Localization of Central Angiotensin II Receptors with [¹²⁵I]-sar¹, ile⁸-Angiotensin II: Periventricular Sites of the Anterior Third Ventricle

Abstract

The radiolabeled angiotensin II (ANG II) antagonist, [¹²⁵I]-sar¹, ile⁸-ANG II, was used to study brain ANG II receptors by both homogenate binding and in vitro autoradiography. In homogenate preparations of the hypothalamus, thalamus, septum and midbrain (HTSM), [¹²⁵I]-sar¹ ile⁸-ANG II bound to a single class (Hill slope 0.84 ± 0.05) of high affinity binding sites (K_D 0.42 ± 0.03 nM, B_max 5.98 ± fmol/mg protein). Competition for the [¹²⁵I]-sar¹ ile⁸-ANG II binding site in HTSM membranes demonstrated a rank order potency characteristic of binding to the ANG II receptor, with the unlabeled antagonist being slightly more potent than ANG II (Ki 0.22 ± 0.03 vs. 0.95 ± 0.06 nM, respectively). Brain slices from the region of the rostral third ventricle were incubated with 0.5 nM [¹²⁵I]-sar¹, ile⁸-ANG II in the presence or absence of 1 µM ANG II and exposed to LKB Ultrofilm. Autoradiographic images of [¹²⁵I]-sar’, ile⁸-ANG II binding revealed that structures situated within the anterior wall of the third ventricle, i.e. the lamina terminalis, were heavily labeled; including the subfornical organ, median preoptic nucleus and organum vasculosum laminae terminalis. These results show the utility of [¹²⁵I]-sar¹, ile⁸-ANG II as a probe to study brain ANG II receptors and provides pharmacological evidence for the rostral third ventricle as a possible site for central ANG II actions.

Comparison of 3 AT1 Receptor Binding Assays: Filtration Assay, ScreenReady™ Target, and WGA Flashplate®

“…The K d values obtained for [ 125 I]-Sar are in agreement with results previously reported in literature. 11,12,[15][16][17] The obtained K i values for losartan and angiotensin II also fit within the range of K i values reported in the literature. 11,[17][18][19][20] A saturation curve and displacement curve are shown in Figure 1.…”

Brain Angiotensin II: New Developments, Unanswered Questions and Therapeutic Opportunities

“…With the development of autoradiographic methods (Mendelsohn et al, 1984;Gehlert et al, 1986;Healy et al, 1986;Saavedra et al, 1986;, it was possible to precisely determine the location of all Ang II receptors in the brain. The comparison of the distribution of brain Ang II receptors and brain Ang II immunoreactivity revealed that most of the brain Ang II, and most of the brain Ang II receptors, were localized in neurons (Lind et al, 1985;.…”

Angiotensin II AT1 receptor antagonists inhibit the angiotensin‐CRF‐AVP axis and are potentially useful for the treatment of stress‐related and mood disorders

“…1), are present in the brain [Phillips and Sumners, 1998;Saavedra, 1992;Ganten et al, 1984]. The most convincing evidence, however, was the discovery and localization of brain Ang II receptors located inside the blood-brain barrier, inaccessible to circulating Ang II [Mendelsohn et al, 1984;Gehlert et al, 1986;Saavedra et al, 1986d;Healy et al, 1986;, and Ang II immunoreactivity [Lind et al, 1985]. Both brain Ang II receptors and brain Ang II immunoreactivity were localized in neurons Lind et al, 1985].…”