Figure 1.
Prenatal LPS decreased GABAergic interneuron density in the hippocampus.
Representative coronal sections of the whole hippocampus and of the CA3 area of 18-day-old SAL (A,B) and LPS animals (C,D) immunolabeled with anti-GAD67 antibody. Scale bar (A,C) = 2 mm, Scale bar (B,D) = 1 mm. (E) Late gestational LPS injection significantly decreased GAD67+ cell density in the hippocampus and notably in the CA3 area. Quantification of GAD67+ neurons has been performed as described in “Materials and Methods” from 4 animals per group. Data are presented as means ± SEM. *p<0.05, for the comparison of SAL (open bars) vs LPS (black bars) using two-way ANOVA followed by Bonferroni's post-test. HC, hippocampus; DG, dentate gyrus.
Figure 2.
Prenatal LPS induced deficits in evoked inhibitory synaptic transmission in CA1 neurons.
(A) I/O relationships of eIPSCs were established for both SAL and LPS rats. The amplitudes of eIPSCs were normalized to the capacitance of the recorded cell and plotted against corresponding stimulus intensities. The duration of the stimuli was fixed at 100 µs. Hyperbolic fitting was performed on 127 experimental points obtained from SAL (open circles; N = 9) animals and 120 experimental points obtained from LPS (filled squares; N = 9) animals. Stippled zones represent the 95% Confidence Intervals of the curves. Each point represents the mean amplitude for the corresponding intensity of stimulation. For readability, SEMs were not represented. * p<0.05, for the comparison of LPS vs SAL animals, using two-way ANOVA followed by Holm-Sidak post-hoc analysis. (B) Representative traces of the eIPSCs recorded in SAL and LPS animals at two different intensities.
Figure 3.
Prenatal LPS-induced deficits in spontaneous and miniature GABAergic activities in CA1 pyramidal cells.
(A) Spontaneous GABAergic activity recorded in CA1 pyramidal cells. The recapitulative graphs plot mean sIPSC frequency (left) and amplitude (right). The boxes define the median, 25th and 75th percentiles. The whiskers represent 10th and 90th percentiles. ** p<0.01, Mann and Whitney rank sum test, N = 8 animals per group. (B) Miniature GABAergic activity recorded in CA1 pyramidal cells in the presence of TTX (100 nM). The recapitulative graphs plot mean mIPSC frequency (left) and amplitude (right). The boxes define the median, 25th and 75th percentiles. The whiskers represent the 10th and 90th percentiles. ** p<0.01, Mann and Whitney rank sum test, N = 5 animals per group. Representative traces of GABAergic spontaneous activity (top) and GABAergic miniature activity (bottom) are shown in individual neurons from SAL (gray) and LPS (black) rats.
Figure 4.
Tiagabine restored LTD via the activation of GABAB receptors in LPS animals.
Tiagabine (20 µM) and/or CGP55845 (1 µM) were applied in the perfusate during both the recording of baseline activity and LFS (1 Hz stimulation, 15 min) delivery. (A) Time-course and recapitulative graph depicting LTD induction in control (SAL) animals. LFS induced an LTD of fEPSP amplitude in control animals (SAL; filled circles; N = 8), which was significantly blocked by the GABAB receptor antagonist CGP55845 (SAL+CGP; open circles; N = 5; * p<0.05 vs SAL group). Tiagabine had no significant effect on LTD level (SAL+T; filled triangles; N = 8). (B) Time-course and recapitulative graph depicting LTD induction in LPS animals. LFS did not trigger LTD in LPS animals (LPS; filled circles, N = 8), while tiagabine restored the LTD occurrence (LPS+T; filled triangles; N = 7; ** p<0.01 vs LPS group). The effect of tiagabine was not observed when co-applied with CGP55845 (LPS+T+CGP; open circles; N = 5; ** p<0.01 vs LPS+T group). Time-courses are illustrated by representative traces of fEPSP recorded in each group and extracted both before (1) and 45 min following (2) the stimulation train as indicated by the time-points on the graphs. On the right, histograms recapitulate mean fEPSP amplitudes (± SEM) recorded under different conditions. Data are fEPSP amplitudes averaged between time 20 min and time 50 min for each animal (see A or B graph legends for respective N). The statistical significance of the differences between groups was assessed by performing pairwise multiple comparison Holm Sidak analysis.
Figure 5.
Tiagabine normalized aberrant synaptic plasticity phenomena in young LPS animals.
Tiagabine (20 µM) and/or CGP55845 (1 µM) were applied in the perfusate during both the recording of baseline activity and ppulse-LFS (paired-pulse 1 Hz stimulation, 50 ms inter-pulse interval, 15 min) delivery. (A) Time-course and recapitulative graph depicting LTD induction in control (SAL) animals. Ppulse-LFS induced an LTD of fEPSP amplitude in control animals (SAL; filled circles; N = 7), which was partially blocked by the GABAB receptor antagonist CGP55845 (SAL+CGP; open circles; N = 9). Tiagabine had no significant effect on LTD level either in the absence (SAL+T; filled triangles; N = 8) or in the presence of CGP55845 (N = 6). (B) Time-course and recapitulative graph depicting LTD induction in LPS animals. Ppulse LFS application led to the occurrence of a slow-onset LTP in LPS offspring (LPS; filled circles; N = 13), which was not observed in the presence of either tiagabine (LPS+T; filled triangles; N = 11; ** p<0.01 vs LPS group) or CGP55845 (LPS+C; open circles; N = 6; * p<0.05 vs LPS group). LTD could be obtained after ppulse-LFS in the concurrent presence of tiagabine and CGP (LPS+T+CGP; filled diamonds; N = 6; ** p<0.01 vs LPS group). Under these latter conditions, the average of fEPSP amplitudes was significantly different the one obtained in the presence of either tiagabine (# p<0.05 vs LPS+T group) or CGP (# p<0.05 vs LPS+C group) after ppulse-LFS application. Time-courses are illustrated by representative traces of fEPSP recorded in each group and extracted both before (1) and 45 min following (2) the stimulation train as indicated by the time-points on the graphs. On the right, histograms recapitulate mean fEPSP amplitudes (± SEM) recorded under different conditions. Data are fEPSP amplitudes averaged between time 20 min and time 50 min for each animal (see A or B graph legends for respective N). The statistical significance of the differences between groups was assessed by performing pairwise multiple comparison Holm Sidak analysis.