P2Y1 and cysteinyl-leukotriene receptors mediate purine and cysteinyl-leukotriene co-release in primary cultures of rat microglia.

Inflammation is widely recognized as contributing to the pathology of acute and chronic neurodegenerative
conditions. Microglial cells are pathologic sensors in the brain and activated microglia have been viewed as
detrimental. Leukotriene, including cysteinyl leukotrienes (CysLTs) are suggested to be involved in brain
inflammation and neurological diseases and ATP by its receptors is a candidate for microglia activation.
A23187(10μM) stimulated microglia to co-release CysLTs and [3H]adenine based purines ([3H]ABPs), mainly
ATP. The biosynthetic production of CysLTs was abolished by 10μM MK-886, an inhibitor of 5-lipoxygenaseactivating
protein activity. RT-PCR analysis showed that microglia expressed both CysLT1 / CysLT2 receptors,
P2Y1 ATP-receptors and several members of the ATP binding cassette (ABC) transporters including MRP1,
MRP4 and P-gp. The increase in [Ca2+]i elicited by LTD4 (0.1 μM) and 2MeSATP(100mM), agonists for CysLTand
P2Y1-receptors, was abolished by the respective antagonists, BAYu9773 (0.5 μM) and suramin (50 μM). The
stimulation of both receptor subtypes, induced a concomitant increase in the release of both [3H]ABPs and CysLTs
that was blocked by the antagonists and significantly reduced by a cocktail of ABC transporter inhibitors,
BAPTA/AM (intracellular Ca2+ chelator) and staurosporine (0.1 μM, PKC blocker). P2Y antagonist was unable
to antagonise the effects of LTD4 and BAYu9773 did not reduce the effects of 2MeSATP. These data suggest that:
i) the efflux of purines and cysteinyl-leukotrienes is specifically and independently controlled by the two receptor
types, ii) calcium, PKC and the ABC transporter system can reasonably be considered common mechanisms
underlying the release of ABPs and CysLTs from microglia. The blockade of P2Y1 or CysLT1/CysLT2 receptors
by specific antagonists that abolished the raise in [Ca2+]i and drastically reduced the concomitant efflux of both
compounds, as well as the effects of BAPTAand staurosporine support this hypothesis. In conclusion, the data of
the present study suggest a cross talk between the purine and leukotriene systems in a possible autocrine/paracrine
control of the microglia-mediated initiation and progression of an inflammatory response.