Abstract
Previously, we demonstrated that sphingosine 1-phosphate (S1P) increased the excitability of small-diameter sensory neurons, in part, through activation of S1P receptor 1 (S1PR(1)), suggesting that other S1PRs can modulate neuronal excitability. Therefore, studies were undertaken to establish the expression profiles of S1PRs in the intact dorsal root ganglion (DRG) and in defined single isolated sensory neurons. To determine mRNA expression of S1PRs in the DRG, SYBR green quantitative PCR (qPCR) was used. To determine the expression of S1PR mRNAs in single neurons of defined diameters, a preamplification protocol utilizing Taqman primer and probes was used to enhance the sensitivity of detection. The preamplification protocol also permitted detection of mRNA for two hallmark neuronal receptor/ion channels, TRPV1 and P(2)X(3). Expression profiles of S1PR mRNA isolated from lung and brain were used as positive control tissues. In the intact DRG, the order of expression of S1PRs was S1PR(3)>>R(1)≈R(2)>R(5)≈R(4). In the single neurons, the expression of S1PRs was quite variable with some neurons expressing all five subtypes, whereas some expressing only one subtype. In contrast to the DRG, S1PR(1) was the highest expressing subtype in 10 of the 18 small-, medium-, and large-diameter sensory neurons. S1PR(1) was the second highest expressor in -50% of those remaining neurons. Overall, in the single neurons, the order of expression was S1PR(1)>>R(3)≈R(5)>R(4)>R(2). The results obtained from the single defined neurons are consistent with our previous findings wherein S1PR(1) plays a prominent but not exclusive role in the enhancement of neuronal excitability.