cl− transport

The effect of gamma-aminobutyric acid (GABA) on intracellular Ca2+ concentration ([Ca2+]i) in cultured prenatal rat cortical neurons was investigated using fluorescence imaging. GABA or muscimol, but not baclofen, increased [Ca2+]i in a dose-dependent manner. The GABAA receptor antagonists, bicuculline and picrotoxin, inhibited the GABA response. Furosemide, an inhibitor of the Na+/K+/2Cl- cotransporter, inhibited the GABA response in a noncompetitive manner. Ethacrynic acid, an inhibitor of an ATP-dependent Cl- pump, also inhibited the GABA-induced increased in [Ca2+]i. These results suggest a role for Cl- transport processes in the GABA response. The coapplication of GABA and high K+ led to a non-additive increase in the GABA response. The GABA response was also inhibited by nifedipine, a voltage-gated Ca2+ channel blocker, and abolished by the absence of extracellular Ca2+. Results indicate that the GABA response shares a common pathway of Ca2+ movement with the high K(+)-induced response. These observations suggest that the stimulation with GABA results in Ca2+ influx through voltage-gated Ca2+ channels, and that these effects are dependent on Cl- transport systems.