Fig. 3.

Hippocampal astrocytic gap junction coupling is impaired in Tsc1^GFAPCKO mice. (A, B) Representative hippocampal sections showing the extent of biocytin-stained astrocytes from control mice and Tsc1^GFAPCKO mice following injection of a single astrocyte in stratum radiatum. Scale bars, 50 μm. (C, D) Representative hippocampal sections showing density of GFAP-positive astrocytes in control and Tsc1^GFAPCKO mice. Scale bars, 20 μm. (E) Typical “passive” current profile of recorded astrocytes. A range of depolarizing and hyperpolarizing voltage steps between −160 to +70 mV (holding potential −70mV) activated large time- and voltage- independent membrane currents. Scale bars, 500 pA, 10 ms. (F)Summarized data from all experiments showed reduced numbers of biocytin stained astrocytes in hippocampal slices from Tsc1^GFAPCKO mice mice. ***p < 0.001, by t-test (n = 20 injections, Tsc1^GFAPCKO mice; n=16 injections, control mice). (G) After injection of a single astrocyte in the stratum radiatum along the superficial surface of the hippocampal slice, 40 μm thick sections obtained from the original, injected 400 μm slice were processed and counted individually for biocytin-positive cells. The number of biocytin-positive cells decreased in sections more distant from the injected astrocyte (in Section 1 or 2) in both Tsc1^GFAPCKO and control mice and was lower in Tsc1^GFAPCKO mice in all sections. Dark triangle, control mice; open triangle, Tsc1^GFAPCKO mice.(H) When comparing the distribution of biocytin-positive cells in different strata within the hippocampus after injection of a single astrocyte in the stratum radiatum, control mice exhibited an expected decrease in coupled cells in the more distal stratum lacunosum-moleculare compared to the stratum radiatum. In contrast, Tsc1^GFAPCKO mice had equal numbers of coupled cells in these two strata. Thus, compared to control mice, the number of coupled cells in Tsc1^GFAPCKO mice was only decreased in the stratum radiatum, not the stratum lacunosum-moleculare. Con – control mice, KO – Tsc1^GFAPCKO mice, s.r – stratum radiatum, s.l.m – stratum lacunosum-moleculare. ***p < 0.001 by ANOVA.