Expression of the voltage-gated chloride channel ClC-2 in rod bipolar cells of the rat retina.
Voltage-gated chloride channels (ClC) are highly conserved during evolution and appear to participate in a variety of physiological functions. Recently, ClC-2 was proposed to play a role in stabilizing the chloride equilibrium potential near or below the resting membrane potential in neurons expressing ligand-gated chloride channels. Because rod bipolar cells in mammalian retina express three forms of inhibitory ligand-gated chloride channels, we decided to study ClC-2 localization and function in the rat retina. RNA encoding ClC-1, -2, -3, -4, and -5 was detected by reverse transcription-PCR in the rat retina. ClC-2-specific antibodies identified protein on cell bodies and in synaptic layers. Double-immunofluorescence staining revealed that intense ClC-2 immunoreactivity colocalized with PKC-stained rod bipolar cells. Patch-clamp experiments performed with individual rod bipolar cells demonstrated the presence of a time-dependent, inwardly rectified current activated at hyperpolarizing membrane potentials. This current demonstrated selectivity for different anions (Cl(-) > I(-) > gluconate), was inhibited by Cd(2+), and was minimally reduced by 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid. These features are consistent with currents generated by ClC-2 channels. Our data indicate that functional ClC-2 channels are present in retinal rod bipolar cells and support a role for ClC-2 in maintaining Cl(-) homeostasis in neurons with ligand-gated chloride channels.
Enz
R
R
Institut für Biochemie, Friedrich-Alexander-Universität Erlargen-Nürnberg, 91054 Erlangen, Germany.
Ross
B J
BJ
Cutting
G R
GR
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D000838:Anions / Q000494:pharmacology; D002104:Cadmium / Q000494:pharmacology; D018118:Chloride Channels / Q000032:analysis / Q000235:genetics* / Q000502:physiology; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects; D011493:Protein Kinase C / Q000032:analysis; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D017948:Retinal Rod Photoreceptor Cells / Q000166:cytology / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
D000838:Anions; D018118:Chloride Channels; D012333:RNA, Messenger; D002104:Cadmium; D011493:Protein Kinase C; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid