Pressure Pulsatility Links Cardio-Respiratory and Brain Rhythmicity.

This article presents evidence indicating that intracranial pressure (ICP) pulsatility, associated with the heartbeat and breathing, is not just a source of mechanical artefact in electrical recordings, but is "sensed" and plays a role in the brain's information processing. Patch-clamp recording of pressure-activated channels, and detection of Piezo2-protein channel expression in brain neurons, suggest that these channels provide neurons with an intrinsic resonance to ICP pulsatility, which acts to synchronize remote neural networks. Direct measurements in human patients indicate that heartbeat and breathing rhythms generate intracranial forces of tens of millinewtons, exceeding by orders of magnitude the localized forces shown by atomic force microscopy and optical tweezers to activate Piezo channels in isolated neocortical and hippocampal neurons. Additionally, many human touch and proprioceptors, which are also transduced by Piezo channels, show spiking that is phase-locked to heartbeat- and breathing-induced extracranial pressure pulsations. Finally, based on the observation that low-frequency oscillations modulate the phase and amplitude of high-frequency oscillations, body and brain oscillations are proposed to form a single hierarchical system in which the heartbeat is the basic frequency and scaling factor for all other oscillations. Together, these results support the idea that ICP pulsatility may be elemental in modulating the brain's electrical rhythmicity.

Piezo2-peripheral baroreceptor channel expressed in select neurons of the mouse brain: a putative mechanism for synchronizing neural networks by transducing intracranial pressure pulses.

Here we use immunohistochemistry to examine the expression of Piezo2 in neurons of the mouse dorsal root ganglia and brain. Whereas Piezo2 is expressed in the large majority (≥ 90%) of dorsal root ganglia neurons, Piezo2 expression is restricted to select neuron types in specific brain regions, including neocortical and hippocampal pyramidal neurons, cerebellar Purkinje cells and mitral cells of the olfactory bulb. Given the well-established role of Piezo2 as a low-threshold pressure sensor (i.e., ≤5 mmHg) in peripheral mechanosensation, including the regulation of breathing and blood pressure, its expression in central neurons has interesting implications. In particular, we hypothesize that Piezo2 provides neurons with an intrinsic resonance that promotes their entrainment by the normal intracranial pressure pulses (~5 mmHg) associated with breathing and cardiac cycles. The pressure-induced change in neural activity need only be very subtle to increase, for example, the robustness of respiration-entrained oscillations reported previously in widely distributed neuronal networks in both rodent and human brains. This idea of a "global brain rhythm" first arose from the effect of nasal airflow in activating mechanosensitive olfactory sensory neurons, which then synaptically entrain mitral cells within the olfactory bulb and through their projections, neural networks in other brain regions, including the hippocampus and neocortex. Our proposed, non-synaptic, intrinsic mechanism, where Piezo2 tracks the highly predictable and "metronome-like" intracranial pressure pulses-to date generally considered epiphenomena-would have the advantage that a physical force rapidly transmitted throughout the brain also contributes to this synchronization.

Arterial pulses link heart-brain oscillations.

A central baroreceptor monitors arterial pressure to modulate brain activity.

Single mechanosensitive and Ca²âº-sensitive channel currents recorded from mouse and human embryonic stem cells.

Cell-attached and inside-out patch clamp recording was used to compare the functional expression of membrane ion channels in mouse and human embryonic stem cells (ESCs). Both ESCs express mechanosensitive Ca²âº permeant cation channels (MscCa) and large conductance (200 pS) Ca²âº-sensitive K⁺ (BK(Ca²âº)) channels but with markedly different patch densities. MscCa is expressed at higher density in mESCs compared with hESCs (70 % vs. 3 % of patches), whereas the BK(Ca²âº) channel is more highly expressed in hESCs compared with mESCs (~50 % vs. 1 % of patches). ESCs of both species express a smaller conductance (25 pS) nonselective cation channel that is activated upon inside-out patch formation but is neither mechanosensitive nor strictly Ca²âº-dependent. The finding that mouse and human ESCs express different channels that sense membrane tension and intracellular [Ca²âº] may contribute to their different patterns of growth and differentiation in response to mechanical and chemical cues.

TRPC1 forms the stretch-activated cation channel in vertebrate cells.

The mechanosensitive cation channel (MscCa) transduces membrane stretch into cation (Na(+), K(+), Ca(2+) and Mg(2+)) flux across the cell membrane, and is implicated in cell-volume regulation, cell locomotion, muscle dystrophy and cardiac arrhythmias. However, the membrane protein(s) that form the MscCa in vertebrates remain unknown. Here, we use an identification strategy that is based on detergent solubilization of frog oocyte membrane proteins, followed by liposome reconstitution and evaluation by patch-clamp. The oocyte was chosen because it expresses the prototypical MscCa (>or=10(7)MscCa/oocyte) that is preserved in cytoskeleton-deficient membrane vesicles. We identified a membrane-protein fraction that reconstituted high MscCa activity and showed an abundance of a protein that had a relative molecular mass of 80,000 (M(r) 80K). This protein was identified, by immunological techniques, as the canonical transient receptor potential channel 1 (TRPC1). Heterologous expression of the human TRPC1 resulted in a >1,000% increase in MscCa patch density, whereas injection of a TRPC1-specific antisense RNA abolished endogenous MscCa activity. Transfection of human TRPC1 into CHO-K1 cells also significantly increased MscCa expression. These observations indicate that TRPC1 is a component of the vertebrate MscCa, which is gated by tension developed in the lipid bilayer, as is the case in various prokaryotic mechanosensitive (Ms) channels.

PIEZO1 Is Selectively Expressed in Small Diameter Mouse DRG Neurons Distinct From Neurons Strongly Expressing TRPV1.

Using a high resolution in situ hybridization technique we have measured PIEZO1, PIEZO2, and TRPV1 transcripts in mouse dorsal root ganglion (DRG) neurons. Consistent with previous studies, PIEZO2 transcripts were highly expressed in DRG neurons of all sizes, including most notably the largest diameter neurons implicated in mediating touch and proprioception. In contrast, PIEZO1 transcripts were selectively expressed in smaller DRG neurons, which are implicated in mediating nociception. Moreover, the small neurons expressing PIEZO1 were mostly distinct from those neurons that strongly expressed TRPV1, one of the channels implicated in heat-nociception. Interestingly, while PIEZO1- and TRPV1- expressing neurons form essentially non-overlapping populations, PIEZO2 showed co-expression in both populations. Using an in vivo functional test for the selective expression, we found that Yoda1, a PIEZO1-specific agonist, induced a mechanical hyperalgesia that displayed a significantly prolonged time course compared with that induced by capsaicin, a TRPV1-specific agonist. Taken together, our results indicate that PIEZO1 should be considered a potential candidate in forming the long sought channel mediating mechano-nociception.

TRPCs as MS Channels.

This chapter reviews recent evidence indicating that canonical or classical transient receptor potential (TRPC) channels are directly or indirectly mechanosensitive (MS) and can therefore be designated as mechano-operated channels (MOCs). The MS functions of TRPCs may be mechanistically related to their better known functions as store-operated and receptor-operated channels (SOCs and ROCs). Mechanical forces may be conveyed to TRPC channels through the "conformational coupling" mechanism that transmits information regarding the status of internal Ca(2+) stores. All TRPCs are regulated by receptors coupled to phospholipases that are themselves MS and can regulate channels via lipidic second messengers. Accordingly, there may be several nonexclusive mechanisms by which mechanical forces may regulate TRPC channels, including direct sensitivity to bilayer mechanics, physical coupling to internal membranes and/or cytoskeletal proteins, and sensitivity to lipidic second messengers generated by MS enzymes. Various strategies that can be used for separating out different MS-gating mechanisms and their possible role in specific TRPCs are discussed.

MscCa Regulation of Tumor Cell Migration and Metastasis.

The acquisition of cell motility is a required step in order for a cancer cell to migrate from the primary tumor and spread to secondary sites (metastasize). For this reason, blocking tumor cell migration is considered a promising approach for preventing the spread of cancer. However, cancer cells just as normal cells can migrate by several different modes referred to as "amoeboid," "mesenchymal," and "collective cell." Under appropriate conditions, a single cell can switch between modes. A consequence of this plasticity is that a tumor cell may be able to avoid the effects of an agent that targets only one mode by switching modes. Therefore, a preferred strategy would be to target mechanisms that are shared by all modes. This chapter reviews the evidence that Ca(2+) influx via the mechanosensitive Ca(2+)-permeable channel (MscCa) is a critical regulator of all modes of cell migration and therefore represents a very good therapeutic target to block metastasis.

Single mechanically-gated cation channel currents can trigger action potentials in neocortical and hippocampal pyramidal neurons.

The mammalian brain is a mechanosensitive organ that responds to different mechanical forces ranging from intrinsic forces implicated in brain morphogenesis to extrinsic forces that can cause concussion and traumatic brain injury. However, little is known of the mechanosensors that transduce these forces. In this study we use cell-attached patch recording to measure single mechanically-gated (MG) channel currents and their affects on spike activity in identified neurons in neonatal mouse brain slices. We demonstrate that both neocortical and hippocampal pyramidal neurons express stretch-activated MG cation channels that are activated by suctions of ~25mm Hg, have a single channel conductance for inward current of 50-70pS and show weak selectivity for alkali metal cations (i.e., Na(+)

ATP suppresses activity in slowly adapting but not rapidly adapting mechanoreceptors in toad skin.

To test the possible role of ATP in transducing or modulating touch sensation, an isolated skin-nerve preparation from the toad, Bufo marinus, perfused on the inner side, was used to examine the effects of ATP on slowly adapting (SA) and rapidly adapting (RA) mechanoreceptors, identified by ramp and hold indentation of the skin. ATP (1-50 mM) did not generate impulse activity in either SA or RA mechanoreceptors nor increase their responses to mechanical stimulation. Instead, ATP (> or =5 mM) reversibly and selectively suppressed impulse discharge from SA mechanoreceptors. Our results indicate that while ATP may modulate the responses of specific touch receptors, it is not involved in transducing touch into nerve impulses.

Mechanosensitive Ca(2+) permeant cation channels in human prostate tumor cells.

The acquisition of cell motility plays a critical role in the spread of prostate cancer (PC), therefore, identifying a sensitive step that regulates PC cell migration should provide a promising target to block PC metastasis. Here, we report that a mechanosensitive Ca(2+)-permeable cation channel (MscCa) is expressed in the highly migratory/invasive human PC cell line, PC-3 and that inhibition of MscCa by Gd(3+) or GsMTx-4 blocks PC-3 cell migration and associated elevations in [Ca(2+)](i). Genetic suppression or overexpression of specific members of the canonical transient receptor potential Ca(2+) channel family (TRPC1 and TRPC3) also inhibit PC-3 cell migration, but they do so by mechanisms other that altering MscCa activity. Although LNCaP cells are nonmigratory, they also express relatively large MscCa currents, indicating that MscCa expression alone cannot confer motility on PC cells. MscCa in both cell lines show similar conductance and ion selectivity and both are functionally coupled via Ca(2+) influx to a small Ca(2+)-activated K(+) channel. However, MscCa in PC-3 and LNCaP cell patches show markedly different gating dynamics--while PC-3 cells typically express a sustained, non-inactivating MscCa current, LNCaP cells express a mechanically-fragile, rapidly inactivating MscCa current. Moreover, mechanical forces applied to the patch, can induce an irreversible transition from the transient to the sustained MscCa gating mode. Given that cancer cells experience increasing compressive and shear forces within a growing tumor, a similar shift in channel gating in situ would have significant effects on Ca(2+) signaling that may play a role in tumor progression.

Revisiting TRPC1 and TRPC6 mechanosensitivity.

This article addresses whether TRPC1 or TRPC6 is an essential component of a mammalian stretch-activated mechano-sensitive Ca(2+) permeable cation channel (MscCa). We have transiently expressed TRPC1 and TRPC6 in African green monkey kidney (COS) or Chinese hamster ovary (CHO) cells and monitored the activity of the stretch-activated channels using a fast pressure clamp system. Although both TRPC1 and TRPC6 are highly expressed at the protein level, the amplitude of the mechano-sensitive current is not significantly altered by overexpression of these subunits. In conclusion, although several TRPC channel members, including TRPC1 and TRPC6, have been recently proposed to form MscCa in vertebrate cells, the functional expression of these TRPC subunits in heterologous systems remains problematic.

Gramicidin A channels switch between stretch activation and stretch inactivation depending on bilayer thickness.

The patch clamp-liposome technique was used to examine the stretch sensitivity of a model membrane ion channel, gramicidin A, in membrane patches of different bilayer thickness. We found that small changes in phospholipid acyl chain length (i.e., PC-20 to PC-18) can switch gramicidin A from a stretch-activated to a stretch-inactivated channel. The demonstration that subnanometer changes in bilayer thickness can reverse the response polarity of a model channel has implications for other signaling proteins that may experience local changes in bilayer thickness as a consequence of dynamic targeting to lipid microdomains, electrocompression, or chemical modification of the bilayer.