Tracking glutamate-receptor dynamics in the mouse somatosensory cortex by in vivo two-photon photo-bleaching and imaging

We are using in vivo 2-photon photo-bleaching recovery to study the dynamics of receptor movement.

Fast excitatory transmission in the mammalian central nervous system is mainly mediated by α-amino-3-hydroxy-5 methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptors (AMPA receptors or GluRs). Movement of these receptors in and out of the synapse appears to be involved in the modulation of synaptic strength and thus in the plasticity of neuronal circuits.

Our knowledge on glutamate receptor trafficking mainly comes from studies in dissociated hippocampal neurons and acute brain slices. Most of the studies have been conducted with stimulation conditions known to long-term potentiation, a form of plasticity strongly implicated in learning and memory functions but little is known about receptor trafficking in a living animals. For tracking glutamate receptor dynamics in vivo, we are using transgenic mice that express GFP-tagged GluR-A receptor subunits in the forebrain. We are able to track receptor dynamics on fast (a few seconds) and slow (a few hours) time scales at different cortical depths of the mouse somatosensory cortex (Hasan et al., Society for Neuroscience, Program # 50.3, 2004 and Meyer zum Alten Borgloh et al., Society for Neuroscience, Program # 827.13, 2007).

To selectively visualize receptor inserted into the plasma membrane, we have also generated recombinant adeno-associated viruses to express pHluorin-tagged AMPA receptors, which are non-fluorescent as long as they are inside high acidity intracellular compartments. Bleach-recovery data from neurons infected with those viruses will allow us to distinguish whether receptor insertion into synapses is via membrane fusion or from insertion via lateral diffusion within the plasma membrane.

Future directions

The next major goal is to follow over time dynamic changes in GluR-A redistribution in neural networks in the same animals before and after sensory experience is altered. With sparse labeling of neuron with AAVs, it should be possible to study with high spatial and temporal resolution the rate of receptor mobility and its re-distribution in single compartments (dendrites and spines). With two-photon photo-bleaching recovery method, it should also be possible to study other candidate "synaptic plasticity molecules" involved in organizing the postsynapitc apparatus. These studies are likely to help in elucidating the crucial role of neuronal signaling systems in map plasticity.