Selective astrocytic gap junctional trafficking of molecules involved in the glycolytic pathway: impact on cellular brain imaging.
To assess the specificity of metabolite trafficking among gap junction-coupled astrocytes, we developed novel, real-time, single-cell enzymatic fluorescence assays to assay cell-to-cell transfer of unlabeled glycolytic intermediates and report (i) highly restricted transfer of glucose-6-phosphate (P) and two analogs, deoxyglucose (DG)-6-P, and 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-DG-6-P, compared with DG and 2- and 6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-DG, (ii) extensive junctional diffusion of glyceraldehyde-3-P, NADH, and NADPH plus three anionic fluorescent dyes used as internal standards for transfer assays, and (iii) stimulation of gap junctional communication by increased intracellular Na(+) that also evokes metabolic responses in nearby coupled astrocytes. Thus, dye transfer does not predict gap junctional permeability of endogenous metabolites. Intracellular retention of flux-regulating compounds (e.g. glucose-6-P) may be necessary for local metabolic control, whereas 'syncytial sharing' may dissipate the work load on peri-synaptic astrocytes. Imaging of brain functional activity depends on local accumulation of exogenous or endogenous signals, and DG-6-P is trapped in the cell where it is phosphorylated, whereas rapid dispersal of cytoplasmic NAD(P)H and labeled glucose metabolites throughout the astrocytic syncytium can interfere with cellular assessment of neuron-astrocyte relationships in autoradiographic, fluorescence microscopic, and magnetic resonance spectroscopic studies.
Gandhi
Gautam K
GK
Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
Cruz
Nancy F
NF
Ball
Kelly K
KK
Theus
Sue A
SA
Dienel
Gerald A
GA
D000818:Animals; D001253:Astrocytes / Q000166:cytology* / Q000378:metabolism*; D001345:Autoradiography; D001921:Brain / Q000166:cytology* / Q000378:metabolism*; D002478:Cells, Cultured; D017629:Gap Junctions / Q000378:metabolism*; D006019:Glycolysis / Q000502:physiology*; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D008856:Microscopy, Fluorescence; D021381:Protein Transport / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar