ETA and ETB receptor antagonists synergistically increase extracellular endothelin-1 levels in primary rat astrocyte cultures

Astrocytes produce and bind endothelins (ETs), suggesting that these cells have ET autoregulatory and eliminatory functions. To further investigate these functions in primary rat astrocytes, ET-1 levels in the cell culture media (RIA/HPLC) and intracellular content of ET-1 mRNA (RT PCR) were measured under basal and stimulated (thrombin, 2.2 U/ml) conditions in the presence and absence of ET_A and ET_B selective antagonists (BQ123 or LU135252, and BQ788, respectively). Neither basal nor stimulated ET-1 levels in astrocyte media were influenced by ET_A or ET_B antagonists alone, but were significantly increased by a combination of both. ir ET-3 levels were not affected by antagonist treatment. Exogenous ET-1, added to the cultures, was rapidly cleared from the supernatant; this clearance was markedly inhibited by a combination of BQ123 and BQ788. ET-1 mRNA levels were not altered by any treatment. To conclude, in primary rat astrocyte cultures, extracellular ET-1 is cleared by binding to ET-receptors, apparently involving both, ET_A and ET_B sites. Thus, a blockade of the astrocytic ET eliminatory function as a consequence of the in vivo application of non-selective ET receptor antagonists may lead to increased extracellular ET levels in the brain.