Layer-specific changes of KCC2 and NKCC1 in the mouse dentate gyrus after entorhinal denervation

Abstract

The cation-chloride cotransporters KCC2 and NKCC1 regulate the intracellular Cl− concentration and cell volume of neurons and/or glia. The Cl− extruder KCC2 is expressed at higher levels than the Cl− transporter NKCC1 in mature compared to immature neurons, accounting for the developmental shift from high to low Cl− concentration and from depolarizing to hyperpolarizing currents through GABA-A receptors. Previous studies have shown that KCC2 expression is downregulated following central nervous system injury, returning neurons to a more excitable state, which can be pathological or adaptive. Here, we show that deafferentation of the dendritic segments of granule cells in the outer (oml) and middle (mml) molecular layer of the dentate gyrus via entorhinal denervation in vivo leads to cell-type- and layer-specific changes in the expression of KCC2 and NKCC1. Microarray analysis validated by reverse transcription-quantitative polymerase chain reaction revealed a significant decrease in Kcc2 mRNA in the granule cell layer 7 days post-lesion. In contrast, Nkcc1 mRNA was upregulated in the oml/mml at this time point. Immunostaining revealed a selective reduction in KCC2 protein expression in the denervated dendrites of granule cells and an increase in NKCC1 expression in reactive astrocytes in the oml/mml. The NKCC1 upregulation is likely related to the increased activity of astrocytes and/or microglia in the deafferented region, while the transient KCC2 downregulation in granule cells may be associated with denervation-induced spine loss, potentially also serving a homeostatic role via boosting GABAergic depolarization. Furthermore, the delayed KCC2 recovery might be involved in the subsequent compensatory spinogenesis.

Bidirectional Regulation of GABAA Reversal Potential in the Adult Brain: Physiological and Pathological Implications

“…For example, in a pharmacological model of cognitive impairment associated with schizophrenia, NKCC1 is increased in the ventral but not in the dorsal subregion of the medial prefrontal cortex (mPFC) [51], and in a rodent model of neuropathic pain, KCC2 expression is increased in the mPFC but not in the neighboring anterior cingulate cortex [16]. NKCC1 and KCC2 can also be differentially regulated in different layers within the same brain areas, as shown in mouse granule cells and outer and middle molecular layers following entorhinal denervation [52].…”

Maintenance of Lognormal‐Like Skewed Dendritic Spine Size Distributions in Dentate Granule Cells of TNF, TNF‐R1, TNF‐R2, and TNF‐R1/2‐Deficient Mice

“…As hippocampal neurons receive a highly layer-specific input, differences in pathway activity could result in differences in spine head sizes and their distributions in each layer. Along this line, layer-specific stimulation of granule cells, such as high-frequency stimulation of the perforant path (Jungenitz et al 2014(Jungenitz et al , 2018 or layer-specific denervation of granule cells (del Turco et al 2023;Deller and Frotscher 1997;Deller et al 2007;Vuksic et al 2011), could theoretically shift the spine head size distribution away from lognormality-or change its skewness or mean. Indeed, alterations in the mean and skewness of spine size distributions have been detected following layer-specific stimulation of perforant path synapses in vivo (Rößler et al 2023), but without a loss of lognormality.…”

Neuronal K+-Cl- cotransporter KCC2 as a promising drug target for epilepsy treatment

“…Nevertheless, spatially resolved studies have investigated the changes in KCC2 and NKCC1 induced by lesions, providing a clear understanding of whether precise, anatomically well-defined lesions can elicit spatially restricted, layer-specific alterations in the expression of these proteins. Turco et al recently demonstrated that entorhinal denervation induces specific changes in the expression of KCC2 and NKCC1 in the molecular layers of the dentate gyrus, specifically the oml/mml layers [ 69 ]. Through the use of laser microdissection, microarray analysis, and RT-qPCR, they identified a decrease in KCC2 mRNA and reduced levels of KCC2 protein in denervated granule cell dendrites.…”