PMC

Pre-treatment of RGCs with pertussis toxin (PTX; Gi/o inhibitor) does not prevent the inhibitory actions of L-803,087 on calcium channel currents. (A) Calcium channel current in an isolated RGC pre-treated (p/t) with PTX. Administration of L-803,087 (gray, 10 nM) reduced calcium channel current compared to control (black trace). (B) The mean data show that L-803,087 significantly reduced calcium channel current(n = 10; *P < 0.05, 2-way RM ANOVA vs. PTX p/t).

Pre-treatment of isolated RGCs with anti-Gβγ peptide reduced calcium channel suppression by L-803,087. (A) Calcium channel currents recorded from an isolated RGC pretreated with a combination of anti-Gβγ peptide (1 μM) in the absence (black) and presence of L-803,087 (gray, 10 nM). (B) The mean data show that L-803,087 reduced calcium channel current in cells pre-treated with anti-Gβγ peptide (n = 10, *P < 0.05, 2 way RM ANOVA).

Calcium channel currents reduced by L-803,087 in isolated adult retinal ganglion cells (RGCs). (A) Calcium channel currents recorded during a voltage ramp from an isolated RGC in the absence (black) and presence of L-803,087 (gray, 10 nM). (B) The mean data show that L-803,087 produced a significant reduction of calcium channel current (n = 27; *P < 0.05, 2-way RM ANOVA vs. control).

Pre-treatment of isolated RGCs with anti-Gßγ peptide and GFX blocked L-803,087 induced suppression of calcium channel current. (A) Calcium channel currents recorded from an isolated RGC pre-treated with a combination of anti-Gßγ peptide (1 μM) and GFX (5 μM) in the absence (black) and presence of L-803,087 (gray, 10 nM). (B) The mean data show that L-803,087 had no effect on calcium channel current in pre-treated cells (n = 6, NS 2 way RM ANOVA).

Pre-treatment of RGCs with PKC antagonist GF109203X reduced the actions of L-803, 087. (A) Calcium channel currents in isolated RGCs pretreated (p/t) with GFX (left: 1 μM, right: 5 μM). Administration of L-803,087 (left: gray; right: bold) reduced calcium channel currents compared to pre-treatment conditions (regular trace). (B) The mean data show that after cells were pre-treated with 1 μM GFX, L-803,087 reduced calcium channel current (n = 11; *P < 0.05, 2-way RM ANOVA vs. 1 μM GFX p/t). C. Pre-treatment of cells with 5 μM GFX reduced the effect of L-803,087 on calcium channel current (n = 8; *P < 0.05, 2-way RM ANOVA vs. 5 μM GFX p.t).

Depolarizing pre-pulse facilitates calcium channel current but has no influence on the suppression by L-803,087. (A) Calcium channel current recorded using a pre-pulse facilitation voltage command protocol (shown below current traces) in an isolated RGC in the presence of L-803. (B) Pre-pulse facilitation was measured as the percentage increase in calcium channel current after the depolarizing step (120 mV, 50 ms). The mean facilitation in the absence (control) and presence of L-803 was the same showing that pre-pulse facilitation did not reverse the suppression of calcium channel current caused by L-803 (n = 5; 8.4 +/− 1.0 vs. 8.8 +/− 3.2% ctrl vs. L-803, paired t-test).

Summary of effects of signaling pathway modulators administered with L-803,087. L-803 reduced I_Ca in isolated RGCs (n = 27; 41.2 +/− 2.7% vs. control). Pre-treatment with pertussis toxin (PTX) had no effect on the L-803-mediated calcium channel current reduction (n = 10; 34.7 +/− 3.8%). The non-selective PKC antagonist, GF109203X had a concentration dependent effect on L-803-mediated current reduction (1 μM: n = 11; 34.3 +/− 4.4%; 5 μM: n = 8; 14.6 +/− 3.2%; *P < 0.05 vs. L-803 alone). Pre-treatment of cells with anti-β gamma (Anti-Gßγ) peptide impaired the ability of L-803 to reduce calcium channel current (n = 10; 23.0 +/− 4.8%; *P < 0.05 ANOVA vs. L-803 alone). Combination treatment with 5 μM GFX and Anti-Gßγ peptide abolished the effects of L-803 (n = 6; 1.8 +/− 6.0%; *P < 0.05 ANOVA vs. L-803 alone).