TRPC1, stretch, adipocytes, and PEMF

This post proposes that PEMF may do something holistic for adipocytes. Many PEMF users may feel gentle mechanical vibrations. The TRPC1 and PEMF post analyzed the hypothesis that TRPC1 may be regulated by reactive oxygen species from PEMF. This post proposes the gentle vibration we feel with PEMF may do something for this mechano-sensitive channel called TRPC1, transient receptor potential C1, a Ca²⁺ and other cation channel.

A link to the companion YouTube video.

TRPC1 stretch sensitivity derived from membranes

Patch clamping is a technique that uses an electrode to measure the conductance through an ion channel on a patch of a cell membrane. The voltage (V) across the ion channel is clamped at a given value and the current (I) measured. The resistance of tehi ion channel (R) is calculated with this equation: V=IR.

The Moroto 2005 study identified a stretch activated non specific cation channel in Xenopus (frog) oocytes. There was a prior notion that this channel was TRPC1. Membranes were divided into fractions with the stretch activated conductance residing in the fraction containing TRPC1 antibody reactive protein. The interesting twist to this study is that the authors used the patch clamp electrode to apply the pressure:

The patch pipette tip was gently touched against unilamellar blisters, which arise spontaneously from the liposomes in the high MgCl₂ solution, and suction applied. Seals (>20 GΩ) either formed immediately or after application of a brief pulse of negative pressure (<50 mmHg) applied to the interior of the patch pipette. To record single-channel currents, inside-out patches were formed by passing the pipette tip briefly through the solution–air interface.

Applying pressure to the membrane is part of the patch clamping technique. Channels were activated by passing suction pulses of –10 to –200 mmHg by mouth or syringe. Applied pressure was measured with a piezoelectric device.

Patch clamp stressing TRPC1 in adipocytes

The El Hachmane 2018 study was based on the observation that white adipocytes rapidly taking up fats may experience membrane tension that lets in Ca²⁺. El Hachmane and coauthors cited prior work stating that the release of leptin and adiponectin is a a Ca²⁺ dependent process. Previous work from this group had implicated ORAI1 and TRPC1. ORAI1 is a hexamer that is opened when intracellular Ca²⁺ stores in the endoplasmic reticulum are depleted. The stromal interacting molecule 1 (STIM1) communicates this Ca²⁺ depletion.

Summary of patch clamp results

1. Extracellular ATP stimulates store depletion operated Ca²⁺ entry (SOCE) through ORAI1 with contribution of TRPC1. The ATP part is suggestive of P2Y2 purinergic receptor G protein coupled receptor signaling.
2. Perforated whole cell patch clamping, which involves mechanical insertion of a glass electrode into the cell membrane, was used to measure currents with the TRPC1 inhibitor 2-ABP and store operated Ca²⁺ inhibitor Gd³⁺ suggested that the mechanics of insertion opened up TRPC1 channels.
3. Store operated Ca²⁺ channels are influenced by phospholipase C. Inhibiting this enzyme decreases the current.
4. A Ca²⁺ fluorescent dye was used to prove that the act of patch clamping increased intracellular Ca²⁺.
5. The direction of Ca²⁺/Na⁺ conductance through TRPC1 is about 0 mV. The more negative reversal potential was suggestive of and proven to be due to chloride channels running in parallel.

This is a beautiful study yet focused on just one job of adipocytes: storing fat and expanding in the process.

Fibroblasts, precursors to adipocytes, and caveolin

Adipocytes also take up glucose via glucose transporter glut4. (Kanzaki 2006) This publication has a fluorescent microscope image of lipid raft domain protein caveolin associating with cytoskeleton protein F-actin for the purpose of trafficking glut4 containing lipid vesicles to the cell membranes.

Caveolin binds TRPC1 and the IP3 receptor

And the story gets far more complicated. Cantu 2021 exposed 3T3 fibroblasts to pulsed electric fields of ~16.2 kV/cm at the cell. Pulses were 0 to 150 pulses at an amplitude of 10 kV/cm and a 1 Hz repetition rate. The expression of caveolin 1 was silenced with siRNA. A construct of caveolin-green fluorescence protein was used to visualize the effect of pulsed electric fields on membrane localization of the caveolin in response to pulsed electric fields. The working hypothesis developed in this study is that caveolin binds both TRPC1 and the IP₃ receptor of the endoplasmic reticulum such they cannot interact. Mechanical influences of the pulsed electric fields causes both to dissociate from caveolin and interact with each other. Cantu 2021 further state in their review of the literature that both the C and N-terminus of TRPC1 interacts with caveolin 1. Figure 6 of this publication illustrates TRPC1 and the IP3R binding to opposite ends of the Cav1 hairpin, both of which ends are on the cytoplasmic side of the cell membrane.

The Cantu 2021 discussion also mentioned literature stating that mechanical stretching inhibited G protein-coupled receptor—Gaq (GPCR-G_aq) Ca²⁺ signaling by flattening the caveolae.

Thought Experiment summary

A Wikimedia Commons image of a Haboob dust storm is combined with images of some adipocytes. PEMF and nanosecond pulsed electric fields may set off intermittent waves of, not dust particles, but charged ions. In the photo there is the framework of a fence and some trees. Let the metal fence be the actin cytoskeleton that is connected to the caveolin complex. Let the trees be surface glycosylation and any other external parts of proteins that are out there catching the charged particle breeze.

The suction action of patch clamping opens TRPC1 channels. Is there a “suction” of pulsed flow of charged particles? The mechanical aspects of nsPEF causes TRPC1 and the IP₃ receptor from caveolin complexes so that they might associate with one another. Can very low frequency PEMF also serve this function? The Ca²⁺ signaling leads to release of healthy adipokines leptin and adipokine. So much remains to be proven about the timing of these waves of charged particles when it comes to this signaling.

What about the actin that forms a “highway” for glut4 vesicles that enable adipocytes to take up glucose? Can nsPEF and/or PEMF cause the caveolin complex to release actin and the membrane insertion machinery? If this is the case, what could it mean for adipocyte function?

References

• Cantu JC, Tolstykh GP, Tarango M, Beier HT, Ibey BL. (2021) Caveolin-1 is Involved in Regulating the Biological Response of Cells to Nanosecond Pulsed Electric Fields. J Membr Biol. 2021 Apr;254(2):141-156. Sci-Hub free paper
• El Hachmane MF, Olofsson CS. (2018) A mechanically activated TRPC1-like current in white adipocytes. Biochem Biophys Res Commun. 2018 Apr 15;498(4):736-742. Sci-hub free paper
• Kanzaki M. (2006) Insulin receptor signals regulating GLUT4 translocation and actin dynamics. Endocr J. 2006 Jun;53(3):267-93. free paper
• Maroto, R et al (2005) TRPC1 forms the stretch-activated cation channel in vertebrate cells. Nat. Cell Biol. 7, 179 –185 PMC free paper