Elevated levels of
free fatty acids (FFA) have been implicated in the pathogenesis of neuronal injury and death induced by
cerebral ischemia. This study evaluated the effects of
immunosuppressants agents,
calcineurin inhibitors and blockade of endoplasmic reticulum (ER)
calcium channels on
free fatty acid formation and efflux in the ischemic/reperfused (I/R) rat brain. Changes in the extracellular levels of arachidonic, docosahexaenoic, linoleic, myristic, oleic and
palmitic acids in cerebral cortical superfusates during four-vessel occlusion-elicited global
cerebral ischemia were examined using a cortical cup technique. A 20-min period of
ischemia elicited large increases in the efflux of all six FFAs, which were sustained during the 40 min of reperfusion.
Cyclosporin A (CsA) and
trifluoperazine, which reportedly inhibit the I/R elicited opening of a mitochondrial permeability transition (MPT) pore, were very effective in suppressing
ischemia/reperfusion evoked release of all six FFAs.
FK506, an
immunosuppressant which does not directly affect the MPT, but is a
calcineurin inhibitor, also suppressed the I/R-evoked efflux of FFAs, but less effectively than CsA.
Rapamycin, a derivative of
FK506 which does not inhibit
calcineurin, did not suppress I/R-evoked FFA efflux.
Gossypol, a structurally unrelated inhibitor of
calcineurin, was also effective, significantly reducing the efflux of docosahexaenoic, arachidonic and
oleic acids. As previous experiments had implicated elevated Ca(2+) levels in the activation of
phospholipases with FFA formation, agents affecting endoplasmic reticulum stores were also evaluated.
Dantrolene, which blocks the
ryanodine receptor (RyR) channel of the ER, significantly inhibited I/R-evoked release of docosahexaenoic, arachidonic, linoleic and
oleic acids.
Ryanodine, which can either accentuate or block Ca(2+) release, significantly enhanced
ischemia/reperfusion-elicited efflux of
linoleic acid, with non-significant increases in the efflux of myristic, arachidonic, palmitic and
oleic acids.
Xestospongin C, an inhibitor of the
inositol triphosphate (IP(3)R) channel, failed to affect I/R-evoked FFA efflux.
Thapsigargin, an inhibitor of the Ca(2+)-
ATPase ER uptake pump, elicited significant elevations in the efflux of myristic, arachidonic and
linoleic acids, in the absence of
ischemia. Collectively, the data suggest an involvement of both ER and mitochondrial Ca(2+) stores in the chain of events which lead to PLA(2) activation and FFA formation.