Mechanisms of Ion Transport Across the Mouse Retinal Pigment Epithelium Measured In Vitro

PURPOSE: To examine ion transport across the mouse retinal pigment epithelium (RPE), measured by the short-circuit current (I(SC)) and transepithelial resistance (TER). METHODS: Sheets of RPE from mice (C57BL6/J) with retina, choroid, and sclera attached were mounted in Ussing chambers (0.031-cm(2) aperture) and Krebs solution. The I(SC) and TER were recorded with voltage clamps. Receptors implicated in ion transport were blocked or stimulated by ligands applied to both sides. RESULTS: The mean initial I(SC) was −12.0 ± 3.9 µA/cm(2) (basolateral negative), and mean TER was 67.1 ± 8.0 ohm·cm(2). RPE preparations remained stable for 3 hours, with I(SC) decreasing by 0.078 ± 0,033 µA/cm(2)/hr. Adenosine triphosphate (100 µM) increased I(SC) by 2.22 ± 0.41 µA/cm(2) (P = 0.003). Epinephrine (100 µM) increased I(SC) by 1.14 ± 0.19 µA/cm(2) (P = 0.011). Bumetanide (100 µM) reduced I(SC) by 1.72 ± 0.73 µA/cm(2) (P = 0.027). Ouabain (1 mM) induced a biphasic response: an I(SC) increase from −7.9 ± 2.4 to −15.49 ± 2.12 µA/cm(2) and then a decrease to −3.7 ± 2.2 µA/cm(2). Ouabain increased TER by 15.3 ± 4.8 ohm·cm(2). These compounds were added sequentially. Apical [K(+)](o) at zero mM transiently increased I(SC) by 3.36 ± 1.06 µA/cm(2). Ba(++) decreased I(SC) from −10.4 ± 3.1 to −6.6 ± 1.8 µA/cm(2) (P = 0.01). Ba(++) reversed the K(+)-free response, with I(sc) decreasing further from −5.65 ± 1.24 to −3.37 ± 0.79 µA/cm(2) (P = 0.029). CONCLUSIONS: The I(SC) and TER can be recorded from the mouse RPE for 3 hours. Adrenergic and purinergic receptors affect murine RPE ion transport. Sodium–potassium adenosine triphosphatase plays a role in net ion transport across mouse RPE, and Na-K-2Cl cotransporter activity partly accounts for transepithelial ion transport. Mimicking light-induced changes, low subretinal [K(+)](o) increases ion transport transiently, dependent on K(+) channels.