Alpha2delta-mediated control of neuronal signaling

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PROJECT SUMMARY Intracellular calcium drives neuronal signaling and excitability, and appropriate spatial and temporal control of neuronal calcium signals are required to prevent dysregulated control of brain activity, which can lead to epileptic seizures. One of the primary sources of calcium in neurons is entry via voltage-gated calcium channels on the cell membrane, which is then coupled to various effector mechanisms through co-localization of channels and calcium-responsive proteins within the neuron. Two specific, but distinct, calcium-dependent processes critical for maintaining brain homeostasis involve retrograde endocannabinoid signaling and the action potential afterhyperpolarization. Endocannabinoids are produced postsynaptically by neurons in a calcium-dependent manner, and then diffuse to presynaptic terminals where they bind to CB1 receptors and powerfully inhibit vesicle release at numerous synapses. Separately, voltage-dependent calcium entry activates nearby coupled calcium-dependent potassium channels, mediating the action-potential afterhyperpolarization which accelerates cell repolarization and controls neuronal firing rates. For each of these phenomena, tight functional coupling of calcium entry to these disparate effectors is critical in maintaining brain function. While the specific cellular mechanisms responsible for controlling the appropriate localization of voltage-gated calcium channels remain largely unknown, our preliminary data indicate a critical role for the alpha2delta proteins in the functional coupling of calcium entry to effectors. These auxiliary calcium channel subunits help traffic voltage-gated calcium channels to the neuronal surface membrane, but have otherwise remained enigmatic despite clear association with neurologic diseases in humans and mice. We hypothesize that the alpha2delta proteins are critical mediators of functional coupling between calcium entry and calcium- dependent signaling throughout the brain. We propose to use genetically modified mice and electrophysiological assays to define the roles of alpha2delta isoforms in 1) calcium-dependent retrograde signaling from cerebellar Purkinje cells to their various synaptic inputs, 2) the control of excitability and endocannabinoid signaling in the hippocampus, and 3) the molecular mechanisms underlying these phenomena using molecular replacement strategies. Together, these experiments will lead to a greatly enhanced appreciation of the function of this important class of calcium channel subunits, which we believe can be leveraged to improve our ability to control runaway excitability in the brain in conditions such as epileptic seizures.