Cell-specific differential modulation of human trabecular meshwork cells by selective adenosine receptor agonists

Abstract

Activation of A 1 and A 2A subtype adenosine receptors (AR) likely exert opposing effects on outflow of aqueous humor, and thereby, on intraocular pressure. Selective agonists of adenosine receptor (AR) subtypes have previously been applied to trabecular meshwork (TM) and Schlemm's canal (SC) cells to identify the site(s) of differential purinergic modulation. However, the apparent changes in volume monitored by previously measuring projected cell area might have partially reflected cell contraction and relaxation. In addition, whole-cell current responses of the TM cells previously described were highly variable following application of selective A 1 , A 2A and A 3 agonists. The complexity of the electrophysiologic responses may have reflected cell heterogeneity of the populations harvested from collagenase digestion of TM explants. We now report measurements of TM-cell volume using calcein fluorescence quenching, an approach independent of contractile state. Furthermore, we have applied selective AR agonists to a uniform population of human TM cells, the hTM5 cell line. A 1 , but not A 2A or A 3 , AR agonists triggered TM-cell shrinkage. Both A 1 and A 2A AR agonists produced reproducible increases in TM-cell whole-cell currents of similar magnitude.The results suggest that previous measurements of explant-derived TM cells may have reflected a range of responses from phenotypically different cell populations, and that the opposing effects of A 1 and A 2A agonists on outflow resistance are not likely to be mediated by actions on a single population of TM cells. These opposing effects might reflect AR responses by two or more subpopulations of TM cells, by TM and SC cells or by inner-wall SC cells, alone.

Regulatory Roles of Anoctamin-6 in Human Trabecular Meshwork Cells

“…As previously described, 41 , 42 TM cells were trypsinized, resuspended, and allowed to settle on glass coverslips. Ruptured-patch, whole-cell currents in TM cells were measured with a MultiClamp 700B amplifier (Axon Instruments, Foster City, CA, USA) coupled to an external Bessel filter (model 900; Frequency Devices, Haverhill, MA, USA).…”

Novel ocular antihypertensive compounds in clinical trials

“…It produced early dose-related IOP increases that were followed by later IOP decreases, an increase in total outflow facility (trabecular pathway), and modest changes in aqueous humor flow in conscious ocular normotensive cynomolgus monkeys 58. The mechanisms by which adenosine receptor agonists increase conventional outflow have been hypothesized to involve shrinkage of cell volume and remodeling of the extracellular matrix following secretion of matrix metalloproteinases in human trabecular meshwork cells 59–62. Adenosine A 3 antagonists may potentially influence aqueous humor formation by preventing adenosine-induced activation of chloride channels in human nonpigmented ciliary epithelial cells 63,64.…”

Nucleoside-derived antagonists to A3 adenosine receptors lower mouse intraocular pressure and act across species

“…Cell volume was monitored continually by measuring both calcein fluorescence quenching (Do et al, 2006; Karl et al, 2007) and cell area (Do et al, 2004; Mitchell et al, 2002). Coverslips containing the cells were placed in a chamber connected to a Nikon Diaphot microscope.…”