Figure 1.

PKC and PKA regulate mGluR1α signaling. (A) Effect of PMA and bisindolylmaleimide I (Bis) on mGluR1α-dependent InsP accumulation. For each experiment, HEK 293 cells were transfected with 10 μg of mGluR1α and 10 μg of carrier DNA and seeded into six wells of a 24-well cluster. Stimulation was carried out in Hanks' saline solution (containing divalents and glucose) after clearing glutamate from the media with glutamic-pyruvic transaminase/pyruvate (9). The cells were treated with either Me₂SO (vehicle) or test reagents and then stimulated with Glu. Results represent means ± SEM of duplicate or triplicate determinations obtained from at least two independent experiments (*, P ≤ 0.05; **, P ≤ 0.01; two-tailed t test). (B) Effect of PMA on mGluR1α-dependent cAMP accumulation. HEK 293 cells were transfected with 5 μg of mGluR1α along with 5 μg of G_s and 10 μg of carrier DNA and seeded into four wells of a 24-well cluster. The cells were preincubated with either Me₂SO or PMA for 30 min and then stimulated with Glu for 20 min in the continued presence of the test reagents. Results represent means ± SEM of duplicate determinations obtained from three independent experiments. (C) Effect of forskolin on mGluR1α-dependent InsP accumulation. Cell transfection and treatment were conducted as described in A; in control experiments, 4 × 10⁵ untransfected cells were plated for each well. Results represent means ± SEM of triplicate determinations obtained from three independent experiments; for untransfected cells, results shown are means ± SD of triplicate determinations, representative of two independent experiments.