Bidirectional Regulation of DARPP-32 Phosphorylation by Dopamine

Abstract

Dopamine has been shown to stimulate phosphorylation of DARPP-32, a phosphoprotein highly enriched in medium-sized spiny neurons of the neostriatum. Here, we investigated the contribution of D1-like and D2-like dopamine receptors in the regulation of DARPP-32 phosphorylation in mouse striatal slices. D1-like and D2-like receptors had opposing effects on the state of DARPP-32 phosphorylation. The D1 receptor agonist SKF82526 increased DARPP-32 phosphorylation. In contrast, the D2 receptor agonist quinpirole decreased basal as well as D1 agonist-, forskolin-, and 8-bromo-cAMP-stimulated phosphorylation of DARPP-32. The ability of quinpirole to decrease D1-stimulated DARPP-32 phosphorylation was calcium-dependent and was blocked by the calcineurin inhibitor cyclosporin A, suggesting that the D2 effect involved an increase in intracellular calcium and activation of calcineurin. In support of this interpretation, Ca 2ϩ -free/EGTA medium induced a greater than 60-fold increase in DARPP-32 phosphorylation and abolished the ability of quinpirole to dephosphorylate DARPP-32. The antipsychotic drug raclopride, a selective D2 receptor antagonist, increased phosphorylation of DARPP-32 under basal conditions and in D2 agonist-treated slices. The results of this study demonstrate that dopamine exerts a bidirectional control on the state of phosphorylation of DARPP-32.Key words: DARPP-32; dopamine; D2 receptor; phosphorylation; neostriatum; calcineurin; raclopride DARPP-32, a dopamine-and cAM P-regulated phosphoprotein of M r 32,000, is a cytosolic protein that is selectively enriched in medium-sized spiny neurons in neostriatum (Ouimet et al., 1984;Walaas and Greengard, 1984). DARPP-32 is phosphorylated by cAM P-dependent protein kinase (PK A) on a single threonine residue, thr 34 , resulting in its conversion into a potent inhibitor of protein phosphatase-1 . DARPP-32 can be dephosphorylated and inactivated in vitro by the calcium/ calmodulin-dependent protein phosphatase calcineurin (King et al., 1984). Dopamine has been shown to stimulate the phosphorylation of DARPP-32 in neostriatum by activation of a biochemical cascade involving stimulation of D1 receptors, activation of adenylyl cyclase, increased cAM P formation, and increased activity of PK A . The selective enrichment of DARPP-32 in dopaminoceptive neurons and its regulation by dopamine strongly indicate that DARPP-32, by regulating protein phosphatase-1 activity, plays a key role in mediating the effects of dopamine on these cells. The control of protein phosphatase-1 activity by DARPP-32 is likely to have a significant role in the regulation of neuronal excitability. For instance, in neostriatum, dopamine-mediated effects on the function of calcium channels (Surmeier et al., 1994), voltagedependent sodium channels (Surmeier et al., 1992;Schiffman et al., 1994), and Na ϩ ,K ϩ -ATPase (Aperia et al., 1991) are regulated directly or indirectly by protein phosphatase-1.Medium-sized spiny neurons of the neostriatum and nucleus accumbens receive dopaminergic input fro...

Glutamate regulation of DARPP-32 phosphorylation in neostriatal neurons involves activation of multiple signaling cascades

“…8. Slices were treated with drugs as specified in each experiment, and the following drugs were obtained: glutamate, MK-801, and SCH23390 from Sigma; 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), LY367385, 2-methyl-6-(phenylethynyl)pyridine (MPEP), 7-nitroindazole monosodium salt (7-NINA), 1H- [1,2,4]oxadiazolo [4,3-a]quinoxalin-1-one (ODQ), U0126, U73122, and ZM241385 from Tocris Cookson (Bristol, U.K.); and calyculin A, cyclosporin A, and okadaic acid from LC Laboratories (Woburn, MA). After drug treatment, slices were transferred to Eppendorf tubes, frozen on dry ice, and stored at Ϫ80°C until assayed.…”

“…Abbreviations: CK, casein kinase; Cdk5, cyclin-dependent kinase 5; DARPP-32, dopamineand cAMP-regulated phosphoprotein of 32 kDa; ERK, extracellular signal-regulated kinase; mGlu, metabotropic glutamate; CNQX, 6-cyano-7-nitroquinoxaline-2,3-dione; nNOS, neuronal NO synthase; PKA, cAMP-dependent kinase; PKG, cGMP-dependent kinase; PLC, phospholipase C; PP, protein phosphatase; PP-1, PP-2A, and PP-2B, protein phosphatases 1, 2A, and 2B; AMPA, ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; MPEP, 2-methyl-6-(phenylethynyl)pyridine; 7-NINA, 7-nitroindazole monosodium salt; ODQ, 1H- [1,2,4]oxadiazolo [4,3-a]quinoxalin-1-one.…”

“…DARPP-32 phosphorylated at Thr-75 inhibits PKA activity and thereby reduces the efficacy of dopamine signaling. Dopamine, by means of dopamine D1 receptors, activates PKA, which directly stimulates DARPP-32 Thr-34 phosphorylation and indirectly stimulates DARPP-32 Thr-75 dephosphorylation (4,5). By regulating the activity of PKA and PP-1, dopamine controls the state of phosphorylation of various intracellular targets.…”

Banishing the homunculus: Making working memory work

“…In brief, our model leverages the observation that dopamine D1 and D2 receptors are relatively segregated in Go and NoGo cells, respectively (Gerfen, 1992;Wise, Murray, & Gerfen, 1996;Aubert, Ghorayeb, Normand, & Bloch, 2000). This is of interest because dopamine effects on neuronal excitability and synaptic plasticity are dependent on whether it acts via D1 or D2 receptors (Hernandez-Lopez, Bargas, Surmeier, Reyes, & Galarraga, 1997; Hernandez-Lopez, Tkatch, Perez-Garci, Galarraga, Bargas, Hamm, & Surmeier, 2000;Nishi, Snyder, & Greengard, 1997;Centonze, Picconi, Gubellini, Bernardi, & Calabresi, 2001). The net effect is that increases in dopamine during positive reinforcement enhance BG Go firing and learning via simulated D1 receptors, whereas decreases in dopamine during negative reinforcement have the opposite effect, enhancing NoGo firing and learning via simulated D2 receptors.…”

Dopamine modulates attentional control of auditory perception: DARPP-32 (PPP1R1B) genotype effects on behavior and cortical evoked potentials

“…The DARPP-32 protein is phosphorylated by dopamine D1 receptor stimulation, and dephosphorylated by D2 receptor stimulation (Nishi, Snyder, & Greengard, 1997). The protein modulates striatal dopamine cellular excitability and synaptic plasticity related to the dopamine receptors (Calabresi et al, 2000;Fienberg et al, 1998;Gould & Manji, 2005).…”