ATP induces intracellular calcium increases and actin cytoskeleton disaggregation via P2x receptors

The consequences of purinoceptor activation on calcium signalling, inositol phosphate metabolism, protein secretion and the actin cytoskeleton were de monstrated in the WRK-1 cell line. Extracellular ATP was used as a secretag ogue to induce a rise in intracellular Ca(2+)concentration ([Ca2+](i)), act ing via P2x purinergic receptors, which causes actin skeleton disaggregatio n and protein secretion. ATP bound specifically to purinergic receptors, wi th Ki of 0.8 muM. The magnitude order for binding of different nucleotides was alpha,beta -Met-ATP greater than or equal to dATP alphaS > ATP greater than or equal to ADP > UTP > AMP > suramin. No increase in inositol phospha tes (IPs) was observed after ATP application suggesting that the purinergic sites in WRK-1 cells are not of a P2y type. ATP (1-100 muM) caused a conce ntration-dependent increase in [Ca2+](i) (EC50 = 30 muM). The responses wer e reproducible without any desensitization over several applications. The r esponse to ATP was abolished when extracellular calcium ([Ca2+](e)) was red uced to 100 nM. A non-specific purinergic antagonist, suramin, reversibly i nhibited the ATP-response suggesting that ATP is able to bind to P2x purine rgic sites to trigger Ca2+ entry and increase of [Ca2f],. ATP induced a con centration-dependent disaggregation of actin and exocytotic release of prot eins both, which were dependent upon [Ca2+](e). Similarly, alpha,beta -Met- ATP, a potent P2x agonist also stimulated Ca2+ mobilization, actin network destructuration, and protein release. In the isolated rat neurohypophysial nerve terminals, ATP was shown to act as a physiological stimulus for vasop ressin release via Ca(2+)entry through a P2x receptor [6], Here, we show th at in these nerve terminals, ATP is also able to induce actin disaggregatio n by a Ca(2+)dependent mechanism. Thus, actin cytoskeleton alterations indu ced by ATP through activation of P2x receptors could be a prelude to exocyt osis. (C) 2001 Harcourt Publishers Ltd.