The β-arrestin recruitment assay, demonstrated that CHO cells expressing wt- and pm-GPR143 displayed high constitutive activation compared with CHO cells that do not express GPR143, as previously reported.
19 Thus, pm-GPR143 retains functionality despite the mutations that route it to the plasma membrane. L-DOPA was tested in the β-arrestin recruitment assay against wt- and pm-GPR143, and only a slight increase in activation was observed at 10 μM. Because the compound was previously reported to have a low affinity for the receptor,
12,13 higher L-DOPA concentrations were tested. However, the signal did not reach threshold. L-DOPA activation of calcium signaling pathway through G
q/11 protein coupling and cAMP accumulation pathways through G
i and G
s protein coupling were also tested, and no significant differences were observed compared to controls, supporting a previous study.
13 It is possible that participation of a third factor only present in pigmented cells is necessary for GPR143 activation by L-DOPA. Alternatively, the observed calcium release in ARPE-19 cells after L-DOPA treatment is not a GPR143-specific effect, because several dopamine receptor subtypes are endogenously expressed in those cells, such as D1 and D5 subtypes, and their activation induces intracellular calcium increase.
38,39 Recent studies indicated that L-DOPA produced by RPE plays a role in retinal development through activation of dopamine receptors present on neural retina cells; however, dopamine receptor targeting has yet to be confirmed.
17 RPE cells differentiate during the first days of eye formation,
16 and during this time, tyrosinase produces L-DOPA, which is then secreted and taken up by retinal neurons. Production of dopamine from L-DOPA in immature retina cells induces activation of dopamine receptors and plays a role in retinal differentiation from early development stages.