Carbenoxolone modifies spontaneous inhibitory and excitatory synaptic transmission in rat somatosensory cortex

Abstract

Gap junction (GJ) coupling between neocortical GABAergic interneurons plays a critical role in the synchronization of activity in cortical networks in physiological and pathophysiological states, e.g., seizures. Past studies have shown that GJ blockers exert anticonvulsant actions in both in vivo and in vitro models of epilepsy. However, the precise mechanisms underlying these antiepileptic effects have not been fully elucidated. This is due, in part, to a lack of information of the influence of GJ blockade on network activity in the absence of convulsant agents or enhanced neuronal excitation. One key question is whether GJ blockers act on excitatory or inhibitory systems, or both. To address this issue, we examined the effects of the GJ blocker carbenoxolone (CarbX, 150 microM) on spontaneous inhibitory postsynaptic currents (sIPSCs) and excitatory postsynaptic currents (sEPSCs) in acute slices of rat somatosensory cortex. Results showed that CarbX decreased the amplitude and frequency of sIPSCs by 30.2% and 25.7%, respectively. CarbX increased the mean frequency of sEPSCs by 24.1%, but had no effect on sEPSC amplitude. During blockade of GABAA-mediated events with picrotoxin (20 microM), CarbX induced only a small increase in sEPSC frequency that was not statistically different from control, indicating CarbX enhancement of sEPECs was secondary to the depression of synaptic inhibition. These findings suggest that in neocortex, blockade of GJs leads to an increase in spontaneous excitation by uncoupling GABAergic interneurons, and that electronic communication between inhibitory cells plays a significant role in regulating tonic synaptic excitation.

Fig. 1

Actions of CarbX on sIPSCs in layer II-III pyramidal cell. A-C: Traces of sIPSCs recorded under voltage-clamp in control saline (A), CarbX (B), and following wash out of CarbX (C). Panels A2, B2, and C2 show expanded sections of the traces displayed in A1, B1, and C1 (indicated by dots). D-F: Amplitude histograms of sIPSCs recorded in control saline (D), CarbX (E), and following CarbX wash out (F). Inset panels show the averaged traces of 30 randomly selected events recorded under each condition. G: Cumulative probability distributions of sIPSC amplitudes. H-J: Inter-event interval histograms of sIPSCs recorded in control saline (H), CarbX (I), and following CarbX wash out (J). K: cumulative probability distributions of sIPSC inter-event interval values. The figure legend in K applies to both G and K.

Fig. 2

Actions of CarbX on sEPSCs in layer II-III pyramidal cell. A-C: Recordings of sEPSCs obtained under voltage-clamp in control saline (A), CarbX (B) and following CarbX wash out (C). Panels A2, B2 and C2 show expanded segments of the traces displayed in A1, B1 and C1 (indicated by dots). D-F: Amplitude histograms of sEPSCs recorded in control saline (D), CarbX (E), and following wash out of CarbX (F). Inset panels show the averages of 30 randomly selected sEPSCs recorded under each condition. G: Cumulative probability distributions of sEPSC amplitudes. H-J: Inter-event interval histograms of sEPSCs recorded in control saline (H), CarbX (I) and after CarbX wash-out (J). K: Cumulative probability distributions of sEPSC inter-event interval values. The legend in K applies to both G and K.

Fig. 3

Enhancement of sEPSC frequency by CarbX is due to the attenuation of GABA_Aergic inhibition consequent to the blockade of electrotonic coupling. A-B: Summary data pooled from 12 cells (12 slices) of CarbX effects on sIPSC amplitude and frequency in standard saline. C-D: Summary data pooled from 8 cells (8 slices) of CarbX effects on sEPSCs in standard saline. E-F: Effects of CarbX on sEPSCs during blockade of GABA_A-mediated inhibition with 20 μM picrotoxin; data pooled from 15 cells (15 slices). G-H: Exposure of slices to 25 mM sodium propionate (Na-propionate) reduced sIPSC amplitude and frequency, mimicking the actions of CarbX. Data was pooled from 9 cells (6 slices). I-J: Na-propionate also reduced sEPSC amplitude and frequency, which is likely due to the direct suppressive effects of acidosis on excitatory synaptic transmission. Data shown was pooled from 8 cells (5 slices). The legend in E applies to panels A-E, and the legend in I applies to panels G-J. *p<0.05.