NeuropharmacologyNeuroprotective effects of tacrolimus (FK506) in a model of ischemic cortical cell cultures: Role of glutamate uptake and FK506 binding protein 12 kDa
Section snippets
Experimental procedures
The experimental protocols, involving animals and their care, strictly conformed to the guidelines of the French Agriculture and Forestry Ministry (decree 87–848). In all experiments attention was paid to the regulations of local authorities for handling laboratory animals, the European Communities Council Directive (86/609/EEC). Particular efforts were made to minimize animal suffering and to reduce the number of animals used.
Protective effects of tacrolimus against OGD-induced neuronal injury
Neuronal damage was first assessed by cell counting after immunostaining experiments (Table 1). Compared with sham wash, OGD induced a significant decrease in the neuronal count observed in controls 24 h after the injury but did not affect the number of GFAP/DAPI positive cells (underlying glial cells). This decrease was significantly reversed by tacrolimus, MK-801 and GPI1046 but not by rapamycin added during OGD.
LDH release and MTT reduction are closely correlated with cell counting for
Discussion
Using a transient OGD on primary mixed cultures of neurons and astrocytes, we show that tacrolimus reverses OGD-induced neuron death when assessed 24 h after the injury. This protective effect is observed when tacrolimus was added either during OGD or during reoxygenation. Both of these beneficial effects can probably be explained by the binding of tacrolimus to FKBP12. Moreover, the effect of tacrolimus added during OGD may be mediated, at least in part, by the fact that it reverses the
Conclusion
In conclusion, this study demonstrates a neuroprotective effect of tacrolimus in an in vitro model of ischemia/reperfusion. This protective effect was observed both when tacrolimus was added at the time of injury and when added during reoxygenation. Both of these beneficial effects of tacrolimus on neuronal viability may involve the FKBP12 pathway and its preventive action may be mediated, at least in part, by a restoration of the GLT1-mediated activity which is dramatically impaired by OGD,
Acknowledgments
This work was supported by a grant from Fujisawa GmbH (2002) and from DGA (Direction Générale de l’Armement, contract no. 00 34 052 00 470 75 01). We thank Mrs. Marjorie Sweetko for the English version of this manuscript. The institution had the responsibility for maintaining objectivity in research by ensuring that the design, conduct, or reporting of research has not been biased by any conflicting financial interest of investigators responsible for the research.
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