Projections Of Endomorphinergic Neurons From The Nucleus Tract Solitarii To The Parabrachial Nucleus And The Periaqueductal Gray In The Rat

Endomorphins are newly found members of the endogenous opioid peptide family, including endomorphin-1 (EM1) and endomorphin-2 (EM2), which have high affinity and selectivity for theμ-opioid receptors (MOR). In the central nervous system (CNS), the EM-immunoreactive (EM-IR) neuronal cell bodies present in both hypothalamus and nucleus tractus solitarii (NTS), while EM-IR fibers and terminals are found in many special structures, including the hypothalamus, NTS, parabrachial nucleus (PBN), spinal dorsal horn and medullar dorsal horn.NTS is mainly located in the dorsalmedial part of the medulla, and it is a majar site of primary termination for many visceral afferents. NTS can be divided into different subnuclei that are associated with different functions. It receives visceral afferent from most of organs in the neck, check and abdomen through theⅦ,ⅨandⅩcranial nerves. NTS is regarded as an initial regulatory point of peripheral and central autonomic reflexes. There are many connections between NTS and other nuclei in the CNS, including PBN and PAG.EM-IR neuronal cell bodies are localized principally in both hypothalamus and NTS in the central nervous system. In our previous studies, we had demonstrated that EM-IR neurons reciprocally connections between hypothalamus and NTS, and EM-IR neurons in the hypothalamus projected to PBN and PAG. There are dense EM-IR fibers and terminals in PBN and PAG. Whether EM-IR fibers and terminals in PBN and PAG originate from NTS is still obscure. In the present study, we attempted to prove whether EMergic neurons in NTS sent axons to PBN and PAG.By using tetramethyl rhodamine (TMR) fluorescent retrograde tracing and immunofluorescent histochemical staining methods, the present study investigated the EM-IR neurons projecting from NTS to PBN or PAG.1. After injection of TMR into the right PBN (mainly located in medial parabrachial nucleus and lateral parabrachial nucleus), we could observe neurons labeled retrogradely with TMR in the right NTS, especially in the medial, commissural and dorsolateral subnuclei of the NTS with predominance ipsilateral to the TMR injection. EM1-IR neurons were distributed in the medial, commissural, dorsolateral and gelatinous subnuclei of the NTS at the level of the area postrema, while EM2-IR neurons were mainly found in the medial and dorsolateral subnuclei of the NTS at the same level and just rostral to the level of the area postrema. Some of the EM1- and EM2-IR neuronal cell bodies in the right NTS were labeled retrogradely with TMR injected into the PBN. EM1/TMR or EM2/TMR double-labeled neurons were detected averagely in 9.4% or 5.8% to the total number of TMR-labeled neurons, 9.1% or 6.5% to the total numbers of the EM1- or EM2-IR neurons, respectively. A few EM1/TMR double-labeled neurons were also found in the commissural and gelatinous subnuclei of the NTS. Meanwhile, there was more number of EM1/TMR double-labeled neurons than that of EM2/TMR double-labeled neurons in all of those NTS subnuclei.These results indicate that the EMergic neurons in the NTS sent axons to the PBN, which may play an important role to regulate the visceral nociceptive informations.2. After injection of TMR into the defferent columns of the PAG, what we observed were as follows:(1) For TMR injection into the ventrolateral column of the PAG (vlPAG), the number of neurons retrogradely labeled with TMR in the medial subnucleus of NTS was the most, then in the communication subnucleus of NTS, and they were also found in the dorsalateral and intermedial subnuclei of the NTS. 9.62% EM1-IR (8.6±1.14) and 5.54% EM2-IR (3.8±0.84) neurons in the NTS were labeled with TMR.(2) For TMR injection into the lateral column of the PAG (lPAG), we could find neurons retrogradely labeled with TMR in the communication subnucleus of the NTS, the medial subnucleus of the NTS and the dorsalateral subnucleus of the NTS, especially in the communication subnucleus of the NTS and the medial subnucleus of the NTS. But the number of neurons labeled retrogradely with TMR was less than it for TMR injection into the vlPAG. 6.18% EM1-IR (5.2±0.84) and 4.63% EM2-IR (3.0±0.71) neurons in the NTS were labeled with TMR.3) For TMR injection into the dorsolateral column of the PAG (dlPAG) and the dorsomedial column of the PAG (dmPAG), we couldn't find typical EM/TMR double-labeled neurons, and the number of neurons retrogradely with TMR was far less than it for TMR injection into the vlPAG and lPAG.The present results suggested that neurons projecting from the NTS to the PAG were mainly located in the communication subnucleus of the NTS and the medial subnucleus of the NTS, as well as EM/TMR double-labeled neurons. The number of neurons labeled retrogradely with TMR for TMR injection into the vlPAG was the most of all, the number of neurons labeled retrogradely with TMR for TMR injection into the dmPAG was the least. All these results indicate that the EMergic fibers and terminals in the PAG partly came from the NTS, and it existed a roughly topographical correspondence between the different subnuclei of NTS and different parts of PAG. The NTS-PAG projecting EMergic fibers and terminals might be involved in the modulation of the PAG's functions, especially the visceral informations. All results can provide the morphological evidences and reveal the mechanisms of EMs participating in the antinociception of the central endogenous originated pain control system, which are significant.