Sympathoexcitation in Rats With Chronic Heart Failure Depends on Homeobox D10 and MicroRNA-7b Inhibiting GABBR1 Translation in Paraventricular Nucleus

Abstract

Background Chronic heart failure (CHF) increases sympathoexcitation through Angiotensin II (ANG II) receptors (AT1R) in the paraventricular nucleus (PVN). Recent publications indicate down regulation of γ-aminobutyric acid B-type receptor 1 (GABBR1) and has a binding site for microRNA-7b (miR-7b) that is expressed in the PVN. We hypothesized that ANG II regulates sympathoexcitation through homeobox D10 (HoxD10), which regulates miR-7b in other tissues. Methods and Results Ligation of the left anterior descendent coronary artery in rats caused CHF and sympathoexcitation. PVN expression of AT1R, HoxD10 and miR-7b was increased, while GABBR1 was lower in CHF. Infusion of miR-7b in the PVN caused sympathoexcitation in control animals, and enhanced the changes in CHF. Antisense miR-7b infused in PVN normalized GABBR1 expression while attenuating CHF symptoms including sympathoexcitation. A luciferase reporter assay detected miR-7b binding to the 3’-UTR of GABBR1 that was absent after targeted mutagenesis. ANG II induced HoxD10 and miR-7b in NG108 cells, effects blocked by AT1R-blocker losartan (Los) and by HoxD10 silencing. miR-7b transfection into NG108 cells decreased GABBR1 expression, which was inhibited by miR-7b antisense. In vivo PVN knockdown of AT1R attenuated the symptoms of CHF, while HoxD10 overexpression exaggerated them. Finally, in vivo PVN ANG II infusion caused dose dependent sympathoexcitation that was abrogated by miR-7b antisense and exaggerated by GABBR1 silencing. Conclusions There is an ANG II/AT1R/HoxD10/miR-7b/GABBR1 pathway in the PVN that contributes to sympathoexcitation and deterioration of cardiac function in CHF.

c‐Jun N‐terminal Kinase mediates prostaglandin‐induced sympathoexcitation in rats with chronic heart failure by reducing GAD1 and GABRA1 expression

“…, Wang et al . ). Each period was 30 min and each measurement was recorded as the average of three 5‐minute recordings during the period.…”

“…PVN and NG108 cells were lysed in standard sample buffer (10 m m Tris, 1 m m ethylene diamine tetraacetic acid (EDTA), 1% SDS, 0.1% Triton X‐100 and 1 m m PMSF) using ultrasound as described previously (Wang et al . ). The supernatant was collected, and protein concentration was determined using the bicinchoninic acid (BCA) protein assay kit (Pierce Biotechnology, Rockford, IL, USA).…”

“…Sections were studied using a laser confocal microscope Zeiss 710 (Carl Zeiss, Germany) and analysed by investigators blind to the protocol (Wang et al . ). For each rat, NFL‐positive cell in PVN was identified in the neuronal cell.…”

“…A general avidin/biotin/peroxidase complex (ABC) procedure was applied as described previously (Wang et al . ). Immunohistochemical labelling was performed in floating sections to identify Fra‐LI (sc‐253; Santa Cruz Biotechnology, Santa Cruz, CA, USA).…”

“…The most well‐described systems are the fast baroreceptor‐dependent response in the brainstem and spinal cord, and the angiotensin II (ANG II) stimulated response through the hypothalamic paraventricular nucleus (PVN) (Wang et al . ), which is important for maintaining sympathoexcitation over time (Pyner & Coote ). The PVN regulates sympathetic activity in heart failure through pre‐sympathetic neurones that regulate activity in the brainstem (Pyner , Chen et al .…”