: Endothelin-1 (ET-1) and angiotensin II (AngII) are critical hormones necessary for the appropriate function of the cardiovascular system, acting centrally as well as peripherally. Both hormonal systems have been implicated in cardiovascular pathology -- hypertension, endothelial dysfunction, cardiac hypertrophy and failure. Several isoreceptors for both ET-1 and AngII have been recently characterized. Of these, the dual ET-1/AngII receptor, isolated by us via an approach based on and supporting the molecular recognition theory elucidates novel information providing the bases for new hypotheses and questions. The long-term objective of this proposal is to dissect the specific contribution of the ET-l/AngII receptor gene to blood pressure regulation in normal and pathophysiological states like hypertension. In this research proposal, we will test the following hypotheses: 1) Based on our preliminary data showing a) gender-bias o 1I the cosegregation of the ET-II /AngII receptor locus with hypertension in an F2 (Dahl S x Dahl R) intercross, and b) structural differences between the Dahl salt-sensitive and the Dahl salt-resistant rat ET-1/AngII receptors, we hypothesize that the ET-1/AngII receptor acts in a gender specific manner influencing blood pressure in the Dahl rat model. 2) Based on the detected abundance of the ET-l/AngII receptor mRNA in brain and cardiovascular tissues, we hypothesize that this receptor is a prominent physiologic target for ET- 1 and AngII in these organs, and therefore plays an important role in blood pressure regulation. Investigating the following specific aims will test these hypotheses: 1. Gender specific role of the ET-1/AngII receptor in susceptibility to salt-sensitive hypertension. 1A) Functional characterization of the Dahl S and Dahl R ET-1/AngII receptors that differ in two amino acids. 1B) Establishment of three inbred trans genie Dahl S rat lines bearing exogenous Dahl R ET-1/AngII receptor, Tg[RET-1/AngIIr], and three control Dahl S rat lines bearing exogenous Dahl S ET- 1/AngIl receptor, Tg[SET- 1/AnglIr], driven by the cognate Dahl R 5' flanking regulatory region. 1C) Determine the transgenic molecular phenotype, that is the relative amounts of exogenous (transgenic) Dahl R and Dahl S ET- 1/Angll receptor mRNA and endogenous Dahl S ET-1/AngIl receptor mRNA expression levels in Tg[RET-1/AngIIr] and Tg[SET-1/AngIIr} rats respectively. 1D) Determine the physiological effects of the Dahl R and Dahl S ET- 1/AngII receptor transgene by analyzing differences between age-matched non-transgenic inbred Dahl S rats (males and females) and transgenic inbred Tg[RET- 1/AnglIr] and Tg[SET1I AngIIr] Dahl S rat lines with respect to 1) the onset, course and degree of hypertension; 2) lifespan; 3) severity of hypertensive renal disease. 2. Dissection of physiologic role(s) of the ET-1/AngII receptor. 2A) Tissue specific, spatial and temporal expression patterns in development of the ET-1/AngII receptor gene. 2B) Targeted disruption of the ET- 1/AngIl receptor gene and development of """"""""knockout"""""""" mouse model in order to define its physiologic role in an integrated biologic experimental system. Our proposed studies will elucidate the physiological role of the ET- 1 /AngII receptor throughout development. Furthermore, it will demonstrate the gender specific involvement of the ET- 1/AngII receptor in salt-sensitive hypertension in the Dahl rat model, thus providing a validated prioritization scheme to investigate the potential role of this receptor in human essential hypertension.
