Mechanisms for the Modulation of Dopamine D1 Receptor Signaling in Striatal Neurons

In the striatum, dopamine D1 receptors are preferentially expressed in striatonigral neurons, and increase the neuronal excitability, leading to the increase in GABAergic inhibitory output to substantia nigra pars reticulata. Such roles of D1 receptors are important for the control of motor functions. In addition, the roles of D1 receptors are implicated in reward, cognition, and drug addiction. Therefore, elucidation of mechanisms for the regulation of dopamine D1 receptor signaling is required to identify therapeutic targets for Parkinson’s disease and drug addiction. D1 receptors are coupled to Gs/olf/adenylyl cyclase/PKA signaling, leading to the phosphorylation of PKA substrates including DARPP-32. Phosphorylated form of DARPP-32 at Thr34 has been shown to inhibit protein phosphatase-1, and thereby controls the phosphorylation states and activity of many downstream physiological effectors. Roles of DARPP-32 and its phosphorylation at Thr34 and other sites in D1 receptor signaling are extensively studied. In addition, functional roles of the non-canonical D1 receptor signaling cascades that coupled to Gq/phospholipase C or Src family kinase become evident. We have recently shown that phosphodiesterases (PDEs), especially PDE10A, play a pivotal role in regulating the tone of D1 receptor signaling relatively to that of D2 receptor signaling. We review the current understanding of molecular mechanisms for the modulation of D1 receptor signaling in the striatum.


INTRODUCTION
Dopamine plays critical roles in the regulation of psychomotor functions in the brain (Bromberg-Martin et al., 2010;Cools, 2011;Gerfen and Surmeier, 2011). The dopamine receptors are a superfamily of heptahelical G protein-coupled receptors, and are grouped into two categories, D 1 -like (D 1 , D 5 ) and D 2 -like (D 2 , D 3 , D 4 ) receptors, based on functional properties to stimulate adenylyl cyclase (AC) via G s/olf and to inhibit AC via G i/o , respectively (Kebabian and Calne, 1979;Jackson and Westlind-Danielsson, 1994;Missale et al., 1998). In the striatum, dopamine D 1 and D 2 receptor expressions are segregated in two types of medium spiny neurons, striatonigral/direct and striatopallidal/indirect pathway neurons, respectively (Hersch et al., 1995;Surmeier et al., 1996;Valjent et al., 2009;Bertran-Gonzalez et al., 2010). In the striatonigral/direct pathway neurons, D 1 receptors are coupled to G s/olf /AC/PKA signaling, and activation of PKA induces the phosphorylation of PKA substrates such as DARPP-32, a dopamine-and cAMP-regulated phosphoprotein of M r 32 kDa, and a transcription factor, CREB, leading to alterations of neuronal functions Hyman and Malenka, 2001). In this review, the roles of DARPP-32 and its phosphorylation in D 1 receptor signaling and the recent findings on the modulation of D 1 receptor/G s/olf /AC/PKA signaling by phosphodiesterase (PDE) inhibitors are discussed. In addition, the non-canonical D 1 receptor signaling cascades that couple to G q /phospholipase C (PLC) or Src family kinase (SFK) are overviewed.
DARPP-32 is selectively deleted in D 1 receptor-enriched striatonigral or D 2 receptor-enriched striatopallidal neurons, revealed that DARPP-32 expressed in two types of medium spiny neurons differentially regulates striatal motor behaviors . The loss of DARPP-32 in striatonigral neurons decreases basal and cocaine-induced locomotor activities and attenuates l-DOPAinduced dyskinesia in a 6-hydorxydopamine hemi-lesioned model of Parkinson's disease, whereas the loss of DARPP-32 in striatopallidal neurons increases basal and cocaine-induced locomotor activities and abolishes haloperidol-induced catalepsy. These findings support the idea that DARPP-32 enhances D 1 receptor functions in striatonigral neurons, but opposes D 2 receptor functions in striatopallidal neurons. The phosphorylation state of DARPP-32 at Thr34 is regulated by the balance of phosphorylation by PKA and dephosphorylation by protein phosphatase 2B (calcineurin) and 2A (PP-2A). PKA signaling in direct pathway neurons is activated by β 1 -adrenceptors (Hara et al., 2010) and 5-HT 4/6 receptors (Svenningsson et al., , 2007 as well as D 1 receptors Kuroiwa et al., 2008), and is inhibited by adenosine A 1 receptors (Yabuuchi et al., 2006), α 2 -adrenceptors (Hara et al., 2010), and μ-opioid receptors (Lindskog et al., 1999). Dephosphorylation of phospho-Thr34 DARPP-32 is mainly regulated by calcineurin . Activation of NMDA or AMPA receptors increases intracellular Ca 2+ and activates calcineurin, leading to the dephosphorylation of DARPP-32 at Thr34 (Halpain et al., 1990;Nishi et al., 1999). D 2 receptors are known to activate calcineurin via PLC/intracellular Ca 2+ signaling (Hernandez-Lopez et al., 2000), leading to the dephosphorylation of DARPP-32 at Thr34 in striatal neurons where D 1 and D 2 receptors are co-expressed (Nishi et al., 1997). PP-2A also contributes to control the dephosphorylation process of DARPP-32 at Thr34 in a coordinated manner with calcineurin, as inhibition of PP-2A and calcineurin induces the synergistic and robust increase in DARPP-32 Thr34 phosphorylation in Frontiers in Neuroanatomy www.frontiersin.org striatal slices . However, the role of PP-2A in the dephosphorylation of DARPP-32 at Thr34 is not fully understood under physiological conditions.
The importance of DARPP-32 Thr75 phosphorylation is implicated in drug abuse. Chronic administration of psychostimulants such as cocaine induces the accumulation of a transcription factor, FosB, resulting in the induction of a downstream target gene, Cdk5 (Bibb et al., 2001). The induced Cdk5 increases DARPP-32 Thr75 phosphorylation and therefore decreases D 1 receptor/PKA signaling. The attenuation of D 1 receptor/PKA signaling is considered as adaptive changes to cocaine addiction. The role of DARPP-32 at Thr75 is also demonstrated in stimulatory action of caffeine (Lindskog et al., 2002). Caffeine, by antagonizing adenosine A 2A receptors in striatopallidal neurons, attenuates A 2A receptor/PKA signaling and PP-2A activity and subsequently increases DARPP-32 Thr75 phosphorylation, which likely contribute to the stimulatory action of caffeine.

ROLE OF THE CK2 PHOSPHORYLATION-SITE AT Ser97 OF DARPP-32 AND THE CK1 PHOSPHORYLATION-SITE AT Ser130 OF DARPP-32
Phosphorylation of DARPP-32 at Ser97 (in mouse sequence; Ser102 in rat sequence) by CK2 is reported to increase the efficacy of DARPP-32 Thr34 phosphorylation by PKA (Girault et al., 1989). In parallel, phosphorylation of DARPP-32 at Ser130 (in mouse sequence; Ser137 in rat sequence) by CK1 decreases the rate of dephosphorylation of Thr34 by calcineurin (Desdouits et al., 1995a). Thus, DARPP-32 phosphorylation by CK2 or CK1 results in the increase in the phosphorylation states of DARPP-32 at Thr34, suggesting the role of CK2 and CK1 to enhance D 1 receptor/PKA/DARPP-32/PP-1 signaling cascade. However, an opposing action of CK2 to inhibit D 1 receptor/PKA signaling is demonstrated (Rebholz et al., 2009). CK2 directly interacts with G s/olf , and negatively controls the functions of D 1 receptors as well as A 2A receptors by enabling faster internalization.
Recently, the phosphorylation state of DARPP-32 at Ser97 is found to be a key regulator of nuclear export of DARPP-32 (Stipanovich et al., 2008). DARPP-32 has been thought as cytoplasmic protein because majority of DARPP-32 was fractionated in the soluble fraction (Walaas and Greengard, 1984), although the immunoreactivity of DARPP-32 was noticed in some nuclei of medium spiny neurons (Ouimet and Greengard, 1990). Importantly, activation of D 1 receptor/PKA signaling induces the nuclear accumulation of DARPP-32 (Stipanovich et al., 2008). The phosphorylation of DARPP-32 at Ser97 by CK2 functions as nuclear export signal of DARPP-32. Ser97 is highly phosphorylated under basal conditions (Girault et al., 1989), and only small fraction of DARPP-32 is located in the nucleus. When PKA is activated, Ser97 is dephosphorylated by PKA-activated PP-2A/B56δ complex, resulting in the accumulation of DARPP-32 (phospho-Thr34/dephospho-Ser97 form of DARPP-32) in the nucleus. The inhibition of PP-1 by phospho-Thr34 DARPP-32 promotes histone H3 phosphorylation and regulates the nuclear function via mechanisms of chromatin remodeling. These findings provide mechanisms for D 1 receptor/PKA/DARPP-32 signaling to regulate gene expression, especially in conditions of drug addiction.
Dopamine D 1 receptors are known to interact and form hetero-oligomers with other neurotransmitter receptors such as dopamine D 2 receptors (Lee et al., 2004;Rashid et al., 2007b), Frontiers in Neuroanatomy www.frontiersin.org dopamine D 3 receptors (Fiorentini et al., 2008;Marcellino et al., 2008), adenosine A 1 receptors (Gines et al., 2000;Toda et al., 2003), and NMDA receptors (Lee et al., 2002), resulting in alteration of D 1 receptor functions (Figure 2). The role of D 1 receptor heterooligomerization will be discussed in other reviews of this special topic.

D 1 RECEPTOR/G q /PLC/IP 3 SIGNALING
In addition to the dopamine D 1 (D 1A ) receptor that couples to G s/olf /AC, the presence of a dopamine D 1 -like receptor that couples to G q /PLC has been proposed (Felder et al., 1989;Undie and Friedman, 1990;Wang et al., 1995;Beaulieu and Gainetdinov, 2011; Figure 1). D 1 -like receptors are shown to couple to both G s /AC and G q /PLC signaling in the striatum and frontal cortex, but solely to G q /PLC signaling in the hippocampus and amygdala (Undie and Friedman, 1990;Wang et al., 1995;Jin et al., 2001). It has been reported that G q /PLC-coupled D 1 -like receptors are coded in mRNA with different size from that of G s/olf /AC-coupled D 1 receptors , and are functional in dopamine receptor D 1A knockout mice (Friedman et al., 1997;Tomiyama et al., 2002), but not in dopamine D 5 receptor knockout mice (Sahu et al., 2009). However, there is a report showing the lack of G q activation in the striatal membrane from D 1A receptor knockout mice (Rashid et al., 2007b). It still needs to be determined whether the cloned D 1A receptor (Drd1a), different types of D 1like receptors such as the D 5 receptor, or both couple(s) to G q /PLC signaling in the striatum. It has been demonstrated that D 1 receptors form the heterooligomer with D 2 receptors, and that the D 1 -D 2 receptor heterooligomer preferentially couples to G q /PLC signaling (Rashid et al., 2007a,b). The expression of dopamine D 1 and D 2 receptors are largely segregated in direct and indirect pathway neurons in the dorsal striatum, respectively (Gerfen et al., 1990;Hersch et al., 1995;Heiman et al., 2008). However, some proportion of medium spiny neurons are known to expresses both D 1 and D 2 receptors (Hersch et al., 1995). Gene expression analysis using single cell RT-PCR technique estimated that 40% of medium spiny neurons express both D 1 and D 2 receptor mRNA (Surmeier et al., 1996). Recently, analysis using drd1a-EGFP and drd2-EGFP bacterial artificial chromosome (BAC) mice revealed that D 1 and D 2 receptors are co-expressed in medium spiny neurons at 5%

FIGURE 2 | D 1 receptors form hetero-oligomers with other receptors.
Formation of hetero-oligomers with dopamine D 1 receptors and other receptors such as D 2 , D 3 , A 1 , and NMDA receptors is shown. Biding of these receptors induces the changes in D 1 receptor function and/or localization.
in the dorsal striatum, 6% in the NAc core, and 17% in the NAc shell . Therefore, D 1 receptors likely interact with D 2 receptors in some populations of medium spiny neurons, where both D 1 and D 2 receptors are co-expressed. D 1 and D 2 receptors, co-expressed in transfected cells and striatal neurons, form the hetero-oligomer (Lee et al., 2004). The D 1 and D 2 receptor hetero-oligomer couples to G q and activates PLC and intracellular Ca 2+ signaling (Lee et al., 2004;Rashid et al., 2007b). For activation of G q /PLC signaling, both D 1 and D 2 receptors are required, because either D 1 or D 2 antagonist and genetic deletion of either D 1 or D 2 receptors abolish the effect (Rashid et al., 2007b). It is likely that SKF83959, which selectively activate G q /PLC signaling but not G s/olf /PKA signaling, acts as a full agonist for D 1 receptors and a partial agonist for D 2 receptors in the form of hetero-oligomer (Rashid et al., 2007a,b). The functional role of the D 1 and D 2 receptor hetero-oligomer to activate Ca 2+ /calmodulin-dependent kinase IIα, which may lead to the induction of brain-derived neurotrophic factor (BDNF) expression, has been suggested (Hasbi et al., 2009), and therefore the D 1 and D 2 receptor hetero-oligomer is implicated as a potential therapeutic target for schizophrenia and drug addiction (Hasbi et al., 2010).
Behavioral studies revealed that activation of D 1 receptor/ G s/olf /AC signaling by SKF83822 mediates sniffing, locomotion, rearing, stereotypy, and seizure (O'Sullivan et al., 2004), whereas activation of D 1 -like receptor/G q /PLC signaling by SKF83959 as well as SKF38393 mediates vacuous jaw movements and intense grooming (Deveney and Waddington, 1995;Undie et al., 2000). Molecular mechanisms by which G q /PLC-coupled D 1 -like receptors regulate the function of neostriatal neurons, and their interaction with the D 1 receptor/G s/olf /AC/PKA signaling cascades need to be clarified.

D 1 RECEPTORS COUPLED TO Src FAMILY KINASE SIGNALING
Activation of D 1 receptors with simultaneous activation of NMDA receptors by drugs of abuse such as cocaine and d-amphetamine induces extracellular-signal regulated kinase (ERK) activation selectively in striatonigral/direct pathway neurons (Valjent et al., 2005;Bertran-Gonzalez et al., 2008), leading to activation of transcription for genes critical for drug-induced plasticity Bertran-Gonzalez et al., 2008; Figure 1). The role of D 1 receptor/PKA/DARPP-32 signaling is also implicated for ERK activation, since the inhibition of PP-1 by phospho-Thr34 DARPP-32 induces activation of mitogen-activated protein kinase/ERK kinase (MEK), which phosphorylates ERK, and inhibition of striatal-enriched tyrosine phosphatase (STEP), which dephosphorylates ERK (Valjent et al., 2005;Girault et al., 2007). In addition, the Ras-guanine nucleotide-releasing factor 1 (Ras-GRF1), a neuronal specific activator of Ras/ERK signaling, is identified as an upstream of MEK/ERK signaling (Fasano et al., 2009). However, mechanisms for activation of NMDA receptors by drugs of abuse were not clearly understood, because the changes in extracellular glutamate levels induced by drugs of abuse were variable (Reid et al., 1997;Zhang et al., 2001). Recently, Pascoli et al. (2011) reported that D 1 receptors potentiate NMDA receptor function via phosphorylation of NR2B subunit (at Tyr1472) of NMDA receptors by SFK, which is responsible for activation of the Ras-GRF1/MEK/ERK signaling cascade by cocaine (Fasano et al., 2009;Pascoli et al., 2011). The activation of SFK by D 1 receptors is mediated through G βγ subunit. The findings are in agreement with previous reports showing that D 1 receptor-mediated increase in NMDA receptor currents requires NR2B subunit and SFK activity (Wittmann et al., 2005), and that activation of D 1 receptors induces trafficking of NMDA receptors to the postsynaptic membrane mediated through NR2B phosphorylation by SFK, especially Fyn (Dunah and Standaert, 2001;Hallett et al., 2006). The novel D 1 receptor/G βγ /SFK/NR2B signaling cascade, which results in activation of NMDA receptors/Ca 2+ /Ras-GRF1/MEK/ERK signaling, likely plays a critical role in pathological conditions such as drug addiction and l-DOPA-induced dyskinesia in the animal model of Parkinson's disease, in which NR2B tyrosine phosphorylation (Menegoz et al., 1995;Pascoli et al., 2011) and activation of ERK signaling by D 1 receptors (Gerfen et al., 2002;Girault et al., 2007;Santini et al., 2007) are enhanced.

MODULATION OF cAMP/PKA SIGNALING IN D 1 -TYPE DIRECT PATHWAY NEURONS AS WELLS AS D 2 -TYPE INDIRECT PATHWAY NEURONS BY PHOSPHODIESTERASE (PDE) INHIBITION
Activation of G s/olf -coupled dopamine D 1 receptors causes an increase in cAMP synthesis, while the subsequent hydrolysis of cAMP is mediated by PDEs. PDEs are subdivided into 11 families, encoded by 21 genes (Conti and Beavo, 2007). Multiple PDEs with different substrate specificities and subcellular localization are expressed in the striatum. PDE10A, PDE1B, and PDE7B are enriched in the striatum, and PDE4 (A, B, and D isoforms), PDE2A and PDE9A, which are widely distributed in the brain, are also expressed in the striatum .
Several types of PDEs such as PDE10A, PDE4, and PDE1B are expressed in direct and indirect pathway neurons. The inhibition of PDEs can result in activation of cAMP/PKA signaling both in direct and indirect pathway neurons. If the function of the PDE (e.g., PDE1B) is predominant in direct pathway neurons, the inhibition of the PDE and activation of cAMP/PKA signaling results in activation of direct pathway neurons, leading to inhibition of GPi/SNpr inhibitory output neurons and activation of thalamocortical motor circuits. Conversely, if the function of the PDE (e.g., PDE10A and PDE4) is predominant in indirect pathway neurons, the inhibition of the PDE and activation of cAMP/PKA signaling results in activation of indirect pathway neurons, leading to activation of GPi/SNpr inhibitory output neurons and inhibition of thalamocortical motor circuits. Thus, PDE inhibitors that predominantly act in direct pathway neurons work like dopamine D 1 receptor agonists and activate motor function, whereas PDE inhibitors that predominantly act in indirect pathway neurons work like dopamine D 2 receptor antagonists and inhibit motor function (Nishi and Snyder, 2010). The balance of action of each PDE inhibitor in indirect and direct pathway neurons determines the behavioral effects (Figure 3).

FIGURE 3 | Roles of phosphodiesterases (PDEs) in the control of basal ganglia-thalamocortical circuitry.
Output neurons in the striatum are medium spiny neurons (MSNs), which consist of striatonigral/direct pathway and striatopallidal/indirect pathway neurons. Direct pathway neurons are GABAergic, and inhibit tonically active neurons in globus pallidus interna (GPi)/substantia nigra pars reticulata (SNpr). Indirect pathway neurons are also GABAergic, and activate neurons in GPi/SNpr via inhibition of globus pallidus externa (GPe) GABAergic neurons and activation of subthalamic nucleus (STN) glutamatergic neurons. Direct and indirect pathway neurons induce opposing effects on the output neurons in GPi/SNpr, resulting in dis-inhibition and pro-inhibition of output, respectively, to motor areas of the thalamus and cortex. The inhibition of PDEs increases cAMP/PKA signaling in both direct and indirect pathway neurons. PDE inhibitors that predominantly act in direct pathway neurons work like dopamine D 1 receptor agonists and activate motor function, whereas PDE inhibitors that predominantly act in indirect pathway neurons work like dopamine D 2 receptor antagonists and inhibit motor function. SNpc, substantia nigra pars compacta. Reproduced with permission from reference Nishi and Snyder (2010).

ROLE OF PDE10A IN DOPAMINE SIGNALING
PDE10A is a dual substrate PDE that hydrolyzes both cAMP and cGMP, and has a higher affinity for cAMP than for cGMP by ∼20fold (Fujishige et al., 1999;Bender and Beavo, 2006). In the striatum, PDE10A is expressed in two types of medium spiny neurons (direct and indirect pathway neurons), but not in interneurons (Xie et al., 2006;Nishi et al., 2008;Sano et al., 2008). Papaverine, an opium alkaloid primarily used for the treatment of visceral spasm and vasospasm, was found to selectively inhibit PDE10A with an IC 50 of 36 nM (Siuciak et al., 2006). Papaverine was used to explore the physiological role of PDE10A in the regulation of striatal function. Recently, the potent PDE10A inhibitors, TP-10 (IC 50 0.3 nM) and MP-10 (IC 50 0.18 nM), were developed (Schmidt et al., 2008). Using these PDE10A inhibitors, PDE10A was shown to hydrolyze both cAMP and cGMP in the striatum in vivo (Siuciak et al., 2006;Schmidt et al., 2008;Grauer et al., 2009). We examined the effect of papaverine on the phosphorylation of PKA substrates including DARPP-32 using neostriatal slices. Papaverine robustly increased the phosphorylation of DARPP-32 at Thr34 and GluR1 at Ser845 in striatal medium spiny neurons in slices as well as in vivo . The effect of papaverine was mediated through the potentiation of cAMP/PKA signaling, but not cGMP/PKG signaling. Similarly to papaverine, inhibition of PDE10A by TP-10 and/or MP-10 in the striatum in vivo was demonstrated to induce the phosphorylation of DARPP-32, GluR1, and CREB at PKA-sites (Schmidt et al., 2008;Grauer et al., 2009). PDE10A is abundantly expressed in direct and indirect pathway neurons, and the expression levels are similar in the two types of neurons (Xie et al., 2006;Nishi et al., 2008;Sano et al., 2008). In agreement, PDE10A regulates cAMP/PKA signaling  as well as gene expression (Strick et al., 2010) in both direct and indirect pathway neurons. In direct pathway neurons, PDE10A inhibition by papaverine activates cAMP/PKA signaling, leading to the potentiation of dopamine D 1 receptor signaling. In indirect pathway neurons, PDE10A inhibition by papaverine also activates cAMP/PKA signaling by simultaneously potentiating adenosine A 2A receptor signaling and inhibiting dopamine D 2 receptor signaling. Since the balance of cAMP/PKA signaling between the direct and indirect pathways determines the output from the basal ganglia, neuronal type-specific regulation of DARPP-32 Thr34 phosphorylation was studied using neostriatal slices from D 1 R-DARPP-32-Flag/D 2 R-DARPP-32-Myc mice , in which Flag-tagged DARPP-32 and Myc-tagged DARPP-32 are expressed selectively in direct and indirect pathway neurons under the control of D 1 and D 2 receptor promoters, respectively . PDE10A inhibition by papaverine increases Myctagged DARPP-32 phosphorylation sixfold in indirect pathways, whereas Flag-tagged DARPP-32 phosphorylation only twofold in direct pathway neurons. Thus, PDE10A inhibitors activate cAMP/PKA signaling in indirect and direct pathway neurons, but the action of PDE10A inhibitors predominates in indirect pathway neurons. A recent electrophysiological study showing that PDE10A inhibition has greater facilitatory effect on corticostriatal synaptic activity in indirect pathway neurons supports the interpretation (Threlfell et al., 2009). The biochemical features of PDE10A inhibitors resemble those of antipsychotic drugs, which act primarily as D 2 receptor antagonists and increase DARPP-32 phosphorylation in indirect pathway neurons . In agreement, PDE10A inhibition by papaverine, TP-10 and MP-10 and genetic deletion of PDE10A display behavioral phenotypes of antipsychotics with therapeutic potency for negative symptoms and cognitive deficits as well as positive symptoms in schizophrenics (Sano et al., 2008;Grauer et al., 2009;Nishi and Snyder, 2010).

ROLE OF PDE4 IN DOPAMINE SIGNALING
PDE4 is a cAMP-specific PDE with high affinity for cAMP (Km 1-10 μM; Beavo, 1995;Bender and Beavo, 2006). The PDE4 family is encoded by four genes (PDE4A-PDE4D), and each isoform has multiple variants (Houslay and Adams, 2003;McCahill et al., 2008). In the CNS, PDE4A, PDE4B, and PDE4D are widely distributed, but the expression of PDE4C is restricted to the olfactory bulb in rodent brain (Cherry and Davis, 1999;Perez-Torres et al., 2000). Each PDE4 variant has a modular structure consisting of a variant-specific N-terminal domain, regulatory domains termed upstream conserved region 1 (UCR1) and UCR2, and a catalytic domain. The phosphorylation of UCR1 by PKA disrupts the inhibitory interaction of UCR2 with the catalytic domain and activates PDE4 activity (McCahill et al., 2008), leading to the downregulation of cAMP/PKA signaling. The N-terminal domain and UCR1/2 interact with PDE4 variant-specific binding proteins including A-kinase anchor proteins (AKAP) and disrupted in schizophrenia 1 (DISC1), leading to the compartmentalization of cAMP signaling in cells (McCahill et al., 2008;Houslay, 2010). The involvement of PDE4B in the molecular mechanisms of schizophrenia is supported by its interaction with DISC1 (Millar et al., 2005;Clapcote et al., 2007;Murdoch et al., 2007), which is a genetic susceptibility factor for schizophrenia (Chubb et al., 2008).
Function of PDE4 has been analyzed using a selective PDE4 inhibitor, rolipram (IC 50 1 μM; Bender and Beavo, 2006). Inhibition of PDE4 by rolipram robustly increases tyrosine hydroxylase (TH) phosphorylation at Ser40 (PKA-site) at dopaminergic terminals in neostriatal slices and in vivo, leading to the enhancement of dopamine synthesis and metabolism . In addition, the inhibition of PDE4 by rolipram weakly enhances cAMP/PKA signaling in striatal neurons, leading to the phosphorylation of DARPP-32 at Thr34 in neostriatal slices . Rolipram treatment augments adenosine A 2A receptormediated phosphorylation of DARPP-32 at Thr34, but has no effect on dopamine D 1 receptor-mediated phosphorylation. However, in neostriatal slices from D 1 R-Flag/D 2 R-Myc DARPP-32 mice, rolipram induces the phosphorylation of both Flag-and Myc-tagged DARPP-32 in direct and indirect pathway neurons, respectively. Immunohistochemical analysis in D 1 R-Flag/D 2 R-Myc DARPP-32 mice revealed that PDE4B expression is higher in indirect pathway neurons than that in direct pathway neurons. These data suggest that PDE4 preferentially regulates cAMP/PKA signaling coupled to adenosine A 2A receptors in indirect pathway neurons compared to that coupled to dopamine D 1 receptors in direct pathway neurons. Activation of cAMP/PKA signaling in indirect pathway neurons elicited by the PDE4 inhibitor, rolipram, is expected to oppose dopamine D 2 receptor signaling. At the same time, rolipram stimulates dopamine synthesis, indicating that PDE4 inhibition raises dopaminergic tone in the striatum.

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www.frontiersin.org Therefore, rolipram mimics the biochemical effects of dopamine D 2 antagonists and to some extent D 1 agonists. The pharmacological profile of the PDE4 inhibitor, including positive effects on mood and cognition, supports its possible efficacy for the treatment of negative symptoms and cognitive deficits in addition to positive symptoms in schizophrenics (Siuciak, 2008). However, the studies in PDE4B knockout mice generally fail to recapitulate the antipsychotic effects of rolipram, and the discrepancy might be explained by the lack of PDE4B selectivity of rolipram and chronic compensatory mechanisms for PDE4B gene deletion (Siuciak, 2008;Nishi and Snyder, 2010).

ROLE OF PDE1B IN DOPAMINE SIGNALING
PDE1B is a dual substrate PDE with a higher affinity for cGMP (Km 2.4 μM) than for cAMP (Km 24 μM; Bender and Beavo, 2006). PDE1B is activated by Ca 2+ and calmodulin, providing a mechanism for crosstalk between Ca 2+ and cyclic nucleotide signaling. PDE1B is abundantly expressed in the striatum (Polli and Kincaid, 1994), and striatal PDE1B is localized to all DARPP-32-positive medium spiny neurons, indicating the PDE1B expression in both direct and indirect pathway neurons (A. Nishi and M. Kuroiwa, unpublished observations). Biochemical studies in PDE1B knockout mice revealed that the function of dopamine D 1 receptors to stimulate the phosphorylation of DARPP-32 and GluR1 at PKA-sites is potentiated in striatal slices from PDE1B knockout mice (Reed et al., 2002). In behavioral analysis, PDE1B knockout mice exhibited the increase in spontaneous locomotor activity and psychostimulant-and NMDA receptor antagonist-stimulated locomotor activity and the cognitive deficit in Morris water maze test (Reed et al., 2002;Ehrman et al., 2006;Siuciak et al., 2007). In spite of similar expression pattern of PDE1B with PDE10A in the striatum, the behavioral profiles of PDE1B knockout mice are pro-psychotic and completely opposite to those of PDE10A knockout mice. We hypothesize that PDE1B predominantly regulates cyclic nucleotide signaling in direct pathway neurons, whereas a predominant role of PDE10A and PDE4 in indirect pathway neurons.

CONCLUSION
D 1 receptor signaling in striatonigral/direct pathway neurons plays an essential role in motor functions (Gerfen and Surmeier, 2011) as well as reward and cognition (Bromberg-Martin et al., 2010;Cools, 2011). The alterations of D 1 receptor signaling are implicated in drug addiction (Shuto and Nishi, 2011) and l-DOPAinduced dyskinesia . Besides the importance of D 1 receptor signaling under pathophysiological conditions, dopaminergic drugs are mainly acting on D 2 receptors, and therapeutic agents that selectively modulate D 1 receptor signaling are not currently utilized. Functional roles of the canonical and non-canonical D 1 receptor signaling cascades are implicated in the same category of diseases. Further understanding of each D 1 receptor signaling cascade and its regulation is required for the development of therapeutic agents targeting D 1 receptor signaling.