The amino hydroxy methyl isoxazolepropionic acid AMPA recept
The α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic ll-37 (AMPA) receptor is also enriched in postsynaptic striatal neurons (Bernard et al., 1997, Kondo et al., 2000, Reimers et al., 2011). Its activity is subjected to the regulation by a mechanism involving phosphorylation (Lu and Roche, 2012). For instance, protein kinase C (PKC) and Ca2+/calmodulin-dependent protein kinase II phosphorylate GluA1, one of four AMPA receptor subunits (GluA1-4, also known as GluR1-4; Greger et al., 2007), at a specific amino acid, serine 831 (S831), in intracellular C-terminal tails of the subunit. Similarly, PKA phosphorylates GluA1 at a serine 845 (S845) site (Roche et al., 1996, Barria et al., 1997, Mammen et al., 1997, Serulle et al., 2007). Consistent evidence shows that phosphorylation of GluA1 at these sites modulates physiological properties, expression, and trafficking of GluA1/AMPA receptors in neurons and transfected cells (Lu and Roche, 2012, Wang et al., 2014).
AMPA receptors are tightly regulated by dopamine signals as D1 agonists robustly enhanced striatal GluA1 phosphorylation at S845 (Price et al., 1999, Snyder et al., 2000, Swayze et al., 2004, Xue et al., 2017). However, whether M4 receptors regulate constitutive and dopamine-stimulated GluA1 phosphorylation in dopamine responsive striatal neurons is unclear. In this study, this topic was investigated and characterized in the adult rat striatum. A systemically active M4 antagonist tropicamide was used to evaluate the role of M4 receptors in vivo.
Discussion This work investigated the role of M4 receptors in the regulation of AMPA receptor GluA1/A2 subunit phosphorylation in the rat striatum in vivo. We found that the M4 antagonist tropicamide after a systemic injection increased GluA1 S845 phosphorylation in the CPu and NAc. This increase was rapid and reversible and was induced in a dose-dependent fashion. Moreover, tropicamide and a D1 agonist SKF81297 or a dopamine stimulant AMPH when coadministered at their subthreshold doses induced a significant increase in S845 phosphorylation. In contrast to GluA1 S845, GluA1 S831 and GluA2 S880 were insensitive to tropicamide. These results reveal that pharmacological blockade of M4 receptors with an antagonist enhances basal, and augments the dopamine-stimulated, S845 phosphorylation in the striatum. An important subpopulation of interneurons within the striatum is the cholinergic aspiny interneurons. These interneurons, despite their low percentage (1–2% of the total population of striatal neurons) (Bolam et al., 1984, Phelps et al., 1985), actively regulate medium spiny projection neurons via their branched and broad axonal arbors. In addition to these intrinsic interneurons, recent studies discovered extrinsic sources of cholinergic innervation of the striatum originating from the pedunculopontine and laterodorsal tegmental nuclei (Dautan et al., 2014, 2016). Two subtypes of muscarinic receptors (M1 and M4) are expressed in postsynaptic striatal neurons and mediate acetylcholine actions. Of note, M4 receptors are present in one of two equally populated projection neurons, i.e., striatonigral but not striatopallidal projection neurons, in the striatum (Ince et al., 1997, Santiago and Potter, 2001). This allows M4 receptors to primarily regulate striatonigral neurons. Thus, the S845 phosphorylation induced by the M4 blocker tropicamide is believed to mainly occur in striatonigral neurons. Since Gαi/o-coupled M4 receptors are negatively coupled to the cAMP-PKA pathway (Wess, 1996), tropicamide is reasoned to block the inhibitory linkage of M4 receptors to PKA and thereby enhance PKA activity and the PKA-mediated phosphorylation of S845. Notably, the M4-mediated inhibition is tonically active since tropicamide enhanced basal S845 phosphorylation under normal conditions. The M4 receptor not only suppresses the constitutive S845 phosphorylation, but it also works in concert with dopamine signaling to modulate the activity-dependent S845 phosphorylation. In fact, acetylcholine and dopamine have been traditionally viewed as a pair of transmitters essential for forming an intrinsic balance controlling striatal function (Di Chiara et al., 1994). While dopamine usually stimulates motor activities and gene expression in the striatum, acetylcholine inhibits them (Chou et al., 1992, Bernard et al., 1993, Morelli et al., 1993, Wang and McGinty, 1996a, Wang and McGinty, 1996b; Wang and McGinty, 1997). At the receptor level, D1 and M4 receptors are coexpressed in postsynaptic striatonigral neurons (Ince et al., 1997, Santiago and Potter, 2001), establishing a receptor model for functional dopamine-acetylcholine interactions in these neurons. It is likely that antagonistic D1 and M4 receptors in striatonigral neurons converge onto a central signaling molecule PKA to regulate PKA-dependent effectors, including the PKA-catalyzed GluA1 phosphorylation at S845. As a result, simultaneously reducing M4 tone and enhancing D1 tone could lead to a higher PKA activity level and thus a higher S845 phosphorylation rate. This study provides evidence supporting this model. We found that coadministration of subthreshold doses of the M4 antagonist tropicamide and the D1 agonist SKF81297 induced an increase in S845 phosphorylation. This type of an increase in S845 phosphorylation was also seen after tropicamide and AMPH coadministration. Additionally, in contrast to tropicamide, a positive allosteric modulator selective for M4 receptors (VU0152100) reduced the SKF81297-stimulated S845 phosphorylation in the striatum (Xue et al., 2017). Apparently, D1 and M4 receptors differentially regulate GluA1 S845 phosphorylation. Future studies need to clarify the cell-type-specific S845 phosphorylation in response to D1/M4 receptor manipulations using cell-type-specific approaches and to define the role of possible D1-M4 interactions in brain areas outside the striatum in regulating striatal GluA1 phosphorylation in response to a systemic injection of D1 and M4 receptor agents.