Kalirin-7 is necessary for normal NMDA receptor-dependent synaptic plasticity

Abstract

Background: Dendritic spines represent the postsynaptic component of the vast majority of excitatory synapses present in the mammalian forebrain. The ability of spines to rapidly alter their shape, size, number and receptor content in response to stimulation is considered to be of paramount importance during the development of synaptic plasticity. Indeed, long-term potentiation (LTP), widely believed to be a cellular correlate of learning and memory, has been repeatedly shown to induce both spine enlargement and the formation of new dendritic spines. In our studies, we focus on Kalirin-7 (Kal7), a Rho GDP/GTP exchange factor (Rho-GEF) localized to the postsynaptic density that plays a crucial role in the development and maintenance of dendritic spines both in vitro and in vivo. Previous studies have shown that mice lacking Kal7 (Kal7(KO)) have decreased dendritic spine density in the hippocampus as well as focal hippocampal-dependent learning impairments.

Results: We have performed a detailed electrophysiological characterization of the role of Kal7 in hippocampal synaptic plasticity. We show that loss of Kal7 results in impaired NMDA receptor-dependent LTP and long-term depression, whereas a NMDA receptor-independent form of LTP is shown to be normal in the absence of Kal7.

Conclusions: These results indicate that Kal7 is an essential and selective modulator of NMDA receptor-dependent synaptic plasticity in the hippocampus.

Figure 1

Basal AMPA receptor-mediated synaptic transmission is not altered by Kal7 deletion. A) Example traces of evoked field EPSPs (fEPSPs) in WT and Kal7^KO mice. B) Input-output curve showing the relationship between stimulation intensity and presynaptic fiber volley for evoked AMPA receptor-mediated hippocampal fEPSPs. This is a measure of the excitability of the presynaptic axons that are being stimulated (WT: n = 8 slices from 3 animals; Kal7^KO: n = 6 slices from 3 animals). C) Input-output curve showing relationship between presynaptic fiber volley and fEPSP amplitude for the same slices as in B. D) Input-output curve showing relationship between presynaptic fiber volley and fEPSP slope for the same slices as in B. There was no significant difference between WT and Kal7^KO for either amplitude or slope.

Figure 2

NMDA receptor-dependent LTP is impaired in Kal7^KO animals. A) Example traces of evoked fEPSPs before (black) and 60 min after (red) 3 trains of theta burst stimulation (TBS) for a WT and Kal7^KO animal. Each TBS train consisted of 10 bursts delivered at 5 Hz, and each burst consisted of 5 stimuli at 100 Hz. Calibration bars: 0.1 mV, 20 ms. Example traces in this and subsequent figures are averages of 10-15 sweeps. B) Group time courses of fEPSP slope recorded from WT (n = 5 animals) and Kal7^KO mice (n = 5) following 3 trains of TBS delivered at time zero. C) Group data showing magnitude of LTP measured 60 min post-induction for 1 train of TBS (WT: n = 8; Kal7^KO: n = 7), 3 trains of TBS (n = 5 for both WT and Kal7^KO), or high frequency stimulation (HFS; 100 Hz/1 sec; n = 5 for both WT and Kal7^KO). *, p < 0.05 compared to baseline; #, p < 0.05 between condition. D) LTP induced by TBS or HFS requires NR2B-containing NMDA receptors. Group data showing the lack of TBS-induced or HFS-induced LTP in the presence of the NMDA receptor antagonist CPP (3 μM) in both WT and Kal7^KO animals (n = 3-5 slices/condition), and the lack of TBS-induced LTP in WT animals in the presence of the NR2B antagonist ifenprodil (3 μM; n = 5).

Figure 3

Long-term depression (LTD) is disrupted in Kal7^KO mice. A) Example traces of evoked fEPSPs before (black) and 30 min after (red) paired-pulse low frequency stimulation (PP-LFS; 3 stimuli/50 ms intervals delivered at 1 Hz for 15 min) for a WT and Kal7^KO animal. Calibration bars: 0.1 mV, 20 ms. B) Group time courses for WT and Kal7^KO mice showing the fEPSP slope in response to PP-LFS. LTD induction was at time zero (excluded from time course). C) Group data showing the effect of PP-LFS on fEPSP slope 60 min post-induction in WT and Kal7^KO animals (n = 8 animals, 2 slices/animal for each genotype). *, p < 0.05 compared to baseline.

Figure 4

NMDA receptor-independent LTP is unimpaired in Kal7^KO animals. A) Example traces of evoked fEPSPS before (black) and 60 min after (red) LTP induction (200 Hz/2 sec in the presence of 3 μM CPP). Calibration bars: 0.1 mV, 20 ms. B) Representative examples from a WT and a Kal7^KO animal showing initial depression and slowly developing potentiation. LTP induction occurred at time zero. B) Group data showing the magnitude of NMDA receptor-independent LTP in WT and Kal7^KO animals 60 min post-induction (n = 7-8 animals, 2 slices/animal). *, p < 0.05 compared to baseline; ns, no significant difference between genotypes.

Figure 5

Application of NPPD, an inhibitor of the Rho-GEF activity of Kal7, suppressed hippocampal LTP and LTD. A) Input-output relationship for effects of NPPD (100 μM) on fEPSPs in 4 week old CD1 mice. B) Sample sweeps before (black) and 60 min after (red) LTP induction for control and NPPD-treated slices. The LTP induction protocol consisted of 3 trains of TBS. Calibration bars: 0.1 mV, 20 ms. C) Group time course for the effects of NPPD on LTP. D) Group data for the effect of NPPD on LTP (n = 5 slices/4 animals) or LTD (n = 7 slices/3 animals). The LTD induction protocol was the same as in Figure 3. For all experiments, NPPD (100 μM) was present for a 15 min pretreatment and throughout the experiment.