Introduction
G-protein coupled receptors (GPCRs) represent a class of 7-transmembrane
receptors that are involved in numerous physiological processes and
remain the most extensively targeted protein family by approved drugs .
Recent research has unveiled that the activity of certain GPCRs can be
modulated by the membrane voltage (Vm). In neurons,
where Vm undergoes permanent changes, this emerging
property could significantly impact the functioning of
neurotransmitter-activated GPCRs and their role as modulators of
synaptic transmission. Indeed, Vm has been shown to
positively or negatively regulate the function of several GPCRs, about
thirty out of the thousand members of this big family, including
acetylcholine , purine , opioid , dopamine , and prostanoid receptors .
Although the structural mechanisms that underlie the sensitivity of
GPCRs to Vm are still being elucidated , functional
studies using site-directed mutagenesis of the Vm sensor
suggest that GPCR activity can be affected, leading to impaired
neurotransmitter release or synaptic plasticity and behavior .
Glutamate is the primary excitatory neurotransmitter in the brain that
binds to both ionotropic (AMPA, NMDA, Kainate receptors) and
metabotropic glutamate (mGlu) receptors. The mGlu receptor family,
comprising eight G protein-coupled receptors (mGlu1-8), has been
extensively studied for their modulatory role in synaptic transmission
and plasticity . Preclinical and clinical studies have targeted these
receptors in various neurological disorders, such as Autism, Fragile X
syndrom, Schizophrenia, Parkinson’s, and Alzheimer’s disease . So far,
only one study, in the Xenopus oocytes expression system, suggests a
direct sensitivity of some mGlu receptors to Vm (mGlu1
and 3, ). Of note, membrane potential changes seem to play a synergistic
role in neurons on signaling events mediated by mGlu5 and mGlu7
receptors, but without any evidence of a direct action of
Vm on the receptor itself. Yet, due to their synaptic
location, these receptors are permanently exposed to membrane voltage
fluctuations. Therefore, demonstrating the sensitivity of mGlu receptors
to Vm could provide insights into their role depending
on the state of neuronal activity. In this study, we have selected the
postsynaptic mGlu5 receptor as a model receptor, known for its role as
neuromodulator of synaptic transmission. mGlu5 is also a key trigger in
the induction of synaptic plasticity, working in concert with ionotropic
NMDA receptors through physical and functional crosstalk .
Interestingly, NMDA receptors are a prototype of receptors whose
activity is regulated by the membrane potential. As a detector of
neuronal activity coincidence, NMDA activation is limited to synapses
whose pre- and postsynaptic elements are activated simultaneously .
Thus, a sensitivity of the mGlu5 receptor to Vm could
similarly specify the identity of synapses on which mGlu5 exerts its
functional effects depending on the neuron’s activity history.
In this report, we explore the intricate signaling pathways associated
with the conformational change of postsynaptic mGlu5 receptor induced by
glutamate binding. This conformational change leads to the activation of
Gq/11-type proteins, the activation of the PLC-PKC
pathway, and the subsequent release of Ca2+ from
internal stores through IP3 receptors , ultimately fine-tuning synaptic
transmission by gating channels and ionotropic receptors, including NMDA
, AMPA , and transient receptor potential canonical (TRPC) channels .
Specifically, we investigate the impact of Vm on each of
the canonical signaling events of the mGlu5 receptor using a variety of
biosensors and patch clamp recordings. Our results demonstrate that
Vm modulates mGlu5 receptor activation and its
downstream signaling, with a depolarization of the membrane favoring the
inactive conformation of mGlu5 and leading to a decrease in
mGlu5-induced Gq/11 activation, Ca2+signaling, TRPC6 gating, and NMDA receptor facilitation. Interestingly,
our data reveals that the mGlu5 receptor functions optimally at
potentials close to the resting potential of the cells.