Thermosensitive transient receptor potential channels in human corneal epithelial cells

Abstract

Thermosensitive transient receptor potential proteins (TRPs) such as TRPV1-TRPV4 are all heat-activated non-selective cation channels that are modestly permeable to Ca2+. TRPV1, TRPV3 and TRPV4 functional expression were previously identified in human corneal epithelial cells (HCEC). However, the membrane currents were not described underlying their activation by either selective agonists or thermal variation. This study characterized the membrane currents and [Ca 2+]i transients induced by thermal and agonist TRPV1 and 4 stimulation. TRPV1 and 4 expressions were confirmed by RT-PCR and TRPV2 transcripts were also detected. In fura2-loaded HCEC, a TRPV1-3 selective agonist, 100 µM 2-aminoethoxydiphenyl borate (2-APB), induced intracellular Ca2+ transients and an increase in non-selective cation outward currents that were suppressed by ruthenium-red (RuR) (10–20 µM), a nonselective TRPV channel blocker. These changes were also elicited by rises in ambient temperature from 25 °C to over 40 °C. RuR (5 µM) and a selective TRPV1 channel blocker capsazepine (CPZ) (10 µM) or another related blocker, lanthanum chloride (La3+) (100 µM) suppressed these temperature-induced Ca2+ increases. Planar patch-clamp technique was used to characterize the currents underlying Ca2+ transients. Increasing the temperature to over 40 °C induced reversible rises in non-selective cation currents. Moreover, a hypotonic challenge (25 %) increased non-selective cation currents confirming TRPV4 activity. We conclude that HCEC possess in addition to thermo-sensitive TRPV3 activity TRPV1, TRPV2 and TRPV4 activity. Their activation confers temperature sensitivity at the ocular surface, which may protect the cornea against such stress.

Hyperosmotic Stress–Induced TRPM2 Channel Activation Stimulates NLRP3 Inflammasome Activity in Primary Human Corneal Epithelial Cells

“…12 In HCECs, some of the members of the TRPV, TRPC, TRPM, and TRPA subfamilies mediate responses to environmental challenges that affect corneal epithelial layer renewal and function. 13,14 TRPV1, TRPV2, TRPM8, and TRPA1 sense noxious heat, moderate cold, and more extreme cold, respectively, in this tissue.…”

Upregulation of Transient Receptor Potential Vanilloid Type-1 Channel Activity and Ca2+Influx Dysfunction in Human Pterygial Cells

“…In Drosophila, photoreceptors and mammalian photosensitive retinal ganglion cell phototransduction is mediated by G protein-activated phospholipase C (PLC), which generates a force in the lipid-bilayer opening TRP-Canonical channels. 23,24 Transient receptor potential V1 is expressed in human conjunctival epithelial cells (HCjEC), 25 human corneal endothelial cells, 26 corneal fibroblasts, 27 and corneal epithelial cells (HCEC) 28,29 (reviewed in Ref. 30 ).…”

Novel screening techniques for ion channel targeting drugs

“…Cell lines such as HEK and CHO cells are used with equal success rates and experiments can be performed at room or physiological temperature. 8 Additionally, primary cells such as human corneal epithelial cells, 9 stem cellderived cardiomyocytes 10 have been used successfully. As the data recorded is low noise and high resolution, single channel recordings in the cell attached mode on the cell membrane, and experiments recording ion channels reconstituted into lipid bilayers, are also routinely performed using the Port-a-Patch.…”