siRNAs targeted to S1PR2 or R3 specifically knockdown the mRNA levels for their respective receptors. (A) 100 and 200 nM siRNA targeted to S1PR2 significantly reduced the mRNA levels of S1PR2 by 79.8% ± 3.1% and 77.1% ± 6.4%, respectively, compared to the untreated control values. There was no difference between the knockdown values for 100 and 200 nM siRNA treatments. The different treatment groups (Cont - untreated control, Meta - metafectene alone, NC - negative control siRNA, and siRNA targeted to the S1PR) were normalized to their respective untreated control values. (B) 100 and 200 nM siRNA targeted to S1PR3 significantly reduced the mRNA levels of S1PR3 by 64.5% ± 7.3% and 70.2% ± 6.3%, respectively, compared to the untreated control values. There was no difference between the knockdown values for 100 and 200 nM siRNA treatments. (C) siRNA targeted to S1PR2 (200 nM) or S1PR3 (200 nM) does not alter the mRNA levels of S1PR1 (P = 0.88). (A-C) values were obtained from neurons isolated from five different tissue harvests; a Pfaffl analysis  was used to quantify the values of receptor mRNA relative to the reference gene, Arbp, for the different treatments; a Kruskal-Wallis ANOVA with a Tukey post hoc test was used to determine statistical differences between the different groups where the asterisks indicate P < 0.05. S1PR - sphingosine-1-phosphate receptor, siRNA - short-interfering RNA.