Abstract 071: Adam17-mediated Neurogenic Hypertension Requires Neuronal Activation of At1a Receptors

We previously reported that neurogenic hypertension is associated with a reduction of Angiotensin Converting Enzyme 2 (ACE2) activity and an increase in A Disintegrin And Metalloprotease 17 (ADAM17) activity in the hypothalamus. Since ADAM17 is known to be expressed in multiple cell types and can be activated by various receptors, we tested the hypothesis that neuronal AT1a receptors (AT1aR) are necessary for ADAM17-mediated ACE2 shedding during neurogenic hypertension. DOCA-salt treatment (DOCA 1mg/g body weight sc + 1% saline p.o./3 weeks) was given to male neuronal AT1aR knockdown mice (AT1aR floxed crossed with Nefh-cre mice, 14-16 week-old) and their non-transgenic (NT) littermates (n=8/group). Following DOCA-salt treatment, enzyme activity assays were performed in hypothalamus of both DOCA-salt-treated and sham mice. Unlike in NT mice, ADAM17 activity was not increased by DOCA-salt treatment in the hypothalamus of neuronal AT1aR knockdown mice (153.7 ±23.8 vs. 128.5 ±11.6 FU/min, P =0.22). To assess whether impaired ADAM17 activation resulted from down-regulation of neuronal AT1aR, or was due to an attenuated BP rise, wild-type mice were infused with norepinephrine (NE; 4 ug/kg/min/14 d, n=6/group). Contrary to DOCA-salt hypertension, ACE2 activity in the hypothalamus remained intact during NE-induced hypertension and there was no significant increase in ADAM17 activity (59.1 ±4.3 vs. 52.3 ±11.7 FU/min, P =0.29). Using FACS, neurons, astrocytes and non-neuronal/non-astrocyte cells were sorted from the hypothalamus. A significant increase in ADAM17 mRNA resulting from DOCA-salt treatment (+1.72 ±0.19 fold vs. sham, P <0.05) was observed only in the neuronal population of NT mice but not in neuronal AT1aR knockdown mice. ERK phosphorylation, NOX4 expression and ROS production, all of which are involved in both intra- and extracellular mechanisms of ADAM17 activation were then assessed using Western blotting and DHE-staining. All were significantly impaired in the hypothalamus of neuronal AT1aR knock-down mice, compared to NT, after DOCA-salt treatment. Taken together, our data provide strong evidence that activation of neuronal AT1aR is responsible for ADAM17-mediated ACE2 shedding and the maintenance of neurogenic hypertension.