Reelin supplementation enhances cognitive ability, synaptic plasticity, and dendritic spine density - PubMed (original) (raw)

. 2011 Aug 18;18(9):558-64.

doi: 10.1101/lm.2153511. Print 2011 Sep.

Ian Rusiana, Justin Trotter, Lisa Zhao, Erika Donaldson, Daniel T S Pak, Lenard W Babus, Melinda Peters, Jessica L Banko, Pascale Chavis, G William Rebeck, Hyang-Sook Hoe, Edwin J Weeber

Affiliations

Reelin supplementation enhances cognitive ability, synaptic plasticity, and dendritic spine density

Justin T Rogers et al. Learn Mem. 2011.

Abstract

Apolipoprotein receptors belong to an evolutionarily conserved surface receptor family that has intimate roles in the modulation of synaptic plasticity and is necessary for proper hippocampal-dependent memory formation. The known lipoprotein receptor ligand Reelin is important for normal synaptic plasticity, dendritic morphology, and cognitive function; however, the in vivo effect of enhanced Reelin signaling on cognitive function and synaptic plasticity in wild-type mice is unknown. The present studies test the hypothesis that in vivo enhancement of Reelin signaling can alter synaptic plasticity and ultimately influence processes of learning and memory. Purified recombinant Reelin was injected bilaterally into the ventricles of wild-type mice. We demonstrate that a single in vivo injection of Reelin increased activation of adaptor protein Disabled-1 and cAMP-response element binding protein after 15 min. These changes correlated with increased dendritic spine density, increased hippocampal CA1 long-term potentiation (LTP), and enhanced performance in associative and spatial learning and memory. The present study suggests that an acute elevation of in vivo Reelin can have long-term effects on synaptic function and cognitive ability in wild-type mice.

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Figures

Figure 1.

Figure 1.

Reelin injection increased expression throughout the entire hippocampus. Reelin immunoreactivity was detected using the G10 anti-Reelin antibody (left column). Immunohistochemical analysis of Reelin revealed increased Reelin immunoreactivity in the hippocampus 15 min following injection, whereas saline injection resulted in no detectable differences. Reelin levels were found to be uniformly increased 3 h following injection and returned to baseline levels 5 d post-injection (scale bar: 50 µm).

Figure 2.

Figure 2.

Reelin injection enhanced Dab1 phosphorylation throughout the entire hippocampus. Activation of Dab1 was determined with p-Tyr220 specific antibody. Immunohistochemical analysis of Dab1 activation revealed that Reelin injection resulted in increased Tyr220 phosphorylation of Dab1 at the 15-min time point, remained elevated through 3 h and returned to baseline after 5 d post-injection (scale bar: 50 µm).

Figure 3.

Figure 3.

Reelin injection enhanced CREB phosphorylation throughout the entire hippocampus. Activation of CREB was determined with p-Ser133 specific antibody. Immunohistochemical analysis of CREB activation revealed that Reelin injection resulted in increased Ser133 phosphorylation of CREB at the 15-min time point, remained elevated through 3 h, and returned to baseline after 5 d post-injection (scale bar: 50 µm).

Figure 4.

Figure 4.

Reelin increased spine density in pyramidal cells in area CA1. Mouse brains were Golgi stained and CA1 were imaged (n = 4 mice/group). (A) Representative AO dendrites at 3 h and at 5 d per group. (B) Averaged AO spine densities for each group at 3 h (saline: 15.21 ± 0.79, n = 25; Reelin: 15.51 ± 0.79, n = 28) and at 5 d (saline: 13.5 ± 0.60, n = 25; Reelin: 19.2 ± 0.86, n = 34). Reelin significantly increased AO spine density in area CA1 at 5 d but not 3 h post-injection. (C) Representative BS dendrites at 3 h and at 5 d for each group. (D) Averaged BS spine densities for each group at 3 h (saline: 15.75 ± 0.76, n = 25; Reelin: 15.30 ± 0.71, n = 33) and at 5 d (saline: 13.64 ± 0.85, n = 28; Reelin: 17.64 ± 0.70, n = 28). Reelin significantly increased BS spine density in area CA1 at 5 d but not 3 h post-injection. (Data expressed as mean ± SEM; *P < 0.0005.)

Figure 5.

Figure 5.

Reelin significantly increased spine density in wild-type mice compared to ApoER2 KO mice 5 d post-injection in area CA1. Spine increases are expressed as a percentage of experimental saline controls. (A) Representative AO dendrites of both Reelin-treated groups at 5 d post-injection. (B) Averaged AO percent dendritic spine increase for each group (wild-type saline: 100.0 ± 4.4%; wild-type Reelin: 142.3 ± 6.5%; ApoER2 KO saline: 100 ± 4.3%; ER2 KO Reelin: 113.6 ± 3.7%). Reelin significantly increased AO spine density in area CA1 compared to saline- or Reelin-injected ER2 − /− mice. (C) Representative BS dendrites of both Reelin-treated groups at 5 d post-injection. (D) Averaged BS percent dendritic spine increase for each group (wild-type saline: 100.0 ± 6.2%; wild-type Reelin: 130.0 ± 5.1%; ApoER2 KO saline: 100 ± 6.5%; ER2 KO Reelin: 110.8 ± 7.6%). Reelin significantly increased BS spine density in area CA1 compared to saline- or Reelin-injected ER2 − /− mice (*P < 0.05; **P < 0.01; ***P < 0.001).

Figure 6.

Figure 6.

Reelin enhances hippocampal synaptic plasticity. Mice were sacrificed 5 d following single, bilateral injections for electrophysiology experiments. LTP was induced with TBS (five bursts of 200 Hz separated by 200 msec, repeated six times with 10 sec between the six trains; arrow) after 20 min of baseline recording and changes in fEPSP slope are expressed as a percentage of baseline. (A) Representative fEPSP traces from both saline (white) and Reelin (black) injected hippocampi. (B) Reelin injection enhanced LTP in area CA1. The last 5 min of fEPSPs slope recordings were averaged (bar) for both saline (n = 9) and Reelin (n = 10) injected (*P < 0.05). (C) Output field analysis following increasing field stimulation fit with nonlinear regression. There were no significant differences between experimental groups. (D) PPF was induced with the use of paired pulses given with an initial delay of 20 msec and the time to the second pulse was increased 20 msec incrementally until a final delay of 300 msec was reached. There was no significant difference between experimental groups.

Figure 7.

Figure 7.

There is no effect of Reelin on general locomotor or anxiety levels in wild-type mice. (A) After 5 d post-injection of saline (n = 7) or Reelin (n = 6), mice underwent open-field testing as a locomotor and general anxiety control for behavioral testing. Data represent the ratio of time spent (sec) in the open field vs. the perimeter of the field. There were no significant differences between experimental groups. (B) After 5 d post-injection, mice underwent elevated plus maze testing as a general anxiety control. Data represent the ratio of time spent (sec) in the open areas vs. the closed arms of the elevated maze. There were no significant differences between experimental groups.

Figure 8.

Figure 8.

Reelin supplementation enhanced spatial learning and memory. Mice began training in the HPWM 5 d post-injection of saline (white, n = 7) or Reelin (black, n = 5). Mice were trained in the HPWM for 4 d, four trials per day. On day 5, the platform was removed and a 60-sec probe trial was conducted. On trial days 6 and 7, the platform was moved to the opposite quadrant. (A) Reelin significantly reduced latency to the platform during HPWM training. (B) Analysis of day 1 training reveals Reelin reduced latencies to the platform after the second trial compared to the initial trial. Reelin significantly lowered latencies on trials 3 and 4 compared to saline-injected animals. (C) Reelin increased target quadrant entries during the probe trail compared to saline-treated animals (*P < 0.05; #P < 0.01).

Figure 9.

Figure 9.

Reelin enhanced associated learning and memory following contextual fear conditioning. Both Reelin (black)- and saline (white)-injected mice were trained with a standard two-shock protocol. Mouse freezing to the context was assessed at 1 h, 24 h, and 72 h. (A) There were no significant differences in freezing to the context at the 1-h time point between Reelin (n = 6)- and saline (n = 6)-injected mice. (B) Reelin supplementation increased context-dependent freezing at the 24-h time point in Reelin (n = 10)-injected mice when compared to saline (n = 8) control mice. (C) Reelin supplementation increased context-dependent freezing at the 72-h time point in Reelin (n = 8)-injected mice when compared to saline (n = 7) control mice (*P < 0.05).

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