Effect of serum and SphK1 expression on activation of S1P receptors. Membranes were isolated from vector-expressing (□) and SphK1-expressing (▪) RBL-2H3 cells cultured in the presence (A,B) or absence (C,D) of 15% serum for 24 hours and [³⁵S]GTPγS binding was determined without or with the indicated concentrations of S1P for 30 minutes. Aliquots were added to wells precoated with Gα_i3 antibody (A,C) or Gα_q antibody (B,D), incubated for 2 hours at room temperature, and bound radioactivity determined. Data are expressed as percent of controls incubated with [³⁵S]GTPγS and vehicle. Values are means ± SD of triplicate determinations. Similar results were obtained in 2 additional experiments. Statistical significance was determined by Student t test; ^*P < .05.