Cerebellar Long-Term Synaptic Depression Requires PKC-Mediated Activation of CPI-17, a Myosin/Moesin Phosphatase Inhibitor

Abstract

Cerebellar LTD requires brief activation of PKC and is expressed as a functional downregulation of AMPA receptors. Modulation of vascular smooth-muscle contraction by G protein-coupled receptors (called Ca(2+) sensitization) also involves PKC phosphorylation and activation of a specific inhibitor of myosin/moesin phosphatase (MMP). This inhibitor, called CPI-17, is also expressed in brain. Here, we tested the hypothesis that LTD, like Ca(2+) sensitization, employs a PKC/CPI-17 cascade. Introduction of activated recombinant CPI-17 into cells produced a use-dependent attenuation of glutamate-evoked responses and occluded subsequent LTD. Moreover, the requirement for endogenous CPI-17 in LTD was demonstrated with neutralizing antibodies plus gene silencing by siRNA. These interventions had no effect on basal synaptic strength but blocked LTD induction. Thus, a biochemical circuit that involves PKC-mediated activation of CPI-17 modulates the distinct physiological processes of vascular contractility and cerebellar LTD.

Protein phosphatase 2A inhibition induces cerebellar long-term depression and declustering of synaptic AMPA receptor

“…Dephosphocytostatin (30 M), an inactive form of cytostatin, did not produce any marked effect on eEPSC amplitude (91 Ϯ 3%, n ϭ 4). In contrast, infusion of 100 nM fostriecin into immature PCs (12-15 DIV) did not depress eEPSCs (93 Ϯ 5%, n ϭ 8), confirming a recent report using young cerebellar cultures from mouse (15). All subsequent experiments were done on PCs aged 22-35 DIV (average 25.7 DIV).…”

“…Although protein phosphatases (PPs) share with protein kinases the task of regulating protein phosphorylation level, they have long been considered mere housekeepers, with poorly understood substrate specificity and loose regulation. In recent years, however, studies using phosphatase inhibitors revealed that these enzymes exert a major influence on synaptic plasticity in the cerebellum (11)(12)(13)(14)(15) and other brain structures (16,17). In the cerebellum, investigation of AMPAR regulation by PP at the GC-PC synapse represents a challenge as PCs abundantly express at least five major types of serine͞threonine PPs: PP-1, PP-2A, PP-2B, PP-2C, and PP-5 (18)(19)(20)(21)(22).…”

AMPA, not NMDA, activates RhoA GTPases and subsequetly phosphorylates moesin

“…In this context, our result might be a missing puzzle fragment, that is, AMPA activated RhoA leading to polymeriz- ation of actin cytoskeleton and also phosphorylate moesin by RhoA-ROK leading to ligation of AMPA or NMDA receptor to underlying actin cytoskeleton since phopshorylation of moesin has been reported to be crucial in memebrane-cytoskeleton ligation (Bretscher et al, 2002). Recent study that a myosin/moesin phosphatase inhibitor, CPI-17 is crucial for cerebellar long-term depression through AMPA receptor (Eto et al, 2002) might be another clue for the role of moesin in AMPA receptor signaling. In this study, silencing CPI-17 by si-RNA blocked long-term depression (LTD) induction although significance of moesin phosphorylation in LTD is not directly proved.…”

Phosphoprotein inhibitor CPI-17 specificity depends on allosteric regulation of protein phosphatase-1 by regulatory subunits

“…Thus, phosphorylation of CPI-17 suppresses myosin phosphatase activity, resulting in phosphorylation of myosin and contraction of smooth muscle. In addition, specific depletion of endogenous CPI-17 by small interfering RNA or antibody microinjection eliminated the cerebellar long-term synaptic depression of Purkinje cells mediated by PKC, demonstrating involvement of CPI-17 in neuronal signaling (21). Although phospho-CPI-17 inhibits monomeric PP1C in addition to myosin phosphatase, myosin phosphatase was proposed as a preferred target of phospho-CPI-17 in smooth muscle (22), fibroblasts (23), and cerebellar Purkinje cells (21).…”