Dendritic Kv4.2 potassium channels selectively mediate spatial pattern separation in the dentate gyrus

Abstract

The capacity to distinguish comparable experiences is fundamental for the recall of similar memories and has been proposed to require pattern separation in the dentate gyrus (DG). However, the cellular mechanisms by which mature granule cells (GCs) of the DG accomplish this function are poorly characterized. Here, we show that Kv4.2 channels selectively modulate the excitability of medial dendrites of dentate GCs. These dendrites are targeted by the medial entorhinal cortex, the main source of spatial inputs to the DG. Accordingly, we found that the spatial pattern separation capability of animals lacking the Kv4.2 channel is significantly impaired. This points to the role of intrinsic excitability in supporting the mnemonic function of the dentate and to the Kv4.2 channel as a candidate substrate promoting spatial pattern separation.

Keywords: Biological sciences; Cellular neuroscience; Neuroscience.

Graphical abstract

Figure 1

Enhanced excitability of mature GCs of Kv4.2 knockout mice in response to medial perforant path synaptic stimulation (A) Scheme showing the positioning of the extracellular stimulation electrode at the middle of the molecular layer in order to stimulate the medial perforant path while recording electrical activity from single mature GCs through the patch-clamp technique. (B) GCs from Kv4.2 knockouts (n = 48) show, for a given EPSP slope, EPSPs of significantly larger amplitude than controls (n = 47). Insets show illustrative EPSP traces, as well as a statistically significant reduction in the EPSP threshold (EPSP slope needed to fire an action potential with 50% probability) in Kv4.2 knockouts. (C) The EPSP amplitudes were significantly larger for Kv4.2 knockouts around the critical range of EPSP slopes needed to fire an action potential with 50% probability. (D) An illustrative example of the amplitude/slope relation of subthreshold EPSPs for a Kv4.2 knockout and a wild-type cell. For a given EPSP slope, the amplitude of the EPSP is increased in the knockout cell. Solid lines show linear fits for each group (slopes of the fitted lines: wild type 2.511 ± 0.03262 ms and knockout 3.083 ± 0.05248 ms). (E) Example of a plotting of the firing probability versus EPSP slope for a wild-type and a Kv4.2 knockout cell. There is an increased firing probability for a given EPSP slope in the knockout cell. All recordings were made in the presence of bicuculline. Data are represented as mean ± SEM. Scale bars represent 10 mV/10 ms. ∗: p < 0.05, ∗∗∗: p < 0.001, t test. #: p < 0.05, ANOVA.

Figure 2

Kv4.2 knockout does not alter the general somatic excitability of mature dentate GCs (A–D) (A) Representative traces showing the firing of mature GCs in response to a 250-ms-long square pulse current injection to the soma. Scale bars, 20 mV/50 ms. Neither the number of action potentials (B) nor the minimum current needed to elicit an action potential (i.e. rheobase), (C) nor the input resistance or resting membrane potential of the cell, (D) were modified in Kv4.2 knockout mice. (E–H) (E) Representative traces showing the firing of mature GCs in response to a 5-ms-long square pulse current injection to the soma. Scale bars, 20 mV/4 ms. Also shown are the phase plots of the action potentials. Scale bars, 100 mV∗ms⁻¹/50 mV. Neither the action potential amplitude (F) nor the action potential threshold, (G) nor the action potential width, (H) were modified. (Kv4.2 −/− n = 19, Kv4.2 +/+ n = 15). Data are represented as mean ± SEM

Figure 3

Loss of Kv4.2 channels does not affect the general morphology of mature dentate GCs (A) Golgi-stained dentate GCs of wild-type (left) and Kv4.2 knockout mice (right). The spine density in medial dendrites was similar in both genotypes. (B–D) (B).Dendritic branching was examined using Sholl analysis. Neither the number of crossings (C) nor the dendritic length (D) differed between genotypes. Data are represented as mean ± SEM. Scale bars, 50 μm or 20 μm for wide field view of cells or enlarged image of spines, respectively. n = 12 for every genotype.

Figure 4

Mature GCs activated during a short exposure to an enriched environment show an enhanced intrinsic excitability, mimicking the phenotype of the Kv4.2 knockout neurons (A) cFos-GFP reporter mice were used to target mature GCs activated by the enriched environment experience. Electrophysiological recordings were obtained from GFP-positive cells (activated) and GFP-negative (non-activated) neighbor cells. (B) Activated mature GCs show, for a given EPSP slope, EPSPs of significantly larger amplitude than non-activated neighbor cells after medial perforant path stimulation as well as a smaller EPSP threshold compared to controls (n = 24). (C) An illustrative example of the amplitude/slope relation of subthreshold EPSPs for a GFP-positive and a neighbor GFP-negative cell. For a given EPSP slope, the amplitude of the EPSP is increased in the GFP-positive cell. (D) Example of a plotting of the firing probability versus EPSP-slope for a GFP-positive and a neighbor GFP-negative cell. There is an increased firing probability for a given EPSP slope in the GFP-positive cell. All recordings were made in the presence of bicuculline. Scale bars, 10 mV/10 ms.

Figure 5

Kv4.2 knockout results in larger bAPs into medial dendrites of mature GCs (A and B) (A) Scheme showing the backpropagation of the action potential from the soma to dendrites. Neurons were filled with Fluo-5F, and calcium transients were measured at different distances from the soma. Mature GCs of Kv4.2 knockouts (Kv4.2 −/− n = 20, Kv4.2 +/+ n = 14) showed an enhanced action potential backpropagation into medial dendrites (B). The insets in (B) show individual values for Kv4.2 wild-type and knockout cells at 75–100 μm from the soma, as well as representative calcium transients. All recordings were made in the presence of bicuculline. Scale bar in (A) 50 μm. Data are represented as mean ± SEM. ∗: p < 0.05, ANOVA.

Figure 6

Occlusion of plasticity-induced excitability changes in Kv4.2 knockout mice after theta burst stimulation to the medial perforant path (A–C) (A) Scheme showing the positioning of the extracellular stimulation electrode at the middle of the molecular layer in order to stimulate the medial perforant path while recording electrical activity from mature GCs. Time course of field EPSP slope (B) and population spike amplitude (PSA) (C). PSA and fEPSP values were normalized for every slice to the pre-conditioning baseline. The theta burst stimulation protocol was applied at time “0”. One hour after the theta burst stimulation protocol, the test stimulation intensity was reduced to match the baseline fEPSP slope values. The remaining PSA potentiation is due to excitability changes independent of synaptic potentiation. GCs from Kv4.2 knockouts (Kv4.2 −/− n = 12, Kv4.2 +/+ n = 9) had impaired PSA potentiation, despite a normal fEPSP potentiation. Data are represented as mean ± SEM. Scale bars, 10 mV/10 ms. ∗∗: p < 0.01, t test.

Figure 7

Kv4.2 knockout results in an impairment in spatial pattern separation task (A) Scheme of the settings of the experiment for the spatial pattern separation task for both the similar and dissimilar conditions. (B) Kv4.2 knockout mice (n = 13) were impaired in their ability to discriminate highly similar locations, compared to the performance of wild types (n = 11). (C) However, they (n = 13) performed as well as controls (n = 14) when in the dissimilar location condition. (D) Scheme of the settings of the experiment for the non-spatial object pattern separation task for both similar and dissimilar conditions. Kv4.2 knockout mice performed at similar levels than controls when discriminating both (E) similar (Kv4.2 +/+ n = 8, Kv4.2 −/− n = 11) and (F) dissimilar (Kv4.2 +/+ n = 8, Kv4.2 −/− n = 12) objects. Scale bars, 2 cm. Data are represented as mean ± SEM. ∗: p < 0.05, t test.