Nerve cells in the brain communicate by releasing neurotransmitters, chemicals such as glutamate. The electrical firing of one cell leads to a tiny amount of glutamate being released in a brief pulse. This glutamate is picked up by sensors, called receptors, embedded in the membrane walls of neighbouring cells. Generally, these receptors respond by briefly changing their shape, and opening a tiny pore in the membrane. Electrical current flows through this pore into the receiving cell. This electrical current can excite the cell to fire its own nerve impulses, passing on the signal. Nerve cells can fire rapidly, often tens or hundreds of times in a second, and so, in order to pass the message reliably, some receptors must work even faster. They receive the glutamate signal, activate and then release the glutamate, ready for another cycle, in a fraction of a second. One kind of glutamate receptor that behaves like this turns out to be important for all kinds of brain functions, such as hearing sounds or recalling memories. Other types of glutamate receptor have slow activations (lasting up to a second) and these convey an entirely different message, which can make the target cell increase the strength of the specific connection. This is one way that the brain stores memories. We follow the notion that by understanding the receptors, we can understand the properties of the connections between nerve cells, and thus help to unravel how the brain processes and stores information. The receptors themselves are also fascinating microscopic machines that, historically, have been full of interesting surprises.
One avenue of research that we have focused on is how different members of the glutamate receptor family, which derive from very similar genes, are tuned to respond to glutamate in such different ways. AMPA and kainate-type glutamate receptors contain a hotspot which controls both their gating and their ability to follow fast signalling in a highly coupled fashion (Carbone and Plested, 2012). This hotspot lies between the binding pocket for glutamate and the gating apparatus of the membrane ion channel. This coupling was rather unexpected at the molecular level, but it makes sense in terms of signalling in the brain. In AMPA receptors, both effects serve to make synaptic transmission more precise at high frequencies. These properties are important at synapses in the auditory pathway, where synapses can signal in the kilohertz range (Taschenberger and von Gersdorff, 2000). There, AMPA receptor subunit composition changes during development - at the onset of hearing, faster AMPA receptors are recruited (Joshi et al, 2004). More recently, we have extended this analysis to complexes of glutamate receptor subunits with auxiliary proteins (Carbone and Plested 2016). We developed a positive feedback mechanism that we call superactivation (some people call it resensitization), including the molecular details (Riva et al, 2017). This research has led us to investigate whether slow AMPA receptor signalling is present in the brain. We found it to be surprisingly prevalent in the hippocampus (Pampaloni et al, 2021). Therefore we have come full circle. We started by asking how AMPA receptors are so fast, compared to other types of glutamate receptors. Our current core question is how, and why, are AMPA receptors also able to act so slowly.
In recent years, we have become more involved in computational methods. A good example is the simulations from Albert Lau that predicted the surface diffusion of glutamate over the binding domain of the receptor (Yu et al, 2018). We participate actively in the DFG Research Group DynIon to pursue molecular dynamics simulations with several other groups, including a very close collaboration with the group of Han Sun. Using these techniques, we showed the permeation properties of monovalent cations in the AMPA receptor (Biedermann et al 2021).
Another technique that we have employed is to directly crosslink subunits, in order to detect receptor gating motions (Das et al, 2010, Lau et al, 2013, Baranovic et al, 2016, Salazar et al, 2017, Baranovic and Plested, 2018). By crosslinking with either disulphide bonds or engineered metal bridges during receptor gating, we could relate activation states to domain geometry. These techniques is particularly powerful when combined with the high-resolution maps of glutamate receptor structure from X-Ray crystallographic experiments.
In recent years we have become increasingly interested in optical approaches for controlling and measuring ion channel activity. We used unnatural amino acids that are sensitive to UV light to crosslink subunits and inactivate the AMPA receptor (Klippenstein et al, 2014, Poulsen et al, 2019). These crosslinking methods, as well as our techniques for cysteine crosslinking are collected in a Methods in Enzymology article (Plested and Poulsen, 2021). By fusing fluorescent proteins to receptor domains, we could produce a glutamate receptor that reports its activity by FRET (Zachariassen et al, 2016). We also collaborated with Tobias Stauber to measure the activity of volume-regulated ion channels (VRACs) using electrophysiology and fluorescence at the same time (König et al, 2019).
Joshi, I., Shokralla, S., Titis, P., and Wang, L.Y. (2004), JNeurosci
Taschenberger, H. and Gersdorff, H.V. (2000), JNeurosci