Supplementary MaterialsFigure 5-1. PIP2-gated G-protein combined rectifying K+ currents aswell inwardly. Scarcity of KCNQ2-comprising M-channels ablated the M1R-induced enhancement of M-current in DGGCs. Simultaneously, M1R activation in DGGCs induced powerful raises in [Ca2+]i, mostly due to TRPC currents, consistent with, and contributing to, neuronal depolarization and hyperexcitability. CA1 neurons did not display such multimodal signaling, but rather M current was suppressed by M1R stimulation in these cells, similar to the previously described actions of M1R stimulation on M-current in peripheral ganglia that mostly involves PIP2 depletion. Therefore, these results point to a pleiotropic network of cholinergic signals that direct cell-type-specific, precise control of hippocampal function with strong implications for hyperexcitability and epilepsy. SIGNIFICANCE STATEMENT At the neuronal membrane, protein signaling cascades consisting of ion channels and metabotropic receptors govern the electrical properties and neurotransmission of neuronal networks. Muscarinic acetylcholine receptors are G-protein-coupled metabotropic receptors that control the excitability of neurons through CSF2RB regulating ion channels, intracellular Ca2+ signals, and other second-messenger cascades. We have illuminated previously unknown actions of Tandutinib (MLN518) muscarinic stimulation on the excitability of hippocampal principal neurons that include M channels, TRPC (transient receptor potential, canonical) cation channels, and powerful regulation of lipid metabolism. Our results show that these signaling pathways, and mechanisms of excitability, are starkly distinct between peripheral ganglia and brain, and even between different principal neurons in the hippocampus. acetylcholine receptors (mAChRs) in sympathetic ganglia (Brown and Adams, 1980; Constanti and Brown, 1981), where the principal mechanism of action is hydrolysis and depletion of phosphatidylinositol 4,5-bisphosphate (PIP2) by phosphalipase C (PLC) (Zhang et al., 2003; Suh and Hille, 2007). Stimulation of other Gq/11-coupled receptors also depresses test, followed by Tukey’s HSD test 0.05 unless otherwise specified. Results M channels and ERG channels have differential fractional contributions to the threshold K+ current of DGGCs and CA1 pyramidal neurons Hippocampal neurons express multiple classes of slowly deactivating, voltage-gated K+ channels including M channels (KCNQ2/3) and ERG channels (KCNH2, Kv11.1) that activate at potentials near threshold and contribute to regulation of neuronal discharge properties (Saganich et al., 2001; Hu et al., 2002; Jan and Jan, 2012; Mateos-Aparicio et al., 2014; Kim et al., 2016a,b). To determine the contribution of = 12C20 cells per group, = 0.3). Open in a separate window Figure 1. Contribution of = 20C22 cells. * 0.05 vs ACSF. in control ACSF or in the presence of XE-991 (20 m) or E4031 (10 m). The ordinates are normalized current (left) or cumulative probability of decay (right). The averaged currents in the presence of either blocker were fit by a single exponential relation. = 16 cells. * 0.05 versus ACSF. and and = 15 cells), which closely corresponds to the kinetics found previously in sympathetic neurons and Tandutinib (MLN518) in hippocampus (Beech et al., 1991; Gamper et al., 2003; Lawrence et al., 2006a). The total current density was 4.42 0.32 pA/pF and the residual current density was significantly reduced to 0.90 0.08 pA/pF after addition of XE-991, indicating that 78.3 1.8% of the total deactivating current was due to 0.0001), with the rest displaying much slower kinetics (Fig. 1= 0.55) nor the deactivation currents (= 0.65, = 15 cells) in DGGCs (Fig. 1= 0.0045; Fig. 1= 0.0036; Fig. 1 0.0001, = 21 cells). Due to the unanticipated and astonishing Tandutinib (MLN518) nature of these results, we also tested the effects of alternative muscarinic agonists. The muscarinic agonist pilocarpine (10 m) also increased = 0.03, = 4 cells). Although unlikely, it is conceivable that such a result could be due to stimulation of Gi/o-coupled M2/M4 mAChRs via some unknown pathway. Therefore, we also tested the M1R-specific allosteric agonist, 77-LH-28-1 (Langmead et al., 2008; Thomas et al., 2008). Bath-application of 77-LH-28-1 over a range of concentrations produced a similar increase of = 0.91, = 5 cells). Given previous findings of muscarinic suppression of = 5 cells).