title: ClC-2 channels regulate neuronal excitability, not intracellular chloride levels.
abstract: Synaptic inhibition by GABA(A) receptors requires a transmembrane chloride gradient. Hyperpolarization or shunting results from outward current produced by chloride flowing down this gradient, into the cell. Chloride influx necessarily depletes the chloride gradient. Therefore, mechanisms that replenish the gradient (by reducing intracellular chloride concentration, [Cl(-)](i)) are crucial for maintaining the efficacy of GABA(A) receptor-mediated inhibition. ClC-2 is an inward-rectifying chloride channel that is thought to help extrude chloride because inward rectification should, in principle, allow ClC-2 to act as a one-way chloride exit valve. But chloride efflux via ClC-2 nevertheless requires an appropriate driving force. Using computer modeling, we reproduced voltage-clamp experiments showing chloride efflux via ClC-2, but testing the same model under physiological conditions revealed that ClC-2 normally leaks chloride into the cell. The discrepancy is explained by the driving force conditions that exist under artificial versus physiological conditions, and by the fact that ClC-2 rectification is neither complete nor instantaneous. Thus, contrary to previous assertions that ClC-2 helps maintain synaptic inhibition by lowering [Cl(-)](i), our simulations show that ClC-2 mediates chloride influx, thus producing outward current and directly reducing excitability. To test how ClC-2 functions in real neurons, we used dynamic clamp to insert virtual ClC-2 channels into rat CA1 pyramidal cells with and without native ClC-2 channels blocked. Experiments confirmed that ClC-2 reduces spiking independently of inhibitory synaptic transmission. Our results highlight the importance of considering driving force when inferring how a channel functions under physiological conditions.
authors: Department of Neurobiology and the Pittsburgh Center for Pain Research, University of Pittsburgh, Pittsburgh, Pennsylvania 15213, USA.
authors: Stéphanie
authors: S
authors: Ratté
authors: Steven A
authors: SA
authors: Prescott
mesh_terms: D000818:Animals; D055592:Biophysical Phenomena / Q000502:physiology*; D018118:Chloride Channels / Q000502:physiology*; D002712:Chlorides / Q000378:metabolism*; D003198:Computer Simulation; D004558:Electric Stimulation; D007424:Intracellular Fluid / Q000378:metabolism*; D008564:Membrane Potentials / Q000502:physiology; D008959:Models, Neurological; D009474:Neurons / Q000502:physiology*; D018408:Patch-Clamp Techniques; D051381:Rats; D011963:Receptors, GABA-A / Q000502:physiology; D027981:Symporters / Q000378:metabolism; D009435:Synaptic Transmission / Q000502:physiology; D005680:gamma-Aminobutyric Acid / Q000378:metabolism; D000096922:K Cl- Cotransporters
chemical_list: D018118:Chloride Channels; D002712:Chlorides; D011963:Receptors, GABA-A; D027981:Symporters; D005680:gamma-Aminobutyric Acid; D000096922:K Cl- Cotransporters; C115376:ClC-3 channel