Activation of the mineralocorticoid receptor (MR) is implicated in clinical hypertension and target organ damage even in patients who have normal or low plasma aldosterone levels. MR antagonists mitigate chronic heart failure in patients who do not have significant elevations in aldo. MR are abundantly expressed in neurons in specific brain areas and in the heart, however the enzymes that confer MR selectivity for aldo over the more abundant glucocorticoids with similar MR affinity, are limited or absent in the heart and most of the brain. We have shown by selective intracerebroventricular infusions of agonists and antagonists that excessive circulating levels of aldo act through MR of the brain to elevate the blood pressure. Inhibition of MR in brains of Dahl Salt Sensitive hypertensive (SS) rats also mitigates their hypertension, though plasma aldo concentrations in this rat are low or normal. We have shown that the enzyme aldo synthase (AS) is expressed in specific neurons in the normal human and rat and that low levels of aldo are synthesized in the rat brain. We hypothesize that local production of aldo acts in a paracrine or autocrine fashion and that excessive aldosterone synthesis in the brain leads to hypertension and end organ pathology. MR and AS are expressed in many parts of the normal human and rat brain, only a few of which are involved in hemodynamic, fluid and electrolyte homeostasis and neurally modulated inflammatory processes. In fact, the highest concentrations of MR are in hippocampal neurons involved in memory, learning and affect that also express AS. It is important to identify the neurons involved in the hemodynamic control that have steroidogenic capacity to develop prevention and treatment strategies that do not target other systems in the brain. To study endogenous aldo production in in steroidogenic neurons in isolation from the many other systemic changes produced by mineralocorticoid excess and the complex genetic etiologies of the hypertension in the Dahl SS rat, we created a transgenic rat that over-expresses the last and unique enzyme for aldo synthesis, the aldosterone synthase (AS). Aldo synthesis would be restricted to neurons that are normally steroidogenic and express the requisite upstream enzymes and cofactors. The Syn1/AS-Tg rat is hypertensive on standard rat chow without extra salt, its plasma and total body aldo production levels are normal, and it is predisposed to heart and kidney failure.
In Specific Aim 1 we propose to study the expression of AS in relation to the MR and the regulation of aldo biosynthesis in the brain in the SS rat compared to that in the normotensive outbred SD.
In Specific Aim 2 we propose study the cardiovascular effects of the increase in aldo synthesis in the brain in the transgenic Syn1/AS-Tg rat in isolation of confounding genes in the SS rat. An AS shRNA carried by a lentiviral vector will be used to decrease AS expression and identify the neurons in which the synthesis of aldo is critical to blood pressure control and cardiovascular and renal pathology in SS and AS rats in Aims 1 and 2.
Specific Aim 3 is to study the hemodynamic and heart and kidney pathology produced in normotensive SD rats in which an established viral vector has been used to selectively over-express AS in neurons in the hemodynamic control regions of SD rats. Immunohistochemical capabilities unique to our lab will be used to study colocalization of the MR and AS proteins. Telemetry with spectral analysis of wave form to assess baroreceptor function and sympathetic drive, and doppler-echocardiography will be used to asses hemodynamic changes and cardiac dysfunction. Quantitative RT-PCR, will be used to assess changes in message. Quantitative RT-PCR, western blots and ihc will be used to follow AS and MR expression. Plasma and tissue concentrations of adrenal steroids and proinflammatory cytokines associated with mineralocorticoid- induced end-organ damage will be measured by ELISA. Neuronal aldosterone production should only be relevant to blood pressure control in those neurons that have MR responsible for BP homeostasis. Relevance to the VA patient care mission: Hypertension and its associated cardiac and renal disease are primary causes of disability and death in VA patients as well as all of the USA. The RALES, EPHESUS and more recent clinical studies confirm the importance of understanding how and under what combination of circumstances activation of the MR results in cardiovascular pathology, particularly when plasma concentrations of aldosterone are normal. Identifying which MR are inappropriately activated, as well as why is important for the development of specific prevention and therapy. We plan to test the postulate that excessive extra adrenal aldosterone production in specific neurons of the CNS leads to hypertension and cardiovascular and renal damage.
Hypertension and its associated heart, vessel and kidney disease are primary causes of disability and death in VA patients as well as all of the USA. Recent clinical studies confirm the importance of the mineralocorticoid receptor in cardiovascular and renal disease. However these receptors are found in several parts of the brain, only some of which control blood pressure and heart function. Most of the mineralocorticoid receptors in the brain are in areas concerned with memory, learning and mood, parts of the brain that are altered in depression, including post traumatic stress disorder. Understanding under what combination of circumstances activation of the mineralocorticoid receptor in the brain results in heart, vessel and kidney disease is important to develop specific prevention and treatment strategies.