Acetylcholine exocytosis in PC12 cells deficient in SNAP-25.
Stimulus-induced acetylcholine (ACh) exocytosis from presynaptic nerve terminals involves two important steps: fusion of ACh loaded vesicles at presynaptic release sites, followed by release into the synaptic cleft. We studied the role of the putative vesicle fusion protein SNAP-25 in this process. The nerve growth factor-differentiated PC12 cell line was used as an experimental model. A bee venom tetradecapeptide (INLKALAALAKKIL-NH2) phospholipase A2 (PLA2) activator, mastoparan, was used to induce ACh release. Treatment of PC12 cells with appropriate antisense oligonucleotides blocked SNAP-25 expression, as judged by Western blot protein analysis with a specific monoclonal antibody. Despite apparent elimination of SNAP-25, treatment of differentiated PC12 cells with mastoparan and high (80 mM) K+ induced ACh exocytosis. The results indicate that in PC12 cells, ACh exocytosis due to mastoparan plus K+ can occur in the absence of SNAP-25.
Ray
P
P
Biology Department, Walter Reed Army Institute of Research, Washington, DC 20307-5100, USA.
Millard
C B
CB
Petrali
J P
JP
Berman
J D
JD
Ray
R
R
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009498:Neurotoxins / Q000494:pharmacology; D016716:PC12 Cells / Q000378:metabolism*; D010455:Peptides; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D050825:Synaptosomal-Associated Protein 25; D014862:Wasp Venoms / Q000494:pharmacology
D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D009498:Neurotoxins; D010455:Peptides; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D014862:Wasp Venoms; D011189:Potassium Chloride; C037500:mastoparan; D000109:Acetylcholine