Elsevier

Neuroscience

Volume 101, Issue 4, 30 November 2000, Pages 1093-1108
Neuroscience

Localization and regulation of cyclo-oxygenase-1 and -2 and neuronal nitric oxide synthase in mouse spinal cord

https://doi.org/10.1016/S0306-4522(00)00361-4Get rights and content

Abstract

Prostaglandins are important mediators in spinal nociceptive processing. They are produced by cyclo-oxygenase isoforms, cyclo-oxygenase-1 and -2, which are both constitutively expressed in the central nervous system. The present immunohistochemical study details localization and regulation of cyclo-oxygenase-1 and -2 and neuronal nitric oxide synthase in lumbar spinal cord before and after induction of a painful paw inflammation in mice. Cyclo-oxygenase-1 immunoreactivity was found in glial cells of the dorsal and ventral horns, but not in neurons. In unstimulated mice, cyclo-oxygenase-2 immunoreactivity was found in motoneurons of the ventral horns and in lamina X, but not in dorsal horn neurons. After induction of a paw inflammation with zymosan, cyclo-oxygenase-2 immunoreactivity increased dramatically in dorsal horn neurons of laminae I–VI and X, paralleled by a significant increase in prostaglandin E2 release from lumbar spinal cord. Cyclo-oxygenase-2 was co-localized with neuronal nitric oxide synthase immunoreactivity in several neurons in superficial laminae of the dorsal horns and in the area surrounding the central canal. Nitric oxide synthase was distributed in the cytoplasm and extended to processes of some neurons. In contrast, electron microscopy revealed that cyclo-oxygenase-2 immunoreactivity was restricted to the nuclear membrane and rough endoplasmic reticulum.

It is shown in the present study that both cyclo-oxygenase isoforms are constitutively expressed in the spinal cord, cyclo-oxygenase-1 in glial cells of the dorsal and ventral horns and cyclo-oxygenase-2 in motoneurons. After induction of a hindpaw inflammation, several dorsal horn neurons express cyclo-oxygenase-2. Some of them are also positive for neuronal nitric oxide synthase, which is also induced following peripheral inflammation. Intracellularly, cyclo-oxygenase-2 is bound to the membranes of the nucleus and endoplasmic reticulum, whereas neuronal nitric oxide synthase is found in the cytoplasm.

Section snippets

Animals

Experiments were performed using male C57 black/6 mice (18–20 g, Charles River, Sulzfeld, Germany). The animals had free access to food and water. They were maintained in climate- and light-controlled rooms (24±0.5°C, 12 h/12 h dark–light cycle). The study was approved by the local Ethics Committee for animal research, and the guidelines set out by the International Association for the study of pain were obeyed in all experiments. All efforts were made to minimize both the suffering and the number

Spontaneous pain

After the intraplantar injection of 2.5 mg zymosan, mice showed an immediate and intense behavioral response that lasted for up to 30–45 min, and generally consisted of licking, biting and spontaneous flicking of the paw.

Paw edema

Intraplantar zymosan produced an increase in paw thickness. One hundred and sixty-eight hours after zymosan injection into the right hindpaw mice showed a 2.5-fold increase in the right paw volume (0.3±0.05 ml; mean±S.D.) compared to pretreatment values (0.12±0.02 ml), whereas paw

Discussion

The present data confirm and extend the current conception of the localization and regulation of the cyclo-oxygenase isoforms in the spinal cord. In addition to the previously described constitutive expression of both cyclo-oxygenase isoforms in rat spinal cord and up-regulation of cyclo-oxygenase-2 mRNA,5., 21. we could demonstrate (i) that cyclo-oxygenase-1 immunoreactivity in the mouse spinal cord is found exclusively in glial cells of the dorsal and ventral horns, (ii) that there is no

Conclusions

We report the distribution and regulation of cyclo-oxygenase-1-, cyclo-oxygenase-2- and nNOS-immunoreactive cells in the spinal cord in a model of a painful peripheral inflammation in mice. We show that cyclo-oxygenase-1 and -2 are both constitutively expressed in the spinal cord, cyclo-oxygenase-1 in glial cells of the dorsal and ventral horns, and cyclo-oxygenase-2 in motoneurons and astrocytes. Peripheral inflammation results in a dramatic increase in cyclo-oxygenase-2 immunoreactivity in

Acknowledgements

This study was supported by the Deutsche Forschungsgemeinschaft (SFB 553 and SFB 353). We would like to thank Beate Layh, Doris Schreiner, Claudia Labahn and Tanja Mittmann for excellent technical assistance. We are grateful to Dr Bernd Mayer for kindly providing the antibody against nNOS and to Andrea Hilpert for her support with the electron microscopy.

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