Comprehensive studies on Kv4.1 and Kv4.2 in mature granule cells of hippocampal dentate gyrus: distinct roles, subcellular distributions, and pathophysiological implications

Abstract

Background: Dentate gyrus (DG), the main gateway to the hippocampus and where adult neurogenesis occurs, play key roles in pattern separation by forming distinct representation of similar inputs. Sparse action potential generation in dentate gyrus granule cells (DG-GCs) has been thought as a cellular mechanism of pattern separation. The balance between excitatory and inhibitory synaptic inputs was reported to be important for determining low excitability of DG-GCs, but the ion channel mechanisms underlying sparse action potential firing remain unclear. Calbindin-D28K (CB) is a major Ca2+ buffer in DG-GCs of the hippocampus. Reduction of CB expression in DG-GCs is reported in various pathologic conditions that accompany with cognitive dysfunction, including Alzheimers disease (AD). However, the link between intracellular Ca2+ dysregulation and neuronal excitability in DG-GCs has not been investigated. Methods: I investigated the mechanism of sparse action potential firing in DG-GCs at the age of 1-2 months old mice, and examined how this mechanism was altered in transgenic mice (Alzheimers disease model mice (Tg2576) and Calbindin-D28K knock-out mice (CBKO)). Using voltage- and current-clamp techniques, voltage-dependent K+ currents and intrinsic excitability of DG-GCs and CA1 pyramidal cells (PCs) in acute brain slices were analyzed. For molecular analysis, quantitative real-time reverse transcription PCR (qRT-PCR), Western blot, and immunohistochemistry were performed. Anti-Kv4.1 and anti-Kv4.2 antibodies were used for selective blockade of corresponding ion channels as well as molecular analysis. Behavior tests for analyzing pattern separation were performed in control, Tg2576, and CBKO mice. Results: Immunohistochemistry analysis showed that Kv4.1 subunits are strongly expressed in mature DG-GCs, but weak in CA1 in the hippocampus. Whole-cell current clamp analysis showed that Kv4.1 antibody increased firing frequency in mature DG-GCs, but did not affect any parameters of intrinsic excitability in immature DG-GCs and CA1 PCs, suggesting that Kv4.1 play a selective role in limiting firing frequency of mature DG-GCs. Whole-cell voltage-clamp analysis revealed that Kv4.1-mediated currents (IKv4.1) in DG-GCs have distinct properties from A-type K+ channels, showing slow inactivation kinetics and sensitivity to intracellular Ca2+. Reduction of Kv4.1 expression in DG was found in CBKO and Tg2576 mice. Consistently, firing frequency in mature DG-GCs was increased in CBKO and Tg2576 mice. Furthermore, CBKO and Tg2576 mice showed a deficit in pattern separation without impairment in memory acquisition, supporting the link between sparse firing of DG-GCs and pattern separation. Unexpectedly, I found that the endogenous Ca2+ buffer capacity in mature DG-GCs was reduced in Tg2576 mice to a level comparable to CBKO mice. The reduction of Ca2+ buffering in Tg2576 DG-GCs was mimicked by the exogenous application of oligomeric amyloid β (Aβ) 1-42 protein and restored by the antioxidant Trolox, suggesting that Aβ-induced oxidative stress induces CB dysfunction in Tg2576 mice. When Tg2576 mice were treated with Trolox for 1 week, hyperexcitability of DG-GCs and impairment of pattern separation were restored. Conclusion: These data suggest that Kv4.1 channels, selectively expressed in DG-GCs and acting as a regulator to prevent hyperexcitability of DG-GCs, are essential for pattern separation. Reduction of Kv4.1 expression in AD and CBKO mice suggests the pathophysiological significance of Kv4.1 in various conditions causing Ca2+ dysregulation.