Commentary: Chloride Regulation: A Dynamic Equilibrium Crucial for Synaptic Inhibition

摘要

The recent review by Doyon et al. ( 2016 ) is for the main part an excellent description of many important aspects of neuronal chloride regulation and will be of good use to many scientists interested in synaptic function. Nevertheless, some information given on the role of the K~(+)Cl~(?)cotransporter 2 (KCC2) is likely to be misleading. While proposing an explanation for the controversial findings by Glykys et al. ( 2014 ) that blockers of cation-chloride cotransporters did not affect the basal intracellular Cl~(?)concentration ([Cl~(?)]_(i)), contrary to expectations from previous transporter manipulations (see e.g., references in Ben-Ari, 2014 ; Kaila et al., 2014 ), Doyon et al. ( 2016 ) suggest that this may be due to the small degree of inhibitory synaptic activity in the preparation used, with extremely low Cl~(?)load. In their explanation, they give an equation for the equilibrium relation between Cl~(?)flux through an inhibitory (Cl~(?)) conductance (g_(inh)) and the Cl~(?)transported by KCC2. On basis of this equation, the conclusion is that the Cl~(?)equilibrium potential “E_(Cl)is sensitive to changes in KCC2 activity (g_(KCC2)) only when Cl~(?)load (g_(inh)) is large.” This conclusion, however, cannot be justified on basis of the relation between Cl~(?)flux through channels and Cl~(?)transported by KCC2. (The equation given by Doyon et al. is not correctly formulated, although the reason for their claim may not depend on this mistake). For an explanation of our point of view, consider a hypothetical cell with Cl~(?)transport across the outer membrane only via Cl~(?)selective channels and KCC2. At equilibrium, the amount (mol/s) of Cl~(?)transported by the channels must be equal, but opposite, to that transported by KCC2. We thus formulate the relation: (1) I Cl / F? = ?g KCC 2 U KCC 2 where I_(Cl)is Cl~(?)current, F is the Faraday constant and U_(KCC2)is the driving force for transport by KCC2. g_(KCC2)is a proportionality factor that may be thought of as an “apparent conductance,” and should reflect the number of transporters in the membrane as well as the transport rate of the individual transporter molecules at fixed K~(+)and Cl~(?)concentrations, similarly as I_(Cl)depends on the Cl~(?)conductance (g_(Cl)) which reflects the number of Cl~(?)channels as well as the conductance of individual channels. (g_(KCC2)is, however, not a conductance in the usual electrical sense). Equation (1) may be reformulated, in several steps, for clarity: (2) g Cl ? ( V m ? E Cl ) / F = g KCC 2 ? ( RT?ln? ( [ Cl ? ] i / [ Cl ? ] o ) + RT?ln? ( [ K + ] i / [ K + ] o ) ) (3) g Cl ? ( V m ? E Cl ) / F?=?g KCC 2 ?F? ( E Cl ? E K ) (4) E Cl = ( g KCC 2 ? F 2 ? E K + g Cl V m ) / ???????? ( g KCC 2 ? F 2 + g Cl ) where V_(m)is membrane potential, R the gas constant, T temperature (in K), E_(K)the K~(+)equilibrium potential and Cl~(?)and K~(+)concentrations are given within brackets with subscripts i and o for inside and outside, respectively. We may use Equation (4) to illustrate the relation between E_(Cl)and g_(KCC2)at various levels of g_(Cl). (Assume that K~(+)concentrations and membrane potential are fixed, as controlled by other factors, such as the cellular Na~(+)-K~(+)-ATPase, not discussed here). As can be seen in Figure 1A , contrary to the claim by Doyon et al. ( 2016 ), E_(Cl)is only weakly dependent on KCC2 transport capacity at high Cl~(?)conductance, while it depends strongly on KCC2 when Cl~(?)conductance is low. At the extremes, when g_(KCC2)= 0, then E_(Cl)= V_(m)and when g_(Cl)= 0, then E_(Cl)= E_(K). Figure 1 Cl~(?)equilibrium potential — dependence on KCC2 transporter capacity at various Cl~(?)conductance and various membrane potentials. (A) E_(Cl)vs. g_(KCC2)with E_(K)fixed at ?100 mV and V_(m)at ?60 mV. g_(Cl)as indicated. Note that E_(Cl)dependence on g_(KCC2)is reduced with increased g_(Cl). (B) E_(Cl)vs. g_(KCC2)with E_(K)fixed at ?100 mV and g_(Cl)at 1 nS. V_(m)as indicated. Note that E_(Cl)dependence on g_(KCC2)increases when V_(m)changes in positive direction. Note the x-axis break between 0.5 and 1.9 10~(?18)mol~(2)/(V C s), to clearer illustrate the steeply decaying region of the curves. Justification of illustrated parameter ranges: The g_(Cl)range (in A ) was chosen to include cells with a low g_(Cl)as evident from the high membrane resistance (Johansson et al., 1995 ) as well as cells with a high g_(Cl)(very low input resistance dominated by inhibitory conductances; Destexhe et al., 2003 ). The g_(KCC2)range (in A,B ) shown likely covers the capacity for most central neurons: When g_(KCC2)= 1 10~(?18)mol~(2)/(V C s), KCC2-mediated transport modeled as described by Karlsson et al. ( 2011 ) may reduce [Cl~(?)]_(i)from 20 mM to ~5 mM with approximated time constants of 0.85, 6.8, and 55 s for spherical cells of radius 5, 10, and 20 μm, respectively, assuming 50% cytosolic volume and no other Cl~(?)transport/leak. Experimentally observed [Cl~(?)]_(i)recovery is slower or comparable (Ber