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Nat Commun. 2012 Apr 10;3:778. doi: 10.1038/ncomms1777.

Nanodomain Ca²⁺ of Ca²⁺ channels detected by a tethered genetically encoded Ca²⁺ sensor.

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Department of Biomedical Engineering, Calcium Signals Laboratory, Ross Building, Room 713, 720 Rutland Avenue, Baltimore, Maryland 21205-2196, USA.


Coupling of excitation to secretion, contraction and transcription often relies on Ca(2+) computations within the nanodomain-a conceptual region extending tens of nanometers from the cytoplasmic mouth of Ca(2+) channels. Theory predicts that nanodomain Ca(2+) signals differ vastly from the slow submicromolar signals routinely observed in bulk cytoplasm. However, direct visualization of nanodomain Ca(2+) far exceeds optical resolution of spatially distributed Ca(2+) indicators. Here we couple an optical, genetically encoded Ca(2+) indicator (TN-XL) to the carboxy tail of Ca(V)2.2 Ca(2+) channels, enabling near-field imaging of the nanodomain. Under total internal reflection fluorescence microscopy, we detect Ca(2+) responses indicative of large-amplitude pulses. Single-channel electrophysiology reveals a corresponding Ca(2+) influx of only 0.085 pA, and fluorescence resonance energy transfer measurements estimate TN-XL distance to the cytoplasmic mouth at ~55 Å. Altogether, these findings raise the possibility that Ca(2+) exits the channel through the analogue of molecular portals, mirroring the crystallographic images of side windows in voltage-gated K channels.

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