Presynaptic inhibition upon CB 1 or mG lu2/3 receptor activation requires ERK / MAPK phosphorylation of Munc18‐1

Abstract

Presynaptic cannabinoid (CB1R) and metabotropic glutamate receptors (mGluR2/3) regulate synaptic strength by inhibiting secretion. Here, we reveal a presynaptic inhibitory pathway activated by extracellular signal-regulated kinase (ERK) that mediates CB1R-and mGluR2/3-induced secretion inhibition. This pathway is triggered by a variety of events, from foot shock-induced stress to intense neuronal activity, and induces phosphorylation of the presynaptic protein Munc18-1. Mimicking constitutive phosphorylation of Munc18-1 results in a drastic decrease in synaptic transmission. ERK-mediated phosphorylation of Munc18-1 ultimately leads to degradation by the ubiquitin-proteasome system. Conversely, preventing ERK-dependent Munc18-1 phosphorylation increases synaptic strength. CB1R-and mGluR2/3-induced synaptic inhibition and depolarization-induced suppression of excitation (DSE) are reduced upon ERK/MEK pathway inhibition and further reduced when ERK-dependent Munc18-1 phosphorylation is blocked. Thus, ERK-dependent Munc18-1 phosphorylation provides a major negative feedback loop to control synaptic strength upon activation of presynaptic receptors and during intense neuronal activity.

Atypical Endocannabinoid Signaling Initiates a New Form of Memory-Related Plasticity at a Cortical Input to Hippocampus

“…We confirmed CB 1 R localization to glutamatergic (i.e., VGluT2 immunopositive [+]) terminals in the rat LPP field and then compared the effects of cannabinoid receptor agonist WIN 55,212-2 (WIN) on synaptic physiology in the S-C and LPP projections. In accord with prior work (Paton et al 1998;Schmitz et al 2016), WIN (5 μM) caused a rapid and pronounced depression of S-C fEPSPs in CA1 (P < 0.0001; Fig. 1a) that was accompanied by an increase in paired-pulse facilitation (P = 0.0067; Fig.…”

“…Recent studies demonstrated that presynaptic signaling through ERK1/2 to the vesicular protein Munc18-1 is critical for CB 1 R-mediated depression of release (Schmitz et al 2016). Here, we show that CB 1 R agonists readily activate this signaling cascade at S-C synapses but not in the LPP.…”

“…CB 1 R signaling through ERK1/2 effects phosphorylation and degradation of the vesicular protein Munc18-1 leading to reductions in transmitter release (Schmitz et al 2016). In accord with this, using dual immunofluorescence and FDT (Fig.…”

“…Conversely, the release depression function of the CB 1 R was poorly developed in the LPP and not involved in the potentiation effect. ECB-mediated release suppression is reportedly sensitive to locally synthesized pregnenolone (Vallee et al 2014) and involves phosphorylation of the vesicular protein Munc18-1 (Schmitz et al 2016). These effects were evident in S-C projections to CA1 but not in the LPP.…”

“…The microscope slide mounted tissue was fixed in 4% paraformaldehyde and processed for immunostaining as described (Seese et al 2014). Immunofluorescence procedures used cocktails of primary antisera (Table 1) including rabbit antisera to pMunc18-1 Ser241 (Schmitz et al 2016), phosphorylated proline-rich tyrosine kinase 2 (pPyk2) Tyr402 (Zhao et al 2000), phosphorylated RhoA associated coiled coil containing kinase 2 (pROCK2) Ser1366 (Chuang et al 2012), or pFAK Y397 (Bock and Herz 2003), used in combination with mouse antisera to synaptophysin (SYN) (Wen et al 2014) or postsynaptic density protein-95 (PSD-95) (Takahashi et al 2011); or guinea pig anti-vGluT2 used in combination with rabbit anti-CB 1 (Matyas et al 2006) that was generously provided by Dr Ken Mackie of Indiana University. Secondary antisera from ThermoFisher Scientific included Alexa Fluor 594 antirabbit IgG (A21207), Alexa Fluor 488 anti-mouse IgG (A21202) and Alexa Fluor 488 anti-guinea pig IgG (A-11073) all used at 1:1000 dilutions.…”