4,4‐diisothiocyanatostilbenedisulphonic acid (DIDS) is cytotoxic and does not provide protection against ischemic insults in cultured neurons or astrocytes

Abstract

DIDS is a putatively cytoprotective compound with numerous cellular targets. Previously our laboratory reported that DIDS provided > 90% cytoprotection in hippocampal slices during ischemic insults (IS); however, recent observations suggest that DIDS‐treated cells may not be viable. To address this inconsistency we examined the effect of DIDS on cellular viability during normoxia and IS in neurons and astrocytes using molecular and biochemical viability assays, electrophysiology, and time‐lapse confocal and transmission electron microscopy (TEM). We found that IS induced membrane depolarization, apoptotic annexin V and necrotic propidium iodide (PI) staining, loss of [ATP], and organelle and DNA damage. DIDS perfusion caused morphological changes in all cells, including rounding off of the cell body and loss of processes. Cellular membrane integrity was better maintained in DIDS‐treated cells than in IS cells; however, DIDS perfusion did not prevent depolarization of cellular membrane potential or [ATP] depletion. Furthermore, in both control and IS cells, annexin V and PI staining were observed within 2 hours of DIDS treatment and were homogenous at 24 hours, while DNA breaks were observed by TUNEL assay. We conclude that DIDS is not cytoprotective against ischemic insults and is cytotoxic to neurons and astrocytes. Previous results implicating DIDS‐mediated cytoprotection should be re‐assessed.