Amitriptyline up-regulates connexin43-gap junction in rat cultured cortical astrocytes via activation of the p38 and c-Fos/AP-1 signalling pathway.

BACKGROUND AND PURPOSE: Intercellular communication via gap junctions, comprised of connexin (Cx) proteins, allow for communication between astrocytes, which in turn is crucial for maintaining CNS homeostasis. The expression of Cx43 is decreased in post-mortem brains from patients with major depression. A potentially novel mechanism of tricyclic antidepressants is to increase the expression and functioning of gap junctions in astrocytes. EXPERIMENTAL APPROACH: The effect of amitriptyline on the expression of Cx43 and gap junction intercellular communication (GJIC) in rat primary cultured cortical astrocytes was investigated. We also investigated the role of p38 MAPK intracellular signalling pathway in the amitriptyline-induced expression of Cx43 and GJIC. KEY RESULTS: Treatment with amitriptyline for 48 h significantly up-regulated Cx43 mRNA, protein and GJIC. The up-regulation of Cx43 was not monoamine-related since noradrenaline, 5-HT and dopamine did not induce Cx43 expression and pretreatment with α- and ß-adrenoceptor antagonists had no effect. Intracellular signalling involved p38 MAPK, as amitriptyline significantly increased p38 MAPK phosphorylation and Cx43 expression and GJIC were significantly blocked by the p38 inhibitor SB 202190. Furthermore, amitriptyline-induced Cx43 expression and GJIC were markedly reduced by transcription factor AP-1 inhibitors (curcumin and tanshinone IIA). The translocation of c-Fos from the cytosol and the nucleus of cortical astrocytes was increased by amitriptyline, and this response was dependent on p38 activity. CONCLUSION AND IMPLICATION: These findings indicate a novel mechanism of action of amitriptyline through cortical astrocytes, and further suggest that targeting this mechanism could lead to the development of a new class of antidepressants.

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.

IL-4-producing CD8(+) T cells may be an immunological hallmark of chronic GVHD.

Chronic graft-versus-host disease (cGVHD) occurs in approximately 60-80% of those who survive over 100 days after allogeneic hematopoietic stem cell transplantation (allo-HSCT). However, the pathophysiology of cGVHD is poorly understood. To gain more insight into the immunological mechanism of cGVHD, we examine cytokine production of peripheral blood T cells from 19 patients in the chronic phase of allo-HSCT. The percentage of IFN-gamma-producing CD8(+) T cells among CD8(+) T cells was significantly higher in patients with or without cGVHD than in normal control subjects (P<0.001). On the other hand, the percentage of IL-4-producing CD8(+) T cells among CD8(+) T cells was significantly higher in patients with cGVHD (mean 3.3%; range 1.3-8.2%) than in patients without cGVHD (mean 1.2%; range 0.8-1.7%) and normal control subjects (mean 1.1%; range 0.1-1.6%) (both P<0.001). By contrast, the percentage of IL-4-producing CD4(+) T cells was not different among patients with and without cGVHD and normal controls. These findings suggest that IL-4-producing CD8(+) T cells may be an immunological marker of cGVHD.

A perspective on the new mechanism of antidepressants: neuritogenesis through sigma-1 receptors.

Sigma receptors were first described as one of the opiate receptor subtypes. Now it is well established that sigma receptors, existing as subtypes sigma-1 and sigma-2, are unique non-opioid receptors which are implicated in higher-ordered brain functions. Sigma-1 receptors have high to moderate affinities for (+)benzomorphans and also many psychotrophic drugs and neurosteroids. Sigma-1 receptor agonists and certain neurosteroids such as dehydroepiandrosterone sulfate (DHEA-S) have antidepressant-like effects in animal behavioral models of depression. The antidepressant-like effect induced by sigma-1 receptor agonists may involve intracellular Ca (2+) mobilization such that sigma-1 receptor agonists modulate Ca (2+) release from endoplasmic reticulum (ER) in a cytoskeletal protein-dependent manner. In addition, growth factor-induced neurite outgrowth is mediated through sigma-1 receptors, suggesting a role of antidepressants in neuroplasticity. Igmesine (JO1783), OPC-14 523 and SA4503, have recently been developed as sigma-1 agonists and are found to have antidepressant-like activity perhaps with fewer side effects. This article reviews the new potential use of sigma-1 receptor ligands in the treatment of mood disorder.

Blood dendritic cells are decreased in acute graft-versus-host disease.

The recipients of allogeneic hematopoietic stem cell transplantation (allo-HSCT) often develop acute graft-versus-host disease (aGVHD), which is closely related to morbidity and mortality. However, the essential part of the immune responses elicited in aGVHD remains largely unknown. We attempt to determine if peripheral blood dendritic cells (PBDCs) are altered in aGVHD, and find that the number of PBDCs (both myeloid and lymphoid DCs) is significantly decreased. Immunohistochemical staining of the biopsied skin from patients with aGVHD demonstrates that a number of fascin(+) cells with dendritic projections infiltrate the dermis of the skin. Based on these findings, we hypothesize that the PBDCs are recruited to the affected tissues and may thus play important roles in immune responses elicited in aGVHD.

Effect of beta-estradiol on voltage-gated Ca(2+) 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 concentrations of interleukin-1beta, interleukin-6, soluble interleukin-2 receptor and tumor necrosis factor alpha 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)-1beta, IL-6, soluble IL-2 receptor (sIL-2R) and tumor necrosis factor (TNF)-alpha 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-1beta, IL-6, sIL-2R and TNF-alpha 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-alpha levels of the depressed patients increased, without any relationship between the change in the HAM-D or the POMS and the change in TNF-alpha. These results suggest that the plasma sIL-2R concentration is associated with mood state, and that the plasma TNF-alpha concentration is increased after pharmacotherapy in Japanese depressed patients.

Three sets of diagnostic criteria for major depression and correlations with serotonin-induced platelet calcium mobilization in cancer patients.

OBJECTIVE: Enhanced serotonin-induced platelet calcium mobilization has been proposed to be a biological marker for the pathophysiology of major depression in physically healthy patients. To determine the most appropriate method of diagnosing major depression in cancer patients, we compared serotonin-induced platelet calcium mobilization between patients with and without major depression diagnosed according to three different sets of diagnostic criteria (inclusive, substitutive and exclusive). METHODS: Among the cancer patients referred to our institution between June 1997 and March 1998, 24 patients diagnosed as having major depression according to the inclusive approach (in which the nine traditional symptoms of major depression contribute towards the diagnosis of depression regardless of its presumed etiology) participated in the study. Serotonininduced platelet calcium mobilization was examined in these patients and in the same number of non-depressed controls matched for age, sex, cancer stage and cancer site. The depressed patients were then re-evaluated using substitutive and exclusive criteria, and calcium mobilization comparisons with the relevant controls were repeated. RESULTS: Compared with the controls, an enhanced serotonin-induced platelet calcium response was only observed in the patients with major depression according to the exclusive criteria. No significant enhancement was observed when the inclusive or substitutive approaches were used. CONCLUSION: These findings, based on the use of enhanced serotonin-induced platelet calcium mobilization as a biological marker, suggest that the exclusive approach might be the most valid and appropriate method of diagnosing major depression in cancer patients, while the inclusive and substitutive approaches might overestimate the occurrence of major depression in these patients.

Effects of antidepressants on gamma-aminobutyric acid- and N-methyl-D-aspartate-induced intracellular Ca(2+) concentration increases in primary cultured rat cortical neurons.

We investigated the effects of antidepressants on the intracellular Ca2+ concentration ([Ca2+]i) increases induced by gamma-aminobutyric acid (GABA) or N-methyl-D-aspartate (NMDA) in primary cultured rat cortical neurons using fluorescence imaging. Acute treatment with imipramine inhibited GABA- and NMDA-induced increases in [Ca2+]i in a concentration-dependent manner. Doses of 30 microM clomipramine, desipramine, amoxapine and maprotiline also inhibited both the GABA- and NMDA-induced [Ca2+]i increases significantly. Both inhibitory effects of the five major antidepressants on the GABA- or the NMDA-induced [Ca2+]i increases were well-correlated. Imipramine could inhibit significantly high-K+-induced [Ca2+]i increases. Our previous study has already shown that the GABA-induced [Ca2+]i increase involves a similar pathway to high-K+-induced Ca2+ influx. In conclusion, imipramine and several other antidepressants have acute inhibitory effects on the GABA-, NMDA- and high-K+-induced [Ca2+]i increases, suggesting that these inhibitory effects are not related to specific receptors. One possibility is that these effects may be commonly mediated via part of the high-K+-induced [Ca2+]i pathway.

Effect of heat stress on serotonin-2A receptor-mediated intracellular calcium mobilization in rat C6 glioma cells.

This study was conducted to investigate an effect of heat stress at 44 degrees C for 30 min on intracellular Ca2+ signaling system and on heat shock protein (HSP)-70 expression. 5-HT-induced Ca2+ mobilization was reduced 1, 3 and 6 hrs after heat stress, and recovered to the control level 12 and 24 hrs after heat stress. One hr after heat stress, Ca2+ rise was significantly decreased when the cells were stimulated by any concentration of 5-HT. Thrombin-induced Ca2+ increase was also markedly reduced 1 hr after heat stress. HSP-70 level was increased 6 and 9 hr after heat stress. In HSP synthesis inhibitor quercetin-treated cells, HSP-70 expression was not enhanced after heat stress, and Ca2+ rise in response to 5-HT did not return to the control level. However, the Ca2+ rise induced by 5-HT was not restored to the control level after stress in Ac-Asp-Glu-Val-Asp-H (DEVD)-exposed cells while DEVD had little effect on heat stress-induced synthesis of HSP-70. Dexamethasone did not alter the change in HSP-70 expression or Ca2+ response after heat stress. These results indicate that heat stress attenuated 5-HT-induced Ca2+ mobilization and that HSP-70 expression played an important role in recovery from Ca2+ impairment, possibly via protease activity in C6 cells.

Chronic electroconvulsive shock decreases (+/-) 1-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane hydrochloride (DOI)-induced wet-dog shake behaviors of dexamethasone-treated rats.

Electroconvulsive shock (ECS) therapy is considered to be an effective treatment for depression, but its mechanism of action is still unknown. We investigated the effect of chronic ECS in rats treated for 14 days with dexamethasone (Dex), a glucocorticoid receptor agonist. Chronic injection of sesame oil decreased body weight change and increased serotonin (5-HT)-2A receptor number and DOI (5-HT-2A, 2C receptor agonist)-induced wet-dog shake (WDS) behaviors. Dex treatment for 14 days decreased body weight of rats, but repeated ECS did not reverse this decrease. Dex also abolished plasma corticosterone levels, and ECS failed to restore these levels. These results indicate that chronic ECS does not antagonize the effect of Dex. The treatment with Dex increased 5-HT-2A receptor binding density of rat frontal cortex and the number of DOI-induced WDS behaviors. Chronic ECS reduced the enhanced WDS behaviors by Dex but had little effect on receptor density. These results suggest that chronic ECS might suppress 5-HT-2A receptor function at the postreceptor signaling level rather than at the receptor itself, without changing HPA axis function in Dex-treated rats.

Lithium and clonazepam treatment of two cases with cluster headache.

Patients with cluster headache are often treated with lithium. However, there are some patients who can not be fully treated with lithium alone. Two patients with cluster headache were treated with clonazepam, one of the most potent benzodiazepines. Lithium prolonged the period of remission, and the addition of clonazepam further prolonged it in case 1. Treatment with clonazepam reduced the symptoms in case 2, and when combined with lithium, the disorder went into remission after 6 months. These findings suggest that the combination of lithium and clonazepam may be effective in patients with cluster headache.

Effect of acute and chronic administration of dehydroepiandrosterone on (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane-induced wet dog shaking behavior in rats.

It has been reported that dehydroepiandrosterone (DHEA) or dehydroepiandrosterone sulfate (DHEA-S) is associated with affective disorders and that pathology of affective disorders are related with dysfunction of serotonin(5-HT)-2A receptor-mediated responses. In this study, we investigated the effect of DHEA on (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2 aminopropane (DOI), 5-HT-2A receptor agonist, -induced wet dog shaking behavior (WDS) in rats. Acute treatment with DHEA inhibited the DOI-induced WDSs dose dependently. This inhibition was recovered by opioid receptor antagonist, naltrexone. 5-HT-2A receptor-mediated WDSs were desensitized after chronic treatment with DOI, however chronic treatment with DHEA had no effect on this desensitization. Chronic treatment with DHEA had no facilitating effect of chronic dexamethasone treatment on DOI-induced WDSs. These findings may lead the possibility that DHEA has the inhibitory effect of 5-HT-2A mediated signaling pathway via non-genomic action.

Plasma dehydroepiandrosterone sulfate in unipolar major depression. Short communication.

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.

Positive influence of a liaison program on the rate of psychiatric consultation referrals for cancer patients.

To improve access to psychiatric consultation for cancer patients as well as non-cancer patients with psychiatric disorders, a psychiatric liaison programme to communicate closely with physicians and ward staff regarding anticipated psychiatric morbidity in patients, was introduced in each ward of a general hospital. The rate of psychiatric consultation referrals for cancer patients was significantly higher after the psychiatric liaison programme was established. The programme had a greater impact on the rate of psychiatric consultation in a unit with cancer patients who were informed of their diagnoses. The greater consultation rates in cancer patients after the liaison programme might be, in part, associated with the physicians' attitude toward the more open disclosure of the cancer diagnosis.

Differential regulation by pregnenolone sulfate of intracellular Ca2+ increase by amino acids in primary cultured rat cortical neurons.

We investigated the effects of pregnenolone sulfate (PS) on the [Ca2+]i increase induced by gamma-aminobutyric acid (GABA) and N-methyl-D-aspartate (NMDA) using fluorescence imaging. PS inhibited the 50 microM GABA-induced increase in [Ca2+]i in a dose-dependent manner with an IC50 of 30 microM. The inhibitory effect of PS was apparent within 5 min and was in a non-competitive manner, suggesting that PS may act directly to the membrane level but indirectly to the GABA binding sites. Our previous study has already shown that the GABA-induced Ca2+ increase involves GABAA receptors and the similar pathway to a high K(+)-induced Ca2+ response (Takebayashi et al., 1996). Because 50 microM of PS could not inhibit a 25 mM K(+)-induced Ca2+ increase, it seems likely that the site of the inhibitory action of PS on the GABA-induced Ca2+ increase may be independent of the pathway of the high K(+)-induced Ca2+ response, but rather at GABAA receptor complex. In contrast, PS potentiated the 50 microM NMDA-induced increase in [Ca2+]i in a dose-dependent manner. The magnitude of the NMDA response was approximately doubled in the presence of 100 microM of PS. However, PS did not affect the acetylcholine(Ach)-induced increase in [Ca2+]i. Furthermore, corticosterone had little effect on the GABA- and NMDA-induced Ca2+ increases, indicating that the alteration of the Ca2+ response is specific for PS. In conclusion, it is suggested that PS modulates differentially [Ca2+]i increase induced by GABA and NMDA.

Plasma levels of cyclic GMP, immune parameters and depressive status during interferon therapy. A prospective study in Japan.

In this report, we investigated the relationship between depressive symptoms and plasma interferon (IFN)-alpha-like immunoreactivity, cyclic GMP (cGMP) and soluble interleukin-2 receptor (sIL-2R) levels during IFN therapy. An altered mood state was observed in 5 of 26 patients. IFN-alpha-like immunoreactivity in the depressed group tended to be elevated. cGMP levels of depressed patients were significantly greater than those of control subjects before and 6 weeks after IFN therapy. However, sIL-2R levels were not different between the two groups. These results suggest that a number of patients suffered from depression during IFN therapy and that patients had greater concentrations of cGMP levels.

Effect of dantrolene on KCl- or NMDA-induced intracellular Ca2+ changes and spontaneous Ca2+ oscillation in cultured rat frontal cortical neurons.

Dantrolene has been known to affect intracellular Ca2+ concentration ([Ca2+]i) by inhibiting Ca2+ release from intracellular stores in cultured neurons. We were interested in examining this property of dantrolene in influencing the [Ca2+]i affected by the NMDA receptor ligands, KCl, L-type Ca2+ channel blocker nifedipine, and two other intracellular Ca2(+)-mobilizing agents caffeine and bradykinin. Effect of dantrolene on the spontaneous oscillation of [Ca2+]i was also examined. Dantrolene in microM concentrations dose-dependently inhibited the increase in [Ca2+]i elicited by NMDA and KCl. AP-5, MK-801 (NMDA antagonists), and nifedipine respectively reduced the NMDA and KCl-induced increase in [Ca2+]i. Dantrolene, added to the buffer solution together with the antagonists or nifedipine, caused a further reduction in [Ca2+]i to a degree similar to that seen with dantrolene alone inhibiting the increase in [Ca2+]i caused by NMDA or KCl. At 30 microM, dantrolene partially inhibited caffeine-induced increase in [Ca2+]i whereas it has no effect on the bradykinin-induced change in [Ca2+]i. The spontaneous oscillation of [Ca2+]i in frontal cortical neurons was reduced both in amplitude and in base line concentration in the presence of 10 microM dantrolene. Our results indicate that dantrolene's mobilizing effects on intracellular Ca2+ stores operate independently from the influxed Ca2+ and that a component of the apparent increase in [Ca2+]i elicited by NMDA or KCl represents a dantrolene-sensitive Ca2+ release from intracellular stores. Results also suggest that dantrolene does not affect the IP3-gated release of intracellular Ca2+ and that the spontaneous Ca2+ oscillation is, at least partially, under the control of Ca2+ mobilization from internal stores.

Design and synthesis of C-linked fucosides as inhibitors of E-selectin.

Two series of C-linked fucosides as mimetics for the tetrasaccharide sialyl Lewis X have been synthesized and tested as inhibitors of E-Selectin. The fucopeptides have been prepared from three key intermediates, including alpha-C-allyl fucose, natural and unnatural amino acids bearing hydroxyl groups and an alpha, omega-diacid moiety for the imitation of the essential three parts of SLex, i.e., the Fuc, Gal, and NeuAc. The nature and distance of the linkage of the fucose moiety to the amino acids as well as the distance between the amino acids and the terminal carboxylic acid group turned out to be crucial for the biological activity. In addition the necessity of both OH groups (4- and 6-OH) in the Gal part could be confirmed. Conformational NMR study of the most active mimetic supports the structure-activity relationship. A second series of mimetics was prepared, where Fuc and Gal moieties were purely C-linked. In the synthesis of beta-C-allyl galactose an intramolecular 1,2-hydride shift led to an interesting side product. However, the substituted glycosidic oxygens led to a substantial loss of conformational constrain, which could not be compensated and resulted in low activity.

Differential regulation of intracellular signaling systems by sodium fluoride in rat glioma cells.

We investigated the rapid and slow effects of NaF on intracellular signaling systems such as Ca2+ homeostasis and cyclic GMP (cGMP) generation in rat glioma C6 cells, using the Ca2+-sensitive dye fura-2 and cGMP enzyme immunoassay. We found that the following: (a) NaF enhanced cGMP generation in a concentration-dependent manner. This enhancement was abolished by pretreatment with 100 microM BAPTA tetraacetoxymethal ester or in the presence of W-7 in a concentration-dependent manner. N G-Monomethyl-L-arginine (NMMA), a competitive inhibitor or nitric oxide synthase (NOS), also inhibited the NaF-induced generation of cGMP. These results suggest that NaF-induced cGMP generation occurs via a calcium/calmodulin- and NOS-dependent pathway. (b) The basal intracellular Ca2+ concentration ([Ca2+]i) was transiently greater at 1 and 3 h after pretreatment with NaF. W-7 and W-13 antagonized the increase in [Ca2+]i, whereas NMMA had little effect. This suggests that the NaF-induced change in basal [Ca2+] was mediated by a calmodulin-dependent pathway but was independent of a NOS-sensitive pathway. (c) The serotonin (5-HT)-induced intracellular mobilization of Ca2+ was reduced by pretreating the cells with NaF. The reduction in Ca2+ mobilization was antagonized by genistein, a tyrosine kinase inhibitor. W-7, W-5, and H-8 had no effect. Results suggest that NaF differentially regulated the cGMP generation. basal [Ca2+]i, and 5-HT2A receptor function in C6 glioma cells.