The sensitivity of catecholamine release to botulinum toxin C1 and E suggests selective targeting of vesicles set into the readily releasable pool.
The impact of syntaxin and SNAP-25 cleavage on [3H]noradrenaline ([3H]NA) and [3H]dopamine ([3H]DA) exocytotic release evoked by different stimuli was studied in superfused rat synaptosomes. The external Ca2+-dependent K+-induced [3H]catecholamine overflows were almost totally abolished by botulinum toxin C1 (BoNT/C1), which hydrolyses syntaxin and SNAP-25, or by botulinum toxin E (BoNT/E), selective for SNAP-25. BoNT/C1 cleaved 25% of total syntaxin and 40% of SNAP-25; BoNT/E cleaved 40% of SNAP-25 but left syntaxin intact. The GABA uptake-induced releases of [3H]NA and [3H]DA were differentially affected: both toxins blocked the former, dependent on external Ca2+, but not the latter, internal Ca2+-dependent. BoNT/C1 or BoNT/E only slightly reduced the ionomycin-evoked [3H]catecholamine release. More precisely, [3H]NA exocytosis induced by ionomycin was sensitive to toxins in the early phase of release but not later. The Ca2+-independent [3H]NA exocytosis evoked by hypertonic sucrose, thought to release from the readily releasable pool (RRP) of vesicles, was significantly reduced by BoNT/C1. Pre-treating synaptosomes with phorbol-12-myristate-13-acetate, to increase the RRP, enhanced the sensitivity to BoNT/C1 of [3H]NA release elicited by sucrose or ionomycin. Accordingly, cleavage of syntaxin was augmented by the phorbol-ester. To conclude, our results suggest that clostridial toxins selectively target exocytosis involving vesicles set into the RRP.
Stigliani
Sara
S
Section of Pharmacology and Toxicology, Department of Experimental Medicine, University of Genova Center of Excellence for Biomedical Research, Genova, Italy.
Raiteri
Luca
L
Fassio
Anna
A
Bonanno
Giambattista
G
D000818:Animals; D001905:Botulinum Toxins / Q000494:pharmacology*; D001923:Brain Chemistry / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002395:Catecholamines / Q000032:analysis / Q000378:metabolism*; D004298:Dopamine / Q000032:analysis / Q000378:metabolism; D005089:Exocytosis / Q000187:drug effects; D015759:Ionomycin / Q000494:pharmacology; D008297:Male; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000187:drug effects / Q000378:metabolism; D009638:Norepinephrine / Q000032:analysis / Q000378:metabolism; D011188:Potassium / Q000494:pharmacology; D050765:Qa-SNARE Proteins; D051381:Rats; D013395:Sucrose / Q000494:pharmacology; D013572:Synaptic Vesicles / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism*; D050825:Synaptosomal-Associated Protein 25; D013574:Synaptosomes / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
D002395:Catecholamines; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050765:Qa-SNARE Proteins; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D005680:gamma-Aminobutyric Acid; D015759:Ionomycin; D013395:Sucrose; D001905:Botulinum Toxins; C096324:botulinum toxin type C; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium; D002118:Calcium; C096326:botulinum toxin type E; D004298:Dopamine; D009638:Norepinephrine