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Calcium current inactivation rather than pool depletion explains reduced exocytotic rate with prolonged stimulation in insulin-secreting INS-1 832/13 cells.

Pedersen MG, Salunkhe VA, Svedin E, Edlund A, Eliasson L - PLoS ONE (2014)

Bottom Line: We studied exocytosis, measured as increase in membrane capacitance (ΔCm), as a function of calcium entry (Q) in insulin secreting INS-1 832/13 cells using patch clamp and mixed-effects statistical analysis.The latter is attenuated by the calcium-buffer EGTA, while IRP is unaffected.These findings suggest that most insulin release occurs away from Ca2+-channels, and that pool depletion plays a minor role in the decline of exocytosis upon prolonged stimulation.

View Article: PubMed Central - PubMed

Affiliation: Islet Cell Exocytosis, Lund University Diabetes Centre, Department of Clinical Sciences Malmö, Lund University, Malmö, Sweden.

ABSTRACT
Impairment in beta-cell exocytosis is associated with reduced insulin secretion and diabetes. Here we aimed to investigate the dynamics of Ca2+-dependent insulin exocytosis with respect to pool depletion and Ca2+-current inactivation. We studied exocytosis, measured as increase in membrane capacitance (ΔCm), as a function of calcium entry (Q) in insulin secreting INS-1 832/13 cells using patch clamp and mixed-effects statistical analysis. The observed linear relationship between ΔCm and Q suggests that Ca2+-channel inactivation rather than granule pool restrictions is responsible for the decline in exocytosis observed at longer depolarizations. INS-1 832/13 cells possess an immediately releasable pool (IRP) of ∼10 granules and most exocytosis of granules occurs from a large pool. The latter is attenuated by the calcium-buffer EGTA, while IRP is unaffected. These findings suggest that most insulin release occurs away from Ca2+-channels, and that pool depletion plays a minor role in the decline of exocytosis upon prolonged stimulation.

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Related in: MedlinePlus

Mixed-effects analysis of pulse-length data in the control and EGTA groups.The panels show capacitance data plotted against Ca2+ influx measured as charge (Q) from individual cells with single-cell fits indicated by solid lines, while group fits (fixed-effects) are given by the dashed lines. Ctrl (panel 1–18), EGTA (panel 19–34).
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pone-0103874-g002: Mixed-effects analysis of pulse-length data in the control and EGTA groups.The panels show capacitance data plotted against Ca2+ influx measured as charge (Q) from individual cells with single-cell fits indicated by solid lines, while group fits (fixed-effects) are given by the dashed lines. Ctrl (panel 1–18), EGTA (panel 19–34).

Mentions: Likelihood ratio test confirmed that the data did not show evidence of differences between the two EGTA groups with ∼60 nM or ∼460 nM free [Ca2+] (p = 0.89). Moreover, the data could be fitted with a model with common intercept (ΔCm0) for the control and EGTA groups. This intercept reflects exocytosis in the limit of zero Ca2+ entry. The random effects describe cell-deviation from the group estimates for the intercept and Ca2+ current sensitivity (Fig. 2).


Calcium current inactivation rather than pool depletion explains reduced exocytotic rate with prolonged stimulation in insulin-secreting INS-1 832/13 cells.

Pedersen MG, Salunkhe VA, Svedin E, Edlund A, Eliasson L - PLoS ONE (2014)

Mixed-effects analysis of pulse-length data in the control and EGTA groups.The panels show capacitance data plotted against Ca2+ influx measured as charge (Q) from individual cells with single-cell fits indicated by solid lines, while group fits (fixed-effects) are given by the dashed lines. Ctrl (panel 1–18), EGTA (panel 19–34).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4126658&req=5

pone-0103874-g002: Mixed-effects analysis of pulse-length data in the control and EGTA groups.The panels show capacitance data plotted against Ca2+ influx measured as charge (Q) from individual cells with single-cell fits indicated by solid lines, while group fits (fixed-effects) are given by the dashed lines. Ctrl (panel 1–18), EGTA (panel 19–34).
Mentions: Likelihood ratio test confirmed that the data did not show evidence of differences between the two EGTA groups with ∼60 nM or ∼460 nM free [Ca2+] (p = 0.89). Moreover, the data could be fitted with a model with common intercept (ΔCm0) for the control and EGTA groups. This intercept reflects exocytosis in the limit of zero Ca2+ entry. The random effects describe cell-deviation from the group estimates for the intercept and Ca2+ current sensitivity (Fig. 2).

Bottom Line: We studied exocytosis, measured as increase in membrane capacitance (ΔCm), as a function of calcium entry (Q) in insulin secreting INS-1 832/13 cells using patch clamp and mixed-effects statistical analysis.The latter is attenuated by the calcium-buffer EGTA, while IRP is unaffected.These findings suggest that most insulin release occurs away from Ca2+-channels, and that pool depletion plays a minor role in the decline of exocytosis upon prolonged stimulation.

View Article: PubMed Central - PubMed

Affiliation: Islet Cell Exocytosis, Lund University Diabetes Centre, Department of Clinical Sciences Malmö, Lund University, Malmö, Sweden.

ABSTRACT
Impairment in beta-cell exocytosis is associated with reduced insulin secretion and diabetes. Here we aimed to investigate the dynamics of Ca2+-dependent insulin exocytosis with respect to pool depletion and Ca2+-current inactivation. We studied exocytosis, measured as increase in membrane capacitance (ΔCm), as a function of calcium entry (Q) in insulin secreting INS-1 832/13 cells using patch clamp and mixed-effects statistical analysis. The observed linear relationship between ΔCm and Q suggests that Ca2+-channel inactivation rather than granule pool restrictions is responsible for the decline in exocytosis observed at longer depolarizations. INS-1 832/13 cells possess an immediately releasable pool (IRP) of ∼10 granules and most exocytosis of granules occurs from a large pool. The latter is attenuated by the calcium-buffer EGTA, while IRP is unaffected. These findings suggest that most insulin release occurs away from Ca2+-channels, and that pool depletion plays a minor role in the decline of exocytosis upon prolonged stimulation.

Show MeSH
Related in: MedlinePlus