title: Effects of ethanol on glycinergic synaptic currents in mouse spinal cord neurons.
abstract: Ethanol increased the frequency of miniature glycinergic currents [miniature inhibitory postsynaptic currents (mIPSCs)] in cultured spinal neurons. This effect was dependent on intracellular calcium augmentation, since preincubation with BAPTA (an intracellular calcium chelator) or thapsigargin [a sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) pump inhibitor] significantly attenuated this effect. Similarly, U73122 (a phospholipase C inhibitor) or 2-aminoethoxydiphenyl borate [2-APB, an inositol 1,4,5-trisphosphate (IP₃) receptor (IP3R) inhibitor] reduced this effect. Block of ethanol action was also achieved after preincubation with Rp-cAMPS, inhibitor of the adenylate cyclase (AC)/PKA signaling pathway. These data suggest that there is a convergence at the level of IP₃R that accounts for presynaptic ethanol effects. At the postsynaptic level, ethanol increased the decay time constant of mIPSCs in a group of neurons (30 ± 10% above control, n = 13/26 cells). On the other hand, the currents activated by exogenously applied glycine were consistently potentiated (55 ± 10% above control, n = 11/12 cells), which suggests that ethanol modulates synaptic and nonsynaptic glycine receptors (GlyRs) in a different fashion. Supporting the role of G protein modulation on ethanol responses, we found that a nonhydrolyzable GTP analog [guanosine 5'-O-(3-thiotriphosphate) (GTPγS)] increased the decay time constant in ∼50% of the neurons (28 ± 12%, n = 11/19 cells) but potentiated the glycine-activated Cl(-) current in most of the neurons examined (83 ± 29%, n = 7/9 cells). In addition, confocal microscopy showed that α1-containing GlyRs colocalized with Gβ and Piccolo (a presynaptic cytomatrix protein) in ∼40% of synaptic receptor clusters, suggesting that colocalization of Gβγ and GlyRs might account for the difference in ethanol sensitivity at the postsynaptic level.
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile; PhD Program in Pharmacology, University of Chile, Santiago, Chile; and.
authors.forename: Trinidad A
authors.initials: TA
authors.lastname: Mariqueo
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile;
authors.forename: Adolfo
authors.initials: A
authors.lastname: Agurto
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile;
authors.forename: Braulio
authors.initials: B
authors.lastname: Muñoz
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile;
authors.forename: Loreto
authors.initials: L
authors.lastname: San Martin
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile;
authors.forename: Cesar
authors.initials: C
authors.lastname: Coronado
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile;
authors.forename: Eduardo J
authors.initials: EJ
authors.lastname: Fernández-Pérez
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile;
authors.forename: Pablo
authors.initials: P
authors.lastname: Murath
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile;
authors.forename: Andrea
authors.initials: A
authors.lastname: Sánchez
authors.affiliation: Department of Anesthesiology and Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, Pennsylvania.
authors.forename: Gregg E
authors.initials: GE
authors.lastname: Homanics
authors.affiliation: Department of Physiology-Laboratory of Neurophysiology, University of Concepción, Concepción, Chile; laguayo@udec.cl.
authors.forename: Luis G
authors.initials: LG
authors.lastname: Aguayo
mesh_terms: D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002492:Central Nervous System Depressants / Q000494:pharmacology*; D018118:Chloride Channels / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000431:Ethanol / Q000494:pharmacology*; D005998:Glycine / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D053444:Inhibitory Postsynaptic Potentials / Q000187:drug effects; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism; D008810:Mice, Inbred C57BL; D055369:Miniature Postsynaptic Potentials / Q000187:drug effects; D009474:Neurons / Q000187:drug effects* / Q000502:physiology; D017729:Presynaptic Terminals / Q000187:drug effects / Q000502:physiology; D018009:Receptors, Glycine / Q000378:metabolism; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D013116:Spinal Cord / Q000187:drug effects* / Q000502:physiology; D009435:Synaptic Transmission / Q000187:drug effects* / Q000502:physiology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D000067956:Adenylyl Cyclase Inhibitors; D002492:Central Nervous System Depressants; D018118:Chloride Channels; D053496:Inositol 1,4,5-Trisphosphate Receptors; D018009:Receptors, Glycine; D000431:Ethanol; D006160:Guanosine Triphosphate; D017868:Cyclic AMP-Dependent Protein Kinases; D010738:Type C Phospholipases; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000262:Adenylyl Cyclases; D002118:Calcium; D005998:Glycine
keywords: adenylate cyclase; calcium; ethanol; forskolin; glycinergic miniature IPSCs; spinal neurons; strychnine