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Cl^– Influx into Rat Cortical Lens Fiber Cells Is Mediated by a Cl^– Conductance That Is Not ClC-2 or -3

¹From the Department of Physiology, School of Medical Sciences and the ²School of Biological Sciences, University of Auckland, Auckland, New Zealand.

PURPOSE. Exposure of organ-cultured lenses to Cl^– channel blockers under isotonic conditions induces a localized cortical zone of extracellular space dilations. The purpose of this study was to investigate whether elongated lens fiber cells from this zone contain an anion conductance that mediates Cl^– influx and whether two chloride channel isoforms known to be expressed in the lens (ClC-2 and -3) are responsible.

METHODS. Fiber cells were isolated by enzymatic dissociation in the presence of Gd³⁺ and Co²⁺ and their electrical properties analyzed by whole-cell patch clamping. Cells from the zone of extracellular space dilations were selected for analysis on the basis of cell length. RT-PCR and immunocytochemistry were used to determine whether ClC-2 or -3 channel isoforms are expressed in fiber cells located in the zone of extracellular space dilations.

RESULTS. Cells from the zone of extracellular space dilations were typically >120 µm in length and exhibited an outwardly rectifying Cl^– conductance that was blocked by DIDS (4,4'-diisothiocyanostilbene-2,2'-disulfonic acid) and displayed an anion selectivity sequence of I^– > Cl^– >> gluconate. ClC-2 and -3 were found to be expressed at the transcript and protein level in lens fiber cells, but subsequent immunocytochemical studies indicated that expressed proteins did not colocalize with cell membranes in the zone of extracellular space dilations, being predominately cytoplasmic in nature.

CONCLUSIONS. Taken together, the data indicate that extracellular space dilations are due to the inhibition of a Cl^– channel(s) that normally mediates Cl^– influx into cortical lens fiber cells under isotonic conditions. The molecular identity of this channel remains to be determined.

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