A role for the volume regulated anion channel in volume regulation in the murine CNS cell line, CAD.
OBJECTIVE
The role of the volume regulated anion channel (VRAC) in a model CNS neuronal cell line, CAD, was investigated.


METHODS
Changes in cell volume following hypotonic challenges were measured using a video-imaging technique. The effect of the Cl(-) channel antagonists tamoxifen (10 microm) and 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS; 100 microm) on regulatory volume decrease (RVD) were measured. The whole-cell voltage-clamp technique was used to characterize ICl(swell), the current underlying the VRAC.


RESULTS
Using the video-imaging technique, CAD cells were found to swell and subsequently exhibit RVD when subjected to a sustained hypotonic challenge from 300 mOsmol kg(-1) H(2)O to 210 mOsmol kg(-1) H(2)O. In the presence of tamoxifen (10 microm) or DIDS (100 microm) RVD was abolished, suggesting a role for the VRAC. A hypotonic solution (230 mOsmol kg(-1) H(2)O) evoked ICl(swell), an outwardly rectifying current displaying time-independent activation, which reversed upon return to isotonic conditions. The reversal potential (E(rev)) for ICl(swell) was -14.7 + or - 1.4 mV, similar to the theoretical E(rev) for a selective Cl(-) conductance. ICl(swell) was inhibited in the presence of DIDS (100 microm) and tamoxifen (10 microm), the DIDS inhibition being voltage dependent.


CONCLUSIONS
Osmotic swelling elicits an outwardly rectifying Cl(-) conductance in CAD cells. The ICl(swell) observed in these cells is similar to that observed in other cells, and is likely to provide a pathway for the loss of Cl(-) which leads to water loss and RVD. As ischaemia, brain trauma, hypoxia and other brain pathologies can cause cell swelling, CAD cells represent a model cell line for the study of neuronal cell volume regulation.
Harvey
V L
VL
Neuroscience, Pharmacology & Physiology, University College London, Gower Street, London WC1E 6BT, UK. viclharvey@googlemail.com
Saul
M W
MW
Garner
C
C
McDonald
R L
RL
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D000838:Anions / Q000378:metabolism / Q000494:pharmacology*; D002460:Cell Line; D048429:Cell Size / Q000187:drug effects*; D002490:Central Nervous System / Q000166:cytology*; D018118:Chloride Channels / Q000378:metabolism / Q000502:physiology*; D004594:Electrophysiology; D006801:Humans; D008564:Membrane Potentials / Q000502:physiology*; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D009997:Osmotic Pressure / Q000187:drug effects / Q000502:physiology*; D013629:Tamoxifen / Q000494:pharmacology
D000838:Anions; D018118:Chloride Channels; D013629:Tamoxifen; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid