Developmental regulation of CREB phosphorylation by dopamine and cyclic AMP in the avian retina

Background: Distinct signaling pathways regulate specific functions during Central Nervous System development such as neuron survival and differentiation. The developing chicken retina is a useful model to study the role of these pathways. Previous work showed that the levels of the ubiquitous second messenger cyclic AMP are regulated by dopamine since early stages of chicken retina development. We have now studied the effects of direct activation of adenylyl cyclase by forskolin or dopamine on CREB phosphorylation during retinal development.

Methods: Retinas from 10-day-old chicken embryos (E10), an early developmental stage, and E16, a more developed stage when most synaptic contacts and retinal layers are already formed, were dissected and stimulated in Hank’s saline with dopamine or the adenylyl cylcase stimulator forskolin. After the incubation period, retinas were lysed and analysed by western blotting.

Results: Dopamine or the D1 receptor agonist SKF 38393 stimulated CREB phosphorylation in E10 but not in E16, effects blocked by the D1 antagonist SCH 23390 or the transmembrane adenylyl cyclase inhibitor SQ 22536. Forskolin strongly stimulated CREB in E10 in a concentration and time-dependent manner, but a much smaller effect was observed in E16. Interestingly, CREB stimulation by forskolin was blocked by the EPAC inhibitor ESI-09 or the soluble adenylyl cyclase inhibitor LRE1, but not by SQ 22536 or the PKA inhibitor KT5720. The stimulation of CREB phosphorylation by forskolin was also observed in rat retina and hippocampus but not in frontal cortex or striatum.

Conclusions: Our results indicate that cyclic AMP increase stimulated by forskolin, or dopamine, differentially controls different signaling pathways leading to CREB phosphorylation during retinal development.