Relationship between tonic inhibitory currents and phasic inhibitory activity in the spinal cord lamina II region of adult mice

Abstract

Phasic and tonic inhibitions are two types of inhibitory activities involved in inhibitory processing in the CNS. In the spinal cord dorsal horn, phasic inhibition is mediated by both GABAergic and glycinergic inhibitory postsynaptic currents. In contrast to phasic inhibitory currents, using patch-clamp recording technique on spinal cord slices prepared from adult mice we revealed that tonic inhibitory currents were mediated by GABAA receptors but not by glycine receptors in dorsal horn lamina II region. We found that there was a linear relationship (r = 0.85) between the amplitude of tonic inhibitory currents and the frequency of GABAergic inhibitory postsynaptic currents. Analysis of charge transfer showed that the charges carried by tonic inhibitory currents were about 6 times of charges carried by phasic inhibitory currents. The prominent charge transfer by tonic inhibitory currents and their synaptic activity dependency suggest a significant role of tonic inhibition in sensory processing.

Figure 1

Revealing tonic inhibitory currents in lamina II neurons of adult mice. A). Traces on the left side show sIPSCs recorded in normal bath solution (control), following the applications of either 2 μM strychnine (Stry) or 20 μM bicuculline (Bic). Traces on the right side are the average of 100 sIPSCs. τ is time constant for averaged trace. B). Sample trace (top panel) shows the shift of baseline holding current following the application of 20 μM bicuculline. The lower traces show at an expanded scale the baselines before and during bicuculline application as well after wash off bicuculline. C). Sample trace shows that application of strychnine (2 μM) inhibited some sISPCs but did not affect baseline holding current. D). Bar graph shows pooled results of baseline shift following the application of either 20 μM bicuculline (n = 30) or 2 μM strychnine (n = 6). Bicuculline or strychnine was applied for 3 min in each experiment.

Figure 2

Lack of correlation between tonic inhibitory currents and resting membrane potentials as well as between tonic inhibitory currents and total sIPSC activity. A). Graph shows a plot of tonic inhibitory currents against membrane potentials for each recorded neuron (n = 30). B). Graph shows a plot of tonic inhibitory current against sIPSC frequency (n = 30). Tonic inhibitory currents were revealed by the applications of 20 μM bicuculline. Both resting membrane potentials and sIPSCs were measured before the application of bicuculline.

Figure 3

Correlation between tonic inhibitory currents and GABAergic inhibitory synaptic activity. A). Sample trace shows bicuculline-induced baseline shift in a lamina II neuron with high GABAergic inhibitory synaptic activity. B). Sample trace shows bicuculline-induced baseline shift in another lamina II neuron with low GABAergic inhibitory synaptic activity. C). Plot of GABAergic tonic inhibitory currents against frequency of GABAergic sIPSCs (n = 14). Linear regress coefficient (r) = 0.8515. All experiments were performed in the presence of 2 μM strychnine.

Figure 4

Charge transfers mediated by phasic inhibitory currents and by tonic inhibitory currents. A). Diagram illustrates the measurement of phasic inhibitory current charge transfer (Q_PC) and tonic inhibitory current charge transfer (Q_TC). B). Bar Graph shows the comparison between Q_PC and Q_TC (n = 32). Tonic currents were revealed by the application of 20 μM bucuculline. Strychnine was not included in bath solution.