Pathways signalling the regulatory volume decrease of cultured non-pigmented ciliary epithelial cells

Purpose. The authors identify the signaling pathways tor the regulatory volume decrease (RVD) of nonpigmented ciliary epithelial (NPE) cells. The RVD is a regulatory response triggered by swelling and reflecting K. C1 release by NPE cells. Methods. The cell volumes of human nonpigmented ciliary epithelial cells were measured in suspension by electronic cell sorting. Measurements were conducted in test solutions of constant ionic strength, but osmolality was varied by sucrose. Results. Cyclic AMP (cAMP), forskolin, PGE 2 , the PKC-inhibitor staurosporine, and increasing cytoplasmic Ca' 2+ activity with thapsigargin all enhanced the RVD. Leukotrienes A 4 , D 4 , E 4 , and the protein phosphatasc inhibitor okadaic acid had no detectable effect under the current experimental conditions. The cyclooxygenase inhibitor indomelhacin, the epoxygenase inhibitors ketoconazole and SKF 525A, and the PKC activator DiC K all downregulatcd the RVD. The addition of the cation ionophore, gramicidin, increased the RVD. In the presence of gramicidin, cAMP, PGE 2 , and indomethacin did not affect the RVD, but ketoconazole, DiC K , and the calcium-calmodulin blocker trilluoroperazine still inhibited-and staurosporine still enhanced-the RVD. Many of these observations are strikingly different from results reported with other cells. Anisosinotic swelling did not increase intraccllular cAMP concentration. Conclusions. The pathways signaling the regulatory responses to swelling are unique for each cell type. The authors propose that hypotonic swelling of NPE cells stimulates arachidonic acid turnover, triggering PGE^-mediated upregulation of K + channels and epoxide-mediatcd upregulation of Cl~ channels. Swelling may also reduce endogenous PKC activity, further upregulating Cl~ channels. Calcium-calmodulin plays a permissive role in upregulating the CI" channels. Invest Ophthalmol Vis Sci. 1994;35:2876-2886 v>iell volume regulation is of major importance in the normal functioning of both absorptive and secretory epithelia. For example, the intracellular fluid of the ciliary epithelium is replaced every 3 minutes in humans and rabbits. 1 Without volume regulation, small discrepancies between the rates of fluid entry from the stroma and fluid exit into the posterior chamber could result in disastrous swelling or shrinkage of the transporting cells. Furthermore, some of the mechanisms subserving cell volume regulation are probably impor

Nuclear Magnetic Resonance of Sodium-23 Linoleate-Water: Basis for an Alternative Interpretation of Sodium-23 Spectra within Cells

The (23)Na spectrum from liquid crystals of sodium linoleate in water has been studied by nuclear magnetic resonance (NMR) techniques. The integrated intensity of the visible central spectral line was 34-39% of the intensity of a reference sample containing an equal quantity and concentration of (23)Na nuclei. Since satellite signals were clearly demonstrable, the effect reflected a nuclear quadrupolar interaction rather than a splitting of the (23)Na into two populations of bound and free nuclei. It is proposed that a similar quadrupolar effect may be the basis for the apparent binding of the (23)Na observed in biological systems

(17)O Nuclear Magnetic Resonance Spectrum of H(2)(17)O in Frog Striated Muscle

Whole striated muscles from the frog Rana esculenta were bathed in Ringer's solution enriched with H(2)(17)O; the muscle water was subsequently collected by vacuum distillation. The integrated intensity of the nuclear magnetic resonance (NMR) signal of (17)O in the muscle was measured to be approximately ¾ of the signal observed in the distilled water. The phenomenon may arise either from immobilization of a population of the water molecules which may be a very small fraction or as much as ¼ of the total, or may reflect tumbling of ⅓ of the water molecules in a compartment containing an anisotropic medium. Such an effect was demonstrated for H(2)(17)O using the model system of sodium linoleate in water