Endothelin 1 impairs norepinephrine neutornal transporter in PC12 cells: Role of the endoplasmic reticulum stress

Abstract

Neuronal norepinephrine transporter (NET) removes norepinephrine from the synaptic cleft. Several studies show that NET impairment is associated with cardiovascular diseases like hypertension (HT) whereas recent works link endoplasmic reticulum stress (ERE), triggered by misfolded proteins accumulation in the reticulum, with the development of HT. We recently reported that endothelin receptors (ETA/ETB) blockade in rats with salt-dependent HT decreases blood pressure and markers of ERE and ERE-dependent apoptosis as well as misfolded NET accumulation in the adrenal medulla. In the present study we aimed to unveil the mechanisms underlying these findings. PC12 cells were exposed to 10mM endothelin 1 (ET1), 10nM thapsigargin (TG) or 0.1ug/ml tunicamycin (TN) (TG and TN were used as ERE positive controls) and NET and ERE markers expression were assessed by Western blot. Results were expressed as the mean ±SD, and p<0.05 was considered significant. Results showed that ET1, TG and TN increased the expression of non-glycosylated NET vs. glycosylated NET (126%, 172% and 130%, respectively, p<0.05. In addition, ET1 increased the expression of chaperone Bip (binging immunoglobulin protein) (205±47% vs. control, p <0.05), proteins of the PERK (protein kinase-like ER kinase) pathway (pPERK: 92±3% vs control, p <0.05; peIF2α/eIF2α: 172± 63% vs. control, p <0.05) and ATF6 α (activating transcription factor 6α) (187±39% vs. control, p <0.05). Similar findings were observed with TG and TN. These findings suggest that the increase in non-glycosylated NET induced by ET-1 would trigger ERE by the accumulation of this misfolded and non-functional protein and likely others in PC12 cells.