Synaptic connections of the neurokinin 1 receptor-like immunoreactive neurons in the rat medullary dorsal horn

Abstract

The synaptic connections between neurokinin 1 (NK1) receptor-like immunoreactive (LI) neurons and γ-aminobutyric acid (GABA)-, glycine (Gly)-, serotonin (5-HT)- or dopamine-β-hydroxylase (DBH, a specific marker for norepinephrinergic neuronal structures)-LI axon terminals in the rat medullary dorsal horn (MDH) were examined under electron microscope by using a pre-embedding immunohistochemical double-staining technique. NK1 receptor-LI neurons were observed principally in laminae I and III, only a few of them were found in lamina II of the MDH. GABA-, Gly-, 5-HT-, or DBH-LI axon terminals were densely encountered in laminae I and II, and sparsely in lamina III of the MDH. Some of these GABA-, Gly-, 5-HT-, or DBH-LI axon terminals were observed to make principally symmetric synapses with NK1 receptor-LI neuronal cell bodies and dendritic processes in laminae I, II and III of the MDH. The present results suggest that neurons expressing NK1 receptor within the MDH might be modulated by GABAergic and glycinergic inhibitory intrinsic neurons located in the MDH and 5-HT- or norepinephrine (NE)-containing descending fibers originated from structures in the brainstem.

Figure 1. Immunofluorescent double labeling of γ-aminobutyric acid (GABA)-, glycine (Gly)-, serotonin (5-HT)- or dopamine-β-hydroxylase (DBH) with neurokinin 1 (NK1) receptor in the MDH of adult rats.

NK1 receptor immunoreactivity is mainly distributed along the cellular and dendritic membrane of the MDH neurons (green), GABA, Gly, 5-HT or DBH - ir boutons (red) are in close apposition to NK1R-ir somas and dendrite (arrows). Scale bars, 10 µm ( A–D, A′–D′, A″–D″).

Figure 2. The density of the different types of inputs contacted with NK1 receptor-LI neurons ( /mm).

Histogram showing input to NK1 receptor-LI somata and dendrites obtained with the confocal microscope.

Figure 3. Electron micrographs of two adjacent sections, showing a GABA-immunoreactive axon terminal (GABA) visualized by peroxidase immunoreaction products that makes symmetric synapses with a NK1R-immunopositive dendrite (NK1R) revealed by silver grains in the MDH.

Arrowheads indicate symmetric post-synaptic dense substance (PSD).

Figure 4. Electron micrographs showing the Gly-immunoreactive axon terminals (Gly) visualized by peroxidase immunoreaction products making symmetric synapses with NK1R-immunopositive cell body (NK1R; A) or dendrite (NK1R; B and C) revealed by silver grains in the MDH, respectively.

Arrowheads indicate symmetric post-synaptic dense substance (PSD).

Figure 5. Electron micrographs showing a serotonin -immunoreactive axon terminal (5-HT; A) and a dopamine-β-hydroxylase-immunopositive axon terminal (DBH; B) visualized by peroxidase immunoreaction products making symmetric synapses with two NK1R-immunopositive dendrites (NK1R) revealed by silver grains in the MDH, respectively.

Arrowheads indicate symmetric post-synaptic dense substance (PSD).

Figure 6. Electron micrographs showing a serotonin -immunoreactive axon terminal (5-HT; A) and a dopamine-β-hydroxylase-immunopositive axon terminal (DBH; B) visualized by peroxidase immunoreaction products making asymmetric synapses with two NK1R-immunopositive dendrites (NK1R) revealed by silver grains in the MDH, respectively.

Arrowheads indicate synaptic active zones.