Background and purpose: Stress on the endoplasmic reticulum (ER) can trigger rescuer responses such as the unfolded protein response (UPR). However, pharmacological modulators of these ER-regulated stress responses are not well understood. In the present study, we found that amiloride, a potassium-sparing diuretic, has unique properties relating to such stress. Experimental approach: We treated mouse primary cultured glial cells with amiloride, in the absence and presence of the ER stress-inducing reagents tunicamycin (Tm) or dithiothreitol, and measured UPR and ER stress-induced cell death. IRE1a phosphorylation, eIF2a phosphorylation, X-box binding protein 1 (XBP1) splicing, glucose regulated protein 78 (GRP78) and CCAAT/enhancer-binding protein homologous protein (CHOP) expression by reverse transcription-polymerase chain reaction and Western blotting were used to assess UPR and lactate dehydrogenase activity was determined to measure ER stress-induced cell death. Key results: Amiloride completely inhibited ER stress-induced activation of IRE1a, an ER-localized stress sensor protein, splicing of XBP1, and subsequent expression of GRP78 at the mRNA and protein levels. ER stress induces the phosphorylation of eIF2a, leading to the expression of CHOP or an attenuation of translation in cells. Surprisingly, treatment with amiloride alone markedly promoted the phosphorylation but actually inhibited ER stress-induced CHOP expression. Finally, we found that amiloride (200 mM) synergistically enhanced ER stress-induced cell death, which was mediated through caspases. On the other hand, a low dose of amiloride (20 mM) significantly prevented Tm-induced cell death. Conclusions and implications:These results suggest that amiloride can modulate UPR. They also suggest amiloride to be an important pharmacological agent and provide basic information for understanding and preventing ER stress-related diseases.

The disruption of endoplasmic reticulum function leads to an accumulation of unfolded proteins, which results in endoplasmic reticulum stress. In the present study, we investigated the effect of vanadate on such stress. Endoplasmic reticulum stress increased glucose-regulated protein 78 (GRP78) and CCAAT/enhancer-binding protein homologous protein (CHOP) expressions in glial cell cultures. We found that vanadate inhibited the endoplasmic reticulum stress-induced increase in GRP78 and CHOP expressions at both mRNA and protein levels. Thus, these results suggest that vanadate modulates endoplasmic reticulum stress responses and that novel vanadate-responsive protein(s) might be involved in these processes.