Cl− concentration changes and desensitization of GABAA and glycine receptors

Abstract

Desensitization of ligand-gated ion channels plays a critical role for the information transfer between neurons. The current view on γ-aminobutyric acid (GABA)A and glycine receptors includes significant rapid components of desensitization as well as cross-desensitization between the two receptor types. Here, we analyze the mechanism of apparent cross-desensitization between native GABAA and glycine receptors in rat central neurons and quantify to what extent the current decay in the presence of ligand is a result of desensitization versus changes in intracellular Cl− concentration ([Cl−]i). We show that apparent cross-desensitization of currents evoked by GABA and by glycine is caused by changes in [Cl−]i. We also show that changes in [Cl−]i are critical for the decay of current in the presence of either GABA or glycine, whereas changes in conductance often play a minor role only. Thus, the currents decayed significantly quicker than the conductances, which decayed with time constants of several seconds and in some cells did not decay below the value at peak current during 20-s agonist application. By taking the cytosolic volume into account and numerically computing the membrane currents and expected changes in [Cl−]i, we provide a theoretical framework for the observed effects. Modeling diffusional exchange of Cl− between cytosol and patch pipettes, we also show that considerable changes in [Cl−]i may be expected and cause rapidly decaying current components in conventional whole cell or outside-out patch recordings. The findings imply that a reevaluation of the desensitization properties of GABAA and glycine receptors is needed.

Electrophysiological Signature of Homomeric and Heteromeric Glycine Receptor Channels

“…However, these data are not in good agreement with previous studies that reported that splicing of the long insert into GlyR ␣3 mRNA impacts on desensitization kinetics (12, 39 -41). Notably, desensitization of GlyR channels may be highly variable (20,41) and influenced by many parameters, such as receptor density (42), membrane potential (43), intracellular and extracellular pH (44,45), ion concentration (46,47), the membrane lipid composition, and phosphorylation of the intracellular TM3-TM4 domain or the N-glycosylation of the receptor N terminus (20). Technical features, such as slow or fast ligand application or the ligand concentration (16,48), duration of ligand administration, or the recording mode may also account for these differences.…”

Contribution of Resting Conductance, GABAA-Receptor Mediated Miniature Synaptic Currents and Neurosteroid to Chloride Homeostasis in Central Neurons

“…Cells were continuously (between test applications of agonists) perfused with gravity-fed extracellular solution provided by a custom-made pipette positioned 100-200 µm from the studied cell. Computer-controlled exchange to solutions containing agonist via solenoid valves occurred with a time constant of ∼50 ms, as measured by the change in offset potential on changing between high and low K + concentrations in the perfusate (Karlsson et al, 2011). …”

“…Experiments to estimate resting Cl – conductance, which depend on estimates of [Cl – ] i but are compatible with slow Cl – equilibration with the patch pipette, were made using the quicker amphotericin B-perforated patch technique (Rae et al, 1991). Both perforated-patch techniques are compatible with rapid Cl – loading (Karlsson et al, 2011). The majority of experiments were made under voltage-clamp conditions, but a few control experiments were made under current-clamp conditions at zero current.…”

“…To improve the electrical recording conditions and to avoid possible [Cl – ] i redistribution between neuronal compartments, we used acutely isolated neurons from which neurites were largely removed, but which retain attached presynaptic nerve terminals that enable analysis of spontaneous GABA release (Haage et al, 1998). Cells were loaded with Cl – by combining GABA or glycine application with a depolarized voltage and [Cl – ] i was obtained from measurements of E Cl by gramicidin-perforated patch recordings with GABA or glycine application in combination with voltage ramps (Karlsson et al, 2011; Yelhekar et al, 2016). …”

Synaptic Inhibition in Avian Interaural Level Difference Sound Localizing Neurons

“…The superior olivary nucleus (SON) is a potential source of glycinergic input, as it has been shown to project to the LLDp in zebra finch (Wild et al, 2010), and possible SON projections to LLDp neurons in chicken have been reported (Westerberg and Schwarz, 1995). The role of glycine could be to maintain neural inhibition in the circuit if GABA is depleted (Fischl et al, 2014; Nerlich et al, 2014), or it could modulate GABAergic inhibition through cross-suppression due to changes in the Cl − driving force (Grassi, 1992; Karlsson et al, 2011) or other signaling cascades (Li et al, 2003). …”