Aqueous humor is a clear fluid that circulates around the anterior segment of the eye. It is secreted by the pigmented and non-pigmented epithelial (NPE) cells of the ciliary body; these cells produce it actively and it is also passively filtered from plasma. The basal aspect of the NPE cells possesses ion channels, pumps and exchangers that are regulated by signalling pathways involving protein kinases, calcium ions (Ca²⁺) and intracellular pH (pHi). In other secretory epithelia steroid hormones, including aldosterone, have a rapid non-genomic effect on the activity and expression of components of the signalling cascades resulting in changes in the secretory or absorptive capacity of the cell. Some ion transporters, for example the sodium-proton exchanger, are directly influenced by intracellular calcium concentrations, which in turn are dependant upon activity of protein kinases.AIM: The aim of this study was to identify rapid signalling responses to aldosterone in human NPE cells, in particular, changes in protein kinase activity and intracellular Ca²⁺ and pH_i.

Protein kinase C α (PKCα) expression was examined in cultured human NPE cells using Western Blotting techniques. A non-radioactive fluorescent assay (Promega) was used to identify Protein Kinase A (PKA) expression. Intracellular calcium and pH changes were monitored by spectrofluorescence imaging of cells loaded with the fluorescent dyes Fura2 and BCECF, respectively.

PKA expression was significantly increased after one min of incubation with physiological concentrations of aldosterone (1 nM). This increase was maximal between one and five min with expression levels of PKA returning to basal levels at ten min. Aldosterone (10 nM) also caused a rapid, transient change in PKCα expression; an increase in PKCα occurred after five min of incubation. This increase had diminished after ten min and was not evident after twenty min. Preliminary results indicate that there is an increase in intracellular Ca²⁺ and pH_i induced within min of treatment with aldosterone (1 nM).

These data demonstrate for the first time rapid (non-genomic) responses to aldosterone in human ocular ciliary body epithelium. The signalling pathways affected, PKC, intracellular Ca²⁺ and pH_i are known regulators of epithelial electrolyte secretion and may have important implications in rapid regulation of aqueous humor production by a local Renin Angiotensin Aldosterone System in the human eye (Cullinane et al. 2002).