Figure 5: Paired recordings disclose changes in inhibitory synaptic strength, efficacy and connectivity following rMS.

(a) Overview of a slice culture prepared from GAD65-GFP mice. Note GFP-expressing interneurons in the stratum radiatum (rad) and lacunosum moleculare (la-mol) of CA1. Occasionally, GFP-expressing interneurons are found in the stratum pyramidale (pyr) and in stratum oriens (oriens). Blue, TO-PRO nuclear stain. Scale bar, 100 μm. (b) Example of simultaneously patched neurons filled with Alexa568 (10 μM). GFP-expressing interneurons are patched in rad or at the border between rad and la-mol. Asterisks indicate positions of the tips of Alexa568-filled patch pipettes. Scale bar, 50 μm. (c) Averaged responses of successfully evoked time-locked postsynaptic currents from CA1 neurons in control (black trace) and stimulated slice cultures (grey trace). Up to 50 action potentials (45±3) were induced at 0.1 Hz in presynaptic GFP-expressing interneurons while recording from postsynaptic CA1 neurons. (d–f) A decrease in the percentage of connected pairs and evoked GABA_AR-mediated IPSC amplitudes was observed, while the percentage of transmission failure rate was markedly increased after rMS (2–4 h pms; control; n=6 connected pairs in four cultures, one value (1,196 pA with 0% failure rate) excluded from analysis; 7 of 12 probed pairs connected; rMS, n=7 connected pairs in four cultures; 7 of 27 probed pairs connected; Mann–Whitney test). (g) Sample traces illustrating paired-pulse protocols at different interpulse intervals (left). No significant difference between non-stimulated and stimulated cultures in paired-pulse depression is observed (control, n=5 cells in four cultures; rMS (2–4 h), n=5 cells in four cultures, linear regression fit, Kruskal–Wallis test followed by Dunn’s post hoc test; six data points outside the axis limits). (h) Sample traces illustrating short-term plasticity (five action potentials at 20 Hz). The amplitude of each consecutive inhibitory postsynaptic current was normalized to the amplitude of the first response. No significant difference between non-stimulated and stimulated cultures (control, n=5 cells in four cultures; rMS (2–4 h), n=5 cells in four cultures, linear regression fit, Kruskal–Wallis test followed by Dunn’s post hoc test; one data points outside the axis limits). Individual data points are indicated by grey dots. Values represent mean±s.e.m. (*P<0.05; *P<0.01; ns, not significant differences).