β-Endorphin-containing memory-cells and μ-opioid receptors undergo transport to peripheral inflamed tissue

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Abstract

Immunocyte-derived β-endorphin can activate peripheral opioid receptors on sensory neurons to inhibit pain within inflamed tissue. This study examined μ-opioid receptors (MOR) on sensory nerves and β-endorphin (END) in activated/memory CD4+ cells (the predominant population homing to inflamed tissue). We found an upregulation of MOR in dorsal root ganglia, an increased axonal transport of MOR in the sciatic nerve and an accumulation of MOR in peripheral nerve terminals in Freund's adjuvant-induced hindpaw inflammation. A large number of CD4+ cells containing β-endorphin, but very few naive cells (CD45RC+), were observed in inflamed tissue, suggesting that this opioid is mainly present in activated/memory cells (CD4+/CD45RC). Taken together, our results indicate an enhanced transport of both MOR and of the endogenous ligand β-endorphin to injured tissue. This unique simultaneous upregulation of both receptors and ligands may serve to prevent excessive and/or chronic inflammatory pain.

Introduction

Opioid receptors on peripheral sensory neurons are involved in pain control, especially under inflammatory conditions (Stein, 1995). These receptors can be activated by endogenous opioid peptides, which are produced by immunoocytes within peripheral inflamed tissue Stein et al., 1990b, Przewlocki et al., 1992, Cabot et al., 1997, Machelska et al., 1998. A prominent opioid peptide involved in endogenous peripheral opioid analgesia is β-endorphin (END) Stein et al., 1990a, Stein et al., 1990b, Schäfer et al., 1994. END is present in immune cells Przewlocki et al., 1992, Cabot et al., 1997, which secrete the opioid to reduce pain Cabot et al., 1997, Machelska et al., 1998 and possibly inflammation (Barber and Gottschlich, 1992). To date, these immunocytes have not been characterized within the subcutaneous tissue. Typically, activated/memory CD4+ (as opposed to naive) cells are the major population homing to peripheral sites of inflammation (Mackay et al., 1992). In the rat, CD4+ cell subsets are discernible by the monoclonal antibody (mAb) MRC OX22 (anti-CD45RC), which distinguishes resting (naive) CD4 cells (CD45RC+) from activated/memory cells (CD45RC) Bell et al., 1995, Lawrence et al., 1996. In the present study, we hypothesized that END is contained in resident CD4+ cells and not in CD45RC cells within inflamed subcutaneous tissue. Since END binds to the μ-opioid receptor (MOR) and this receptor has not yet been identified by specific antisera on peripheral sensory nerve terminals, we also examined the expression and the axonal transport of MOR in dorsal root ganglia (DRG), in the sciatic nerve and on peripheral nerve terminals in normal and inflamed tissue.

Section snippets

Animals

Adult male Wistar rats weighing 200–250 g were briefly anesthetized with halothane (Halocarbon Laboratories), and 150 μl of complete Freund's adjuvant (CFA) (Calbiochem) was injected into the plantar surface (i.pl.) of the right hindpaw. Control animals were anesthetized and injected with saline. Animals were kept individually in cages at an ambient temperature of 20–25°C under a 12/12 h light/dark cycle and had free access to standard rodent chow and water. The NIH guidelines for the Care and

Inflammation

The i.pl. injection of CFA led to swelling, erythema, hyperthermia and hyperalgesia in the treated hindpaw. These signs were obvious several hours after inoculation and persisted for the entire observation period. No inflammation was observed in the contralateral paw. These observations are consistent with our previous studies (Stein et al., 1988). Since in those studies, many biochemical and behavioral measurements were examined at 4 days after CFA injection, we chose the same time point in

Discussion

We studied the anatomical substrates of an interaction between the peripheral nervous system and immunocytes in a model of painful inflammation. Our findings indicate that the expression of MOR in the DRG is enhanced, that these receptors are transported into the peripheral sensory nerve terminals, and that an endogeneous ligand (END) is carried to the surrounding tissue by CD4+ cells. We have previously shown that this peptide is released and can activate μ-opioid receptors localized on

Conclusions

Our findings have shown that inflammation in the rat hindpaw not only increases the MOR-IR in DRG neurons but also enhances the transport of MOR along the sciatic nerve and leads to an accumulation of MOR-IR on the peripheral nerve terminals. Activated/memory cells contain END and also migrate to the inflamed subcutaneous tissue. Thus, inflammation apparently induces a unique simultaneous upregulation of peripheral opioid receptors and of their endogenous ligands. This is in contrast to the

Acknowledgments

We thank M. Schäfer, Drs. S.R. Goldberg and P. Cabot for continuous support and critical discussions; Drs. Robert Elde (Department of Cell Biology and Neuroanatomy, University of Minnesota, Minneapolis), Stefan Schulz and Volker Höllt (Department of Pharmacology and Toxicology, Otto-von-Guericke University, Magdeburg, Germany) for the gifts of the μ-receptor antibodies.

Supported by NIH (RO1NS32466) and by Deutsche Forschungsgemeinschaft (SFB 507/B8).

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