Continuous stress promotes expression of VGF in melanotroph via suppression of dopamine.

Prolonged exposure to stress elicits profound effects on homeostasis that may lead to cryptogenic disorders such as chronic fatigue syndrome. To investigate the pathophysiology associated with the syndrome, we used a rat continuous stress (CS) model where the pituitary represents one of the most affected organs. Here we found that mRNA for VGF (non-acronymic), a member of the granin family, was induced specifically in the intermediate lobe (IL). This was matched by a concomitant increase at the peptide/protein level assessed by C-terminal antibody. Furthermore, the up-regulation of VGF was confirmed by immunohistochemistry in a subset of melanotrophs. VGF expression was altered in the IL of rats receivingthe dopamine D2 receptor agonist bromocriptine or the antagonist sulpiride. In vitro, dopamine dose-dependently decreased the mRNA levels in cultured melanotrophs. These findings suggest that VGF expression under CS is negatively regulated by dopaminergic neurons projecting from the hypothalamus.

Continuous stress-induced dopamine dysregulation augments PAP-I and PAP-II expression in melanotrophs of the pituitary gland.

Under continuous stress (CS) in rats, melanotrophs, the predominant cell-type in the intermediate lobe (IL) of the pituitary, are hyperactivated to secrete α-melanocyte-stimulating hormone and thereafter degenerate. Although these phenomena are drastic, the molecular mechanisms underlying the cellular changes are mostly unknown. In this study, we focused on the pancreatitis-associated protein (PAP) family members of the secretory lectins and characterized their expression in the IL of CS model rats because we had identified two members of this family as up-regulated genes in our previous microarray analysis. RT-PCR and histological studies demonstrated that prominent PAP-I and PAP-II expression was induced in melanotrophs in the early stages of CS, while another family member, PAP-III, was not expressed. We further examined the regulatory mechanisms of PAP-I and PAP-II expression and revealed that both were induced by the decreased dopamine levels in the IL under CS. Because the PAP family members are implicated in cell survival and proliferation, PAP-I and PAP-II secreted from melanotrophs may function to sustain homeostasis of the IL under CS conditions in an autocrine or a paracrine manner.

The increase of alpha-melanocyte-stimulating hormone in the plasma of chronic fatigue syndrome patients.

BACKGROUND: Despite extensive research, no reliable biological marker for chronic fatigue syndrome (CFS) has yet been identified. However, hyperactivation of melanotrophs in the pituitary gland and increased levels of plasma alpha-melanocyte-stimulating hormone (alpha-MSH) have recently been detected in an animal model of chronic stress. Because CFS is considered to be caused partly by chronic stress events, increased alpha-MSH plasma levels may also occur in CFS patients. We therefore examined alpha-MSH levels in CFS patients. METHODS: Fifty-five CFS patients, who were previously diagnosed within 10 years of with the disease, were enrolled in this study. Thirty healthy volunteers were studied as controls. Fasting bloods samples were collected in the morning and evaluated for their plasma levels of alpha-MSH, adrenocorticotropic hormone (ACTH), serum cortisol and dehydroepiandrosterone sulfate (DHEA-S). Mean levels of alpha-MSH were compared between the CFS and control groups using Welch's t test. RESULTS: The mean plasma alpha-MSH concentration in the CFS group (17.9 +/- 1.0 pg/mL) was significantly higher than that in healthy controls (14.5 +/- 1.0 pg/mL, p = 0.02). However, there was a wide range of values in the CFS group. The factors correlated with the plasma alpha-MSH values were analyzed using Spearman's rank correlation. A negative correlation was found between the duration of the CFS and the plasma alpha-MSH values (p = 0.04, rs = -0.28), but no correlations with ACTH, cortisol or DHEA-S levels were identified (p = 0.55, 0.26, 0.33, respectively). The CFS patients were divided into two groups: patients diagnosed for

Chronic stress elicits prolonged activation of alpha-MSH secretion and subsequent degeneration of melanotroph.

Prolonged stress affects homeostasis in various organs and induces stress-associated disorders. We examined the cellular changes of pituitary gland under the continuous stress condition using a rat model in which rats were kept in a cage filled with water to a height of 1.5 cm for up to 5 days. Among the pituitary hormone mRNAs, proopiomelanocortin mRNA was up-regulated specifically in the intermediate lobe (IL) of this rat model. Additionally, the peripheral blood levels of alpha-melanocyte stimulating hormone (alpha-MSH), a major product of proopiomelanocortin in IL were increased. The alpha-MSH secreting cells, melanotrophs, showed a markedly developed endoplasmic reticulum and Golgi apparatus in the early phase of the experiment. Subsequent continuous stress caused remarkable dilation of the endoplasmic reticulum, disruption of the Golgi structure, and the degeneration of some melanotrophs. In addition the dopaminergic nerve fibers from hypothalamus were markedly decreased in IL. A dopamine antagonist elicited the similar morphologic changes of melanotroph in normal rat. These findings suggest that prolonged stress suppressed hypothalamus-derived dopamine release in IL, which elicited over-secretion of alpha-MSH from the melanotrophs. The present study also suggests that prolonged hyperactivation of endocrine cells could lead to disorder of secretion mechanisms and eventual degeneration.

Neuronal injury-inducible gene is synergistically regulated by ATF3, c-Jun, and STAT3 through the interaction with Sp1 in damaged neurons.

Nerve injury requires the expression of large ensembles of genes. The key molecular mechanism for this gene transcription regulation in injured neurons is poorly understood. Among many nerve injury-inducible genes, the gene encoding damage-induced neuronal endopeptidase (DINE) showed most marked expression response to various kinds of nerve injuries in central and peripheral nervous system neurons. This unique feature led us to examine the promoter region of the DINE gene and clarify both the injury-responsive element within the promoter and its related transcriptional machinery. This study showed that DINE promoter was activated by leukemia inhibitory factor and nerve growth factor withdrawal, which were pivotal for the up-regulation of DINE mRNA after nerve injury. The injury-inducible transcription factors such as activating transcription factor 3 (ATF3), c-Jun, and STAT3, which were located at the downstream of leukemia inhibitory factor and nerve growth factor withdrawal, seemed to be involved in the activation of the DINE promoter. Surprisingly, these transcription factors did not bind to the DINE promoter directly. Instead, the general transcription factor, Sp1, bound to a GC box within the promoter. ATF3, c-Jun, and STAT3 interacted with Sp1 and are associated with the GC box region of the DINE gene in injured neurons. These findings suggested that Sp1 recruit ATF3, c-Jun, and STAT3 to obtain the requisite synergistic effect. Of these transcription factors, ATF3 may be the most critical, because ATF3 is specifically expressed after nerve injury.

Altered expression of neprilysin family members in the pituitary gland of sleep-disturbed rats, an animal model of severe fatigue.

Alterations of the expression of some peptidases in the pituitary gland of a fatigued rat model were identified. Rats were kept in a cage filled with water to a height of 1.5 cm to disturb deep sleep. After 24-h sleep disturbance, expression of neutral endopeptidase 24.11 (neprilysin) mRNA was increased in the intermediate lobe of the pituitary gland, whereas the mRNA expression of another family member, damage-induced neuronal endopeptidase, which is normally expressed in a subgroup of anterior pituitary cells, was significantly suppressed. These alterations were demonstrated by RT-PCR, northern blotting and in situ hybridization. Other family members, such as neprilysin 2 and endothelin converting enzyme-1, did not show any change in mRNA expression. An increase of neprilysin mRNA expression was not seen in any other tissues of the sleep-disturbed rats. The enzymatic activity of neprilysin was also increased in the pituitary. The augmentation of neprilysin expression and activity was prolonged as long as the sleep disturbance continued (up to 5 days), and returned to the basal level when rats were allowed to sleep freely. These results suggest that peptide processing and degradation in the pituitary may be an influential factor in fatigued states such as sleep disturbance.

Three-dimensional electron microscopic studies of the transitional oligodendrocyte associated with the initial stage of myelination in developing rat hippocampal fimbria.

We identified the transitional oligodendrocyte and their processes of rat hippocampal fimbria associated with the initial stage of myelination in both the morphological and functional classifications by means of three-dimensional ultrastructural analysis. Transitional oligodendrocytes appeared around P7, and their cell bodies were morphologically an intermediate form between the light and medium oligodendrocytes described by Mori and Leblond [J. Comp. Neurol. 139 (1970) 1]. Three phenotypes of the transitional oligodendrocytic processes were recognized. Spiral wrapping processes were ensheathing processes, club-like processes were nonensheathing processes, and sheet-like processes were possibly the transmuting form between the nonensheathing and ensheathing processes. Club-like processes were the major part of the nonensheathing processes, and most likely function as sensors to perceive axon maturation and find target axons. Multivesicular bodies that appeared to be associated with the initial ensheathment were observed in the transitional oligodendrocytic processes, suggesting that their roles are crucial in myelinogenesis.