Currently, our understanding of the regulation of the renin angiotensin system (RAS) during fetal life is limited. This system is crucial in the fetus, having been implicated in a diverse series of roles from the regulation of angiogenesis and kidney growth to modulation of cardiovascular and body fluid homeostasis under basal and stressful conditions. In the present application, we wish to test the unifying hypothesis that the renal nerves are essential for the increases in renin gene expression and activity in the RAS which occur over the last third of gestation and that these changes result, at least in part, from an increase in gene responsiveness to stimulation by the intracellular messenger cyclic adenosine monophosphate. To test this hypothesis, we propose to: a) measure renin gene responsiveness to stimulation at two stages of gestation; b) determine if the renal nerves modulate renin gene responsiveness to stimulation at two stages of gestation; c) determine if chronic activation of the RAS system will increase gene responsiveness to stimulation and if the renal nerves are essential for such an effect; and d) assess if the changes in renin gene responsiveness are associated with an increase in the number of cells which express and secrete renin in the fetal kidney, an increase in the amount of renin secreted per cell or both. Finally, we will correlate changes observed in vitro with results of in vivo studies on renin secretion in the fetus. We will use the fetal sheep for these experiments because we have defined the period in gestation when activation of the RAS occurs, and we can manipulate and study this model in utero. In addition, this model will provide adequate amounts of tissue for the in vitro studies with the use of relatively small numbers of animals. To accomplish our goal, we will employ RNase protection assays for the measurement of renin mRNA, nuclear run-on assays for transcription (i.e., polymerase loading) determinations and immunoblot techniques for studies of renin secretion from single cells. To our knowledge, these will be the first studies to examine the developmental changes in the fetal RAS from a molecular, cellular and whole animal basis simultaneously. Thus, the proposed studies will provide important new information on the role of the renal nerves in the developmental regulation of renin gene expression, the mechanisms involved in increasing activity in the RAS in late gestation and the mechanisms by which a chronic stimulus to renin secretion activates the RAS during development. Such information will be useful in understanding the adaptations to stress which occur in the perinatal period and may provide valuable clues about how alterations in the RAS occur in adults under certain pathological conditions.
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