A novel mechanism for glial signaling studied by two-photon imaging

The project aimed at elucidating activity-induced signaling cascades in astrocytes, focusing on the roles of aquaporin-4 (AQP4) and its anchoring complex. We found that AQP4 reduced peak extracellular [K+] in stratum radiatum of acute hippocampal slices during 20 Hz stimulation of Schaffer collaterals. Using the genetically encoded fluorescent Ca2+ indicator GCaMP5E we demonstrated that the same stimulation protocol elicited robust Ca2+ signals in all astrocytic compartments, including endfeet ensheathing blood vessels. In a mouse model for mesial temporal lobe epilepsy (MTLE) with hippocampal sclerosis, characterized by loss of perivascular AQP4 and the AQP4 anchoring complex, we found that the endfoot Ca2+ signals in response to Schaffer collateral stimulation were augmented. These findings suggest that pathological Ca2+ signals in glial endfeet may underlie vascular dysregulation and perturbed tissue ion and water homeostasis. This conclusion is supported by an observation in cortical spreading depression, in which a pronounced increase in endfoot Ca2+ level was followed by arteriolar constriction. Taken together, our results have identified Ca2+ signals in glial endfeet as a possible target for future therapies.

A number of neurological diseases for which current therapies are nonexisting or inadequate involve deficiencies in glial function. Examples are stroke, Alzheimers disease and epilepsy. The recent Nobel-recognized discovery of aquaporins - the channels th at mediate rapid and regulated transport of water - opened up a new field in molecular medicine. The applicant pioneered the research on aquaporins in the CNS and disclosed that aquaporin-4 (AQP4), the predominant brain aquaporin, is concentrated in speci alized glial membranes at the brain-blood interface. Despite a decade of intense research the roles of glial water channels are still an enigma. The project utilizes two-photon imaging to unravel novel roles of AQP4 in brain. The new concepts launched are that AQP4 facilitates glial Ca2+ signaling and gliotransmitter release, thus promoting the neuronal hyperexcitability charachteristic of epileptic disorders. To delineate mechanisms underlying glial dysfunction in epilepsy we will combine gene knockout s trategies with electrophysiological recordings and two-photon microscopy. Further, the project will assess the role of AQP4 in cerebral blood flow regulation. The project is multidisciplinary and draws on the expertise of a national and international comp etence network including the EMBL Partnership for Molecular Medicine, an US group that is among the leading groups in the world on in vivo two-photon imaging, and Oslo University Hospital.