Effect of novel GPCR ligands on blood pressure and vascular homeostasis

Abstract

Maintenance of normal blood pressure under conditions of drug treatment is a measure of system-wide neuro-hormonal controls and electrolyte/fluid volume homeostasis in the body. With increased interest in designing and evaluating novel drugs that may functionally select or allosterically modulate specific GPCR signaling pathways, techniques that allow us to measure acute and long-term effects on blood pressure are very important. Therefore, this chapter describes techniques to measure acute and long-term impact of novel GPCR ligands on blood pressure regulation. We will use the angiotensin type 1 receptor, a powerful blood pressure regulating GPCR, in detailing the methodology. Normal blood pressure maintenance depends upon dynamic modulation of angiotensin type 1 receptor activity by the hormone peptide angiotensin II. Chronic activation of angiotensin type 1 receptor creates hypertension and related cardiovascular disease states which are treated with angiotensin type 1 receptor blockers (ARBs). Thus, a prototype for evaluation of blood pressure control under experimental evaluation of novel drugs.

Distinct Mechanisms of β-Arrestin–Biased Agonist and Blocker of AT1R in Preventing Aortic Aneurysm and Associated Mortality

“…The concept behind this strategy is that puromycin has an analog structure of aminoacyl-tRNAs; thus, it is incorporated into the nascent polypeptide chain of synthesized proteins, reflecting the rate of mRNA translation in vivo. [49][50][51] Thus, puromycin was injected in vivo to quantify protein synthesis. 150 μL of puromycin (catalog no.…”

“…A previous report showed that AT1R/G-protein and βarrestin signaling increase protein synthesis in smooth muscle and HEK293 cells. 51 We evaluated de novo protein synthesis using the surface sensing of translation method as described earlier. [49][50][51] Minimal amount of puromycin injected into peritoneum and allowed to incorporate into actively synthesized protein pool for 30 min.…”

“…51 We evaluated de novo protein synthesis using the surface sensing of translation method as described earlier. [49][50][51] Minimal amount of puromycin injected into peritoneum and allowed to incorporate into actively synthesized protein pool for 30 min. Puromycin incorporated protein profile in the suprarenal segment of the aorta visualized by immunoblot is shown in Figure 5.…”