Phosphodiesterase 10A inhibitor, MP-10 (PF-2545920), produces greater induction of c-Fos in dopamine D2 neurons than in D1 neurons in the neostriatum
Introduction
The medium spiny neurons (MSN) of the neostriatum are the primary target of antipsychotic and anti-parkinsonian medications. Specifically, these drugs target the Gαi-coupled dopamine D2 family of G protein-coupled receptors on the MSNs (Perreault et al., 2011). Morphological and functional studies have shown that the D2 receptor-expressing MSNs constitute the striatopallidal pathway whereas the Gαs-coupled D1 expressing cells form the striatonigral pathway (Gerfen, 1992). Classical antipsychotic drugs are thought to produce their efficacy by antagonizing the D2 receptors (Kapur and Mamo, 2003). However, these drugs also induce extrapyramidal motor side effects by reducing basal ganglia output (Parr-Brownlie and Hyland, 2005). Although the newer, so-called atypical antipsychotics have a reduced liability to trigger extrapyramidal side effects, they induce intolerable metabolic side effects (Manu et al., 2012). Therefore, it remains critical to identify additional therapeutic mechanisms for treatment of psychotic symptoms associated with schizophrenia and other psychiatric disorders.
Phosphodiesterases (PDEs) are a family of enzymes that cleave cyclic nucleotides and thereby regulate second messenger signaling casacades (Sharma et al., 2013). PDE10A, like several other PDEs, cleaves both cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) to AMP and GMP, respectively (Wilson and Brandon, 2015). PDE10A is particularly relevant to the basal ganglia system because it is highly enriched in MSNs (Kleiman et al., 2011, Seeger et al., 2003). MP-10 (also known as PF-2545920; 2-((4-(1-methyl-4-(pyridin-4-yl)-1H-pyrazol-3-yl)phenoxy)methyl)quinoline) is a potent and selective PDE10A inhibitor (PDE10i) both in vitro and in vivo (Schulke et al., 2014). Treatment with MP-10 increases intracellular concentrations of cAMP/cGMP, augmenting dopamine D2 receptor antagonist- and dopamine D1 receptor agonist-mediated effects (Grauer et al., 2009, Schmidt et al., 2008). Given that dopamine D2 receptor inhibition is a pharmacological activity shared by all antipsychotic drugs, PDE10i have been proposed as a new class of therapeutics to treat psychoses (Strick et al., 2010).
The PDE10A knockout mice show reduced locomotor activity (Siuciak et al., 2006, Siuciak et al., 2008). Hence, we hypothesize that although PDE10i will affect activity of all MSNs, it may induce relatively greater activation of D2(+) MSNs. The present set of studies tested this hypothesis using MP-10 as a pharmacological probe. We assessed c-Fos immunoreactivity as a marker of increased neuronal activity in two sub-regions of the neostriatum, the dorsolateral and dorsomedial striatum, preferentially regulated by dopamine D2 and D1 receptors, respectively (Merchant et al., 1994). The regional pattern of c-Fos induction by MP-10 was compared to that produced by acute treatment with, prototypical D2 antagonist, haloperidol and, D1 agonist, SKF82958. Finally, to directly confirm this effect, we utilized the bacterial artificial chromosome (BAC) transgenic Drd1a-tdTomato mice line 6 (Ade et al., 2011) to assess co-localization of c-Fos immunoreactivity in MSN populations that express the D1 receptor or those without the D1 receptor. In these mice, the expression of the fluorescent reporter tdTomato is under the regulation of the Drd1a gene and the reporter is expressed with high selectivity and specificity only in D1(+) MSNs (Ade et al., 2011), thereby providing a direct method to assess the induction of c-Fos in both the D1 or D2-expressing sub-populations of MSNs.
Section snippets
Animals/dosing
Experiments were carried out in accordance with the Declaration of Helsinki and the Guide for the Care and Use of Laboratory Animals by the U.S. National Institutes of Health and the EU Directive 86/609/EEC. All studies were performed on male rodents. Sprague Dawley (SD) rats (Charles River Laboratories, Margate, UK), C57BL/6 mice (Taconic Biosciences, Cambridge City, IN) and BAC transgenic Drd1a-tdTomato mice (The Jackson Laboratory, Bar Harbor, ME) were acclimatized for at least 3 days
MP-10 dose-dependently increased c-Fos expression in the rat striatum
Fig. 1A shows the c-Fos immunopositive cells at two different magnifications following MP-10 at 30 mg/kg, PO, to rats. A statistically significant, dose-dependent increase in c-Fos immunopositive cells was observed in both the DLS and DMS at 10 and 30 mg/kg of MP-10 (Fig. 1B). Notably, the c-Fos signal density was higher in the DLS than the DMS at both 10 and 30 mg/kg doses. The c-Fos dominance index derived by subtracting the DMS signal from that in the DLS confirmed that MP-10 treatment at 10
Discussion
The present studies utilized a combination of pharmacological and genetic tools to test the hypothesis that PDE10A inhibition produces greater activation of D2(+) striatopallidal pathway than D1(+) striatonigral pathway. Using c-Fos as a marker of neuronal activation and a selective PDE10i tool, MP-10, we provide two major but complementary lines of evidence in support of this hypothesis. First, like the effects of the D2 inhibitory ligand, haloperidol, the effects of MP-10 were greater in the
Disclosures
J.M.W., A.M.L.O., S.L., G.G. and K.M.M. were all employees of Eli Lilly & Company during this study.
A.J.B. and T.L.B-A have no conflicts to disclose.
Acknowledgments
Expert technical assistance of Rebecca Wright is gratefully acknowledged. We thank Dr. Hong Wang for critically reading the manuscript. Funding for this study was provided by Eli Lilly & Company. The funding source had no further role in study design; in the collection, analysis and interpretation of data; in the writing of the report; and in the decisions to submit the paper for publication.
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