Patch-clamp capacitance analysis of the effects of alpha-SNAP on exocytosis in adrenal chromaffin cells.
We have examined the effect of alpha-SNAP on exocytosis in adrenal chromaffin cells by direct assay of exocytosis using patch-clamp capacitance analysis. Cells were recorded using the whole cell patch-clamp configuration and the cells dialysed with control pipette solution or with a pipette solution containing alpha-SNAP or the deletion mutant alpha-SNAP(41-295). The deletion mutant was found to be unable to bind to syntaxin allowing a test of the requirement for syntaxin-binding for any effect of alpha-SNAP on exocytosis. Following cell dialysis for 10 minutes, cells were depolarised five times at 2 minute intervals. At each depolarisation step cells dialysed with alpha-SNAP showed a significant increase in both the initial rate and extent of exocytosis which was seen as a rise in membrane capacitance. This increase in exocytosis was not observed with alpha-SNAP(41-295) which instead produced some inhibition of the extent but had no effect on the initial rate of exocytosis. These results show directly that alpha-SNAP has a specific and marked stimulatory effect on exocytosis in chromaffin cells.
Kibble
A V
AV
Physiological Laboratory, University of Liverpool, UK.
Barnard
R J
RJ
Burgoyne
R D
RD
D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D002417:Cattle; D019439:Chromaffin Cells / Q000187:drug effects* / Q000502:physiology*; D017931:DNA Primers / Q000235:genetics; D005089:Exocytosis / Q000187:drug effects*; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D018408:Patch-Clamp Techniques; D050765:Qa-SNARE Proteins; D017384:Sequence Deletion; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D033921:Vesicular Transport Proteins
D002352:Carrier Proteins; D017931:DNA Primers; D008565:Membrane Proteins; D050765:Qa-SNARE Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D033921:Vesicular Transport Proteins