Figure 4. Spinal neuron excitability increased during the continuous blockade of GABARs in vitro. Motoneuron excitability was measured in isolated spinal cords using a ramp protocol from 0 to 200 pA in 1.2 s to assess rheobase current and voltage threshold in the absence (control, 33 cells; 12 cords) or in the continuous presence of gabazine 10 μM at three different blockade periods: 0–2 h (13 cells, four cords cells), 2–4 h (nine cells, three cords), and 4–6 h (10 cells, four cords). Representative traces of motoneuron firing in a control motoneuron or after 4 h in the continuous presence of gabazine in the bath. Lower trace shows the ramp protocol applied to evoke motoneuron firing (A). Box and dot plots showing quartile distribution and individual values for of the rheobase (B1), threshold (B2), RMP (B3), and input resistance (B4) in control; 0–2, 2–4, and 4–6 h in gabazine. Inset of B3 shows results for cords that were never treated with gabazine, but cells were recorded in the first, second, or third 2-h periods. C, D, Interneuron excitability was measured in isolated spinal cords using the same protocol as above to assess rheobase and threshold in the absence (control, 22 cells; eight cords) or in the continuous presence of gabazine (10 μM) at three different blockade periods: 0–2 h (14 cells, five cords), 2–4 h (nine cells, three cords), and 4–6 h (six cells, three cords). Representative traces of firing in a control interneuron or after 4 h in the continuous presence of gabazine in the bath. Lower trace shows the ramp protocol applied to evoke interneuron firing (C). Box and dot plots showing the value of the rheobase (D1), threshold (D2), RMP (D3), or input resistance (D4) in control; 0- to 2-, 2- to 4-, and 4- to 6-h periods in the presence of gabazine. Inset of D3 shows results for cords that were never treated with gabazine but interneurons were recorded in the first, second or third 2-h period. Kruskal–Wallis method followed by a post hoc Dunn test was used to asses statistical significance; *p < 0.05, **p < 0.01, ***p < 0.001. For all plots, the continuous line represents the median and the dotted line represent the mean of the sample.