The contribution of TWIK-1 channels to astrocyte K+ current is limited by retention in intracellular compartments

Abstract

TWIK-1 two-pore domain K+ channels are expressed abundantly in astrocytes. In the present study, we examined the extent to which TWIK-1 contributes to the linear current-voltage (I–V) relationship (passive) K+ membrane conductance, a dominant electrophysiological feature of mature hippocampal astrocytes. Astrocytes from TWIK-1 knockout mice have a more negative resting potential than those from wild type animals and a reduction in both inward rectification and Cs+ permeability. Nevertheless, the overall whole-cell passive conductance is not altered significantly in TWIK-1 knockout astrocytes. The expression of Kir4.1 and TREK-1, two other major astrocytic K+ channels, or of other two-pore K+ channels is not altered in TWIK-1 knockout mice, suggesting that the mild effect of TWIK-1 knockout does not result from compensation by these channels. Fractionation experiments showed that TWIK-1 is primarily localized in intracellular cytoplasmic fractions (55%) and mildly hydrophobic internal compartment fractions (41%), with only 5% in fractions containing plasma membranes. Our study revealed that TWIK-1 proteins are mainly located in the intracellular compartments of hippocampal astrocyte under physiological condition, therefore a minimal contribution of TWIK-1 channels to whole-cell currents is likely attributable to a relatively low level presence of channels in the plasma membrane.

Electrophysiological properties of NG2 + cells: Matching physiological studies with gene expression profiles

“…One possible explanation for the discrepancy between mRNA expression data and physiological data is that channels may not be heavily trafficked to the cell membrane, despite high levels of transcription and/or translation. This was found to be the case for TWIK1 in passive astrocytes, where the channel is largely sequestered in the intracellular compartment (Feliciangeli et al, 2010; Wang et al, 2013). Better pharmacological and genetic tools will enable a more thorough investigation of the contribution of K 2P channels to the passive conductance of NG2 + cells.…”

mGluR3 Activation Recruits Cytoplasmic TWIK-1 Channels to Membrane that Enhances Ammonium Uptake in Hippocampal Astrocytes

“…TWIK-1 knockout mice were created from mice of C57BL/6J genetic background, where the exon 2 gene was deleted [3, 8]. All the experiments were performed from the littermates that contained wild-type (WT), TWIK-1 heterozygous (TWIK-1 +/− ), and knockout (TWIK-1 −/− ) male and female mice at postnatal day (P) 21–28 unless indicated otherwise.…”

“…All the experiments were performed from the littermates that contained wild-type (WT), TWIK-1 heterozygous (TWIK-1 +/− ), and knockout (TWIK-1 −/− ) male and female mice at postnatal day (P) 21–28 unless indicated otherwise. The genotypes of the used mice were individually confirmed by a polymerase chain reaction (PCR) genotype procedure we have reported [3]. …”

“…However, astrocytes in TWIK-1 gene knockout mice show only a mildly altered membrane potential and K + conductance attributable to retention of large amount of channels in the intracellular compartments [3]. Interestingly, the membrane TWIK-1 exhibits unique ion permeability.…”

“…Interestingly, the membrane TWIK-1 exhibits unique ion permeability. First, the membrane TWIK-1 is a nonselective monovalent cation channel with high permeability to Na + in astrocytes, kidney tubular, and pancreatic β cells [3–5]. Second, the TWIK-1 preferentially conducts inward NH 4 + currents that is 40-fold higher than the K + currents in heterologous systems [6]; this implies that TWIK-1 may function as a highly efficient NH 4 + uptake channel in astrocytes.…”