Electrical and optical monitoring of alpha-latrotoxin action at Drosophila neuromuscular junctions.
Electrophysiological recording demonstrates that alpha-latrotoxin, a 125,000 mol. wt component of black widow spider venom, promotes high frequency quantal discharges at larval neuromuscular junctions of Drosophila. Concomitantly, fluorescence imaging of presynaptic calcium ion activity reveals that this toxin qualitatively elevates cytosolic ionized calcium in this preparation. These activities of alpha-latrotoxin are selectively antagonized by a monoclonal antibody, 4C4.1, that was previously shown to inhibit the action of this toxin in PC-12 cells. However, 4C4.1 does not block the release-promoting activity of gel-filtered extracts of black widow spider venom. This indicates that black widow spider venom has multiple components that promote quantal transmitter secretion in invertebrates. This investigation demonstrates that alpha-latrotoxin is among the active principles in black widow spider venom that enhance transmitter release and raise cytosolic ionized calcium in Drosophila. These results suggest that Drosophila, because of the relative ease of genetic manipulation, may be useful to study the target protein(s) that mediate the binding and action of alpha-latrotoxin at nerve endings. Moreover, the procedure that we report for loading Drosophila nerve terminals with the calcium ion-sensing dye, Calcium Crimson, may have utility for studying calcium dynamics in mutant alleles with alterations in synapse development and function in this organism.
The Department of Molecular and Medical Pharmacology, UCLA School of Medicine, Los Angeles, CA 90095, USA.
J A
JA
Umbach
A
A
Grasso
S D
SD
Zurcher
H I
HI
Kornblum
A
A
Mastrogiacomo
C B
CB
Gundersen
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects*; D002850:Chromatography, Gel; D004396:Coloring Agents; D004331:Drosophila melanogaster / Q000187:drug effects* / Q000254:growth & development; D005071:Evoked Potentials / Q000187:drug effects; D005089:Exocytosis / Q000187:drug effects; D005456:Fluorescent Dyes; D019476:Insect Proteins / Q000187:drug effects / Q000502:physiology; D007814:Larva; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000187:drug effects / Q000502:physiology; D009469:Neuromuscular Junction / Q000187:drug effects* / Q000502:physiology; D009930:Organic Chemicals; D018000:Receptors, Peptide / Q000187:drug effects / Q000502:physiology; D013111:Spider Venoms / Q000037:antagonists & inhibitors / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology*; D009435:Synaptic Transmission / Q000187:drug effects* / Q000502:physiology
D000911:Antibodies, Monoclonal; D004396:Coloring Agents; D005456:Fluorescent Dyes; D019476:Insect Proteins; D009419:Nerve Tissue Proteins; D009930:Organic Chemicals; D018000:Receptors, Peptide; D013111:Spider Venoms; C034800:alpha-latrotoxin receptor; C071334:calcium crimson; C020763:alpha-latrotoxin