Priming of long-term potentiation in mouse hippocampus by corticotropin-releasing factor and acute stress: implications for hippocampus-dependent learning

Abstract

In the present experiments, we characterized the action of human/rat corticotropin-releasing factor (h/rCRF) and acute stress (1 hr of immobilization) on hippocampus-dependent learning and on synaptic plasticity in the mouse hippocampus. We first showed that h/rCRF application and acute stress facilitated (primed) long-term potentiation of population spikes (PS-LTP) in the mouse hippocampus and enhanced context-dependent fear conditioning. Both the priming of PS-LTP and the improvement of context-dependent fear conditioning were prevented by the CRF receptor antagonist [Glu(11,16)]astressin. PS-LTP priming and improved learning were also reduced by the protein kinase C inhibitor bisindolylmaleimide I. Acute stress induced the activation of Ca2+/calmodulin-dependent kinase II (CaMKII) 2 hr after the end of the stress session. The CaMKII inhibitor KN-62 antagonized the stress-mediated learning enhancement, however, with no effect on PS-LTP persistence. Thus, long-lasting increased neuronal excitability as reflected in PS-LTP priming appeared to be essential for the enhancement of learning in view of the observation that inhibition of PS-LTP priming was associated with impaired learning. Conversely, it was demonstrated that inhibition of CaMKII activity reduced contextual fear conditioning without affecting PS-LTP priming. This observation suggests that priming of PS-LTP and activation of CaMKII represent two essential mechanisms that may contribute independently to long-term memory.

Fig. 1.

h/rCRF-mediated facilitation of PS-LTP persistence is prevented by the inhibition of CRF receptors and PKC.A, Left, Representative recordings performed before (1), during (2), and after (3) h/rCRF application and 1 hr after tetanus (4).Traces represent the average of six recordings.Right, A 20 min h/rCRF application (125 nm; ●) transiently increased population spike amplitudes and subsequently enhanced the persistence of PS-LTP induced by TBS compared with controls (○). The selective PKC inhibitor BIS-I (1,2 μm; ■) was bath-applied for the rest of the experiment. This treatment had no effect on PS-LTP persistence. B, Preincubation of slices with BIS-I (1,2 μm; ■) for 1 hr markedly prevented the h/rCRF-mediated increase (125 nm; ●) of population spike amplitudes and subsequent priming of hippocampal PS-LTP. Preincubation of slices with the CRF receptor antagonist [Glu^11,16]astressin (500 nm; ▵) for 40 min completely blocked the h/rCRF-mediated increase of population spike amplitudes and the facilitation of TBS-induced PS-LTP. Data are presented as mean ± SEM.

Fig. 2.

Inhibition of PKC and CRF receptors but not of CaMKII prevents stress-mediated facilitation of LTP maintenance in the hippocampal CA1 area. A, TBS-induced PS-LTP in slices prepared from nonstressed animals (●) and in slices prepared 2 hr after exposure of the animal to 1 hr of immobilization (○).B, TBS-induced PS-LTPs from animals that were injected intracerebroventricularly with KN-62 (⋄), BIS-I (■), or [Glu^11,16]astressin (▴) immediately before 1 hr of immobilization. Hippocampal slices were prepared 2 hr after the end of the stress session. Data are presented as mean ± SEM.

Fig. 3.

Acute stress induces activation of CaMKII in the hippocampal CA1 area. A, CA1 homogenates dissected from animals at several time points after 1 hr of immobilization were probed with an antibody specific for Thr²⁸⁶-phosphorylated CaMKII or an antibody recognizing total CaMKII. Nonstressed mice are shown as controls. Results are representative of four independent Western blots. B, The bar graph summarizes Western blot data of four experiments and shows the levels of Thr²⁸⁶-phosphorylated CaMKII expressed as a percentage of nonstressed controls. The dashed linerepresents the normalized average of nonstressed controls (set to 100%). Statistically significant differences: *p< 0.05 versus nonstressed controls.

Fig. 4.

Inhibition of CRF receptors, PKC, and CaMKII in the dorsal hippocampus prevents stress-mediated enhancement of context-dependent fear conditioning. A, Mice were injected intrahippocampally (i.h.) or intracerebroventricularly (i.c.v.) with aCSF, KN-62, or BIS-I immediately before the start of the stress session and trained at several time points after the end of the stress session.B, Mice were injected intrahippocampally or intracerebroventricularly with [Glu^11,16]astressin immediately before the stress session and trained 2 hr after immobilization. Freezing was measured in the memory test performed 24 hr after training. Statistically significant differences: *p < 0.05 versus aCSF; **p < 0.05 versus control.