Angiotensin II‐Mediated AT1R‐TLR4 Crosstalk Promotes Blood‐Brain Barrier Disruption, Microglial Activation, and Sympathoexcitation in Hypertension

Abstract

Previous efforts to define the mechanisms underlying neurogenic hypertension implicate angiotensin II (AngII) as a critical force behind the blood‐brain barrier (BBB) disruption observed in this pathology. We have further shown that exposure of hypothalamic microglia to AngII stimulates Toll‐like receptor 4 (TLR4) via AngII type 1 receptor (AT1R), resulting in microglial activation. We propose that abnormal AngII‐mediated microglial activation through AT1R‐TLR4 crosstalk drives a vicious feed‐forward loop comprised of BBB disruption, neuroinflammation, and sympathoexcitation. Spontaneously hypertensive rats (SHRs) were treated with either Losartan (AT1R antagonist; 20mg/kg/day; oral gavage; 4 weeks) or TAK‐242 (TLR4 inhibitor; 2mg/kg/day; i.p.; 14 days) and age‐matched with sham SHRs and Wistar Kyoto (WKY) rats. Indirect mean arterial pressure (MAP) readings were recorded with a tail‐cuff volume‐pressure system (CODA‐6, Kent Scientific). TLR4 protein density and microglial activation status were examined by immunofluorescence assays with primary antibodies against TLR4 and IBA1. In the TAK‐242 cohort, extravasation of a low molecular weight dextran‐conjugated dye was quantified as an index of BBB permeability. Sympathetic nervous system activity was determined by ganglionic blockade (hexamethonium bromide; 20mg/kg; i.v.) in conscious rats during continuous direct MAP recording via femoral catheterization. MAP was normalized in Losartan‐treated SHRs (104±2 vs 100±3mmHg in WKYs), and significantly reduced compared to control SHRs with TLR4 inhibition (129±3 vs 154±2mmHg in SHRs). TLR4 density in the hypothalamic paraventricular nucleus (PVN) and rostral ventrolateral medulla (RVLM) of SHRs was increased by 104% and 113%, respectively, compared to WKYs, and normalized following Losartan treatment. Analysis of microglial morphology showed significant enhancement of activation in SHRs, demonstrated by a decrease in total end‐points (EP) and branch length (BL) per frame versus WKYs in the PVN (EP: −20.8%; BL: −34.3%) and RVLM (EP: −41.6%; BL: −33.4%). In both nuclei, AT1R inhibition halted excess microglial activation. Within the PVN of TAK‐242‐treated animals, a similar normalization of microglial activation with TLR4 antagonism was observed (EP: +4.7%; BL: +3.9% vs WKYs). BBB permeability analysis in the PVN and RVLM revealed complete restoration after TAK‐242 treatment, whereas dye leakage was significantly elevated in SHRs (PVN: 132.3%; RVLM: 135.4% vs WKYs). Ganglionic blockade induced a greater pressor response in SHRs (−65.1±3.1) compared to WKYs (−46.7±4.2), reflecting sympathoexcitation in the hypertensive animals that was abolished with TLR4 inhibition (−42.9±4.3mmHg). Our data suggest a pro‐hypertensive cycle involving BBB disruption, microglial activation, and sympathoexcitation driven by AngII‐induced microglial AT1R‐TLR4 crosstalk. In combination with prior work, our findings suggest a substantial contribution of AngII's interactions with the innate immune system to the pathophysiology of neurogenic hypertension.Support or Funding InformationThis work was funded by AHA14SDG20400015 to VCB.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.