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

ABSTRACT: 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.

SUBMITTER: Tay LH 

PROVIDER: S-EPMC3615648 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature