title: The lysosome or lysosome-related organelle may serve as a Ca2+ store in the boutons of hippocampal pyramidal cells.
abstract: Boutons are specialised presynaptic compartments that lie along the axons of central neurons. Release of neurotransmitter from boutons is tightly regulated by the level of intracellular calcium [Ca2+]i. A rise in Ca2+ level may be generated in several ways; entry of extracellular Ca2+ via voltage gated calcium channels (VGCCs), entry via ligand-operated channels (LOCs) or the release of Ca2+ from intracellular Ca2+ stores. The role of Ca2+ stores in boutons remains poorly understood, despite recent work indicating that the release of Ca2+ from the endoplasmic reticulum (ER) may contribute to transmitter release. In this study we assess whether the lysosome or a closely related organelle functions as a Ca2+ store in the boutons of hippocampal pyramidal neurones. Lysosomes are small acidic organelles more commonly known for their role in degrading redundant cellular constituents. Using a fluorescent lysosomal marker, we show that lysosomes are located in the axons of hippocampal CA3 neurones. Selective pharmacological lysis of the lysosomes with glycyl-phenylalanine 2-naphthylamide (GPN) generates rapid, highly focal Ca2+ transients within the axon and increases the frequency of spontaneous miniature excitatory post-synaptic currents (mEPSCs), revealing that the organelle contains Ca2+ at a concentration sufficient to evoke transmitter release. Confocal laser scanning microscopy, combined with electrophysiology is used to monitor the action potential evoked increases in [Ca2+]i in boutons. We show that disruption of lysosomes compromises action potential evoked [Ca2+]i but this effect is occluded if the ER is discharged. Conversely, disruption of the lysosome does not appear to impact on the capacity of the ER to release Ca2+. These results suggest that the lysosome may serve as a Ca2+ store within hippocampal boutons, with a Ca2+ signalling role that is unique from that of the ER.
authors.affiliation: University of Oxford, Department of Pharmacology, Mansfield Road, Oxford OX1 3QT, UK.
authors.forename: Lindsay
authors.initials: L
authors.lastname: McGuinness
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Bardo
authors.forename: Nigel J
authors.initials: NJ
authors.lastname: Emptage
mesh_terms: D000588:Amines / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism*; D004151:Dipeptides / Q000494:pharmacology; D004558:Electric Stimulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D006624:Hippocampus / Q000166:cytology*; D066298:In Vitro Techniques; D008247:Lysosomes / Q000187:drug effects / Q000502:physiology*; D008297:Male; D018613:Microscopy, Confocal / Q000379:methods; D015388:Organelles / Q000187:drug effects / Q000502:physiology; D017729:Presynaptic Terminals / Q000187:drug effects / Q000378:metabolism*; D017966:Pyramidal Cells / Q000378:metabolism / Q000648:ultrastructure*; D051381:Rats; D017208:Rats, Wistar; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors
chemical_list: D000588:Amines; D004151:Dipeptides; D004791:Enzyme Inhibitors; C493330:Red DND-99; D019284:Thapsigargin; C039234:glycyl-proline-1-naphthylamide; D002118:Calcium