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Analysis of neuronal Ca 2+ handling properties by combining perforated patch clamp recordings and the added buffer approach

Abstract : Ca 2+ functions as an important intracellular signal for a wide range of cellular processes. These processes are selectively activated by controlled spatiotemporal dynamics of the free cytosolic Ca 2+ . Intracellular Ca 2+ dynamics are regulated by numerous cellular parameters. Here, we established a new way to determine neuronal Ca 2+ handling properties by combining the ‘added buffer’ approach (Neher and Augustine, 1992) with perforated patch-clamp recordings (Horn and Marty, 1988). Since the added buffer approach typically employs the standard whole-cell configuration for concentration-controlled Ca 2+ indicator loading, it only allows for the reliable estimation of the immobile fraction of intracellular Ca 2+ buffers. Furthermore, crucial components of intracellular signaling pathways are being washed out during prolonged whole-cell recordings, leading to cellular deterioration. By combining the added buffer approach with perforated patch-clamp recordings, these issues are circumvented, allowing the precise quantification of the cellular Ca 2+ handling properties, including immobile as well as mobile Ca 2+ buffers.
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https://hal.archives-ouvertes.fr/hal-03031583
Contributor : Christophe Pouzat <>
Submitted on : Monday, November 30, 2020 - 3:19:54 PM
Last modification on : Tuesday, December 1, 2020 - 9:18:33 AM

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Simon Hess, Christophe Pouzat, Lars Paeger, Andreas Pippow, Peter Kloppenburg. Analysis of neuronal Ca 2+ handling properties by combining perforated patch clamp recordings and the added buffer approach. 2020. ⟨hal-03031583⟩

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