Gilberto Fisone

Loss of bidirectional striatal synaptic plasticity in L-DOPA–induced dyskinesia

Long-term treatment with the dopamine precursor levodopa (L-DOPA) induces dyskinesia in Parkinsons disease (PD) patients. We divided hemiparkinsonian rats treated chronically with L-DOPA into two groups: one showed motor improvement without dyskinesia, and the other developed debilitating dyskinesias in response to the treatment. We then compared the plasticity of corticostriatal synapses between the two groups. High-frequency stimulation of cortical afferents induced long-term potentiation (LTP) of corticostriatal synapses in both groups of animals. Control and non-dyskinetic rats showed synaptic depotentiation in response to subsequent low-frequency synaptic stimulation, but dyskinetic rats did not. The depotentiation seen in both L-DOPA–treated non-dyskinetic rats and intact controls was prevented by activation of the D1 subclass of dopamine receptors or inhibition of protein phosphatases. The striata of dyskinetic rats contained abnormally high levels of phospho[Thr34]-DARPP-32, an inhibitor of protein phosphatase 1. These results indicate that abnormal information storage in corticostriatal synapses is linked with the development of L-DOPA–induced dyskinesia.

Critical Involvement of cAMP/DARPP-32 and Extracellular Signal-Regulated Protein Kinase Signaling in l-DOPA-Induced Dyskinesia

The molecular basis of l-3,4-dihydroxyphenylalanine (l-DOPA)-induced dyskinesia (LID), one of the major hindrances in the current therapy for Parkinsons disease, is still unclear. We show that attenuation of cAMP signaling in the medium spiny neurons of the striatum, achieved by genetic inactivation of the dopamine and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32), reduces LID. We also show that, in dyskinetic mice, sensitized cAMP/cAMP-dependent protein kinase/DARPP-32 signaling leads to phosphorylation/activation of the extracellular signal-regulated protein kinases 1 and 2 (ERK1/2). The increase in ERK1/2 phosphorylation associated with dyskinesia results in activation of mitogen- and stress-activated kinase-1 (MSK-1) and phosphorylation of histone H3, two downstream targets of ERK involved in transcriptional regulation. In line with these observations, we found that c-Fos expression is abnormally elevated in the striata of mice affected by LID. Persistent enhancement of the ERK signaling cascade is implicated in the generation of LID. Thus, pharmacological inactivation of ERK1/2 achieved using SL327 (α-[amino[(4-aminophenyl)thio]methylene]-2-(trifluoromethyl)benzeneacetonitrile), an inhibitor of the mitogen-activated kinase/ERK kinase, MEK, during chronic l-DOPA treatment counteracts the induction dyskinesia. Together, these results indicate that a significant proportion of the abnormal involuntary movements developed in response to chronic l-DOPA are attributable to hyperactivation in striatal medium spiny neurons of a signaling pathway including sequential phosphorylation of DARPP-32, ERK1/2, MSK-1, and histone H3.

Increased D1 dopamine receptor signaling in levodopa-induced dyskinesia

Involuntary movements, or dyskinesia, represent a debilitating complication of levodopa therapy for Parkinsons disease. Although changes affecting D1 and D2 dopamine receptors have been studied in association with this condition, no causal relationship has yet been established. Taking advantage of a monkey brain bank constituted to study levodopa‐induced dyskinesia, we report changes affecting D1 and D2 dopamine receptors within the striatum of normal, parkinsonian, nondyskinetic levodopa‐treated parkinsonian, and dyskinetic levodopa‐treated parkinsonian animals. Whereas D1 receptor expression itself is not related to dyskinesia, D1 sensitivity per D1 receptor measured by D1 agonist‐induced [35S]GTPγS binding is linearly related to dyskinesia. Moreover, the striata of dyskinetic animals show higher levels of cyclin‐dependent kinase 5 (Cdk5) and of the dopamine‐ and cAMP‐regulated phosphoprotein of 32kDa (DARPP‐32). Our data suggest that levodopa‐induced dyskinesia results from increased dopamine D1 receptor–mediated transmission at the level of the direct pathway. Ann Neurol 2004

Caffeine as a psychomotor stimulant: mechanism of action

The popularity of caffeine as a psychoactive drug is due to its stimulant properties, which depend on its ability to reduce adenosine transmission in the brain. Adenosine A1 and A2A receptors are expressed in the basal ganglia, a group of structures involved in various aspects of motor control. Caffeine acts as an antagonist to both types of receptors. Increasing evidence indicates that the psychomotor stimulant effect of caffeine is generated by affecting a particular group of projection neurons located in the striatum, the main receiving area of the basal ganglia. These cells express high levels of adenosine A2A receptors, which are involved in various intracellular processes, including the expression of immediate early genes and regulation of the dopamine- and cyclic AMP-regulated 32-kDa phosphoprotein DARPP-32. The present review focuses on the effects of caffeine on striatal signal transduction and on their involvement in caffeine-mediated motor stimulation.

Distinct subclasses of medium spiny neurons differentially regulate striatal motor behaviors

The direct and indirect pathways of the basal ganglia have been proposed to oppositely regulate locomotion and differentially contribute to pathological behaviors. Analysis of the distinct contributions of each pathway to behavior has been a challenge, however, due to the difficulty of selectively investigating the neurons comprising the two pathways using conventional techniques. Here we present two mouse models in which the function of striatonigral or striatopallidal neurons is selectively disrupted due to cell type–specific deletion of the striatal signaling protein dopamine- and cAMP-regulated phosphoprotein Mr 32kDa (DARPP-32). Using these mice, we found that the loss of DARPP-32 in striatonigral neurons decreased basal and cocaine-induced locomotion and abolished dyskinetic behaviors in response to the Parkinsons disease drug L-DOPA. Conversely, the loss of DARPP-32 in striatopallidal neurons produced a robust increase in locomotor activity and a strongly reduced cataleptic response to the antipsychotic drug haloperidol. These findings provide insight into the selective contributions of the direct and indirect pathways to striatal motor behaviors.

Galanin and galanin antagonists: molecular and biochemical perspectives

The neuropeptide galanin potently inhibits insulin release, hippocampal acetylcholine release and firing of locus coeruleus cells, and stimulates feeding and release of growth hormone. Galanin regulates K+ channels, adenylyl cyclase and phospholipase C by acting at Gi/Go protein-coupled high-affinity receptors. Galanin receptor agonists such as the N-terminal fragment galanin1-16 act synergistically with morphine in the somatosensory system and have potential analgetic application. Galanin antagonists may be useful therapeutic agents in endocrinology, neurology and psychiatry. The enhancing effect of such agents on hippocampal cholinergic function would be useful in treatment of Alzheimers disease. Recent synthesis of a series of high-affinity galanin antagonists, reviewed, along with galanins actions, by Tamas Bartfai and colleagues, opens the possibility of examining the functions of endogenous galanin and test the pharmacological usefulness of antagonism of galanin function in the endocrine, somatosensory and central nervous systems.

Inhibition of mTOR Signaling in Parkinson’s Disease Prevents l-DOPA–Induced Dyskinesia

Dyskinetic side effects of a Parkinson’s disease medication may involve dopamine D1 receptor–mediated activation of mTORC1. Dyskinesia Relief In its role as a regulator of cell growth, the mammalian complex of rapamycin (mTOR) phosphorylates several proteins involved in protein synthesis, such as 4E-BP (eukaryotic initiation factor 4E binding protein) and S6K (p70 S6 kinase), in response to growth factors and nutrient availability. Santini et al. show that l-DOPA, the most commonly used medication to alleviate the immobility and rigidity (akinesia) characteristic of Parkinson’s disease (PD), also stimulates the rapamycin-sensitive mTOR complex 1 (mTORC1). In a mouse model of PD, l-DOPA treatment increased phosphorylation of several direct and indirect mTOR targets, including S6K, its substrate ribosomal protein S6 (S6), 4E-BP, and eukaryotic initiation factor 4E (eIF4E). These phosphorylation increases required the activity of dopamine D1 receptors and extracellular signal–regulated kinase (ERK). Furthermore, increased phosphorylation of S6K, S6, 4E-BP, and eIF4E correlated with stronger abnormal involuntary movements (AIMs), a measure of dyskinesia (a side effect of l-DOPA that limits its clinical use). Administration of rapamycin, which predominantly inhibits mTORC1, decreased the severity of AIMs without affecting the ability of l-DOPA to reduce akinesia. Thus, the mTORC1 signaling pathway could be targeted in PD patients suffering from the dyskinesia associated with l-DOPA treatment. Parkinson’s disease (PD), a disorder caused by degeneration of the dopaminergic input to the basal ganglia, is commonly treated with l-DOPA. Use of this drug, however, is severely limited by motor side effects, or dyskinesia. We show that administration of l-DOPA in a mouse model of Parkinsonism led to dopamine D1 receptor–mediated activation of the mammalian target of rapamycin (mTOR) complex 1 (mTORC1), which is implicated in several forms of synaptic plasticity. This response occurred selectively in the GABAergic medium spiny neurons that project directly from the striatum to the output structures of the basal ganglia. The l-DOPA–mediated activation of mTORC1 persisted in mice that developed dyskinesia. Moreover, the mTORC1 inhibitor rapamycin prevented the development of dyskinesia without affecting the therapeutic efficacy of l-DOPA. Thus, the mTORC1 signaling cascade represents a promising target for the design of anti-Parkinsonian therapies.

Involvement of DARPP-32 phosphorylation in the stimulant action of caffeine

Caffeine has been imbibed since ancient times in tea and coffee, and more recently in colas. Caffeine owes its psychostimulant action to a blockade of adenosine A2A receptors, but little is known about its intracellular mechanism of action. Here we show that the stimulatory effect of caffeine on motor activity in mice was greatly reduced following genetic deletion of DARPP-32 (dopamine- and cyclic AMP-regulated phosphoprotein of relative molecular mass 32,000). Results virtually identical to those seen with caffeine were obtained with the selective A2A antagonist SCH 58261. The depressant effect of the A2A receptor agonist, CGS 21680, on motor activity was also greatly attenuated in DARPP-32 knockout mice. In support of a role for DARPP-32 in the action of caffeine, we found that, in striata of intact mice, caffeine increased the state of phosphorylation of DARPP-32 at Thr 75. Caffeine increased Thr 75 phosphorylation through inhibition of PP-2A-catalysed dephosphorylation, rather than through stimulation of cyclin-dependent kinase 5 (Cdk5)-catalysed phosphorylation, of this residue. Together, these studies demonstrate the involvement of DARPP-32 and its phosphorylation/dephosphorylation in the stimulant action of caffeine.

Distinct roles of dopamine D2L and D2S receptor isoforms in the regulation of protein phosphorylation at presynaptic and postsynaptic sites.

Dopamine D2 receptors are highly expressed in the dorsal striatum where they participate in the regulation of (i) tyrosine hydroxylase (TH), in nigrostriatal nerve terminals, and (ii) the dopamine- and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32), in medium spiny neurons. Two isoforms of the D2 receptor are generated by differential splicing of the same gene and are referred to as short (D2S) and long (D2L) dopamine receptors. Here we have used wild-type mice, dopamine D2 receptor knockout mice (D2 KO mice; lacking both D2S and D2L receptors) and D2L receptor-selective knockout mice (D2L KO mice) to evaluate the involvement of each isoform in the regulation of the phosphorylation of TH and DARPP-32. Incubation of striatal slices from wild-type mice with quinpirole, a dopamine D2 receptor agonist, decreased the state of phosphorylation of TH at Ser-40 and its enzymatic activity. Both effects were abolished in D2 KO mice but were still present in D2L KO mice. In wild-type mice, quinpirole inhibits the increase in DARPP-32 phosphorylation at Thr-34 induced by SKF81297, a dopamine D1 receptor agonist. This effect is absent in D2 KO as well as D2L KO mice. The inability of quinpirole to regulate DARPP-32 phosphorylation in D2L KO mice cannot be attributed to decreased coupling of D2S receptors to G proteins, because quinpirole produces a similar stimulation of [35S]GTPγS binding in wild-type and D2L KO mice. These results demonstrate that D2S and D2L receptors participate in presynaptic and postsynaptic dopaminergic transmission, respectively.

Looking BAC at striatal signaling: cell-specific analysis in new transgenic mice

Understanding how molecular signaling pathways participate in behavioral responses requires determining precisely in which neuronal populations they are activated. The recent development of bacterial artificial chromosome (BAC) transgenic mice expressing a variety of reporters, epitope tagged-proteins or Cre recombinase driven by specific promoters, is a significant step forward in this direction. These mice help overcome the limitations of traditional approaches that examine an average of signaling events occurring in mixed populations of cells. Here, we review how recent studies using such tools have revisited the regulation of striatal signaling pathways, demonstrating the striking segregation between neurons expressing dopamine D1 and D2 receptors and significantly extending our overall knowledge of striatal neurons. Thus, BAC transgenic mice are changing the way to conceive experiments and provide an opportunity to fill the gaps between molecular and systems neurosciences.