Targeting the Cation-Chloride Co-Transporter NKCC1 to Re-Establish GABAergic Inhibition and an Appropriate Excitatory/Inhibitory Balance in Selective Neuronal Circuits: A Novel Approach for the Treatment of Alzheimer's Disease

Abstract

GABA, the main inhibitory neurotransmitter in the adult brain, depolarizes and excites immature neurons because of an initially higher intracellular chloride concentration [Cl^-]i due to the delayed expression of the chloride exporter KCC2 at birth. Depolarization-induced calcium rise via NMDA receptors and voltage-dependent calcium channels is instrumental in shaping neuronal circuits and in controlling the excitatory (E)/inhibitory (I) balance in selective brain areas. An E/I imbalance accounts for cognitive impairment observed in several neuropsychiatric disorders. The aim of this review is to summarize recent data on the mechanisms by which alterations of GABAergic signaling alter the E/I balance in cortical and hippocampal neurons in Alzheimer's disease (AD) and the role of cation-chloride co-transporters in this process. In particular, we discuss the NGF and AD relationship and how mice engineered to express recombinant neutralizing anti-NGF antibodies (AD11 mice), which develop a neurodegenerative pathology reminiscent of that observed in AD patients, exhibit a depolarizing action of GABA due to KCC2 impairment. Treating AD and other forms of dementia with bumetanide, a selective KCC2 antagonist, contributes to re-establishing a proper E/I balance in selective brain areas, leading to amelioration of AD symptoms and the slowing down of disease progression.

Keywords: AD11 transgenic mice; Alzheimer’s disease; KCC2 dysfunction; NGF; bumetanide treatment; cation-chloride co-transporters; depolarizing GABAA-mediated neurotransmission.

Figure 1

Altered chloride homeostasis in AD. In the adult brain (top), the intracellular levels of chloride, are maintained at very low levels by the cation-chloride importer and exporter NKCC1 and KCC2, respectively. GABA binds to its receptor and opens the channel, giving rise to an inward flux of chloride, which inhibits targeted cells by hyperpolarizing the membrane. In this way, it contributes to preserve the appropriate E/I balance in selective neuronal circuits. In AD, the APP-induced downregulation of GABA_B Rs, the age-dependent impact of APP on KCC2, or neutralization of NGF in AD11 mice allows GABA to shift from the hyperpolarizing to the depolarizing direction, as in immature neurons, giving rise to an outwardly directed flux of chloride and E/I imbalance with consequent enhancement of network excitability (down left), which is an effect that can be rescued by blocking NKCC1 with bumetanide (down right).

Figure 2

Depolarizing action of GABA in six-month-old AD11 mice. (A) Cell-attached recordings of single NMDA and GABA_A receptor channels obtained at two different pipette potentials in neurons from WT (green) and AD11 (violet) mice. (B) Summary plot of single NMDA (I_NMDA) and GABA (I_GABA) currents versus pipette potentials (Vp). Single NMDA currents reversed at −75.3 mV and −76.2 mV in WT and AD11 mice, respectively. Assuming a reversal of NMDA currents equal to 0 mV, we estimated a mean resting membrane potential of −75mV and −76 mV, respectively. In contrast, single GABA_A currents reversed at −76 mV and at −56 mV in WT and AD11 mice, respectively. (C) Left: Perforated patch recordings of GPSCs evoked in CA3 principal cells by local stimulation of GABAergic interneurons in the presence of DNQX (20 μM) and D-AP-5 (50 μM) at three different holding potentials in hippocampal slices from WT (green), AD11 (violet), and AD11 mice exposed to bumetanide (yellow). Right: Synaptic currents (I_GPSCs) shown in A are plotted versus membrane potentials (Vm). (D) Left: individual RMPs and E_GPSCs values in hippocampal slices obtained from WT (green), AD11 (violet), and AD11 exposed to bumetanide (yellow). Larger symbols on the left and right refer to mean ± SEM values. Right: plot of the driving force for GABA_A (E_GABA–RMP) in individual experiments from the three different groups of mice. ** p < 0.001 (Modified from [153]).

Figure 3

Interaction network in AD11 mice for NKCC1, KCC2, GABA_A, and GABA_B receptor genes. Interaction network extracted from the STRING database (https://string-db.org/, high interaction confidence level) for the NKCC1, KCC2 cation-chloride co-transporter genes (encircled in red), and the GABA_A, GABA_B receptor channel subunits genes in mouse. The node color corresponds to the Log2-fold change-relative expression ratio in 6-month-old AD11 mouse compared to control mouse in three brain areas (basal forebrain, BFB; cortex, CTX; and hippocampus, HP). The thickness of grey edges between nodes corresponds to the interaction reliability provided by STRING.