Figure 4. Medial entorhinal cortex-layer Vb (MEC-LVb) neurons preferentially target LII/III pyramidal neurons.

(A) Image of a representative horizontal slice showing expression of oChIEF-citrine in LVb neurons (green) and recorded neurons labeled with biocytin (magenta) in MEC. Inset shows a low-power image of the section indicating the position of the higher power image. Scale bars represent 500 μm (inset) and 100 μm. (B) Voltage responses to injected current steps recorded from neurons shown in (A): i, pyramidal cell in layer Va (LVa); ii, pyramidal cell in LIII; iii, pyramidal cell in LII; iv, stellate cell in LII. Inset in (iii) and (iv) shows the depolarizing afterpotential (DAP) in expanded voltage- and time scale. Note that LII stellate cells (iv) show a clear sag potential and DAP compared to LII pyramidal cells (iii). (C) Voltage responses to light stimulation (light blue line) recorded from neurons shown in (A). Average traces (blue) are superimposed on the individual traces (gray). (D–G) The proportion of responding cells (D), excitatory postsynaptic potential (EPSP) amplitude (E), the normalized EPSP based on LIII response (F, one-way ANOVA, F_3,47 = 33.29, ***p<0.0001, Bonferroni’s multiple comparison test, **p<0.01, ***p<0.001), and the input resistance (G, one-way ANOVA, F_3,82 = 21.99, ***p<0.0001, Bonferroni’s multiple comparison test, ***p<0.001) was examined for each cell type (error bars: mean ± standard errors). (H) Latency of EPSP onset for MEC neurons to optical activation (F, one-way ANOVA, F_3,47 = 11.65, ***p<0.0001, Bonferroni’s multiple comparison test, *p<0.05, ***p<0.001). LIIs: LII stellate cell; LIIp: LII pyramidal cell. Figure 4—source data 1. See also Figure 4—figure supplement 1, Figure 4—figure supplement 2.

Figure 4—figure supplement 1. Representative patch-clamp recordings after optical stimulation of layer Vb (LVb) fibers in medial entorhinal cortex (MEC).

(A–C) Horizontal MEC slice showing expression of oChIEF-citrine in LVb neurons (green) and biocytin-filled recorded layer II/III neurons (magenta, A). Voltage responses to injected current steps (B) and to light stimulation (light blue line, C) are shown for all three neurons (i, pyramidal neuron in LIII; ii, iii, stellate cells in LII). Average traces (blue) are superimposed on the individual traces (gray). (D–F) Images of recorded MEC neurons (D) and their voltage responses (E, F; iv, v pyramidal cells in LVa; vi, pyramidal cell in LIII; vii, stellate cell in LII). (G, H) Images of recorded MEC neurons (G) in LVa (Viii–x) and LIII (xi–xiii), and their voltage responses (H). This slice is from the animal shown in Figure 4, but it is more ventral than the one shown in Figure 4. Scale bars represent 500 μm for insets and 100 μm for higher-power images.

Figure 4—figure supplement 2. Responses of medial entorhinal cortex-layer Va (MEC-LVa) neurons at different dorsoventral levels.

(A) Schematic drawing and representative images showing three dorsoventral levels of slices in which the LVa neurons were recorded (D, dorsal level; M, medial level; V, ventral level). D (blue) corresponds to the sample shown in Figure 4—figure supplement 1D-F, and M (green) and V (red), which are taken from the same animal, correspond to the sample shown in Figure 4 and Figure 4—figure supplement 1G, H, respectively. (B, C) Normalized excitatory postsynaptic potentials (EPSPs) of LVa neurons (B) are significantly larger in more ventral slices (one-way ANOVA, F_2,15 = 9.82, ***p=0.0019, Bonferroni’s multiple comparison test, **p<0.01), whereas EPSPs of LIII neurons recorded at different dorsoventral level are indifferent (C) (error bars: mean ± standard errors, one-way ANOVA, F_2,14 = 1.045, p=0.3777). (D) Normalized EPSPs of LVa neurons in MEC (left panel) and lateral entorhinal cortex (LEC) (right panel) are shown against the maintained length of their apical dendrites. Note that severing the dendrites does not impact the presence or absence of responses in dorsal MEC, and that LEC LVa neurons respond even with having severed apical dendrites. Voltage responses recorded from the slice in (A) are shown in (BD) as filled-color dots (filled-blue dots in D, filled-green dots in M, and filled-red dots in V). Figure 4—figure supplement 2—source data 1.