Figure 1.
KIF13A is required for LTP. (A) Quantification of KIF13A mRNA levels from dissociated hippocampal neurons infected with the shKIF13A lentivirus (red) or control lentivirus (lacking the shRNA sequence; black) relative to the uninfected condition. Cultures were lysed after 15 DIV and 7 d after infection. Bars show mean ± SEM, together with individual values from each experiment. n = 9 independent experiments; P = 0.008, Wilcoxon test. (B) Left: Representative Western blot result from protein lysates from conditions described in A. Apparent protein size was lower than expected, perhaps as a consequence of proteolytic degradation. Right: Quantification of KIF13A protein levels relative to uninfected condition. Bars show mean ± SEM, together with individual values from each experiment; n = 4 independent experiments. (C) Left: Time-course of the averaged AMPAR-mediated evoked synaptic currents (mean ± SEM per condition). LTP was induced (pairing protocol of 300 pulses at 3 Hz) in organotypic hippocampal slices (7‒10 DIV) infected in the CA1 layer with a lentivirus expressing shKIF13A and analyzed at 7‒10 d after infection. Uninfected (black) and infected (red) cells were recorded. Amplitude of the synaptic responses is normalized to a 3-min baseline. Right: Average of AMPAR-mediated responses from the last 5 min of the recording and normalized to the baseline. Bar plots show mean ± SEM, together with individual values for each experiment. Left bars (LTP, paired) correspond to the stimulation pathway in which postsynaptic depolarization (0 mV) was paired to presynaptic stimulation (3 Hz, 300 pulses). Right (shaded) bars (control, unpaired) correspond to the pathway that was not stimulated during depolarization. Inset: Representative traces from uninfected (black lines) and shKIF13A (red lines), averaged from baseline (dark lines) or from the last 5 min of the recording (light lines). Scale bars: vertical, 10 pA; horizontal, 10 ms. n = 10 uninfected cells, and n = 11 shKIF13A-expressing cells. Uninfected neurons are significantly potentiated from baseline (P = 0.01, Wilcoxon test). Uninfected and shKIF13A-expressing neurons were significantly different (P = 0.03, Mann–Whitney test). (D) Basal synaptic responses mediated by AMPARs (left) and NMDARs (right) from adjacent uninfected and shKIF13A-infected CA1 pyramidal neurons at −60 mV and at +40 mV (at 65 ms), respectively. Gray symbols represent each individual pair while red symbols show the mean ± SEM of the measured currents. n = 26 cell pairs for AMPAR currents and n = 19 cell pairs for NMDAR currents; P value according to the Wilcoxon test; n.s., not significantly different. Insets: Representative traces for uninfected (black lines) and shKIF13A (red lines) cells; scale bars: vertical, 50 and 10 pA respectively; horizontal, 10 ms. (E) RT-qPCR quantification of KIF13B mRNA levels from dissociated hippocampal neurons infected with shKIF13B (burgundy) or control (lacking shRNA sequence; black) lentivirus, relative to uninfected condition. Bars show mean ± SEM, together with individual values for each experiment. n = 6 independent experiments; P = 0.03, Wilcoxon test. (F) Similar to C, but with shKIF13B-expressing (burgundy) neurons. Inset: scale bars: vertical, 10 pA; horizontal, 20 ms. n = 9 uninfected cells and n = 11 shKIF13B-expressing cells for the LTP (paired) pathway, and n = 5 uninfected cells and n = 8 shKIF13B-expressing cells for the control (Unpaired) pathway. Uninfected and shKIF13B-expressing neurons are significantly different from baseline (P = 0.02 and P = 0.03, respectively) according to the Wilcoxon test.

KIF13A is required for LTP. (A) Quantification of KIF13A mRNA levels from dissociated hippocampal neurons infected with the shKIF13A lentivirus (red) or control lentivirus (lacking the shRNA sequence; black) relative to the uninfected condition. Cultures were lysed after 15 DIV and 7 d after infection. Bars show mean ± SEM, together with individual values from each experiment. n = 9 independent experiments; P = 0.008, Wilcoxon test. (B) Left: Representative Western blot result from protein lysates from conditions described in A. Apparent protein size was lower than expected, perhaps as a consequence of proteolytic degradation. Right: Quantification of KIF13A protein levels relative to uninfected condition. Bars show mean ± SEM, together with individual values from each experiment; n = 4 independent experiments. (C) Left: Time-course of the averaged AMPAR-mediated evoked synaptic currents (mean ± SEM per condition). LTP was induced (pairing protocol of 300 pulses at 3 Hz) in organotypic hippocampal slices (7‒10 DIV) infected in the CA1 layer with a lentivirus expressing shKIF13A and analyzed at 7‒10 d after infection. Uninfected (black) and infected (red) cells were recorded. Amplitude of the synaptic responses is normalized to a 3-min baseline. Right: Average of AMPAR-mediated responses from the last 5 min of the recording and normalized to the baseline. Bar plots show mean ± SEM, together with individual values for each experiment. Left bars (LTP, paired) correspond to the stimulation pathway in which postsynaptic depolarization (0 mV) was paired to presynaptic stimulation (3 Hz, 300 pulses). Right (shaded) bars (control, unpaired) correspond to the pathway that was not stimulated during depolarization. Inset: Representative traces from uninfected (black lines) and shKIF13A (red lines), averaged from baseline (dark lines) or from the last 5 min of the recording (light lines). Scale bars: vertical, 10 pA; horizontal, 10 ms. n = 10 uninfected cells, and n = 11 shKIF13A-expressing cells. Uninfected neurons are significantly potentiated from baseline (P = 0.01, Wilcoxon test). Uninfected and shKIF13A-expressing neurons were significantly different (P = 0.03, Mann–Whitney test). (D) Basal synaptic responses mediated by AMPARs (left) and NMDARs (right) from adjacent uninfected and shKIF13A-infected CA1 pyramidal neurons at −60 mV and at +40 mV (at 65 ms), respectively. Gray symbols represent each individual pair while red symbols show the mean ± SEM of the measured currents. n = 26 cell pairs for AMPAR currents and n = 19 cell pairs for NMDAR currents; P value according to the Wilcoxon test; n.s., not significantly different. Insets: Representative traces for uninfected (black lines) and shKIF13A (red lines) cells; scale bars: vertical, 50 and 10 pA respectively; horizontal, 10 ms. (E) RT-qPCR quantification of KIF13B mRNA levels from dissociated hippocampal neurons infected with shKIF13B (burgundy) or control (lacking shRNA sequence; black) lentivirus, relative to uninfected condition. Bars show mean ± SEM, together with individual values for each experiment. n = 6 independent experiments; P = 0.03, Wilcoxon test. (F) Similar to C, but with shKIF13B-expressing (burgundy) neurons. Inset: scale bars: vertical, 10 pA; horizontal, 20 ms. n = 9 uninfected cells and n = 11 shKIF13B-expressing cells for the LTP (paired) pathway, and n = 5 uninfected cells and n = 8 shKIF13B-expressing cells for the control (Unpaired) pathway. Uninfected and shKIF13B-expressing neurons are significantly different from baseline (P = 0.02 and P = 0.03, respectively) according to the Wilcoxon test.

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