TREK-1 and Best1 Channels Mediate Fast and Slow Glutamate Release in Astrocytes upon GPCR Activation

Abstract

Astrocytes release glutamate upon activation of various GPCRs to exert important roles in synaptic functions. However, the molecular mechanism of release has been controversial. Here, we report two kinetically distinct modes of nonvesicular, channel-mediated glutamate release. The fast mode requires activation of G(αi), dissociation of G(βγ), and subsequent opening of glutamate-permeable, two-pore domain potassium channel TREK-1 through direct interaction between G(βγ) and N terminus of TREK-1. The slow mode is Ca(2+) dependent and requires G(αq) activation and opening of glutamate-permeable, Ca(2+)-activated anion channel Best1. Ultrastructural analyses demonstrate that TREK-1 is preferentially localized at cell body and processes, whereas Best1 is mostly found in microdomains of astrocytes near synapses. Diffusion modeling predicts that the fast mode can target neuronal mGluR with peak glutamate concentration of 100 μM, whereas slow mode targets neuronal NMDA receptors at around 1 μM. Our results reveal two distinct sources of astrocytic glutamate that can differentially influence neighboring neurons.

A disulphide-linked heterodimer of TWIK-1 and TREK-1 mediates passive conductance in astrocytes

“…The specificity of the TREK-1 shRNA has been documented previously 20 . Treatment with TREK-1 shRNA caused a dramatic reduction in cell surface expression of the endogenous TWIK-1 channel proteins, measured by surface biotinylation, in comparison to control Sc shRNA treatment, without affecting the total TWIK-1 protein levels ( Fig.…”

“…2c) and TREK-1 (ref. 20) was extensively documented. Co-IP experiments also demonstrated a strong association between native TWIK-1 and TREK-1 in astrocytes (Fig.…”

“…Astrocytes rapidly release glutamate via a TREK-1-containing K2P channel upon the activation of GPCRs such as proteaseactivated receptor-1 (PAR1) 20 . This glutamate release is initiated when the C terminus of the G g subunit, GNG4, binds to the N terminus of TREK-1 to open the glutamate-permeable, TREK-1-containing K2P channel 20 . Of interest to note, is that data from Y2H screening using cytosolic N-and C termini of TWIK-1 showed that GNG4 is also bound to both cytosolic parts of this channel.…”

“…In addition, we have recently shown that TREK-1 is also expressed in astrocytes within the brain and mediates rapid release of the gliotransmitter glutamate upon activation of GPCRs such as the cannabinoid receptor 1 (CB1), due to direct binding of G bg to the N terminus of TREK-1 (ref. 20). Astrocytes have been reported to express TWIK-1 along with TREK-1 (ref.…”

Astrocytes in endocannabinoid signalling

“…Indeed, astrocytes respond with intracellular calcium elevations to neurotransmitters released from synaptic terminals [14 -18]. These calcium elevations have been shown to stimulate the release of gliotransmitters through different mechanisms [19][20][21][22][23][24], which still remain debated (for recent reviews, see [25][26][27]). …”