Gating the mechanical channel Piezo1

Abstract

Piezo1 is a eukaryotic cation-selective mechanosensitive ion channel. To understand channel function in vivo, we first need to analyze and compare the response in the whole cell and the patch. In patches, Piezo1 inactivates and the current is fit well by a 3-state model with a single pressure-dependent rate. However, repeated stimulation led to an irreversible loss of inactivation. Remarkably, the loss of inactivation did not occur on a channel-by-channel basis but on all channels at the same time. Thus, the channels are in common mechanical domain. Divalent ions decreased the unitary conductance from ~68 pS to ~37 pS, irrespective of the cation species. Mg and Ca did not affect inactivation rates, but Zn caused a 3-fold slowing. CytochalasinD (cytoD) does not alter inactivation rates or the transition to the non-inactivating mode but does reduce the steady-state response. Whole-cell currents were similar to patch currents but also had significant differences. In contrast to the patch, cytoD inhibited the current suggesting that the activating forces were transmitted through the actin cytoskeleton. Hypotonic swelling that prestressed the cytoskeleton and the bilayer greatly increased the sensitivity of both control and cytoD cells so there are two pathways to transmit force to the channels. In contrast to patch, removing divalent ions decreased the whole-cell current. The difference between whole cell and patch properties provide new insights into our understanding of the Piezo1 gating mechanisms and cautions against generalization to in situ behavior.

The biophysics of piezo1 and piezo2 mechanosensitive channels

“…In cell-attached patches, divalent ions reduced the single channel conductance by about 50% for Mg 2+ , Ca 2+ and Zn 2+ (Coste et al, 2012;Gottlieb et al, 2012). In addition, divalent cations slowed the time course of inactivation (Gottlieb et al, 2012). Moreover, in whole-cell recording, extracellular divalent ions were reported to be required for channel activity evoked by cell indentation (Gottlieb et al, 2012).…”

“…The supramolecular organization of the functional piezo unit is expected to be tightly linked with its ability to respond to external mechanical stimuli, but the details of this mechanism have not yet been fully disclosed. In general, the activation and inactivation of the currents seem to be associated with the presence of whole tetramers, but two different perspectives have been proposed (Gottlieb et al, 2012;Bae et al, 2013a). One possible picture is that one central pore is formed at the boundary of the monomers.…”

“…In cell-attached patches, divalent ions reduced the single channel conductance by about 50% for Mg 2+ , Ca 2+ and Zn 2+ (Coste et al, 2012;Gottlieb et al, 2012). In addition, divalent cations slowed the time course of inactivation (Gottlieb et al, 2012).…”

“…In addition, divalent cations slowed the time course of inactivation (Gottlieb et al, 2012). Moreover, in whole-cell recording, extracellular divalent ions were reported to be required for channel activity evoked by cell indentation (Gottlieb et al, 2012).…”

“…From a more detailed kinetic analysis, Gottlieb et al (2012) proposed that piezo1 gating kinetics could be well fit by a 3 states model: closed, open and inactivated. However, repeated cell stimulations by high pressure (positive or negative) and high voltage led to a gradual decrease of the degree of inactivation (Gottlieb et al, 2012).…”