Elsevier

Neuroscience

Volume 125, Issue 2, 2004, Pages 427-439
Neuroscience

Inhibition of calpains, by treatment with leupeptin, improves motoneuron survival and muscle function in models of motoneuron degeneration

https://doi.org/10.1016/j.neuroscience.2004.01.046Get rights and content

Abstract

The effect of treatment with leupeptin, a calpain inhibitor, on motoneuron survival and muscle function was examined in in vitro and in vivo models of motoneuron degeneration. Exposure of primary rat motoneurons to α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) is an established in vitro model of excitotoxic motoneuron death. Here we show that leupeptin treatment improved motoneuron survival following exposure to AMPA (50 μM). Application of leupeptin (100 μM) to AMPA treated cultures rescued many motoneurons so that 74% (±3.4 S.E.M., n=5) survived compared with only 49% (±2.4 S.E.M., n=5) in untreated cultures.

The effect of treatment with leupeptin on motoneuron survival and muscle function was also examined in vivo. In 3 day-old rats, the sciatic nerve was crushed and at the time of injury, a silicon implant containing leupeptin was inserted onto the lumbar spinal cord. The effect on long-term motoneuron survival and muscle function was assessed 12 weeks after injury. The results showed that there was long-term improvement in motoneuron survival in the leupeptin treated group. Thus, in untreated animals 12 weeks after nerve crush only 30% (±2.8. S.E.M., n=3) of sciatic motoneurons survived compared with 43% (±1.5 S.E.M., n=3) in the leupeptin-treated group. This improvement in motoneuron survival was reflected in a significant improvement in muscle function in the leupeptin-treated group. For example in the soleus muscle of treated rats 20.8 (±1.40 S.E.M., n=5) motor units survived compared with only 14.6 (±1.21 S.E.M., n=5) in untreated animals.

Thus, treatment with leupeptin, a calpain inhibitor, rescues motoneurons from cell death and improves muscle function following nerve injury.

Section snippets

Experimental procedures

All animals used in this study were bred and housed either in the Biological Services Unit at University College London or the Denny Brown Laboratories at the Institute of Neurology. Throughout this study all efforts were made to minimise the number of animals used and their suffering. All experiments described in this study were carried out under licence from the U.K. Home Office (Animals (Scientific Procedures) Act 1986) and following approval by the Ethical Review Panel of the Institute of

Results

The effect of leupeptin, a calpain inhibitor, on motoneuron survival was examined in vitro in an excitotoxic model of motoneuron degeneration, and also in vivo in a model of injury-induced motoneuron death.

Discussion

The results of this study show that treatment with leupeptin, an inhibitor of calpain proteases, rescues motoneurons in vitro following exposure to glutamate agonists and improves long-term motoneuron survival and muscle function in an in vivo model of injury-induced motoneuron degeneration.

Calpains have been described as cellular mediators of both necrotic (Wang, 2000) and apoptotic (Nath et al., 1996a,b) cell death. Early studies looking at the role of calpains in apoptosis showed that they

Acknowledgements

L.G. is the Graham Watts Senior Research Fellow. We are grateful to the Brain Research Trust for their support, D.K. is a BRT prize student. We would also like to thank Mr. James Dick, Dr. Jo Dekkers and Dr. Bernadett Kalmar for their assistance and expert advice.

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