The widespread use of Glutamate (Glu) as the major excitatory neurotransmitter (NT) in the mammalian brain is both critical for normal physiology and a source of predicaments: a) As seen in stroke and a range of neurodegenerative diseases, any disruption in Glu clearance causes its accumulation, leading to over- excitation of postsynaptic cells and excitotoxic neurodegeneration. b) The use of the same NT in so many adjacent synapses can cause signaling to ?bleed over? between neuronal circuits, and the loss of processing fidelity. An idealized view of the brain describes synapses as well insulated from each other, enveloped by glia that expresses high levels of Glu clearing transporters (GluTs). However, a more realistic examination reveals that some particularly-important brain areas (e.g., hippocampus) show severely deficient glial isolation, with estimated 2/3 of released Glu seeping out of the original synapse. How sufficient Glu clearance is achieved in glia-deficient brain areas remains unclear. To overcome the limitation of current techniques we will study Glu clearance in the glia-deficient synaptic hub of the C. elegans nerve ring. We are aided by the availability of information on the precise identification of individual neurons, the exact location of their synapses, the circuits that they participate in, and the sensory inputs and behavioral outputs of these circuits. Together with animal transparency and the wide availability of optogenetic tools, this is an ideal system to study Glu clearance without perturbing interstitial fluids. In our recent studies we have discovered that specific synapses fall into watershed territories of Glu clearance, and that synapses might be affected by the agitation of body fluids. We therefore propose a novel concept, where Glu clearance in a glia-deficient synaptic hub can be robust enough to allow functional synaptic isolation. Such robust clearance depends on division of labor between proximal and distal GluTs, and is facilitated by agitation and perfusion of interstitial fluids. To provide further support to this model we will use genetically-encoded florescent Ca2+ reporters (GCaMP) to follow synaptic activity and assign additional synapses and circuits to GluT drainage territories; we will stimulate one circuit and record responses from an adjacent one to detect spillover; we will use genetically-encoded fluorescent detectors to study the flow of Glu in the interstitial space; we will study the effect of paralysis on neuronal responses and Glu flow; We will correlated the differences between the structure of proximal and distal GluTs to potential differences transport in affinity and capacity. These studies will provide novel insights to mechanisms of robust Glu clearance in the absence of glia, and highlight the significance of agitation of interstitial fluids in synaptic areas that are deficient in glia insulation, a feature shared between nematodes and some areas of the mammalian brain. These insights will aid in the design of future therapeutic interventions to prevent excitotoxicity (seen in stroke and a range of neurodegenerative diseases), and highlight the significance of vascular pulsatility in CNS physiology.
Novel strategies for glutamate clearance in a glia-deprived synaptic hub: Lessons from C. elegans Narrative Clearance of the neurotransmitter glutamate is a fundamental issue in the physiology of the nervous system, while glutamate accumulation causes neurodegeneration in prevalent diseases like stroke. An idealized view of glutamate clearance by glia cells falls short of providing a satisfactory explanation of its removal strategies in a number of particularly important brain areas, where glia cells do not completely insulate synaptic connections. We propose a novel mechanism for glutamate clearance (by division to proximal and distal drainage and by extra-cellular fluid perfusion), and use a particularly powerful animal model (where we image neuronal activity at identified synapses in the intact animal), to reveal basic principles that will help us develop strategies to combat improper glutamate clearance in critical neurological diseases.
