Injections of angiotensin-converting enzyme 2 inhibitor MLN4760 into nucleus tractus solitarii reduce baroreceptor reflex sensitivity for heart rate control in rats

Abstract

Injections of the angiotensin(1-7) [Ang(1-7)] antagonist [d-Ala7]-Ang(1-7) into the nucleus of the solitary tract (NTS) of Sprague-Dawley rats reduce baroreceptor reflex sensitivity (BRS) for control of heart rate by approximately 40%, whereas injections of the angiotensin II (Ang II) type 1 receptor antagonist candesartan increase BRS by 40% when reflex bradycardia is assessed. The enzyme angiotensin-converting enzyme 2 (ACE2) is known to convert Ang II to Ang(1-7). We report that ACE2 activity, as well as ACE and neprilysin activities, are present in plasma membrane fractions of the dorsomedial medulla of Sprague-Dawley rats. Moreover, we show that BRS for reflex bradycardia is attenuated (1.16 +/- 0.29 ms mmHg-1 before versus 0.33 +/- 0.11 ms mmHg-1 after; P < 0.05; n = 8) 30-60 min following injection of the selective ACE2 inhibitor MLN4760 (12 pmol in 120 nl) into the NTS. These findings support the concept that within the NTS, local synthesis of Ang(1-7) from Ang II is required for normal sensitivity for the baroreflex control of heart rate in response to increases in arterial pressure.

Figure 1. Activity for ACE, ACE2 and neprilysin (NEP) in membrane homogenates from rat brain dorsal medulla

Activities of ACE and neprilysin were determined with Ang I, and activity of ACE2 was determined with Ang II as the substrate. Of the two Ang(1–7) forming enzymes, ACE2 activity is sixfold greater than NEP. Values for each enzyme were greater than zero, indicating that significant activity was detected for all three enzymes.

Figure 2. Effects of ACE2 inhibition on mean arterial pressure and heart rate at time points coincident with reflex testing

There was a significant reduction in mean arterial pressure within the first 30–60 min of the response to NTS injection of MLN4760 and this persisted for 90–180 min. There were no differences in heart rate throughout the study. *P < 0.05 versus before.

Figure 3. Baroreceptor reflex sensitivity for control of heart rate in response to increases in pressure produced by phenylephrine before and after NTS injections of MLN4760

There is a significant reduction in BRS within the first 30–60 min following injection of the ACE2 inhibitor MLN4760 relative to values before treatment (upper panel), which recovered to control levels 90–180 min later. In 5 rats, the reflex was tested after MLN4760 (bottom panel) and again ~50 min later within 5 min of [d-Ala⁷]-Ang(1–7) (DALA), the Ang(1–7) receptor antagonist, injected bilaterally into the NTS. The BRS after [d-Ala⁷]-Ang(1–7) was not different from that after MLN4760 alone. *P < 0.05 versus before.