Pharmacological protection against the cytotoxicity induced by 6-hydroxydopamine and H2O2 in chromaffin cells

Abstract

We present in this report the characteristics of the damage induced by 6-hydroxydopamine and H₂O₂ on bovine chromaffin cells in primary culture. Cytotoxicity was quantified using catecholamine cell contents, lactate dehydrogenase (LDH) release, trypan blue exclusion and morphological appearance. An excellent correlation between these four parameters was found. The cytotoxic effects of 6-hydroxydopamine were Ca²⁺-independent. In spite of this, the Ca²⁺ channel antagonists R56865 (N-[1-(4-(fluorophenoxy)butyl)]-4-piperidinyl-N-methyl-2-benzo-thiazolamine) and lidoflazine exhibited marked cytoprotective effects against both 6-hydroxydopamine and H₂O₂. The selective dopamine uptake blocker, bupropion, increased the viability of 6-hydroxydopamine and H₂O₂-treated cells from 20% to around 80%. Catalase drastically protected against the cytotoxic effects of 6-hydroxydopamine and H₂O₂. In contrast, desferrioxamine gave better protection against H₂O₂ cytotoxicity; glutathione and N-acetylcysteine only afforded substantial protection against 6-hydroxydopamine. Three main conclusions emerge from this study. (1st) 6-Hydroxydopamine causes chromaffin cell damage via a mechanism probably related to the production of free radicals, but unrelated to Ca²⁺ ions. Cytoprotection afforded by R56865 and lidoflazine must be unrelated to their Ca²⁺ antagonist properties. This suggests a novel component in the cytoprotective mechanism of action of these drugs. (2nd) The strong cytoprotective effects of bupropion seem to be unrelated to its ability to block the plasmalemmal dopamine carrier. (3rd) Bovine adrenal chromaffin cells in primary cultures are a suitable model for adult neurons to study the basic mechanism of cell damage, and to screen new drugs with putative neuroprotective properties.