Jin-an Li

Expression of vesicular glutamate transporters, VGluT1 and VGluT2, in axon terminals of nociceptive primary afferent fibers in the superficial layers of the medullary and spinal dorsal horns of the rat.

We examined immunohistochemically whether the vesicular glutamate transporters (VGluTs), VGluT1 and VGluT2, might be expressed in synaptic terminals of nociceptive primary afferent fibers within laminae I and II of the medullary and spinal dorsal horns of the rat. VGluT1 immunoreactivity (IR) was intense in the inner part of lamina II but weak in lamina I and the outer part of lamina II. VGluT2‐IR was most intense in lamina I and the outer part of lamina II. Expression of VGluTs in synaptic terminals was confirmed by dual immunofluorescence histochemistry for VGluTs and synaptophysin. Expression of VGluTs in axon terminals of primary afferent fibers terminating in laminae I and II was also confirmed immunohistochemically after unilateral dorsal rhizotomy. The dual immunofluorescence histochemistry indicated expression of VGluTs in substance P (SP)‐containing axon terminals in lamina I and the outer part of lamina II. Electron microscopy confirmed the coexpression of VGluTs and SP in axon terminals within laminae I and II; VGluTs was associated with round synaptic vesicles at the asymmetric synapses. It was further observed that isolectin IB4, a marker for unmyelinated axons, often bound with VGluT2‐immunopositive structures but rarely with VGluT1‐immunopositive structures in lamina II. Thus, the results indicated in laminae I and II of the medullary and spinal dorsal horns that both VGluT1 and VGluT2 were expressed in axon terminals of primary afferent fibers, including SP‐containing nociceptive fibers and that VGluT in unmyelinated primary afferent fibers terminating in lamina II was primarily VGluT2. J. Comp. Neurol. 457:236–249, 2003.

Immunocytochemical localization of μ-opioid receptor in primary afferent neurons containing substance P or calcitonin gene-related peptide. A light and electron microscope study in the rat

Co-expression of mu-opioid receptor-like immunoreactivity (MOR-LI) with substance P (SP)- or calcitonin gene-related (CGRP)-LI was observed in rat trigeminal and dorsal root ganglion neurons. In particular, MOR-LI was found in axon terminals with SP- or CGRP-LI in laminae I and II of the medullary and spinal dorsal horns. MOR may be implicated in modulation of release of SP and CGRP from primary sensory afferents.

Vesicular Glutamate Transporters, VGluT1 and VGluT2, in the Trigeminal Ganglion Neurons of the Rat, with Special Reference to Coexpression

Vesicular glutamate transporters are responsible for glutamate transport into synaptic vesicles. In the present study, we examined immunohistochemically the expression of vesicular glutamate transporters, VGluT1 and VGluT2, in trigeminal ganglion neurons of the rat. Immunohistochemistry for VGluT1 and VGluT2 indicated that more than 80% of trigeminal ganglion neurons express VGluT1 and/or VGluT2 in their cell bodies. It also indicated that large and small trigeminal ganglion neurons express VGluT2 more frequently than VGluT1. Dual immunofluorescence histochemistry for VGluT1 and VGluT2 indicated that trigeminal ganglion neurons express VGluT2 more frequently than VGluT1 and that more than 80% of VGluT‐expressing trigeminal ganglion neurons express VGluT1 and VGluT2. Many axon terminals in the superficial layers of the medullary dorsal horn also showed VGluT1 and VGluT2 immunoreactivities. Some of these axon terminals were confirmed to form the central core of the synaptic glomerulus. These results indicated that VGluT1 and VGluT2 are coexpressed in the cell bodies and axon terminals in most trigeminal ganglion neurons. J. Comp. Neurol. 463:212–220, 2003.

Distribution of Trigeminohypothalamic and Spinohypothalamic Tract Neurons Displaying Substance P Receptor-Like Immunoreactivity in the Rat

Primary afferent neurons containing substance P (SP) are apparently implicated in the transmission of noxious information from the periphery to the central nervous system, and SP released from primary afferent neurons acts on second‐order neurons with the SP receptor (SPR). In the rat, nociceptive information reached the hypothalamus not only through indirect pathways but also directly through trigeminohypothalamic and spinohypothalamic pathways. Thus, in the present study, the distribution pattern of trigeminohypothalamic and spinohypothalamic tract neurons showing SPR‐like immunoreactivity (SPR‐LI) was examined in the rat by a retrograde tract‐tracing method combined with immunofluorescence histochemistry for SPR.

Substance P receptor (NK1)-immunoreactive neurons projecting to the periaqueductal gray: distribution in the spinal trigeminal nucleus and the spinal cord of the rat

Substance P receptor (SPR)-immunoreactive neurons projecting to the periaqueductal gray (PAG) were examined in the rat spinal trigeminal nucleus and spinal cord by a retrograde tracing method combined with immunofluorescence histochemistry. After injection of Fluoro-gold (FG) into the PAG, SPR-immunoreactive neurons labeled with FG were observed mainly in the lateral spinal nucleus and lamina I of the medullary and spinal dorsal horns and additionally in laminae V and X of the spinal cord.

Distribution of trigeminothalamic and spinothalamic tract-neurons showing substance P receptor-like immunoreactivity in the rat.

Trigeminothalamic and spinothalamic-tract neurons provided with substance P receptor (SPR) were examined in the rat by SPR immunofluorescence histochemistry combined with Fluoro-Gold (FG) fluorescent retrograde labeling. After FG injection in the thalamic regions, FG-labeled cells with SPR-like immunoreactivity were seen mainly in laminae I and III of the medullary and spinal dorsal horns and lateral spinal nucleus. In these regions, about one-fourth to one-third of FG-labeled cells showed SPR-like immunoreactivity.

Immunohistochemical localization of a metabotropic glutamate receptor, mGluR7, in ganglion neurons of the rat; with special reference to the presence in glutamatergic ganglion neurons

Immunoreactivity for the metabotropic glutamate receptor 7 (mGluR7) and that for phosphate-activated glutaminase (PAG) were examined in the trigeminal (TG), dorsal root (DRG), nodose (NG), superior cervical, celiac, and pelvic ganglia of the rat. Virtually all neuronal cell bodies showed mGluR7-like immunoreactivity (mGluR7-LI) in these ganglia. On the other hand, PAG-like immunoreactivity (PAG) was seen in almost all neuronal cell bodies in the TG, DRG and NG, but not in the other ganglia. Co-existence of mGluR7- and PAG-LI in the TG, DRG and NG was confirmed by a double-immunofluorescence immunohistochemical method. The results indicate that virtually all sensory ganglion neurons are glutamatergic and equipped with mGluR7.

Effects of peripheral nerve ligation on expression of μ-opioid receptor in sensory ganglion neurons: an immunohistochemical study in dorsal root and nodose ganglion neurons of the rat

The present study was attempted to examine if mu-opioid receptor (MOR) might be transported by axonal flow peripherally through peripheral axons of somatic sensory ganglion neurons. After unilateral ligation of the sciatic nerve or the vagus nerve distal to the dorsal root ganglion (DRG) or nodose ganglion (NG), MOR-like immunoreactivity (MOR-LI) of neuronal cell bodies in the DRG of the fourth and fifth lumbar nerves, NG, ambiguus nucleus (Amb) and dorsal motor nucleus of the vagus nerve (DMV) on the side of the ligation was apparently reduced within 1 week after the nerve ligation. However, within 24 h after the nerve ligation, a transient enhancement of MOR-LI was observed in cell bodies of DRG neurons, sciatic nerve stump proximal to the ligature, and cell bodies of NG neurons on the side of the ligation; such a transient enhancement of MOR-LI was not detected in the Amb and DMV. The results suggest that MOR undergoes centrifugal axonal flow in peripheral axons of somatic and visceral sensory ganglion neurons, and that MOR synthesis in sensory ganglion neurons is vulnerable to damage of the peripheral axons.

The relationship between neurokinin-1 receptor and substance P in the medullary dorsal horn A light and electron microscopic immunohistochemical study in the rat

The synaptic relationship between substance P (SP) and its receptor, i.e. neurokinin-1 receptor (NK1R), was examined in the superficial laminae of the caudal subnucleus of the spinal trigeminal nucleus (medullary dorsal horn; MDH) of the rat. For confocal laser-scanning microscopy, double-immunofluorescence histochemistry for NK1 and SP was performed. In electron microscopic double-immunolabeling study, immunoreactivity for NK1R was detected with the silver-intensified gold method, while immunoreactivity for SP was detected with peroxidase immunohistochemistry. SP-immunoreactive axon terminals were observed to be in synaptic (mostly asymmetric) contact with NK1R-immunoreactive neuronal profiles in lamina I and lamina IIo. Although some SP-immunoreactive axon terminals were in synaptic contact with NK1R-immunoreactive sites of plasma membranes, NK1R-immunoreactivity was observed at both synaptic and non-synaptic sites of plasma membrane. Thus, SP released from axon terminals might not only act on NK1Rs facing the SP-containing axon terminals, but also diffuse in the extracellular fluid for distances larger than the synaptic cleft to act on NK1Rs at some distances from the synaptic sites.

Association of serotonin-like immunoreactive axons with nociceptive projection neurons in the caudal spinal trigeminal nucleus of the rat.

Serotoninergic projections to the spinal dorsal horn are implicated in the modulation of nociceptive transmission. However, morphological evidence indicating that serotoninergic projection fibers make synapses on nociceptive neurons in the medullary dorsal horn is still meager. Thus, we examined whether axonal varicosities with serotonin (5‐HT)‐like immunoreactivity (5‐HT‐LI) might make synapses on nociceptive projection neurons in the caudal spinal trigeminal nucleus (Vc) of the rat. Projection neurons were retrogradely labeled with tetramethylrhodamine‐dextran amine (TMR‐DA) or wheat germ agglutinin‐horseradish peroxidase (WGA‐HRP) that was injected into the parabrachial or thalamic region. Vc neurons in which c‐fos protein‐like immunoreactivity (Fos‐LI) was induced by subcutaneous injection of formalin into the lip were considered nociceptive. Vc neurons in direct contact with axonal varicosities that bind isolectin I‐B4 were also considered nociceptive.