Spinal cord trunk preparation for analyzing cross-segmental primary afferent signal transmission and modulation

The spinal cord dorsal horn is pivotal for primary afferent signal transmission and modulation. Primary afferent fibers from each dorsal root arrive at the dorsal horn and travel 1-2 segments caudally and rostrally. Usually, in vitro spinal cord slices or in vivo preparations are employed for primary afferent stimulation and patch-clamp recordings to assess input signals. However, the spinal cord slices lose "intact" cross-segmental pathways, and in vivo studies are technically challenging.

Here, we describe the preparation of a spinal cord trunk for analyzing afferent signal cross-segmental transmission in adult rats. By combining patch-clamp recording, Lissauer's tract stimulation, and ambient temperature manipulation, our methods enable accessing primary afferent pathways within several segments.

Our present spinal trunk preparation can be maintained healthy for about 5 h. Lissauer's tract stimulation induced evoked excitatory postsynaptic currents (eEPSCs) recorded in 6-10 mm rostrally in the ipsilateral dorsal horn. The eEPSCs, spontaneous EPSCs (sEPSCs), and neural excitability can be modulated by ambient temperature rise. Neuropharmacological studies can also be conducted on this spinal trunk preparation.

Compared with conventional in vitro spinal cord slices, our present method maintains a relatively intact cross-segment pathway in the dorsal horn; compared with in vivo study, it avoids mechanical vibration and other technical challenges in living animals.

The rodent spinal cord trunk can be maintained for an extended period in a fully submerged chamber; combined with patch clamp recordings, our protocol facilitates the study of primary afferent transmission and modulation in the dorsal horn within adjacent segments.