Ontogenetic peculiarities of regulation of apoptosis of hypothalamic neurosecretory cells in TNF-knockout mice

Tumor necrosis factor (TNF) participates in regulation of many processes, including carcinogenesis and apoptosis. However, at present, there are practically no studies on peculiarities of apoptosis control in tnf-knockout (tnf-/-) mice. These mice develop without morphologic abnormalities, but they seem to have impairment of many biological processes, such as inflammation, programmed cell death, etc. Therefore, the goal of our work was to study possible pathways of regulation of apoptosis in the absence of TNF in neurosecretory cells (NSC) of young and old mice. For this purpose, we determined immunohistochemically expression of apoptosis markers caspase-8, caspase-9, Bax, Bcl-2, Mcl-1, neuropeptide vasopressin and the apoptosis level in hypothalamus of tnf-knockout mice of different ages as compared with mice with unchanged level of TNF synthesis. It was shown that the apoptosis activation observed during aging did not depend on the tnf gene and that apoptosis at aging was caspase-dependent. It was revealed that at aging in mouse NSC the external cell death pathway with participation of caspase-8 is activated. The pathways mediating cell death in different neurosecretory centers at aging are different. Thus, in supraoptic nucleus (SON), in all studied animal groups, an important cause of the NSC apoptosis is Bax. In paraventricular nucleus (PVN), of the greater importance is a decrease of the anti-apoptotic protection. Hence, misbalance of synthesis of proteins of the Bcl-2 family plays an important role in development of senescent apoptosis.     

Ultrastructure of neurosecretory cells of the hypothalamic supraoptic nucleus in white rats following painful stress and during aging

The ultrastructural organization of neurosecretory cells (NSC) belonging to the hypothalamic supraoptic nucleus was investigated in young rats following periods of painful stress differing in duration (of 2 and 20 min). Intact young rats of similar age and others at late stages of development (24–27 months old) served as controls. Short- and longer-acting painful stress was found to intensify and inhibit the functional activity of NSC respectively. Complex ultrastructural changes in the NSC of young rats following protracted painful stress is compared with the ultrastructural organizational pattern in several NSC of aging rats. Findings would imply that aging occurs in rats following prolonged painful stress, resulting from functional hypersecretion and depletion.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 21, No. 3, pp. 389–395, May–June, 1989.     

Antidromic indentification of hypothalamic neurosecretory cells in rats

Characteristics of antidromic action potentials of neurons of the paraventricular and supraoptic nuclei of the rat hypothalamus were studied during stimulation of the hypothalamo-hypophyseal tract by stimuli of varied amplitude and frequency. Step-like changes were found in spike latency in response to an increase in strength (up to 1.5–2.5 thresholds) or frequency (over 100 Hz) of stimulation, as well as cases with variation of the degree of division of the peak into A and B components. Injection of leu-enkephalin analog into the third ventricle or intravenous injection of NaCl solution (1 M) caused reversible changes in the level of excitability of antidromically activated neurons: leu-enkephalin mainly increased the latent period and threshold of action potential generation and reduced the reproducible frequency of stimulation to 10 Hz, whereas NaCl had the opposite effect. The results indicate that when the adopted criteria of antidromic identification of neurosecretory cells are used the level of their excitability must be taken into account.A. A. Ukhtomskii Physiological Research Institute, A. A. Zhdanov Leningrad State University. Translated from Neirofiziologiya, Vol. 14, No. 6, pp. 585–591, November–December, 1982.     

Ultrastructural changes in rat hypothalamic neurosecretory cells and their associated glia during minimal dehydration and rehydration

Summary A quantitative ultrastructural study was performed to determine the changes in the neurosecretory neurons of the supraoptic (SON) and circularis (NC) nuclei following 4–24 h of water deprivation (WD) and subsequent rehydration (12 and 24 h). In both nuclei, the amount of direct soma-somatic contact increased throughout WD, apparently by retraction of fine glial processes from between the cells. Rehydration reversed these changes. The number of smaller (<1600 Å) neurosecretory granules (NSG's) decreased in both nuclei at 4 h of WD but returned to control levels by 24 h of WD and remained so during rehydration. Larger (<1600 Å) NSG's decreased in number at 4 h of WD in SON and then returned to control levels by 24 h of WD and remained the same throughout rehydration. In NC, these NSG's did not change in number with WD, but significantly increased between 12 and 24 h of rehydration. No cells with dilated rough endoplasmic reticulum were seen in NC during this study. In SON, however, the percentage of such cells increased at 4 and 12 h of dehydration only to decrease to control levels at 24 h of dehydration and throughout rehydration. Lysosomes decreased at 4 h of dehydration in SON and returned to control levels thereafter. In NC, lysosomes tended to decrease with dehydration and increase with rehydration. These findings indicate that detectable morphological changes take place in the course of alterations in hydration state that are well within the physiological range.Supported by NIH Grant NS 09140. The use of the electron microscope facility of the College of Osteopathic Medicine is gratefully acknowledged. Thanks are due W.E. Armstrong and W.A. Gregory for helpful comments, and R. Meyers, A. Ridener, and R. Herbold for technical assistance     

Plasticity in the hypothalamic magnocellular neurosecretory system

Over the past decade or so, plasticity has emerged as an important, quantifiable property of the mammalian hypothalamic magnocellular neurosecretory system. This plasticity has turned out to be genuinely related to normal function in the sense that it is a set of responses to physiological stimulation rather than only the sequelae of insult or injury, and it is generally completely reversible. This latter property, of course, distinguishes it further from the plasticity observed after injury. Four features of this magnocellular system that have been shown to display predictable and reversible intercellular plasticity are reviewed: the relationships between neurons and their associated astrocytic glia at various levels (dendritic somatic and terminal) of the magnocellular elements; the extent of terminal and glial contact with the basement lamina in the neurohypophysis; the type and possible efficacy of synaptic input, and the extent of electrotonic coupling among the magnocellular neurons.     

Regulation of brain adrenergic receptors during aging

Recent studies in our laboratory suggest that the synthesis of alpha- and beta-adrenergic receptors in certain tissues from brain and pineal gland may be impaired with age. This decreased ability of the aged brain to synthesize adrenergic receptors may explain the loss of these receptors in selected brain regions during the aging process, as well as the reduced capacity of aged brain tissue to increase or up-regulate the density of these receptors in response to reduced noradrenergic activation of the tissues or to reduced estrogen levels. The reduced adaptability of brain adrenergic receptors, in turn, may account for the decreased ability of aged individuals to adjust their physiological responses to a changing environment.     

Regulation of prolactin in mice with altered hypothalamic melanocortin activity

This study used two mouse models with genetic manipulation of the melanocortin system to investigate prolactin regulation. Mice with overexpression of the melanocortin receptor (MC-R) agonist, α-melanocyte-stimulating hormone (Tg-MSH) or deletion of the MC-R antagonist agouti-related protein (AgRP KO) were studied. Male Tg-MSH mice had lower blood prolactin levels at baseline (2.9±0.3 vs. 4.7±0.7ng/ml) and after restraint stress (68±6.5 vs. 117±22ng/ml) vs. WT (p<0.05); however, pituitary prolactin content was not different. Blood prolactin was also decreased in male AgRP KO mice at baseline (4.2±0.5 vs. 7.6±1.3ng/ml) and after stress (60±4.5 vs. 86.1±5.7ng/ml) vs. WT (p<0.001). Pituitary prolactin content was lower in male AgRP KO mice (4.3±0.3 vs. 6.7±0.5μg/pituitary, p<0.001) vs. WT. No differences in blood or pituitary prolactin levels were observed in female AgRP KO mice vs. WT. Hypothalamic dopamine activity was assessed as the potential mechanism responsible for changes in prolactin levels. Hypothalamic tyrosine hydroxylase mRNA was measured in both genetic models vs. WT mice and hypothalamic dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) content were measured in male AgRP KO and WT mice but neither were significantly different. However, these results do not preclude changes in dopamine activity as dopamine turnover was not directly investigated. This is the first study to show that baseline and stress-induced prolactin release and pituitary prolactin content are reduced in mice with genetic alterations of the melanocortin system and suggests that changes in hypothalamic melanocortin activity may be reflected in measurements of serum prolactin levels.     

Regulation of apoptosis in mast cells

Apoptosis is a physiological process of cell death that occurs in all multicellular organisms. Its dysregulation has been postulated as one of the main causes in the development of diseases such as cancer, AIDS, autoimmune diseases and allergy. Apoptosis has been mainly studied in the inflammatory cells that participate in the late and chronic stages of allergy (eosinophils, neutrophils, lymphocytes and macrophages) as a new way to elucidate the pathogenesis of this disease. Nevertheless, much less it is known about the regulation of apoptosis in the initiators of the allergic process: The Mast Cells. In normal conditions, mast cells are described as long-living cells that keep a constant number of cells in tissues. However, increased numbers of mast cells are observed in the late phase of asthma and in both the inflammatory and in the repair/remodeling stage of various inflammatory/fibrotic disorders. In this report, we discuss the possible mechanisms that regulate the apoptotic process in normal conditions and disease, such as survival factors and death receptors. A link between mast cell activation, during the early stages of the allergic process, and triggering of anti-apoptotic signaling pathways is also suggested as an important contributor to the extended life of mast cells.     

The hypothalamic magnocellular neurosecretory system in developing rats

Studies concerning the development of the magnocellular system are scarce and discordant in literature. We carried out an immunohistochemical study on supraotic and paraventricular hypothalamic nuclei using antivasopressin and antioxytocin antibodies in developing rats between the 15th day of intrauterine life and the 6th day of postnatal life. In addition, we performed RT-PCR experiments to establish the stage at which these hormones appear and neurosecretory activity commences. The results showed that supraoptic and paraventricular nuclei appear, respectively, on the 16th and the 18th day of intrauterine life and both immediately synthetize vasopressin neurohormone. By contrast, synthesis of oxytocin takes place from the 2nd day after birth. Probably, these nuclei synthetize oxytocin in conjunction with the decline of placental maternal oxytocin.     

Ultrastructure of the hypothalamic neurosecretory system of the dog

Summary The hypothalamic neurosecretory system of normal dogs was studied by light and electron microscopy after perfusion-fixation. In the supraoptic nucleus most neurons are loaded with elementary neurosecretory granules having a content of low electron density. Neurons with less neurosecretory material and signs of enhanced synthetic activity, as recognized by the changes in the endoplasmic reticulum, were also observed.The vesiculated neurons ofJewell were studied under the electron microscope and various stages of development were described. It was postulated that they originate by a localized process of cytoplasmic cytolysis which ends in the formation of a large aqueous intracellular cavity limited by a plasma membrane. The possible significance of these vesiculated neurones is discussed. Some few myelinated neurosecretory axons are found in the supraoptic nucleus.The neurons of the paraventricular nucleus are smaller and contain less neurosecretory material. This is abundant and very pale in the axons. The median eminence consists of an inner zone, mainly occupied by the neurosecretory axons of the hypothalamic-neurohypophysial tracts, and an outer zone in which some neurosecretory axons end on the capillary of the portal system. This outer zone contains numerous axons with the axoplasm rich in neurofilaments and some containing granulated and non-granulated synaptic vesicles. Some neurons with granulated vesicles were observed in this region. The adrenergic nature of these neurons and axons is postulated.The infundibular process of the neurohypophysis shows small axons with discrete amounts of elementary granules and vesicles of synaptic type at the endings. Some enlarged axons having, in addition, large polymorphic bodies are observed and related to the Herring bodies.The size and morphology of the granules are analyzed along the entire hypothalamic-neurohypophysial system. The changes in diameter and electron density are related to the maturation of the granules and the possible significance of such evolution.Supported by grants from the Consejo Nacional de Investigaciones Cientificas y Técnicas and by the Air Force Office of Scientific Research (AF-AFOSR 963-66).     

Regulation of Leydig cell steroidogenic function during aging

This article summarizes a talk on Leydig cell aging presented at the 1999 Annual Meeting of the Society for the Study of Reproduction. In the Brown Norway rat, serum testosterone levels decrease with aging, accompanied by increases in serum FSH. The capacity of Leydig cells to produce testosterone is higher in young than in old rats. Binding studies with hCG revealed reduced receptor number in old vs. young Leydig cells. In response to incubation with LH, cAMP production was found to be reduced in old vs. young Leydig cells, indicating that signal transduction mechanisms in the old cells are affected by aging. Steroidogenic acute regulatory protein and mRNA levels are reduced in old Leydig cells, suggesting that there may be deficits in the transport of cholesterol to the inner mitochondrial membrane of aged cells. The activity of P450 side-chain cleavage enzyme is reduced in old vs. young cells, as are the activities of each of 3beta-hydroxysteroid dehydrogenase, 17alpha-hydroxylase/C17-20 lyase, and 17-ketosteroid reductase. Serum LH levels do not differ between young and old rats, and the administration of LH failed to induce old Leydig cells to produce high (young) testosterone levels, suggesting that the cause of age-related reductions in steroidogenesis is not LH deficits. We hypothesized that reactive oxygen, produced as a by-product of steroidogenesis itself, might be responsible for age-related reductions in testosterone production by the Leydig cells. Consistent with this, long-term suppression of steroidogenesis was found to prevent or delay the reduced steroidogenesis that accompanies Leydig cell aging. A possible explanation of this finding is that long-term suppression of steroidogenesis prevents free radical damage to the cells by suppressing the production of the reactive oxygen species that are a by-product of steroidogenesis itself.