Effect of alpha‐latrotoxin on the frog neuromuscular junction at low temperature.

Abstract

SUMMARY1. a-Latrotoxin (a-LTx) was applied to frog cutaneus pectoris muscles bathed at 1-3°C in either Ringer solution, Ca2"-free Ringer solution with 1 mM-EGTA and 4 mM-Mg2+ or Ringer solution plus 4 mM-Mg2+, and its effects on miniature end-plate potential (MEPP) frequency, nerve terminal ultrastructure and uptake of horseradish peroxidase (HRP) were studied.2. Large concentrations (2 ,ug/ml) of a-LTx increased MEPP rates to levels above 100/s at all junctions, but the time course of the increases depended upon the divalent cation content of the bathing solution. However, similar numbers of MEPPs (0-3-407 x 106) were recorded at all junctions during 2 h of secretion.3. Nerve terminals exposed to a-LTx for 2 h lost 60-75 % of their synaptic vesicles and were swollen; their presynaptic membranes were deeply infolded and they often contained many large vesicular structures. Terminals in Ringer solution retained the largest number of synaptic vesicles; terminals in Ringer solution plus Mg2+ swelled the least and contained the largest number of coated vesicles. The average number of synaptic vesicles lost was approximately equal to the average number of MEPPs recorded.4. Few vesicles became loaded with HRP when this extracellular tracer was present in the bathing solution and the muscles were fixed near the peak of secretion.5. When the terminals were warmed to 20°C, those in the Caa2+-free solution with Mg2+ secreted additional quanta and lost almost all their residual vesicles; those in Ringer solution without Mg2+ secreted few additional quanta and retained most of their residual vesicles.6. These results suggest that recycling was blocked at these terminals and that for each quantum secreted a vesicle became permanently incorporated into the axolemma.

Differential effect of alpha-latrotoxin on exocytosis from small synaptic vesicles and from large dense-core vesicles containing calcitonin gene-related peptide at the frog neuromuscular junction.

“…5). Furthermore, in agreement with immunofluorescence findings, a morphometric analysis carried out on electron micrographs of nerve terminals revealed that, in spite of the profound (-75%) depletion of SSVs (26), there was no significant change in the number of LDCVs (Table 1). …”

“…The frog neuromuscular junction is the most thoroughly characterized model for the study of quantal release of classical neurotransmitters via SSVs (26)(27)(28). The detection of CGRP-containing LDCVs at these terminals opens the possibility of obtaining important new information on the relation between secretion of peptides and classical neurotransmitters from the same nerve ending.…”

Regulated delivery of AMPA receptor subunits to the presynaptic membrane

“…The regulated, vesicular recruitment of AMPA receptors to the plasma membrane of growth cones was further supported by immunofluorescence experiments carried out in the presence of α‐latrotoxin, a potent stimulator of synaptic vesicle exocytosis (Ceccarelli et al ., 1988; Schiavo et al ., 2000; Sudhof, 2001). When applied to cultured hippocampal neurons in the absence of extracellular calcium, α‐latrotoxin not only induces synaptic vesicle exocytosis, but also prevents synaptic vesicle endocytosis (Pennuto et al ., 2002).…”

Correlation of Miniature Synaptic Activity and Evoked Release Probability in Cultures of Cortical Neurons

“…Miniature release can be regulated in a [Ca 2ϩ ] i -independent manner by agents that directly affect the presynaptic release machinery, e.g., ␣-latrotoxin (Ceccarelli et al, 1988;Capogna et al, 1996a,b) or ruthenium red (Trudeau et al, 1996). However, under physiological conditions, a strong [Ca 2ϩ ] i dependence of miniature synaptic activity is well documented (Matthews and Wickelgren, 1977;Marcus et al, 1992;Doze et al, 1995;Katz et al, 1995;Scanziani et al, 1995;Capogna et al, 1996aCapogna et al, , 1997Poisbeau et al, 1996;Schoppa and Westbrook, 1997;Bao et al, 1998;Li et al, 1998).…”