A novel alcohol+nicotine co-use self-administration procedure reveals sex differences and differential alteration of mesocorticolimbic TLR- and cholinergic-related neuroimmune gene expression in rats.

Although alcohol and nicotine are two of the most commonly co-used drugs with upwards of 90% of adults with an alcohol use disorder (AUD) in the US also smoking, we don't tend to study alcohol and nicotine use this way. The current studies sought to develop and assess a novel alcohol + nicotine co-access self-administration (SA) model in adult male and female Long-Evans rats. Further, both drugs are implicated in neuroimmune function, albeit in largely opposing ways. Chronic alcohol use increases neuroinflammation via toll-like receptors (TLRs) which in turn increases alcohol intake. By contrast, nicotine produces anti-inflammatory effects, in part, through the monomeric alpha7 receptor (ChRNa7). Following long-term co-access (6 months), rats reliably administered both drugs during daily sessions, however males generally responded for more alcohol and females for nicotine. This was reflected in plasma analysis with translationally relevant intake levels of both alcohol and nicotine, making it invaluable in studying the effects of co-use on behavior and CNS function. Moreover, male rats show sensitivity to alterations in alcohol concentration whereas females show sensitivity to alterations in nicotine concentration. Rats trained on this procedure also developed an anxiogenic phenotype. Finally, we assessed alterations in neuroimmune-related gene expression in the medial prefrontal cortex - prelimbic, (mPFC-PL), nucleus accumbens core (AcbC), and ventral tegmental area (VTA). In the AcbC, where α7 expression was increased and ß2 was decreased, markers of pro-inflammatory activity were decreased, despite increases in TLR gene expression suggesting that co-use with nicotine modulates inflammatory state downstream from the receptor level. By contrast, in mPFC-PL where α7 was not increased, both TLRs and downstream proinflammatory markers were increased. Taken together, these findings support that there are brain regional and sex differences with co-use of alcohol + nicotine SA and suggest that targeting nicotinic α7 may represent a novel strategy for treating alcohol + nicotine co-dependence.

Extrapyramidal Side Effects with Chronic Atypical Antipsychotic Can Be Predicted by Labeling Pattern of FosB and phosphoThr34-DARPP-32 in Nucleus Accumbens.

Extrapyramidal side effects (EPS) can be induced by neuroleptics that regulate the expression of transcription factor FosB and dopaminergic mediator DARPP-32 in the striatum. However, the long-term neurobiological changes in striatal projection neurons resulting from a cumulative dosage of typical and atypical antipsychotics are poorly understood. The present study aimed to determine the differential and long-lasting changes in FosB distribution and DARPP-32 phosphorylation in the striatum and nucleus accumbens (NAc) associated with chronic antipsychotic-induced EPS. Male C57Bl/6J mice received daily injections of Olanzapine (Olz, 15 mg/kg), Clozapine (Clz, 20 mg/kg), or Haloperidol (Hal, 1 mg/kg), for a period of 11 weeks with a 4-day withdrawal period before the last dosage. Catalepsy for detection of EPS, along with open-field and rotarod tests, were assessed as behavioral correlates of motor responses. Additionally, FosB and phosphorylated-DARPP-32 immunohistochemistry were examined in striatal regions after treatment. All antipsychotics produced catalepsy and reduced open-field exploration, such as impaired rota-rod performance after Olz and Hal. The washout period was critical for Clz-induced side effects reduction. Both Olz and Clz increased FosB in NAc Shell-region, and phosphoThr34-DARPP-32 in NAc. Only Clz reduced phosphoThr75-DARPP-32 in the dorsal striatum and showed FosB/phosphoThr34-Darpp-32-ir in the NAc Core region. This study provides evidence that atypical antipsychotics such as Olz and Clz also give rise to EPS effects frequently associated with a cumulative dosage of typical neuroleptics such as Hal. Nevertheless, FosB/phosphoThr34-Darpp-32-ir in the NAc Core region is associated with hypokinetic movements inhibition.

DARPP-32 and t-DARPP in the development of resistance to anti-HER2 agents. Pre-clinical evidence from the STEP study.

The therapeutic scenario of Human Epidermal Growth Factor Receptor 2 positive advanced breast cancer (ABC) has been recently enriched by a number of innovative agents, which are reshaping treatment sequence. While randomized trials have documented an advantage in terms of efficacy, for the newly available agents we lack effectiveness and tolerability evidence from the real-world setting. Similarly, the identification of predictive biomarkers might improve clinical decision. We herein describe the outline of a prospective/retrospective study which aims to explore the optimal sequence of treatment in HER2+, pertuzumab pre-treated ABC patients treated in II line with anti-HER2 agents in clinical practice. As part of the pre-clinical tasks envisioned by the STEP study, in vitro cell models of resistance were exploited to investigate molecular features associated with reduced efficacy of HER2 targeting agents at the transcript level. The aggressive behavior of resistant cell populations was measured by growth assessment in mouse models. This approach led to the identification of DARPP-32 and t-DARPP proteins as possible predictive biomarkers of efficacy of anti-HER2 agents. Biomarkers validation and the clinical goals will be reached through patients' inclusion into two independent cohorts, i.e., the prospective and retrospective cohorts, whose setup is currently ongoing.

The involvement of ADAR1 in chronic unpredictable stress-induced cognitive impairment by targeting DARPP-32 with miR-874-3p in BALB/c mice.

Introduction: Chronic stress exposure is the main environmental factor leading to cognitive impairment, but the detailed molecular mechanism is still unclear. Adenosine Deaminase acting on double-stranded RNA1(ADAR1) is involved in the occurrence of chronic stress-induced cognitive impairment. In addition, dopamine and Adenosine 3'5'-monophosphate-regulated phospho-protein (DARPP-32) gene variation affects cognitive function. Therefore, we hypothesized that ADAR1 plays a key role in chronic stress-induced cognitive impairment by acting on DARPP-32. Methods: In this study, postnatal 21-day-old male BALB/c mice were exposed to chronic unpredictable stressors. After that, the mice were treated with ADAR1 inducer/inhibitor. The cognitive ability and cerebral DARPP-32 protein expression of BALB/c mice were evaluated. In order to explore the link between ADAR1 and DARPP-32, the effects of ADAR1 high/low expression on DARPP-32 protein expression in vitro were detected. Results: ADAR1 inducer alleviates cognitive impairment and recovers decreased DARPP-32 protein expression of the hippocampus and prefrontal cortex in BALB/c mice with chronic unpredictable stress exposure. In vivo and in vitro studies confirm the results predicted by bio-informatics; that is, ADAR1 affects DARPP-32 expression via miR-874-3p. Discussion: The results in this study demonstrate that ADAR1 affects the expression of DARPP-32 via miR-874-3p, which is involved in the molecular mechanism of pathogenesis in chronic unpredictable stress-induced cognitive impairment. The new findings of this study provide a new therapeutic strategy for the prevention and treatment of stress cognitive impairment from epigenetics.

Ferulic Acid Improves Synaptic Plasticity and Cognitive Impairments by Alleviating the PP2B/DARPP-32/PP1 Axis-Mediated STEP Increase and Aß Burden in Alzheimer's Disease.

The burden of Alzheimer's disease, the most prevalent neurodegenerative disease, is increasing exponentially due to the increase in the elderly population worldwide. Synaptic plasticity is the basis of learning and memory, but it is impaired in AD. Uncovering the disease's underlying molecular pathogenic mechanisms involving synaptic plasticity could lead to the identification of targets for better disease management. Using primary neurons treated with Aß and APP/PS1 animal models, we evaluated the effect of the phenolic compound ferulic acid (FA) on synaptic dysregulations. Aß led to synaptic plasticity and cognitive impairments by increasing STEP activity and decreasing the phosphorylation of the GluN2B subunit of NMDA receptors, as well as decreasing other synaptic proteins, including PSD-95 and synapsin1. Interestingly, FA attenuated the Aß-upregulated intracellular calcium and thus resulted in a decrease in PP2B-induced activation of DARPP-32, inhibiting PP1. This cascade event maintained STEP in its inactive state, thereby preventing the loss of GluN2B phosphorylation. This was accompanied by an increase in PSD-95 and synapsin1, improved LTP, and a decreased Aß load, together leading to improved behavioral and cognitive functions in APP/PS1 mice treated with FA. This study provides insight into the potential use of FA as a therapeutic strategy in AD.

DARPP-32 cells and neuropil define striosomal system and isolated matrix cells in human striatum.

The dorsal striatum forms a central node of the basal ganglia interconnecting the neocortex and thalamus with circuits modulating mood and movement. Striatal projection neurons (SPNs) include relatively intermixed populations expressing D1-type or D2-type dopamine receptors (dSPNs and iSPNs) that give rise to the direct (D1) and indirect (D2) output systems of the basal ganglia. Overlaid on this organization is a compartmental organization, in which a labyrinthine system of striosomes made up of sequestered SPNs is embedded within the larger striatal matrix. Striosomal SPNs also include D1-SPNs and D2-SPNs, but they can be distinguished from matrix SPNs by many neurochemical markers. In the rodent striatum the key signaling molecule, DARPP-32, is a exception to these compartmental expression patterns, thought to befit its functions through opposite actions in both D1- and D2-expressing SPNs. We demonstrate here, however, that in the dorsal human striatum, DARPP-32 is concentrated in the neuropil and SPNs of striosomes, especially in the caudate nucleus and dorsomedial putamen, relative to the matrix neuropil in these regions. The generally DARPP-32-poor matrix contains scattered DARPP-32-positive cells. DARPP-32 cell bodies in both compartments proved negative for conventional intraneuronal markers. These findings raise the potential for specialized DARPP-32 expression in the human striosomal system and in a set of DARPP-32-positive neurons in the matrix. If DARPP-32 immunohistochemical positivity predicts differential functional DARPP-32 activity, then the distributions demonstrated here could render striosomes and dispersed matrix cells susceptible to differential signaling through cAMP and other signaling systems in health and disease. DARPP-32 is highly concentrated in cells and neuropil of striosomes in post-mortem human brain tissue, particularly in the dorsal caudate nucleus. Scattered DARPP-32-positive cells are found in the human striatal matrix. Calbindin and DARPP-32 do not colocalize within every spiny projection neuron in the dorsal human caudate nucleus.

p-CREB and p-DARPP-32 orchestrating the modulatory role of cAMP/PKA signaling pathway enhanced by Roflumilast in rotenone-induced Parkinson's disease in rats.

Recently, phosphodiesterases (PDEs) have gained great attention due to their implication in Parkinson's disease (PD) pathogenesis. Noteworthy, the PDE4 enzyme is highly expressed in the striatum and selectively degrades cyclic adenosine monophosphate (cAMP). The cAMP was shown to play a vital role in dopamine (DA) signaling besides maintaining the plasticity of dopaminergic neurons as well as protecting them from inflammation and oxidative stress-mediated death. Thus, PDE4 inhibition could be a promising strategy for treating PD. Accordingly, the present study investigated the neuroprotective efficacy of roflumilast, a PDE4 inhibitor, in abolishing neurodegeneration in the rotenone-induced PD model. Rotenone (1.5 mg/kg, s.c) was delivered via 11 injections on matching days. Roflumilast treatment (0.5 mg/kg, p.o) was given daily after the fifth rotenone injection. Roflumilast significantly reversed rotenone's adverse effects, as it enhanced trophic factors expression and abrogated inflammation as well as oxidative stress. Thus, promoting dopaminergic neuronal plasticity and survival, as well as restoring striatal DA level and function, which resulted in enhanced motor performance. The beneficial effect of roflumilast was mediated through inhibition of striatal PDE4 with consequent activation of cAMP-dependent protein kinase A (PKA) signaling pathways, including the cAMP response element-binding protein (CREB) pathway and dopamine and cAMP-regulated phosphoprotein 32,000 (DARPP-32) pathway that is essential for maintaining dopaminergic function. Therefore, the present work sheds light on the substantial neuroprotective potential of roflumilast in treating PD through the activation of the cAMP/PKA cascade.

DARPP-32/protein phosphatase 1 regulates Rasgrp2 as a novel component of dopamine D1 receptor signaling in striatum.

Dopamine regulates psychomotor function by D1 receptor/PKA-dependent phosphorylation of DARPP-32. DARPP-32, phosphorylated at Thr34 by PKA, inhibits protein phosphatase 1 (PP1), and amplifies the phosphorylation of other PKA/PP1 substrates following D1 receptor activation. In addition to the D1 receptor/PKA/DARPP-32 signaling pathway, D1 receptor stimulation is known to activate Rap1/ERK signaling. Rap1 activation is mediated through the phosphorylation of Rasgrp2 (guanine nucleotide exchange factor; activation) and Rap1gap (GTPase-activating protein; inhibition) by PKA. In this study, we investigated the role of PP1 inhibition by phospho-Thr34 DARPP-32 in the D1 receptor-induced phosphorylation of Rasgrp2 and Rap1gap at PKA sites. The analyses in striatal and NAc slices from wild-type and DARPP-32 knockout mice revealed that the phosphorylation of Rasgrp2 at Ser116/Ser117 and Ser586, but not of Rasgrp2 at Ser554 or Rap1gap at Ser441 or Ser499 induced by a D1 receptor agonist, is under the control of the DARPP-32/PP1. The results were supported by pharmacological analyses using a selective PP1 inhibitor, tautomycetin. In addition, analyses using a PP1 and PP2A inhibitor, okadaic acid, revealed that all sites of Rasgrp2 and Rap1gap were regulated by PP2A. Thus, the interactive machinery of DARPP-32/PP1 may contribute to efficient D1 receptor signaling via Rasgrp2/Rap1 in the striatum.

Lasting mesothalamic dopamine imbalance and altered exploratory behavior in rats after a mild neonatal hypoxic event.

Introduction: Adversities during the perinatal period can decrease oxygen supply to the fetal brain, leading to various hypoxic brain injuries, which can compromise the regularity of brain development in different aspects. To examine the catecholaminergic contribution to the link between an early-life hypoxic insult and adolescent behavioral aberrations, we used a previously established rat model of perinatal hypoxia but altered the hypobaric to normobaric conditions. Methods: Exploratory and social behavior and learning abilities were tested in 70 rats of both sexes at adolescent age. Inherent vertical locomotion, sensory-motor functions and spatial learning abilities were explored in a subset of animals to clarify the background of altered exploratory behavior. Finally, the concentrations of dopamine (DA) and noradrenaline in midbrain and pons, and the relative expression of genes for DA receptors D1 and D2, and their down-stream targets (DA- and cAMP-regulated phosphoprotein, Mr 32 kDa, the regulatory subunit of protein kinase A, and inhibitor-5 of protein phosphatase 1) in the hippocampus and thalamus were investigated in 31 rats. Results: A lesser extent of alterations in exploratory and cognitive aspects of behavior in the present study suggests that normobaric conditions mitigate the hypoxic injury compared to the one obtained under hypobaric conditions. Increased exploratory rearing was the most prominent consequence, with impaired spatial learning in the background. In affected rats, increased midbrain/pons DA content, as well as mRNA levels for DA receptors and their down-stream elements in the thalamus, but not the hippocampus, were found. Conclusion: We can conclude that a mild hypoxic event induced long-lasting disbalances in mesothalamic DA signaling, contributing to the observed behavioral alterations. The thalamus was thereby indicated as another structure, besides the well-established striatum, involved in mediating hypoxic effects on behavior through DA signaling.

Comparison of rule- and ordinary differential equation-based dynamic model of DARPP-32 signalling network.

Dynamic modelling has considerably improved our understanding of complex molecular mechanisms. Ordinary differential equations (ODEs) are the most detailed and popular approach to modelling the dynamics of molecular systems. However, their application in signalling networks, characterised by multi-state molecular complexes, can be prohibitive. Contemporary modelling methods, such as rule- based (RB) modelling, have addressed these issues. The advantages of RB modelling over ODEs have been presented and discussed in numerous reviews. In this study, we conduct a direct comparison of the time courses of a molecular system founded on the same reaction network but encoded in the two frameworks. To make such a comparison, a set of reactions that underlie an ODE model was manually encoded in the Kappa language, one of the RB implementations. A comparison of the models was performed at the level of model specification and dynamics, acquired through model simulations. In line with previous reports, we confirm that the Kappa model recapitulates the general dynamics of its ODE counterpart with minor differences. These occur when molecules have multiple sites binding the same interactor. Furthermore, activation of these molecules in the RB model is slower than in the ODE one. As reported for other molecular systems, we find that, also for the DARPP-32 reaction network, the RB representation offers a more expressive and flexible syntax that facilitates access to fine details of the model, easing model reuse. In parallel with these analyses, we report a refactored model of the DARPP-32 interaction network that can serve as a canvas for the development of more complex dynamic models to study this important molecular system.

Cellular Profiles of Prodynorphin and Preproenkephalin mRNA-Expressing Neurons in the Anterior Olfactory Tubercle of Mice.

The olfactory tubercle (OT) is a striatal region that receives olfactory inputs. mRNAs of prodynorphin (Pdyn) and preproenkephalin (Penk), precursors of dynorphins and enkephalins, respectively, are strongly expressed in the striatum. Both produce opioid peptides with various physiological effects such as pain relief and euphoria. Recent studies have revealed that OT has anatomical and cytoarchitectonic domains that play different roles in odor-induced motivated behavior. Neuronal subtypes of the OT can be distinguished by their expression of the dopamine receptors D1 (Drd1) and D2 (Drd2). Here, we addressed whether and which type of opioid peptide precursors the D1- and D2-expressing neurons in the OT express. We used multiple fluorescence in situ hybridization for mRNAs of the opioid precursors and dopamine receptors to characterize mouse OT neurons. Pdyn was mainly expressed by Drd1-expressing cells in the dense cell layer (DCL) of the OT, whereas Penk was expressed primarily by Drd2-expressing cells in the DCL. We also confirmed the presence of a larger population of Pdyn-Penk-Drd1 co-expressing cells in the DCL of the anteromedial OT compared with the anterolateral OT. These observations will help understand whether and how dynorphins and enkephalins in the OT are involved in diverse odor-induced motivated behaviors.

Neuron types in the primate striatum: Stereological analysis of projection neurons and interneurons in control and parkinsonian monkeys.

AIMS: The striatum is mainly composed of projection neurons. It also contains interneurons, which modulate and control striatal output. The aim of the present study was to assess the percentages of projection neurons and interneuron populations in the striatum of control monkeys and of parkinsonian monkeys. METHODS: Unbiased stereology was used to estimate the volume density of every neuron population in the caudate, putamen and ventral striatum of control monkeys and of monkeys treated with MPTP, which results in striatal dopamine depletion. The various neuron population phenotypes were identified by immunohistochemistry. All analyses were performed within the same subjects using similar processing and analysis parameters, thus allowing for reliable data comparisons. RESULTS: In control monkeys, the projection neurons, which express the dopamine-and-cAMP-regulated-phosphoprotein, 32-KDa (DARPP-32), were the most abundant: ~86% of the total neurons counted. The interneurons accounted for the remaining 14%. Among the interneurons, those expressing calretinin were the most abundant (Cr+: ~57%; ~8% of the total striatal neurons counted), followed those expressing Parvalbumin (Pv+: ~18%; 2.6%), dinucleotide phosphate-diaphorase (NADPH+: ~13%; 1.8%), choline acetyltransferase (ChAT+: ~11%; 1.5%) and tyrosine hydroxylase (TH+: ~0.5%; 0.1%). No significant changes in volume densities occurred in any population following dopamine depletion, except for the TH+ interneurons, which increased in parkinsonian non-symptomatic monkeys and even more in symptomatic monkeys. CONCLUSIONS: These data are relevant for translational studies targeting specific neuron populations of the striatum. The fact that dopaminergic denervation does not cause neuron loss in any population has potential pathophysiological implications.

A Homozygous Missense Variant in PPP1R1B/DARPP-32 Is Associated With Generalized Complex Dystonia.

BACKGROUND: The dystonias are a heterogeneous group of hyperkinetic disorders characterized by sustained or intermittent muscle contractions that cause abnormal movements and/or postures. Although more than 200 causal genes are known, many cases of primary dystonia have no clear genetic cause. OBJECTIVES: To identify the causal gene in a consanguineous family with three siblings affected by a complex persistent generalized dystonia, generalized epilepsy, and mild intellectual disability. METHODS: We performed exome sequencing in the parents and two affected siblings and characterized the expression of the identified gene by immunohistochemistry in control human and zebrafish brains. RESULTS: We identified a novel missense variant (c.142G>A (NM_032192); p.Glu48Lys) in the protein phosphatase 1 regulatory inhibitor subunit 1B gene (PPP1R1B) that was homozygous in all three siblings and heterozygous in the parents. This gene is also known as dopamine and cAMP-regulated neuronal phosphoprotein 32 (DARPP-32) and has been involved in the pathophysiology of abnormal movements. The uncovered variant is absent in public databases and modifies the conserved glutamate 48 localized close to the serine 45 phosphorylation site. The PPP1R1B protein was shown to be expressed in cells and regions involved in movement control, including projection neurons of the caudate-putamen, substantia nigra neuropil, and cerebellar Purkinje cells. The latter cells were also confirmed to be positive for PPP1R1B expression in the zebrafish brain. CONCLUSIONS: We report the association of a PPP1R1B/DARPP-32 variant with generalized dystonia in man. It might be relevant to include the sequencing of this new gene in the diagnosis of patients with otherwise unexplained movement disorders. © 2021 International Parkinson and Movement Disorder Society.

Carnosic Acid Alleviates Levodopa-Induced Dyskinesia and Cell Death in 6-Hydroxydopamine-lesioned Rats and in SH-SY5Y Cells.

The present study investigated the impact of carnosic acid (CA) from rosemary on the levodopa (L-dopa)-induced dyskinesia (LID) in rats treated with 6-hydroxydopamine (6-OHDA). To establish the model of LID, 6-OHDA-lesioned rats were injected intraperitoneally with 30 mg/kg L-dopa once a day for 36 days. Rats were daily administrated with 3 or 15 mg/kg CA by oral intubation prior to L-dopa injection for 4 days. Rats pretreated with CA had reduced L-dopa-induced abnormal involuntary movements (AIMs) and ALO scores (a sum of axial, limb, and orofacial scores). Moreover, the increases of dopamine D1-receptor, p-DARPP-32, ΔFosB, p-ERK1/2, and p-c-Jun ser63, along with the decrease in p-c-Jun ser73, induced by L-dopa in 6-OHDA-treated rats were significantly reversed by pretreatment with CA. In addition, we used the model of SH-SY5Y cells to further examine the neuroprotective mechanisms of CA on L-dopa-induced cytotoxicity. SH-SY5Y cells were treated with CA for 18 h, and then co-treated with 400 µM L-dopa for the indicated time points. The results showed that pretreatment of CA attenuated the cell death and nuclear condensation induced by L-dopa. By the immunoblots, the reduction of Bcl-2, p-c-Jun ser73, and parkin and the induction of cleaved caspase 3, cleaved Poly (ADP-ribose) polymerase, p-ERK1/2, p-c-Jun ser63, and ubiquitinated protein by L-dopa were improved in cells pretreated with CA. In conclusion, CA ameliorates the development of LID via regulating the D1R signaling and prevents L-dopa-induced apoptotic cell death through modulating the ERK1/2-c-Jun and inducing the parkin. This study suggested that CA can be used to alleviate the adverse effects of LID for PD patients.

Subregion-Specific Regulation of Dopamine D1 Receptor Signaling in the Striatum: Implication for L-DOPA-Induced Dyskinesia.

The striatum is the main structure of the basal ganglia. The striatum receives inputs from various cortical areas, and its subregions play distinct roles in motor and emotional functions. Recently, striatal maps based on corticostriatal connectivity and striosome-matrix compartmentalization were developed, and we were able to subdivide the striatum into seven subregions. Dopaminergic modulation of the excitability of medium spiny neurons (MSNs) is critical for striatal function. In this study, we investigated the functional properties of dopamine signaling in seven subregions of the striatum from male mice. By monitoring the phosphorylation of PKA substrates including DARPP-32 in mouse striatal slices, we identified two subregions with low D1 receptor signaling: the dorsolateral portion of the intermediate/rostral part (DL-IR) and the intermediate/caudal part (IC). Low D1 receptor signaling in the two subregions was maintained by phosphodiesterase (PDE)10A and muscarinic M4 receptors. In an animal model of 6-hydroxydopamine (6-OHDA)-induced hemi-parkinsonism, D1 receptor signaling was upregulated in almost all subregions including the DL-IR, but not in the IC. When L-DOPA-induced dyskinesia (LID) was developed, D1 receptor signaling in the IC was upregulated and correlated with the severity of LID. Our results suggest that the function of the striatum is maintained through the subregion-specific regulation of dopamine D1 receptor signaling and that the aberrant activation of D1 receptor signaling in the IC is involved in LID. Future studies focusing on D1 receptor signaling in the IC of the striatum will facilitate the development of novel therapeutics for LID.SIGNIFICANCE STATEMENT Recent progress in striatal mapping based on corticostriatal connectivity and striosome-matrix compartmentalization allowed us to subdivide the striatum into seven subregions. Analyses of D1 receptor signaling in the seven subregions identified two unique subregions with low D1 receptor signaling: the dorsolateral portion of the intermediate/rostral part (DL-IR) and the intermediate/caudal part (IC). Aberrant activation of D1 receptor signaling in the IC is involved in L-DOPA-induced dyskinesia (LID). Previous studies of LID have mainly focused on the DL-IR, but not on the IC of the striatum. Future studies to clarify aberrant D1 receptor signaling in the IC are required to develop novel therapeutics for LID.

PP1, PKA and DARPP-32 in breast cancer: A retrospective assessment of protein and mRNA expression.

Cyclic AMP-dependent protein kinase A (PKA) and protein phosphatase 1 (PP1) are proteins involved in numerous essential signalling pathways that modulate physiological and pathological functions. Both PP1 and PKA can be inhibited by dopamine- and cAMP-regulated phosphoprotein 32 kD (DARPP-32). Using immunohistochemistry, PKA and PP1 expression was determined in a large primary breast tumour cohort to evaluate associations between clinical outcome and clinicopathological criteria (n > 1100). In addition, mRNA expression of PKA and PP1 subunits was assessed in the METABRIC data set (n = 1980). Low protein expression of PKA was significantly associated with adverse survival of breast cancer patients; interestingly, this relationship was stronger in ER-positive breast cancer patients. PP1 protein expression was not associated with patient survival. PKA and PP1 subunit mRNA was also assessed; PPP1CA, PRKACG and PRKAR1B were associated with breast cancer-specific survival. In patients with high expression of DARPP-32, low expression of PP1 was associated with adverse survival when compared to high expression in the same group. PKA expression and PP1 expression are of significant interest in cancer as they are involved in a wide array of cellular processes, and these data indicate PKA and PP1 may play an important role in patient outcome.

cAMP-regulated phosphoproteins DARPP-32, ARPP16/19, and RCS modulate striatal signal transduction through protein kinases and phosphatases.

Decades of research led by Paul Greengard identified protein phosphorylation as a ubiquitous and vital post-translational modification involved in many neuronal signaling pathways. In particular, his discovery that second messenger-regulated protein phosphorylation plays a central role in the propagation and transduction of signals in the nervous system has been essential in understanding the molecular mechanisms of neuronal communication. The establishment of dopamine (DA) as an essential neurotransmitter in the central nervous system, combined with observations that DA activates G-protein-coupled receptors to control the production of cyclic adenosine monophosphate (cAMP) in postsynaptic neurons, has provided fundamental insight into the regulation of neurotransmission. Notably, DA signaling in the striatum is involved in many neurological functions such as control of locomotion, reward, addiction, and learning, among others. This review focuses on the history, characterization, and function of cAMP-mediated regulation of serine/threonine protein phosphatases and their role in DA-mediated signaling in striatal neurons. Several small, heat- and acid-stable proteins, including DARPP-32, RCS, and ARPP-16/19, were discovered by the Greengard laboratory to be regulated by DA- and cAMP signaling, and found to undergo a complex but coordinated sequence of phosphorylation and dephosphorylation events. These studies have contributed significantly to the establishment of protein phosphorylation as a ubiquitous and vital process in signal propagation in neurons, paradigm shifting discoveries at the time. Understanding DA-mediated signaling in the context of signal propagation has led to numerous insights into human conditions and the development of treatments and therapies.

Effects of nicotine on DARPP-32 and CaMKII signaling relevant to addiction.

Paul Greengard brought to neuroscience the idea of, and evidence for, the role of second messenger systems in neuronal signaling. The fundamental nature of his contributions is evident in the far reach of his work, relevant to various subfields and topics in neuroscience. In this review, we discuss some of Greengard's work from the perspective of nicotinic acetylcholine receptors and their relevance to nicotine addiction. Specifically, we review the roles of dopamine- and cAMP-regulated phospho-protein of 32kDa (DARPP-32) and Ca2+/calmodulin-dependent kinase II (CaMKII) in nicotine-dependent behaviors. For each protein, we discuss the historical context of their discovery and initial characterization, focusing on the extensive biochemical and immunohistochemical work conducted by Greengard and colleagues. We then briefly summarize contemporary understanding of each protein in key intracellular signaling cascades and evidence for the role of each protein with respect to systems and behaviors relevant to nicotine addiction.

Modeling the activity of the dopamine signaling pathway by combination of analog electrical circuit and mathematical approaches.

This paper demonstrates the application of the system biology principles on the example of the dopamine signaling pathway in neurons. Presented model is based on two approaches - cytomorphic electronic circuits and mathematical modeling. Transcription and phosphorylation of DARPP-32 was modeled by analog circuit, based on well-known approaches presented in [1]. It was shown that application of circuit helps to receive signal oscillations that close to described ones in real biological systems. This combination on the one hand gives possibility to simplify calculations, on another to show this signaling pathway dynamics. The expected effect of changes in the functioning of calcium channels is considered, and the mathematical model of the interaction of system components is proposed. The average frequency of calcium current oscillations due to the presence of dopamine was 30 Hz in presented model, that is consistent with the literature, where the frequency of such oscillations is up to several tens of Hz. All presented results shows good correlation with known data, which already published today.

Methylglyoxal-Mediated Dopamine Depletion, Working Memory Deficit, and Depression-Like Behavior Are Prevented by a Dopamine/Noradrenaline Reuptake Inhibitor.

Methylglyoxal (MGO) is an endogenous toxin, mainly produced as a by-product of glycolysis that has been associated to aging, Alzheimer's disease, and inflammation. Cell culture studies reported that MGO could impair the glyoxalase, thioredoxin, and glutathione systems. Thus, we investigated the effect of in vivo MGO administration on these systems, but no major changes were observed in the glyoxalase, thioredoxin, and glutathione systems, as evaluated in the prefrontal cortex and the hippocampus of mice. A previous study from our group indicated that MGO administration produced learning/memory deficits and depression-like behavior. Confirming these findings, the tail suspension test indicated that MGO treatment for 7 days leads to depression-like behavior in three different mice strains. MGO treatment for 12 days induced working memory impairment, as evaluated in the Y maze spontaneous alternation test, which was paralleled by low dopamine and serotonin levels in the cerebral cortex. Increased DARPP32 Thr75/Thr34 phosphorylation ratio was observed, suggesting a suppression of phosphatase 1 inhibition, which may be involved in behavioral responses to MGO. Co-treatment with a dopamine/noradrenaline reuptake inhibitor (bupropion, 10 mg/kg, p.o.) reversed the depression-like behavior and working memory impairment and restored the serotonin and dopamine levels in the cerebral cortex. Overall, the cerebral cortex monoaminergic system appears to be a preferential target of MGO toxicity, a new potential therapeutic target that remains to be addressed.