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Widely used for mapping afferent activated brain areas  in vivo , the label-free intrinsic optical signal (IOS) is mainly ascribed to blood volume changes subsequent to glial glutamate uptake. By contrast, IOS imaged  in vitro  is generally attributed to neuronal and glial cell swelling, however the relative contribution of different cell types and molecular players remained largely unknown. We characterized IOS to Schaffer collateral stimulation in the rat hippocampal slice using a 464-element photodiode-array device that enables IOS monitoring at 0.6 ms time-resolution in combination with simultaneous field potential recordings. We used brief half-maximal stimuli by applying a medium intensity 50 Volt-stimulus train within 50 ms (20 Hz). IOS was primarily observed in the  str. pyramidale  and proximal region of the  str. radiatum  of the hippocampus. It was eliminated by tetrodotoxin blockade of voltage-gated Na +  channels and was significantly enhanced by suppressing inhibitory signaling with gamma-aminobutyric acid(A) receptor antagonist picrotoxin. We found that IOS was predominantly initiated by postsynaptic Glu receptor activation and progressed by the activation of astroglial Glu transporters and Mg 2+ -independent astroglial N-methyl-D-aspartate receptors. Under control conditions, role for neuronal K + /Cl −  cotransporter KCC2, but not for glial Na + /K + /Cl −  cotransporter NKCC1 was observed. Slight enhancement and inhibition of IOS through non-specific Cl −  and volume-regulated anion channels, respectively, were also depicted. High-frequency IOS imaging, evoked by brief afferent stimulation in brain slices provide a new paradigm for studying mechanisms underlying IOS genesis. Major players disclosed this way imply that spatiotemporal IOS reflects glutamat ergic  neuronal activation and astroglial response, as observed within the hippocampus. Our model may help to better interpret  in vivo  IOS and support diagnosis in the future.

Neuronal and Astroglial Correlates Underlying Spatiotemporal Intrinsic Optical Signal in the Rat Hippocampal Slice