Antibodies against cysteine string proteins inhibit evoked neurotransmitter release at Xenopus neuromuscular junctions.
Cysteine string proteins (CSPs) are evolutionarily conserved proteins that are associated with synaptic vesicles and other regulated secretory organelles. To investigate the role of CSPs in vertebrate neuromuscular transmission, we introduced anti-CSP antibodies into the cell bodies of Xenopus spinal motor neurons that form synapses with embryonic muscle cells in culture. These antibodies produced a rapid (within 3-6 min), and in most cases complete, inhibition of stimulus-dependent neurotransmitter secretion. However, spontaneous neurotransmitter release was stable (both in frequency and amplitude) throughout the period of antibody exposure. Several control experiments validated the specificity of the anti-CSP antibody effects. First, the anti-CSP antibody actions were not mimicked either by antibodies against another synaptic vesicle protein SV2, or by nonspecific immunoglobins. Second, heat treatment of the anti-CSP antibodies eliminated their effect on evoked secretion. Third, immunoblot experiments showed that the anti-CSP and anti-SV2 antibodies were highly selective for their respective antigens in these Xenopus cultures. We conclude from these results that CSPs are vital constituents of the pathway for regulated neurotransmitter release in vertebrates. Moreover, the selective inhibition of evoked, but not spontaneous transmitter release by anti-CSP antibodies indicates that there is a fundamental difference in the machinery that mediates these secretory processes.
Department of Neuroscience, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, USA.
R E
RE
Poage
S D
SD
Meriney
C B
CB
Gundersen
J A
JA
Umbach
D000200:Action Potentials / Q000502:physiology; D000818:Animals; D018920:Coculture Techniques; D004625:Embryo, Nonmammalian; D005071:Evoked Potentials / Q000502:physiology*; D050956:HSP40 Heat-Shock Proteins; D007074:Immunoglobulin G / Q000494:pharmacology*; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000276:immunology / Q000502:physiology*; D009046:Motor Neurons / Q000166:cytology / Q000502:physiology*; D018482:Muscle, Skeletal / Q000166:cytology / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000276:immunology / Q000502:physiology; D009469:Neuromuscular Junction / Q000502:physiology*; D018377:Neurotransmitter Agents / Q000378:metabolism*; D013116:Spinal Cord / Q000166:cytology / Q000502:physiology; D013569:Synapses / Q000502:physiology; D009435:Synaptic Transmission / Q000502:physiology; D013997:Time Factors; D014982:Xenopus laevis
D050956:HSP40 Heat-Shock Proteins; D007074:Immunoglobulin G; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D018377:Neurotransmitter Agents; C088460:cysteine string protein