Abstract
Ionotropic glutamate receptors are ligand-gated ion channels that mediate excitatory synaptic transmission in the central nervous system. Desensitization of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid subtype after glutamate binding appears critical for brain function and involves rearrangement of the ligand binding domains (LBDs). Recently, several full-length structures of ionotropic glutamate receptors in putative desensitized states were published. These structures indicate movements of the LBDs that might be trapped by cysteine cross-links and metal bridges. We found that cysteine mutants at the interface between subunits A and C and lateral zinc bridges (between subunits C and D or A and B) can trap freely desensitizing receptors in a spectrum of states with different stabilities. Consistent with a close approach of subunits during desensitization processes, the introduction of bulky amino acids at the A-C interface produced a receptor with slow recovery from desensitization. Further, in wild-type GluA2 receptors, we detected the population of a stable desensitized state with a lifetime around 1 s. Using mutations that progressively stabilize deep desensitized states (E713T and Y768R), we were able to selectively protect receptors from cross-links at both the diagonal and lateral interfaces. Ultrafast perfusion enabled us to perform chemical modification in less than 10 ms, reporting movements associated to desensitization on this timescale within LBD dimers in resting receptors. These observations suggest that small disruptions of quaternary structure are sufficient for fast desensitization and that substantial rearrangements likely correspond to stable desensitized states that are adopted relatively slowly on a timescale much longer than physiological receptor activation.
Highlights
Glutamate receptor ion channels mediate most of the fast excitatory synaptic transmission in the vertebrate central nervous system [1]
Placing the A665C mutations in subunits A and C of this structure allows the SG groups to be only 5 Adistant, within striking distance of forming a disulfide bridge, suggesting the TARP-bound desensitized state is quite compact; this concept was discussed at length in our previous work [21]
Because the duration of these measurements was long and the rundown of the current was often substantial, only measurements from four patches could be completed before the patch was lost and had sufficiently good quality for the whole set of records, namely 10–90% rise times of the glutamate response
Summary
Glutamate receptor ion channels mediate most of the fast excitatory synaptic transmission in the vertebrate central nervous system [1]. Glutamate binding initiates the opening of an integral ion pore, permitting cations to flow into the postsynaptic cell. 206 Biophysical Journal 119, 206–218, July 7, 2020 rapidly and profoundly in response to the sustained presence of glutamate for more than $25 ms [2,3]. The number of receptors available to respond to glutamate is reduced during a phase of recovery from desensitization, which in turn can determine the amplitude of postsynaptic responses [4,5]. The timescale of recovery from desensitization, being for AMPA receptors in the order of tens to hundreds of milliseconds, is pertinent during high-frequency release of glutamate (above 10 Hz). Steady-state desensitization may offer protection during pathological glutamate insults that lead to brain damage [6] and during development [7].
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
