Fast synaptic inhibition in the nervous system depends on the transmembrane flux of Cl.sup.- ions based on the neuronal Cl.sup.- driving force. Established theories regarding the determinants of Cl.sup.- driving force have recently been questioned. Here, we present biophysical models of Cl.sup.- homeostasis using the pump-leak model. Using numerical and novel analytic solutions, we demonstrate that the Na.sup.+/K.sup.+-ATPase, ion conductances, impermeant anions, electrodiffusion, water fluxes and cation-chloride cotransporters (CCCs) play roles in setting the Cl.sup.- driving force. Our models, together with experimental validation, show that while impermeant anions can contribute to setting [Cl.sup.-].sub.i in neurons, they have a negligible effect on the driving force for Cl.sup.- locally and cell-wide. In contrast, we demonstrate that CCCs are well-suited for modulating Cl.sup.- driving force and hence inhibitory signaling in neurons. Our findings reconcile recent experimental findings and provide a framework for understanding the interplay of different chloride regulatory processes in neurons. eLife digest Cells called neurons in the brain communicate by triggering or inhibiting electrical activity in other neurons. To inhibit electrical activity, a signal from one neuron usually triggers specific receptors on the second neuron to open, which allows particles called chloride ions to flow into or out of the neuron. The force that moves chloride ions (the so-called 'chloride driving force') depends on two main factors. Firstly, chloride ions, like other particles, tend to move from an area where they are plentiful to areas where they are less abundant. Secondly, chloride ions are negatively charged and are therefore attracted to areas where the net charge (determined by the mix of positively and negatively charged particles) is more positive than their current position. It was previously believed that a group of proteins known as CCCs, which transport chloride ions and positive ions together across the membranes surrounding cells, sets the chloride driving force. However, it has recently been suggested that negatively charged ions that are unable to cross the membrane (or 'impermeant anions' for short) may set the driving force instead by contributing to the net charge across the membrane. Düsterwald et al. used a computational model of the neuron to explore these two possibilities. In the simulations, altering the activity of the CCCs led to big changes in the chloride driving force. Changing the levels of impermeant anions altered the volume of cells, but did not drive changes in the chloride driving force. This was because the flow of chloride ions across the membrane led to a compensatory change in the net charge across the membrane. Düsterwald et al. then used an experimental technique called patch-clamping in mice and rats to confirm the model's predictions. Defects in controlling the chloride driving force in brain cells have been linked with epilepsy, stroke and other neurological diseases. Therefore, a better knowledge of these mechanisms may in future help to identify the best targets for drugs to treat such conditions.