Fig. 6
Pten+/− neurons display depressed ADBE. A-C) Hippocampal synaptosome lysates from either wild-type (WT) or Pten+/− rats were probed for the presence of PTEN (A) and total protein (B). (C) Quantification of PTEN levels in both are displayed, normalised to total protein ± SEM, n = 5 independent synaptosome preparations for both WT and Pten+/−, * p = 0.0178, unpaired t test. D-J) Hippocampal neurons derived from either WT or Pten+/− rat embryos were transfected with synaptophysin-pHluorin (sypHy) after 7 days in vitro (DIV) and imaged at DIV 13–15. D, H) Mean sypHy fluorescence traces of WT and Pten+/− hippocampal neurons normalised to either the total SV pool as revealed by NH4Cl (D) or peak fluorescence during electrical stimulation (H) ± SEM. E) Mean sypHy surface fraction presented as a percentage of the total SV pool ± SEM. F, G) Mean peak sypHy response in response to either 10 Hz (F) or 40 Hz (G) action potential trains ± SEM. I, J) Mean sypHy retrieval time constants (τ) in response to either 10 Hz (I) or 40 Hz (J) action potential trains ± SEM. For D-J, WT n = 15 coverslips, Pten+/−n = 19 from 3 independent cultures. K, L) Primary hippocampal cultures derived either wild-type (WT) or Pten+/− rat embryos were challenged with a train of action potentials (40 Hz, 10 s) in the presence of tetramethylrhodamine (TMR)-dextran (50 µM). TMR-dextran was immediately washed away and the number of TMR-dextran puncta were counted. K) Representative images of TMR-dextran uptake in WT and Pten+/− cultures. Scale bar = 50 μm. L) Mean number of TMR-dextran puncta per field of view normalised to WT ± SEM (WT n = 24 coverslips, Pten+/−n = 20 from 3 independent cultures)