Randall R. Sakai - US grants

DESCRIPTION: The PI was among the first to demonstrate that sodium appetite is normally caused by a synergistic action of the peptide, angiotensin-II (A-II) and the steroid, aldosterone (ALDO), and he has also been instrumental in beginning to elucidate the receptor mechanisms involved. Because of the rapid elicitation of sodium appetite that can be observed by ALDO in some situations, the PI now proposes several experiments to examine both the genomic and the non-genomic effects of ALDO, to determine specific brain sites where adrenal steroids work to cause sodium appetite, and to interfere with the expression of endogenous steroid receptors in the brain by use of molecular biological techniques and determine the effect upon sodium appetite. In addition to a large behavioral and physiological base, the present research is based on the observation that many brain regions which contain corticosteroid receptors also contain enzymes that reduce the A- ring of steroids, thereby creating molecules which are thought to interact with membrane-bound receptors (possibly the GABAa- benzodiazepine receptor) and hence elicit rapid responses. These are called the non-genomic effects because they do not require the formation of new gene products in order to work. In pilot work, the PI has found that A-ring-reduced metabolites of both ALDO and deoxycorticosterone (DOC, another mineralocorticoid) both elicit rapid sodium appetite in rats. In proposed work, the PI will identify specific brain areas where these metabolites are active, compare their activity to that caused by the parent hormone, and correlate the findings with measurements of activity of the enzymes in the same areas. Although several areas will be investigated, the amygdala is hypothesized to be particularly important since ablation there decreases mineralocorticoid-elicited sodium appetite. Other areas include the medial preoptic area and the paraventricular nuclei. The other series of proposed experiments will investigate genomic control over salt appetite by corticosteroids by using intracranially applied antisense oligomers to the mineralocorticoid (Type I) and glucocorticoid (Type II) receptors. An additional series of experiments will relate the genomic and the non-genomic effects of ALDO. Several models of eliciting sodium appetite will be used in these experiments, and extensive controls are proposed.

[unreadable] DESCRIPTION (provided by applicant): The regulation of food intake and body weight is a complex interaction of multiple factors. Among these factors is the social milieu in which the animal lives and in particular the social stress that accompanies dominance hierarchies. We utilize a unique animal model of social stress to study how such social hierarchies influence the regulation of food intake and body weight and additionally to uncover the neuroendocrine mechanisms that underlie social influences on body weight. In particular, we found that when animals are placed into a visible burrow system (VBS), we can identify the subordinate (SUB) and dominant (DOM) animals and that SUB animals have reduced body weight when they are in the burrow. The focus of this proposal is to test several hypotheses concerning the mechanism by which social stress in subordinate animals alters the fundamental neuroendocrine circuits involved in the regulation of food intake and body weight. The proposed experiments are critical to a complete understanding of how food intake and body weight are regulated. Importantly, the proposed model offers a novel approach for exploring the complicated but vital relationship between the stress/HPA system and body weight regulation. Specific Aims are: 1) To determine whether the weight loss that is observed in SUB is a result of reduced energy intake and/or increased energy expenditure. In addition, we will determine several endocrine and neurochemical endpoints that are altered by negative energy balance. With several novel control groups for comparison (dominant rats, ad lib fed, pair-fed and body weight-matched), we will be able to determine which major systems interact to produce the observed body weight changes. 2) To determine whether the reduced food intake and/or increased energy expenditure of SUB is secondary to a homeostatic down regulation of the defended amount of body fat, or to a direct catabolic action of stress hormones and neurotransmitters. 3) To test the hypothesis that multiple cycles of chronic stress in the VBS and recovery will produce more severe and enduring changes in body weight regulatory systems than exposure to a single episode. The health significance of this research is clear. Determining how CNS mechanisms involved in the regulation of energy balance are altered by stress is critical to a complete understanding of the co-morbidity of stress related disease states and obesity. Interestingly both stress and food intake regulation share common neurochemical systems and this proposal attempts to define both the interrelatedness as well as the independence of the mechanisms that govern each system. The execution of this proposal will provide insight into the connections between different levels of this regulatory system and thereby give direction to how multiple interventions might best be used to prevent or treat obesity and associated conditions resulting from stress. [unreadable] [unreadable] [unreadable]

Cloning animals by nuclear transfer of somatic cells is a scientifically important topic that is also politically charged. It is therefore imperative for rigorous experimentation to inform both the scientist and the public at large, and that the work be conducted with the highest standards of care, control and understanding. The goal of this proposal is to investigate an undesirable side effect of the cloning process, obesity, using a mouse model. We have found that the obese phenotype is maintained over successive generations of cloned mice;i.e. clones derived from clones, but it is not passed through the germ line to naturally derived offspring, suggesting that the obese phenotype of cloned mice is an epigenetic rather than a genetic phenomenon. Proposed experiments will begin to explore possible mechanisms for the obesity in cloned mice. Specific Aims are: 1) To determine whether obesity in cloned mice is the result of hyperphagia or differences in metabolic rate during early postnatal development. We hypothesize that cloned mice are hyperphagic and/or have lower energy expenditure and metabolic rate early in development relative to controls;2) To test the hypothesis that obese clones defend their elevated body weight comparably to other obese animals, and to identify components of the neuroendocrine control system of energy homeostasis that are altered in clones;and 3) To test the hypothesis that increased body weight and obesity in cloned (and in vitro-manipulated control mice to a lesser extent) result from in vitro embryo production and/or in vitro mechanical manipulation. Collectively, these experiments will provide detailed information about clones as well as the long-term effects of the process used to generate the clones. To accomplish this, we have put together a team of authorities on the cloning process itself (Yanagimachi and Yamazaki), on obesity (Woods), and on assessment of the requisite behavioral, physiological, endocrine and neurobiological parameters (Sakai). Investigators at the University of Hawaii (UH) who pioneered the nuclear transfer technique will generate the cloned mice as well as the control animal groups. The group at the University of Cincinnati (UC) will conduct behavioral, physiological and neurochemical studies to phenotype the cloned mice and their controls. The proposed research will provide the first set of longitudinal studies in cloned mice that examine the long-term consequences of somatic cell cloning. In addition, the behavioral, physiological, and neurochemical data obtained will enhance our understanding of the mechanisms of obesity, a serious and growing chronic public health issue worldwide.