Local protein synthesis mediates a rapid increase in dendritic elongation factor 1A after induction of late long-term potentiation - PubMed (original) (raw)

Synaptic stimulation that induces protein synthesis-dependent LTP activates the mTOR pathway and increases eEF1A expression in area CA1. A, Rat hippocampal slices were stimulated, at the time indicated by the arrow, with either two trains of HFS (strong stimulation, as described in Materials and Methods; open symbols; n = 5) or a single train delivered at a lower intensity (weak stimulation; filled symbols; n = 5). Strong HFS produced LTP that persisted for at least 2 h, whereas weak stimulation resulted in only a decremental potentiation that returned to baseline within 2 h. The dashed line represents the normalized baseline value of 100%. Inset traces show superimposed sample fEPSPs recorded during the baseline period and 2 h after weak HFS (left traces) or strong HFS (right traces). Calibration:0.5 mV, 5 ms. B, In slices treated with 10 μ

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anisomycin (filled symbols; n = 7), the synaptic potentiation that followed strong HFS (delivered at arrow) was decremental, returning to baseline within 90 min. In contrast, vehicle-treated controls (open symbols; n = 7) showed LTP that was stable for 2 h. C, Strong stimulation, but not weak stimulation, activated the mTOR pathway and increased eEF1A levels in area CA1 in an NMDA receptor-mediated manner. C1, Left, Representative immunoblots from homogenates of CA1 regions from slices that had been frozen 30 min after control stimulation (CON; left lane), strong HFS (center lane), or weak HFS (right lane). Right, Immunoblots from CA1 regions of slices that received either control stimulation (left lane) or strong HFS after preincubation with vehicle (center lane) or 50 μ

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-APV (right lane). Membranes were probed for phospho(Ser2448)-mTOR and actin (top) or for phospho(Thr389)-p70S6K, total eEF1A, and actin (bottom). C2, Summary data showing mean immunoreactivity for each protein as determined by densitometry, normalized to control values within the same immunoblot. Strong stimulation increased immunoreactivity for phospho-mTOR [n = 6; p < 0.001 vs controls (_n_ = 7)], phospho-p70S6K [_n_ = 15; _p_ < 0.001 vs controls (_n_ = 15)], and eEF1A [_n_ = 10; _p_ < 0.001 vs controls (_n_ = 10)], but weak stimulation had no effect on these proteins (all _p_ values > 0.10; for phospho-mTOR, n = 3 and control, n = 7; for phospho-p70S6K, n = 10 and control, n = 15; for eEF1A, n = 7 and control, n = 10). Asterisks indicate significant differences from control, which is represented by the dashed line. Preincubation with

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-APV blocked the ability of strong HFS to increase the phosphorylation of mTOR [for APV treated, n = 5; p > 0.10 vs controls (n = 7)] and the phosphorylation of p70S6K [for APV treated, n = 6; p > 0.10 vs controls (n = 15)], as well as the expression of eEF1A [for APV treated, n = 4; p > 0.10 vs controls (n = 10)]. Asterisks indicates significant differences from control. D, Anisomycin blocked the ability of strong HFS to increase eEF1A expression. Left, Representative immunoblot of CA1 homogenates taken from slices that had received control stimulation (Con; left lane) or strong HFS after preincubation with either vehicle (center lane) or 10 μ

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anisomycin (Aniso). Slices had been frozen 30 min after stimulation. Right, Summary data showing that the activation by strong HFS of the mTOR pathway in area CA1 was intact in anisomycin (ANISO)-treated slices, as shown by the phosphorylation of p70S6K at T389 (left columns). However, anisomycin completely blocked the HFS-induced increase in eEF1A immunoreactivity (right columns), indicating translation-dependent expression of eEF1A in response to strong HFS [in vehicle-treated slices, phospho-p70S6K (n = 3), p < 0.05 vs controls (_n_ = 3); eEF1A (_n_ = 3), _p_ < 0.01 vs controls (_n_ = 3); in anisomycin-treated slices, phospho-p70S6K (_n_ = 3), _p_ < 0.05 vs controls (_n_ = 3); eEF1A (_n_ = 3), _p_ > 0.10 vs controls (n = 3; two-tailed t test)]. Asterisks indicate significant differences from control. Error bars represent SE.