Regulator of calcineurin (Rcan)1-4 protein and mRNA expression increase at sites of transient focal brain ischemia, paralleling increases in inflammatory cytokine mRNA levels. (A), panel (i) Representative immunoblot showing expression of endogenous Rcan1-1 and Rcan1-4 proteins in infarcted cortical tissue (I) and the corresponding region of the contralateral hemisphere (C) of rats subjected to middle cerebral artery occlusion (MCAO) for 1 h followed by reperfusion for the indicated times. Sh = sham operated animals. A total of 100 μg protein was analyzed per lane. Panel (ii), band density quantification from four independent experiments as in (A), normalized to α-tubulin. (B) Histograms showing relative quantification of mRNA expression by TaqMan real time qRT-PCR. Rat mRNAs for Rcan1-4 (i), TNF α (ii), IL-6 (iii) and Cox-2 (iv) were amplified from total RNA from infarcted and corresponding contralateral hemispheres of rats subjected to MCAO (1 h) followed by reperfusion (I/R) as indicated. Transcript amounts are normalized to 18S rRNA, and are expressed relative to contralateral samples from sham-operated animals after reperfusion for 5 h (Sh). Real time qPCR reactionwas conducted in triplicate for each condition, and data are the means ± SD of four experiments. **P < 0.01, *P < 0.05 (analysis of variance (ANOVA)) versus contralateral Sh samples.