Although it is not known if CP AMPAR regulation

Although it is not known if CP-AMPAR regulation is required for LTD, our previous characterization of AKAP150ΔPIX knockin mice that are selectively deficient in CaN anchoring, due to disruption of a PxIxIT-type CaN docking motif, provided important insights. We found that AKAP-CaN signaling dephosphorylates GluA1 S845 and limits synaptic incorporation of CP-AMPARs to constrain LTP and promote LTD (Sanderson et al., 2012). These previous findings suggested that phosphorylation-regulated CP-AMPAR recruitment to synapses may also occur during LTD, but how inhibition of either AKAP-PKA or -CaN signaling and either increases or decreases in S845 Nilvadipine all result in impaired LTD remained unclear. Here, we address this conundrum by using a combination of molecular genetic, pharmacological, and electrophysiological approaches to uncover a novel mechanism whereby phosphorylated GluA1 CP-AMPARs are transiently recruited to synapses during NMDAR-dependent LTD induction before signaling their own removal and being dephosphorylated. Through analysis of AKAP150 mutant mice, we demonstrate that CP-AMPARs are recruited to synapses by anchored PKA during LTD induction but are then rapidly removed by anchored CaN and that blocking CP-AMPAR activity, recruitment, or removal interferes with LTD. Our findings of PKA-dependent synaptic recruitment of CP-AMPARs during LTD reveal remarkable and unexpected parallels with some previously proposed LTP and homeostatic plasticity mechanisms, thus further underscoring the complex signaling crosstalk underlying these distinct, yet interrelated, forms of synaptic regulation.
Results
Discussion Our present findings demonstrate that AKAP-anchored PKA is required during LTD induction to facilitate NMDAR-dependent recruitment of CP-AMPARs to participate in postsynaptic Ca2+ signaling, as depicted in Figure 7. Importantly, CP-AMPAR signaling is then required, along with NMDAR signaling, to rapidly trigger AKAP-CaN mediated removal of the newly recruited GluA2-lacking CP-AMPARs, along with GluA2-containing receptors, to allow for optimal LTD expression. If CP-AMPAR signaling is not engaged, such as in 150ΔPKA mice or with continuous application of NASPM, then LTD expression is substantially reduced but not eliminated. Thus, NMDAR signaling alone is capable of removing GluA2-containing AMPARs to induce some LTD, but additional contributions from CP-AMPARs are required for robust LTD expression. Figure 7 depicts two distinct, sequential stages: (1) NMDAR and PKA-dependent CP-AMPAR recruitment and (2) NMDAR, CP-AMPAR, and CaN-dependent AMPAR removal, accompanied by spine shrinkage (Zhou et al., 2004) and disruption of AKAP150 membrane targeting and PSD-95 interactions that may prevent AMPARs from returning to the synapse (Smith et al., 2006), but in reality, these processes must be occurring almost simultaneously and continuously during the LTD induction stimulus to prevent potentiation and promote depression. Previous studies found that pharmacological inhibition of PKA activity or AKAP-PKA anchoring can lead to depression of both synaptic and extrasynaptic AMPAR activity that occludes subsequent LTD induction (Kameyama et al., 1998, Rosenmund et al., 1994, Snyder et al., 2005, Tavalin et al., 2002). However, even acute inhibition of PKA activity that does not result in depression of basal AMPAR transmission can inhibit LTD (Lu et al., 2008). In addition, LTD is impaired in AKAP150 knockout and PKA anchoring-deficient D36 knockin mice, yet both strains exhibit no decreases in basal AMPAR transmission and at best partial decreases in basal GluA1 S845 phosphorylation (Lu et al., 2007, Lu et al., 2008, Tunquist et al., 2008, Zhang et al., 2013). Likewise, here in 150ΔPKA mice we found impaired LTD but only a ∼40% reduction in basal S845 phosphorylation and no decrease in basal AMPAR transmission. Thus, although prior inhibition of AKAP-PKA signaling can lead to depression that occludes LTD, AKAP-PKA signaling also plays a more dynamic role during LTD induction, which we now show involves the transient recruitment of CP-AMPAR to synapses, a mechanism previously only implicated in AKAP-PKA LTP regulation (Lu et al., 2007).