Norihiko Murakami

Effects of interleukins on plasma arginine vasopressin and oxytocin levels in conscious, freely moving rats

To elucidate whether interleukins are involved in vasopressin or oxytocin release during cytokine-related stressful conditions, we examined the effects of human interleukin-1 beta and interleukin-6 on plasma vasopressin and oxytocin levels in rats. Interleukin-1 beta administrated intravenously stimulated both the vasopressin and oxytocin secretion in dose-dependent manners. Neither hormone release was observed following interleukin-6 administration. Pretreatment with aspirin significantly attenuated the effects of interleukin-1 beta on both the vasopressin and oxytocin levels. SC-19220, a prostaglandin E2 receptor antagonist, did not affect the interleukin-1 beta-induced increase of plasma oxytocin levels, but almost completely abolished its effect on plasma vasopressin levels. These results suggest that under certain stressful conditions which accompany the stimulation of cytokine production, interleukin-1 is involved in the increase of plasma vasopressin and oxytocin levels and, moreover, different kinds of prostaglandins are suggested to participate in these interleukin-1-induced hormone release.

Tumor necrosis factor receptors in the pituitary cells

To clarify the site and mode of action of tumor necrosis factor (TNF) in the pituitary, we studied the effects, binding sites of TNF and its receptor mRNA in the two types of mouse pituitary-derived cell lines, AtT-20, ACTH-producing cells and TtT/GF, folliculo-stellate (FS)-like cells. First, we examined the expression of TNF receptor mRNA in these cells. Using Northern blot analyses with radiolabeled cDNA to murine TNF receptor p60 and p80 mRNAs as probes, we identified both types of mRNA in the poly(A)-containing RNA prepared from AtT-20 cells and p60 TNF receptor mRNA from TtT/GF. The identified mRNA were compatible in size with those detected in the immune-competent cells. Next, we studied the TNF-binding sites on these cells. Scatchard plot analysis of the significant binding of [125I]TNF revealed a single type of binding site with a Kd (dissociation constant) of 210 pM and 131 binding sites/cell on AtT-20. Similarly on TtT/GF, [125I]TNF showed 353 binding sites/cell with a Kd of 900 pM. [125I]TNF binding on both types of cells competed with TNF and lymphotoxin (TNF beta) in an equimolar fashion. Third, TNF stimulates ACTH synthesis in AtT-20 cells, while TNF increases immunoreactive interleukin (IL)-6 release from TtT/GF cells. These findings demonstrate that AtT-20 and TtT/GF cells are equipped with fully functional TNF receptor system, and suggest that ligand of the receptor, TNF alpha and/or TNF beta, can modulate ACTH synthesis and release as a direct hormonal effector on corticotrophs or indirect modulator through another paracrine mediator, such as IL-6 from FS cells.

Adrenocorticotropic hormone-releasing activities of interleukins in a homologous invivo system

We compared adrenocorticotropin-releasing activities of several interleukins in a homologous or heterologous in vivo system. Intravenous injection of rat interleukin-1 alpha significantly increased plasma adrenocorticotropin in conscious, freely-moving rats 30 min after the injection, and the effect was 10 times greater than that of human interleukin-1 alpha. Rat interleukin-2 affected plasma adrenocorticotropin in a much slower manner and increased its levels significantly 120 min after the injection. Human interleukin-2 had no effect on plasma adrenocorticotropin. Thus, species difference in the experimental system should be considered to assess the physiological significance of cytokines in the neuroendocrine system.

Interleukin-1β analogues with markedly reduced pyrogenic activity can stimulate secretion of adrenocorticotropic hormone in rats

We examined the adrenocorticotropic hormone-releasing activities of several human interleukin-1 beta analogues that have markedly reduced pyrogenic activities in rats. Among the analogues tested, [Gly4]-, [Leu93]- and [1-148]-interleukin-1 beta increased the plasma adrenocorticotropic hormone level to almost that induced by authentic human interleukin-1 beta. Modifications of the N-terminus of the authentic molecule, i.e., [7-153]- and [Des-Ala1, Asp4]-interleukin-1 beta, significantly reduced the hormone-releasing activity. These data suggest that the adrenocorticotropic hormone-releasing activity of human interleukin-1 beta resides in the N-terminal structure of the authentic peptide and can be separated from its pyrogenic activity.

Regulation of interleukin-1 receptors on AtT-20 mouse pituitary tumour cells

To study the cellular mechanisms of interleukin‐I (IL‐1) in the pituitary corticotroph, we studied the properties of IL‐1 receptors on a mouse pituitary ACTH‐producing cell line, AtT‐20. Scatchard plot analysis revealed a single type of receptor with a K d (dissociation constant) of 93 pM, and 482 binding sites/cell. [125I]IL‐1α binding competed with IL‐1α and IL‐1β in an equimolar fashion. A 24 h pre‐incubation with either CRH, epinephrine or nor‐epinephrine increased the [125I]IL‐1α binding sites in the AtT‐20 cells and conversely, a similar pre‐incubation with either dexamethasone or tumour necrosis factor‐α (TNFα) decreased them without affecting the affinity of the receptors in either case.

Immunoreactive corticotropin-releasing hormone levels in the hypothalamus of female Wistar fatty rats.

We have studied immunoreactive corticotropin-releasing hormone (CRH) levels in the hypothalamus of female Wistar fatty rats, a strain with the fa gene transferred from the Zucker rat to the Wistar Kyoto rat, in an attempt to understand the role of CRH in the development of obesity. A study was conducted with 5-week- and 12-week-old female Wistar fatty rats and lean littermates. There was no significant difference in hypothalamic CRH levels between lean and obese rats at the age of 5 weeks (1887 +/- 99.6 vs. 1767 +/- 124 pg/tissue; mean +/- S.E.M.). Hypothalamic CRH immunoreactivities, however, were significantly lower in 12-week-old obese rats (2361 +/- 132 pg/tissue) than those in lean littermates (2992 +/- 118 pg/tissue; P less than 0.05). The difference of CRH contents between the lean and obese group becomes apparent as they grow up and develop obesity.

In Vivo and in Vitro Methods for Studying Effects of Cytokines on Adrenocorticotropic Hormone, Arginine Vasopressin, and Oxytocin Secretion

Publisher Summary Cytokines, which were originally isolated from lymphocytes and monocytes and identified as immunomediators, have a broad spectrum of actions as factors involved in various kinds of cell-to-cell communication. Among the cytokines identified so far, interleukin lα (IL-lα), interleukin lβ (IL-1β), interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), and interferons have been reported to be major cytokines induced during the early inflammatory processes and affecting various neuroendocrine functions, especially the stimulation of activities in the hypothalamic-pituitary-adrenal (HPA) axis. These cytokines are, therefore, presumed to be the mediators of acute-phase responses to infectious challenge. In in vivo experiments, a cascade of events induced by cytokines must be carefully considered because neuroendocrine functions may be modified as a second effect. Data acquired by in vitro (artificial) systems should also be assessed carefully as to their physiological significance. This chapter describes the in vivo and in vitro systems that have been used to the study of the effects of several cytokines on pituitary hormone secretion, especially that of adrenocorticotropic hormone (ACTH), arginine vasopressin (AVP), and oxytocin (OT), and also to present some data obtained using these methods as examples.

Drug delivery system. Therapeutic application of hypothalamic hypophysiotropic hormones.

Therapeutic applications of hypothalamic hypophysiotropic hormones were discussed. Pulsatile administration of gonadotropin-releasing hormone (Gn RH) is effective to maintain menstrual cycles and ovulation in female patients with hypothalamic gonadal dysfunction and testicular function in male patients. On the other hand, continuous administration of Gn-RH or its analog is able to suppress gonadal function and thereby gonadal hormonedependent activities effectively. Application of this medical gonadectomy includes treatments for excessive menstrual bleeding, endometriosis, prostatic cancer and benign prostatic hypertrophy. Thyrotropin-releasing hormone has been applied in patients with spinocerebellar degeneration or some kinds of consciousness disturbance and somatostatin analog has benn tried to treat acromegaly. Corticotropin-releasing hormone and growth hormone-releasing hormone have also therapeutic potentials. We believe that many-sided research trials, which include development of new potent analogs, new effective drug delivery systems and further elucidation of physiology and pathophysiology about the functions of the hypothalamic hormones, are inevitable to materialize therapeutic potentials of these hormones.