Ontogenetische Veränderung des Plasmaleptinspiegels und seine Regulation bei 10 Tage alten Ratten
Abstract
The investigations of this study were concerned with the ontogenetic changes in plasma concentration of the hormone leptin in suckling-age rats and its physiological regulations with special reference to the so-called "fa" gene defect of the leptin receptor. Analytical studies proceeded from the evaluation of the time course of plasma leptin and its relationship with body fat content during the developmental period between the first and fifth week of life, concentrating in particular on the phase around the 10th day of life. Based on these studies the influence of the "gene dose", i.e., the homozygous (fa/fa), and heterozygous (+/fa) presence of the gene defect or its total absence in the wildtype animal (+/+) on plasma leptin was analyzed at two defined states of ontogeny. First, the influence of gene dose on plasma leptin was determined during the perinatal phase. Second, the roles of gene dose and of several physiological and cellular parameters known to influence plasma leptin were studied at the age of 10 days, because preceding observations had indicated that the gene dose would become effective at this age. In order to delineate the influence of gene dose against those of various metabolic parameters, control of food supply was established by artificial rearing and control of metabolic expenditure either by imposing a mild cold load or by norepinephrine infusion to activate non-shivering thermogenesis. Measurements included plasma leptin and the mRNA concentrations in the brown (BAT) and white (WAT) adipose tissue, metabolic rate (MR) as an estimate of the state of sympatho-adrenal activation (SAA), net energy storage, determined as the difference between the energy supplied with the milk and metabolic energy dissipation, as an estimate of the global energy flux, and body fat content as an estimate of the global lipid content of the adipocytes. Evaluation of the data by subjecting them to 2-way ANOVA and to simple or multiple regression analysis produced the following results. 1. The earliest evidence for an influence of the gene dose on plasma leptin was found immediately after birth, i.e., prior to the start of triglyceride storage, but could not be discovered immediately before birth. 2. The plasma concentration of leptin in wildtype animals declines during the first to fifth week of life. The decline is most rapid during the first 2 weeks and varies more strongly during this period than subsequently. Rapid sustained changes that might have compromised the measurements made in 10 day old pups did not occur at that age. 3. Artificial rearing under moderate cold load conditions does not eliminate the difference between the high plasma levels of the fa/fa animals in comparison to +/fa and +/+ animals, as they are observed under natural rearing conditions. 4. Artificial rearing for 2 days in thermoneutral conditions leads to comparable minimum energy consumption without changes in body fat storage in both fa/fa and +/fa pups and eliminates the stimulatory effects responsible for the high plasma leptin levels in the fa/fa animals. 5. Under continuous application of norepinephrine for 6 days in artificially reared pups, low doses are sufficient to adjust plasma leptin of the fa/fa pups to the level of +/fa pups, and reduction of body fat content is not a prerequisite for this effect. 6. For plasma leptin levels <10 ng/ml, irrespective of raising the animals naturally or artificially, a strong influence of leptin mRNA concentration in the BAT may be stated which explains 60% of plasma leptin variations, while leptin mRNA concentration in the WAT accounts for only 10% of the plasma leptin variations. In addition, a weak genotype effect could be confirmed. 7. For plasma leptin levels >10 ng/ml, which were observed mainly in fa/fa pups, body fat content was confirmed to be influential and accounted for 50% of the variations in plasma leptin. The influence of gene dose was neither associated with differences in BAT and Wat leptin mRNA concentrations nor with the state of sympathoadrenal activation. 8. Analysis applied to the control of plasma leptin at levels <10 ng/ml, when leaving out of account the influences of leptin mRNA concentrations in the BAT and WAT, revealed a positive relationship with body fat content accounting for 50% of plasma leptin variations and a negative relationship with the state of sympathoadrenal activation accounting for 10% of the plasma leptin variations, while gene-dose influences could not be detected. 9. Leptin mRNA concentrations in the BAT correlated closely with the state of sympathoadrenal activation which accounted for 90% of their variations. Only 50% of the variations in WAT leptin mRNA concentration could be attributed to any one of the investigated variables, with net energy supply emerging as the dominant parameter and both body fat content and state of sympathoadrenal activity being less influential. The difference in the control of leptin mRNA concentration in BAT and WAT may be related to the facts that net energy supply is loosely connected to energy flux in the WAT whereas MR as the estimate for the sympathoadrenal state of activity is closely connected to energy flux in the BAT. The results suggest the following conclusions: (a) Immediately after birth the "fa" gene defect is effective as a factor elevating plasma leptin concentration in rat pups. (b) In 10-day-old rat pups autocrine control of leptin production, due to the action of the circulating hormone on leptin receptors of the fat cell as the source of leptin, is not yet effective. Since a feedback relationship between leptin and its production was shown to exist in the BAT at this age, it is most likely established by central nervous leptin receptors ultimately controlling sympathoadrenal activity. For leptin production in the WAT, the available evidence suggests some kind of feedback control of leptin mRNA expression in which local energy flux seems to be involved. c) Apart from the feedback relationships identified in this study between plasma leptin and leptin mRNA expression, circumstantial evidence suggests the involvement of additional unknown control mechanisms which are effective in the conditions of natural as well as artificial rearing involving some degree of cold stress, but are eliminated in thermoneutral conditions.
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