Elsevier

Neuroscience

Volume 162, Issue 1, 4 August 2009, Pages 31-38
Neuroscience

Cellular Neuroscience
Research Paper
Selective sigma receptor agonist 2-(4-morpholinethyl)1-phenylcyclohexanecarboxylate (PRE084) promotes neuroprotection and neurite elongation through protein kinase C (PKC) signaling on motoneurons

https://doi.org/10.1016/j.neuroscience.2009.03.067Get rights and content

Abstract

Neuronal loss and interruption of axonal pathways are occurring after spinal cord injury. This is initiated by the mechanical damage and propagated by secondary events that include the fast rise of glutamate concentration and the subsequent over-activation of glutamate receptors, triggering noxious processes to the cell. Excitotoxic processes are also observed in degenerative diseases that involve motoneuron loss. Sigma-1 receptors (Sig-1Rs) are expressed in the CNS and their ligands have been shown to prevent neuronal death associated to glutamate toxicity. In the present study, we used organotypic cultures of spinal cord slices and dorsal root ganglia (DRG) explants from 7–8 days old postnatal rats to assess whether the agonist of the Sig-1R, 2-(4-morpholinethyl)1-phenylcyclohexanecarboxylate (PRE084), protects the spinal cord against glutamate excitotoxicity and promotes neurite elongation. The results showed that PRE084 exerted a bell-shape dose-dependent protective response of the motoneurons, with a significant neuroprotection obtained with 10 μM PRE084. PRE084 also caused an increase in the length of neurites in both motoneurons and neurons in DRG explants. Both effects were abrogated with the addition of BD 1063, an antagonist of Sig-1R, and the use of chelerythrine, a protein kinase C (PKC) pan-inhibitor indicating that PKC is implicated in the observed effects. These results suggest the use of PRE084 as a neuroprotective agent for spinal cord damage.

Section snippets

Organotypic spinal cord slice cultures

Organotypic slice cultures were prepared from Sprague–Dawley rats of postnatal day 7 (P7) using a modification (Guzman-Lenis et al., 2008) of previously described methods (Rothstein et al., 1993). Briefly, pups were over-exposed to CO2 and decapitated. The thoracic segment of their spinal cord was removed and placed in ice cold high glucose (6 mg/ml) Hanks' balanced salt solution (HBSS) (Sigma, St. Louis, MO, USA). Under sterile conditions, meninges and roots were removed and the spinal cord

Results

We first analyzed the expression of Sig-1R in the spinal cord organotypic culture by immunochemistry. Sig-1R was present in ventral (Fig. 1A) as well as in dorsal horn neuronal cells as confirmed by co-labeling with the neural marker PGP9.5 (Fig. 1B–D). Within the neurons, Sig-1R was mainly localized in the soma, with the highest density around the nucleus, and in minority along the neurites.

We assessed the potential of PRE084, a specific Sig-1R ligand, as neuroprotector against glutamatergic

Discussion

The results of this study show that the Sig-1R agonist PRE084 reduces the toxicity induced by glutamate to spinal cord organotypic cultures and promotes the elongation of neurites of both MNs and DRG neurons through PKC-mediated signaling.

Numerous studies have demonstrated neuroprotective properties of Sig-1R ligands in animal models of cerebral ischemia (Goyagi et al 2001, Goyagi et al 2003, Harukuni et al 2000, Takahashi et al 1995). Although the mechanisms involved are still unknown, several

Conclusion

In conclusion, the Sig-1R agonist PRE084 seems a promising drug to be tested for the treatment of SCI and other neurodegenerative diseases affecting the spinal cord in which excitotoxicity plays a central role. In addition to its neuroprotective effects, PRE084 has a significant pro-regenerative action, thus offering an interesting combination of actions in order to reduce neuronal damage and also to enhance re-growth of axonal pathways across the lesion site.

Acknowledgments

This research was supported by grants from the Ministerio de Ciencia y Tecnología (SAF2006-08682) and the Ministerio de Sanidad y Consumo (PI060201) of Spain, and funds from CIBERNED and TERCEL, Spain, and FEDER. We thank Dr. José M. Vela for fruitful discussions.

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