Transcranial electrical stimulation of cortico-cortical connections in anesthetized mice.

We developed a technique of transcranial electrical stimulation (TES) to investigate cortico-cortical connections in mice. After the skull was shaved with the blade of a dental bar, a blunt tip of a needle was gently pushed onto the thinned skull. The skull was deformed by the force, and the subarachnoid space between the skull and the cortex was minimized around the needle tip. Under these conditions, stimulus currents applied to the needle directly flowed into the cortex through the thinned skull. Cortico-cortical functional connections stimulated by this method were visualized by transcranial flavoprotein fluorescence imaging. The cortical responses evoked by TES exhibited spatial and temporal activity patterns comparable to those elicited by a conventional method, in which an electrode is directly inserted into superficial cortical layers. A comparison of the two methods revealed that TES required a slightly stronger stimulus intensity and preferentially activated superficial layers of the cortex compared with the conventional method. Using the new method, we revealed the presence of reciprocal cortico-cortical functional connections between lateral and medial parts of higher visual cortices in mice. This new method combined with transcranial flavoprotein fluorescence imaging allowed us to activate cortico-cortical pathways arising from the primary sensory areas and investigate sensory information flow in the mouse cerebral cortex.