P2Y₁₂ but not P2Y₁₃ Purinergic Receptor Controls Postnatal Rat Retinogenesis In Vivo

Adenine nucleotides through P2Y₁ receptor stimulation are known to control retinal progenitor cell (RPC) proliferation by modulating expression of the p57^KIP2, a cell cycle regulator. However, the role of Gi protein-coupled P2Y₁₂ and P2Y₁₃ receptors also activated by adenine nucleotides in RPC proliferation is still unknown. Gene expression of the purinergic P2Y₁₂ subtype was detected in rat retina during early postnatal days (P0 to P5), while expression levels of P2Y₁₃ were low. Immunohistochemistry assays performed with rat retina on P3 revealed P2Y₁₂ receptor expression in both Ki-67-positive cells in the neuroblastic layer and Ki-67-negative cells in the ganglion cell layer and inner nuclear layer. Nonetheless, P2Y₁₃ receptor expression could not be detected in any stratum of rat retina. Intravitreal injection of PSB 0739 or clopidogrel, both selective P2Y₁₂ receptor antagonists, increased by 20 and 15%, respectively, the number of Ki-67-positive cells following 24 h of exposure. Moreover, the P2Y₁₂ receptor inhibition increased cyclin D1 and decreased p57^KIP2 expression. However, there were no changes in the S phase of the cell cycle (BrdU-positive cells) or in mitosis (phospho-histone-H3-positive cells). Interestingly, an increase in the number of cyclin D1/TUNEL-positive cells after treatment with PSB 0739 was observed. These data suggest that activation of P2Y₁₂ receptors is required for the successful exit of RPCs from cell cycle in the postnatal rat retina.