WAVE1--the Wiskott-Aldrich syndrome protein (WASP)--family verprolin homologous protein 1--is a key regulator of actin-dependent morphological processes in mammals, through its ability to activate the actin-related protein (Arp2/3) complex. Here we show that WAVE1 is phosphorylated at multiple sites by cyclin-dependent kinase 5 (Cdk5) both in vitro and in intact mouse neurons. Phosphorylation of WAVE1 by Cdk5 inhibits its ability to regulate Arp2/3 complex-dependent actin polymerization. Loss of WAVE1 function in vivo or in cultured neurons results in a decrease in mature dendritic spines. Expression of a dephosphorylation-mimic mutant of WAVE1 reverses this loss of WAVE1 function in spine morphology, but expression of a phosphorylation-mimic mutant does not. Cyclic AMP (cAMP) signalling reduces phosphorylation of the Cdk5 sites in WAVE1, and increases spine density in a WAVE1-dependent manner. Our data suggest that phosphorylation/dephosphorylation of WAVE1 in neurons has an important role in the formation of the filamentous actin cytoskeleton, and thus in the regulation of dendritic spine morphology.

Dopamine receptor subtypes D1 and D2, and many other seven-transmembrane receptors including adenosine receptor A2A, are colocalized in striatum of brain. These receptors stimulate or inhibit adenylyl cyclases (ACs) to produce distinct physiological and pharmacological responses and interact with each other synergistically or antagonistically at various levels. The identity of the AC isoform that is coupled to each of these receptors, however, remains unknown. To investigate the in vivo role of the type 5 adenylyl cyclase (AC5), which is preferentially expressed in striatum, mice deficient for the AC5 gene were generated. The genetic ablation of the AC5 gene eliminated >80% of forskolin-induced AC activity and 85-90% of AC activity stimulated by either D1 or A2A receptor agonists in striatum. However, D1- or A2A-specific pharmaco-behaviors were basically preserved, whereas the signal cascade from D2 to AC was completely abolished in AC5(-/-), and motor activity of AC5(-/-) was not suppressed by treatment of cataleptic doses of the antipsychotic drugs haloperidol and sulpiride. Interestingly, both haloperidol and clozapine at low doses remarkably increased the locomotion of AC5(-/-) in the open field test that was produced in part by a common mechanism that involved the increased activation of D1 dopamine receptors. Together, these results suggest that AC5 is the principal AC integrating signals from multiple receptors including D1, D2, and A2A in striatum and the cascade involving AC5 among diverse D2 signaling pathways is essential for neuroleptic effects of antipsychotic drugs.

SUMMARY p11, through unknown mechanisms, is required for behavioral and cellular responses to selective serotonin-reuptake inhibitors (SSRIs). Here we have identified SMARCA3, a chromatin-remodeling factor, as a novel target for the p11/annexin A2 heterotetrameric complex. Determination of the crystal structure indicates that SMARCA3 peptide binds to a hydrophobic pocket in the heterotetramer. Formation of this complex increases the DNA binding affinity of SMARCA3 and its localization to the nuclear matrix fraction. In the dentate gyrus, both p11 and SMARCA3 are highly enriched in hilar mossy cells and basket cells. In response to the SSRI, fluoxetine, the expression of p11 is induced in both cell types, and the amount of the ternary complex of p11/annexin A2/SMARCA3 is increased. SSRI-induced neurogenesis and behavioral responses are abolished by constitutive knockout of SMARCA3. Our studies indicate a central role for a chromatin-remodeling factor in the SSRI/p11 signaling pathway, and suggest a novel approach to the development of improved antidepressant therapies.

Opioid drugs produce their pharmacological effects by activating inhibitory guanine nucleotide-binding regulatory protein-linked , ␦, and opioid receptors. One major effector for these receptors is adenylyl cyclase, which is inhibited upon receptor activation. However, little is known about which of the ten known forms of adenylyl cyclase are involved in mediating opioid actions. Here we show that all of the major behavioral effects of morphine, including locomotor activation, analgesia, tolerance, reward, and physical dependence and withdrawal symptoms, are attenuated in mice lacking adenylyl cyclase type 5 (AC5), a form of adenylyl cyclase that is highly enriched in striatum. Furthermore, the behavioral effects of selective or ␦ opioid receptor agonists are lost in AC5 ؊/؊ mice, whereas the behavioral effects of selective opioid receptor agonists are unaffected. These behavioral data are consistent with the observation that the ability of a or ␦ opioid receptor agonist to suppress adenylyl cyclase activity was absent in striatum of AC5 ؊/؊ mice. Together, these results establish AC5 as an important component of and ␦ opioid receptor signal transduction mechanisms in vivo and provide further support for the importance of the cAMP pathway as a critical mediator of opioid action. striatum ͉ opioid receptors ͉ analgesia ͉ addiction ͉ cAMP M orphine and most other opioid drugs are widely used clinically because of their potent analgesic effects, but this use is limited by their addiction liability. Both the analgesic and addicting actions of morphine and related drugs are initiated by their binding to , and to a lesser extent, ␦ opioid receptors (1-5). In contrast, other opioid drugs, such as U69593 and U50488H, activate opioid receptors, which generally produce distinct, and in some cases opposite, behavioral effects (6-8).Activation of all three types of opioid receptors is translated into physiological responses via coupling to inhibitory guanine nucleotide-binding regulatory protein, which then acts through several effectors, including most prominently inhibition of adenylyl cyclase, activation of G protein-linked inwardly rectifying K ϩ channels (GIRKs), and inhibition of voltage-gated Ca 2ϩ channels (2, 9). Regulation of G protein-linked inwardly rectifying K ϩ channels has been most closely related to the acute electrophysiological effects of , ␦, and opioid receptor activation of target cells (10, 11), whereas inhibition of adenylyl cyclase, and subsequent inhibition of the cAMP pathway, has been implicated mostly in longer term adaptations to repeated opiate administration (12, 13). Thus, up-regulation of adenylyl cyclase and other components of the cAMP pathway within specific regions of the central and peripheral nervous system has been shown to contribute to tolerance and dependence, and to changes in reward mechanisms, after repeated opioid administration.Despite the important role for adenylyl cyclase in mediating the actions of opioid drugs, little information is available concerning which type of th...