title: Mechanism of GABA receptor-mediated inhibition of spontaneous GABA release onto cerebellar Purkinje cells.
abstract: gamma-Aminobutyric acid (GABA)(B) receptor-mediated modulation of spontaneous GABA release onto Purkinje cells was investigated in cerebellar slices from 3- to 5-week-old mice. The GABA(B) receptor agonists baclofen and CGP 44533 each reduced the frequency of miniature inhibitory postsynaptic currents (mIPSCs), with no significant effect on mIPSC amplitude; together, consistent with a presynaptic site of action. The GABA(B) receptor antagonist CGP 55845 blocked baclofen-induced inhibition. The sulphydryl alkylating agent N-ethylmaleimide occluded baclofen effects, implicating G(i/o) subunits in mediating a GABA(B) G protein-coupled receptor pathway. Baclofen-induced inhibition persisted in the presence of Ba(2+), a blocker of K(+) channels, and Cd(2+), a blocker of Ca(2+) channel-mediated GABA release. Application of nominally Ca(2+)-free extracellular solutions reduced mIPSC frequency and amplitude; however, baclofen produced a significant inhibition in mIPSC frequency, further suggesting that this pathway was independent of Ca(2+) influx. Spontaneous GABA release was increased by the adenylate cyclase activator, forskolin, and the phorbol ester, phorbol 12,13-dibutyrate. However, baclofen-induced inhibition was not significantly changed in either condition. Baclofen action was also not affected by the adenylate cyclase inhibitor SQ 22536 or the protein kinase C inhibitor chelerythrine chloride. Baclofen still reduced mIPSC frequency in the presence of the polyvalent cation ruthenium red, which acts as a secretagogue here; however, baclofen-induced inhibition was reduced significantly. Furthermore, baclofen produced no clear inhibition during high-frequency mIPSCs bursts induced by the potent secretagogue alpha-Latrotoxin. Together, these results suggest that GABA(B) inhibition occurs downstream of Ca(2+) influx and may be mediated, in part, by an inhibition of the vesicular release mechanism.
authors.affiliation: Department of Pharmacology, University College London, Gower Street, London WC1E 6BT, UK.
authors.forename: Victoria L
authors.initials: VL
authors.lastname: Harvey
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Stephens
mesh_terms: D000200:Action Potentials / Q000187:drug effects / Q000502:physiology; D000367:Age Factors; D000818:Animals; D001418:Baclofen / Q000494:pharmacology; D001464:Barium / Q000494:pharmacology; D002104:Cadmium / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002531:Cerebellum / Q000166:cytology* / Q000187:drug effects; D005576:Colforsin / Q000494:pharmacology; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005098:Exotoxins / Q000494:pharmacology; D018755:GABA Agonists / Q000494:pharmacology; D018756:GABA Antagonists / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D009433:Neural Inhibition / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques / Q000379:methods; D010721:Phosphinic Acids / Q000494:pharmacology; D011412:Propanolamines / Q000494:pharmacology; D011689:Purkinje Cells / Q000187:drug effects / Q000378:metabolism*; D011963:Receptors, GABA-A / Q000502:physiology*; D012430:Ruthenium Red / Q000494:pharmacology; D009435:Synaptic Transmission / Q000187:drug effects / Q000502:physiology; D013779:Tetrodotoxin / Q000037:antagonists & inhibitors / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D005098:Exotoxins; D018755:GABA Agonists; D018756:GABA Antagonists; D010721:Phosphinic Acids; D011412:Propanolamines; D011963:Receptors, GABA-A; C018579:leukotoxin; D002104:Cadmium; D012430:Ruthenium Red; C085075:CGP 55845A; D005576:Colforsin; D001464:Barium; D013779:Tetrodotoxin; D005680:gamma-Aminobutyric Acid; D001418:Baclofen; D002118:Calcium