Antibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells.
SNARE-proteins (soluble NSF-attachment protein receptor) are important for Ca(2+)-dependent exocytosis. We have used capacitance measurements and confocal imaging to dissect the role of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 in rapid exocytosis in insulin-secreting pancreatic beta-c...
| Main Authors: | , , , , |
|---|---|
| Format: | Journal article |
| Language: | English |
| Published: |
2006
|
| _version_ | 1797057937316773888 |
|---|---|
| author | Vikman, J Ma, X Hockerman, G Rorsman, P Eliasson, L |
| author_facet | Vikman, J Ma, X Hockerman, G Rorsman, P Eliasson, L |
| author_sort | Vikman, J |
| collection | OXFORD |
| description | SNARE-proteins (soluble NSF-attachment protein receptor) are important for Ca(2+)-dependent exocytosis. We have used capacitance measurements and confocal imaging to dissect the role of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 in rapid exocytosis in insulin-secreting pancreatic beta-cells. Following immunoneutralization of syntaxin 1 and SNAP-25, exocytosis was strongly reduced and associated with a marked reduction in the size of the readily releasable pool (RRP) by 65% and 86% in the presence of the anti-SNAP-25 and anti-syntaxin 1 antibodies respectively. The size of the immediately releasable pool (IRP), a subset of RRP in close association with the voltage-dependent Ca(2+)-channels, was reduced to an equal extent. The reduction in IRP correlated with slowed release kinetics and the time constant (tau) increased from a control value of 16 to 36 ms and 51 ms after inclusion of anti-SNAP-25 and anti-syntaxin 1 antibodies respectively in the pipette solution. We further show that SNAP-25 and syntaxin 1 aggregate in clusters along the plasma membrane. The size of these clusters was estimated to be approximately 300 nm and every beta-cell contained approximately 400 SNAP-25/syntaxin 1 clusters. Whereas the inhibitory action of the anti-syntaxin 1 antibody on exocytosis could be attributed almost entirely to suppression of the voltage-dependent Ca(2+)-current (-40%), the effect of the anti-SNAP-25 antibody was not mediated by decreased Ca(2+)-entry and is more likely due to a direct interference with the exocytotic machinery. Our data are consistent with the concept that both syntaxin 1 and SNAP-25 are required for rapid exocytosis in beta-cells. |
| first_indexed | 2024-03-06T19:43:27Z |
| format | Journal article |
| id | oxford-uuid:2173ce08-b7a2-4160-ae11-7453a86584fc |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-06T19:43:27Z |
| publishDate | 2006 |
| record_format | dspace |
| spelling | oxford-uuid:2173ce08-b7a2-4160-ae11-7453a86584fc2022-03-26T11:33:32ZAntibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:2173ce08-b7a2-4160-ae11-7453a86584fcEnglishSymplectic Elements at Oxford2006Vikman, JMa, XHockerman, GRorsman, PEliasson, LSNARE-proteins (soluble NSF-attachment protein receptor) are important for Ca(2+)-dependent exocytosis. We have used capacitance measurements and confocal imaging to dissect the role of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 in rapid exocytosis in insulin-secreting pancreatic beta-cells. Following immunoneutralization of syntaxin 1 and SNAP-25, exocytosis was strongly reduced and associated with a marked reduction in the size of the readily releasable pool (RRP) by 65% and 86% in the presence of the anti-SNAP-25 and anti-syntaxin 1 antibodies respectively. The size of the immediately releasable pool (IRP), a subset of RRP in close association with the voltage-dependent Ca(2+)-channels, was reduced to an equal extent. The reduction in IRP correlated with slowed release kinetics and the time constant (tau) increased from a control value of 16 to 36 ms and 51 ms after inclusion of anti-SNAP-25 and anti-syntaxin 1 antibodies respectively in the pipette solution. We further show that SNAP-25 and syntaxin 1 aggregate in clusters along the plasma membrane. The size of these clusters was estimated to be approximately 300 nm and every beta-cell contained approximately 400 SNAP-25/syntaxin 1 clusters. Whereas the inhibitory action of the anti-syntaxin 1 antibody on exocytosis could be attributed almost entirely to suppression of the voltage-dependent Ca(2+)-current (-40%), the effect of the anti-SNAP-25 antibody was not mediated by decreased Ca(2+)-entry and is more likely due to a direct interference with the exocytotic machinery. Our data are consistent with the concept that both syntaxin 1 and SNAP-25 are required for rapid exocytosis in beta-cells. |
| spellingShingle | Vikman, J Ma, X Hockerman, G Rorsman, P Eliasson, L Antibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells. |
| title | Antibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells. |
| title_full | Antibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells. |
| title_fullStr | Antibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells. |
| title_full_unstemmed | Antibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells. |
| title_short | Antibody inhibition of synaptosomal protein of 25 kDa (SNAP-25) and syntaxin 1 reduces rapid exocytosis in insulin-secreting cells. |
| title_sort | antibody inhibition of synaptosomal protein of 25 kda snap 25 and syntaxin 1 reduces rapid exocytosis in insulin secreting cells |
| work_keys_str_mv | AT vikmanj antibodyinhibitionofsynaptosomalproteinof25kdasnap25andsyntaxin1reducesrapidexocytosisininsulinsecretingcells AT max antibodyinhibitionofsynaptosomalproteinof25kdasnap25andsyntaxin1reducesrapidexocytosisininsulinsecretingcells AT hockermang antibodyinhibitionofsynaptosomalproteinof25kdasnap25andsyntaxin1reducesrapidexocytosisininsulinsecretingcells AT rorsmanp antibodyinhibitionofsynaptosomalproteinof25kdasnap25andsyntaxin1reducesrapidexocytosisininsulinsecretingcells AT eliassonl antibodyinhibitionofsynaptosomalproteinof25kdasnap25andsyntaxin1reducesrapidexocytosisininsulinsecretingcells |