The Role of Cdk5 in the Regulation of Dopamine Neurotransmission, Neuronal Excitability, and Reward-Related Behavior
Cyclin-dependent kinase 5 (Cdk5) regulates dopamine neurotransmission and synaptic plasticity and has been implicated as a homeostatic target of chronic psychostimulant exposure. In order to investigate the role of Cdk5 in the modulation of signal transduction pathways that regulate motivation and rewardrelated behavior, we developed several Cre/loxP conditional knock-out systems that allow temporal and spatial control of Cdk5 expression in the adult brain. Loss of Cdk5 in the adult brain increased the psychomotor-activating properties of cocaine and enhanced incentive motivation for food. Behavioral changes were accompanied by increased excitability of medium spiny neurons in the nucleus accumbens (NAc) in conditional Cdk5 knock-out mice. Targeted deletion of Cdk5 in the NAc facilitated cocaine-induced locomotor sensitization and conditioned place preference for cocaine. These results suggest that Cdk5 acts as a negative regulator of neuronal excitability in the NAc and that Cdk5 may govern the behavioral effects of cocaine and motivation for reinforcement. In a candidate search for putative Cdk5 substrates that may regulate dopamine-cAMP dependent kinase (PKA) signaling, we identified, confirmed, and characterized Cdk5-dependent phosphorylation of Thr69 RIIβ. We generated novel phosphorylation state-specific antisera to pThr69 RII-beta and found that pThr69 RII-beta levels were reduced in conditional Cdk5 knock-out mice. Functionally, phosphorylation of Thr69 dramatically enhanced calpain-mediated cleavage of RII-beta in vitro, but did not alter inhibition of PKAcat or cAMP binding. Phosphorylation of Thr69 also increased the interaction of RII-beta with an A-kinase anchoring protein (AKAP) docking motif in vitro. In addition, we generated novel phosphorylation state specific antibodies to regulatory phosphorylation sites on cAMP-specific phosphodiesterase 4 (PDE4), an important modulator of PKA signaling. These antibodies to Ser53 PDE4D3 (PKA site) and pSer573 PDE4D3 (ERK site) provide insight into the crosstalk between PKA and ERK pathways in the regulation of cAMP. Finally, we identified and confirmed Thr292 as the site of Cdk5-dependent phosphorylation of MEK1. In conclusion, Cdk5 likely regulates PKA signaling through numerous pathways, including but not limited to DARPP-32, RII-beta, PDE, and MEK1. In summary, these studies identify novel regulatory mechanisms through which Cdk5 controls PKA signaling as well as the interplay between PKA and ERK signaling pathways that regulate motivation and reward-related behavior.